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Combining AI, Mind Genomics, and Young Medical Minds to Address Communication: Diabetes Patients and Their Desire for Herbal Remedies

DOI: 10.31038/MGSPE.2023314

Abstract

Medical students were given the task to identify situations involving patients with diabetes, the situations developed through AI. The situation became variables or questions about the nature of the patient with diabetes or about the interaction. AI then suggested four answers to each question. The test materials became the four sets of four answers, vignettes which described the nature of the diabetes patient and the nature of the interactions. Each of 50 respondents, foreign medical graduates associated with xxxx, evaluated unique sets of 24 sets of descriptions, vignettes, constructed from combinations of these 16 answers, according to an experimental design. The respondents rated the vignettes on their feelings and the expected feeling of the diabetic patient. Modeling the relation between the answers (elements) and the ratings showed three distinct mind-sets of respondents. These mind-sets, different ways of responding to the elements, were.

Introduction

In today’s world where technology is advancing, and where people have many options for health, it is important to develop a system which can educate the medical professional in the way to work with the patient. The patient is becoming a client of the health care system. It is increasingly recognized that ‘one size does not fit all,’ that people are different, and that medical communications, or more generally communications about the arc of wellness to illness, needs to be fine-tuned for the person. Salespeople know this need for differentiated communication better than just about anyone. People are not convinced by facts, but by how they react both to the facts and to the method that the facts are presented. Ask any salesperson of long-standing experience and that salesperson typically ‘knows’ what to say to prospects. The knowledge may not be formalized, but comes from the years of experience, the trial and error.

In the world of medicine, where technology changes, where the customer is often in an unpleasant situation, and where the medical professional is young, how can a bank of information be developed about what types of communications seem appropriate. We are not talking about a military phrase book with the desire to achieve a single objective, but rather a way of using communication to understand the other person, that understanding moving towards a productive relationship, and restored or maintain health.

Mind Genomics

The present study comes from the effort to create a body of knowledge about how to communicate in the world medicine, and more generally in the arc of life from health and wellness to illness and hospitalization. The approach used here and in previous reports is known as Mind Genomics. Mind Genomics is an emerging science, roughly 30 years old, which deals with the decision making of the everyday world. Whereas many published papers in psychology and behavioral economics deal with unusual situations worthy of note, e.g., counter-intuitive behaviors, Mind Genomics espouses the view that a true relevant opportunity exists through the assessment of the quotidian world, the ordinary world of the everyday, where most people spend most of their time. The origins of Mind Genomics came from interest in applied problems, such as what are the decision rules that people use to buy products? Or what are the types of phrases to which people react when they want to decide? The ordinary, everyday world presents us with uncountable opportunities to understand the rules of decision making and behavior of daily life. The Mind Genomics ‘project’ began with a departure from the conventional ‘surveys’. A survey instructs people to answer questions, forcing the people to think analytically. One need only watch a political pollster ask the interviewer some questions pertaining to hotly contested election to see that the interviewee moves from a person moving through life to a suddenly thoughtful person, one making conscious decision, one trying to figure out the ‘correct answer.; In the words of Nobel laureate Daniel Kahneman, research ends up looking at thinking Slow, thinking 2, whereas the real daily activity is thinking fast, system 1 [1]. The bottom line is that the survey methods end up moving the research to an intellectualized process, often requiring experimental situations out of the ordinary to reveal how we think. Either than, or ask people to intellectualize, even though their daily life consists of automatic behaviors.

Rethinking the Process

Mind Genomics emerged from the combination of three disciplines: Experimental Psychology (and specifically psychophysics), Statistics (specifically experimental design of independent variables), and consumer research (specifically consumer behavior through conjoint analysis [2]; Psychophysics focused on the measurement of percepts. The original world of psychophysics focused on what Harvard’s S.S. Stevens called the ‘outer psychophysics’, viz., the measurement of how strong a physical stimulus was perceived [3]. One could measure the physical sound pressure level, but that did not tell the research how loud the sound felt. Or, moving into the world of communication, one could tell a person about winning or losing a certain amount of money, but that did not tell us the ‘utility’ of winning or losing the amount of money. In simple terms, just knowing the physical strength of a stimulus does not tell us how the stimulus is perceived. When it comes to Mind Genomics, the objective is to measure the strength of perception of an idea, not a simple physical stimulus. Statistics provides a way to help us create combinations of test stimuli. Often, it is the test stimuli, the mixture, which is meaningful for one’s experience, not the individual components. The individual components tested by themselves have no real meaning. But how then does one measure the response attributed to a component of a mixture, when one can only test mixtures. It is that problem which occupies statisticians, namely, to design experiments where the respondent evaluate meaningful or reasonable mixtures, but where the analysis can pull out the contribution of the components. Consumer research, the third part of the foundation, focuses on the world of the consumer, and the world of daily activities involving communication to drive a purchase. For most consumer research, the effort is to understand the parts of the daily process in a way which serves both science/understanding and daily commerce. It is from consumer research that we learn that there are ways to communicate which are effective, ways which are not effective, and how to discover the effective method. The three foregoing fields of knowledge provide both the way to think about problems, and the way(s) to solve the problems. For this study, the objective is to understand what types of messages are felt to ‘work’ among diabetic patients, from the point of view of a young professional in the medical field. The approach of Mind Genomics is straightforward. The strategy is to present respondents (viz., survey takers) with combinations of relevant messages, in our case about diabetes, and measure the response to the combination on one or several scales. Knowing the experimental design undergirding the vignettes allows the researcher to obtain ratings of the combination, and then trace the rating to the particular message or element. The design, analysis, and even some of the interpretation is ‘templated’, allowing any person to become a researcher, or at least follow the steps properly, design an experiment, get the data processed automatically, and emerge with results that often drive new insights.

Applications in the Fields of Wellness and Illness

Mind Genomics enjoys increasing use world-wide. Early work with Mind Genomics focused on the application to evaluating how people felt about different aspects of foods [4]. The Mind Genomics approach quickly found other interested audiences, such as interest in food and good nutrition [5,6]. Finally, interest emerged for applying Mind Genomics to commercial applications in diabetes [7], and in insurance with diabetes [8]. The ability of Mind Genomics to provide a deep understanding of decision emerged in books on social issues [9] and on the practice of law [10].

Those early studies with Mind Genomics revealed that it was quite straightforward and easy to discover how people thought about themselves regarding health and medical experiences, and what would be good language to use. Those topics and discoveries, done by inexperienced young researchers, revealed how easy it was to understand the mind of people regarding health. When the same science was put into the hands of the nurse in charge of hospital discharge for congestive heart failure, the science was able to identify the mind-sets of CHF patients, and prescribe what to say upon discharge, resulting in a large decrease in within 30-day readmission’s [11]. The same approach was used some years later to understand what to say to a low-income catchment area near Philadelphia to encourage colonoscopies, an effort which doubled the number of colonoscopies simply by knowing what to say to people about the topic [12]. One of the stumbling blocks is the need to create questions and answers, and then use combinations of answers in the actual Mind Genomics study/experiment. The study can be only as good as the questions. When the researcher can ask good questions, and create meaningful answers, the Mind Genomics process works well. Often, however, the prospect of creating a set of questions and then answers comes with the daunting prospect of having to think in ways that were never part of education. People can answer questions; we are taught that in school, and it is drilled into the mind of the student. It is the framing of good questions, however, which is the problem. Students are not taught to think critically, to pose a series of questions in a way which drives understanding. One consequence of this weakness in critical thinking affects Mind Genomics. The prospective user is intimidated by the prospect of coming up with a set of four questions, and then coming up with answers to the questions. The prospect is often so anxiety-provoking as to abort the process in the beginning, as the prospective researcher figuratively throws up her or his hands, and in fruition simply aborts the effort. AI, artificial intelligence, is creating an entirely new opportunity for Mind Genomics in wellness and illness, by providing a way to obtain questions and answers in an almost automatic fashion. The use of AI to obtain these questions will be explained in this paper, dealing with the interaction of a doctor and a diabetic patient. The specific topic chosen is: I am a doctor who wants to counsel a difficult diabetic patient who does not want to change her diet plan and wants herbal remedies. The remainder of this paper presents the results of a small-scale study with 50 respondents, medical students from the clinic of Dr. Rizwan Hameed in Brooklyn, NY. The paper shows the depth of information provided by the Mind Genomics approach, information about the granularity of experience of a doctor with a patience regarding a specific condition. It is important to emphasize here the notion of granularity of experience. Rather than looking for broad findings to confirm or falsify a hypothesis in the manner of today’s science, the Mind Geonomics world view is that taken grounded theory. That is, the science evaluates what exists, that which manifests, so in an organized, structured, yet more or less realistic format. The output provides clear information about the mind as it grapples with an everyday issue in the world of medicine, but also in the world of ordinary people faced with a medical decision.

Creating the Ideas Book: AI-generated Questions and Answers

Mind Genomics as presented to the user in the ‘BimiLeap’ app (www.bimileap.com) provides a templated, almost scripted set of steps by which the researcher can begin with virtually no knowledge about a topic yet proceed within 30-60 minutes to understand a topic in depth through the use of artificial intelligence. Figure 1 and Table 1 present the steps, and some output. The Mind Genomics study begins with the selection of a study name (Figure 1, Panel A). The study name should be short. Again, and again novice researchers find it hard to give the study a short name, often because they have been so conditioned to confuse the topic with the method that thy lose sight of the simple overarching topic. The study here is ‘Diabetes’. It comes as a surprise to many novice users that the short name is correct to begin. So many of the users feel compelled to expand the name to the actual study, creating a paragraph out of a word. This comment, while not germane to the actual study, is a cautionary word to the research to keep things simple and focused. The second stage in the Mind Genomics study is to develop four questions or categories of issues, related to the topic, and which ‘tell a story.’ The requirement of ‘story telling’ is not fixed in stone, but the combination of questions should provide different aspects of the general topic. Figure 2, Panel 2, shows the second screen of the BimiLeap program. The screen is empty. Although it might not seem to be daunting, viz., to create four questions which ‘tell a story’, it is at this point that many researchers or want-to-be researchers are gripped with discomfort. In the past, quite a number of budding researchers, as well as a number of professionals, have created an account, logged into their account, named a study, and then given up when confronted with the requirement to create four questions. It was in answer to this need that Dr. Judith Moskowitz Kunstadt, sister of author HRM and herself a child psychologist, suggested that this ‘stumbling block’ might be addressed by providing a guide to creating questions. Author Rappaport, in turn, suggested a pre-set list of questions as a tutorial. Both were meaningful, but during the beginning of 2023, AI emerged in the form of Chat GPT, which allowed the researcher to query the AI. The AI would return answers. The focus of AI in Mind Genomics is to provide questions to the researcher, rather than factual answers. Figure 1 (Panel C) shows the request by BimiLeap for the researcher to provide a ‘squib’ or short description of what is desired. Figure 1 (Panel D) shows four questions which emerge from the AI.

FIG 1

Figure 1: The set-up sets of BimiLeap. Panel A shows the choice of name. Panel B shows the request for four questions which tell a story, and the option to use Idea Coach. Panel C shows the box where the researcher enters a description of the topic to prompt the AI-driven Idea Coach. Panel D shows four questions provided by Idea Coach but edited by the researcher afterwards.

Table 1: Results from the first iteration of Idea Coach to answer the question put into the ‘squib’

TAB 1(1)

TAB 1(2)

TAB 1(3)

When looking at the four questions, it is important to keep in mind that the use of AI is to suggest topics. A confident researcher could do equally well. The role of AI here is to give some ‘tutoring’ or ‘coaching’ to the diffident respondent. In Table 1 we will see the nature of th questions returned by the AI.

Table 1 shows the first set of results from Idea Coach. The top (section A) shows the 15 questions from the first iteration (Results 1). The topic as written in the box for Idea Coach was: : I am a doctor who wants to counsel a difficult diabetic patient who does not want to change her diet plan and wants herbal remedies.

Idea Coach begins by returning 15 questions. These questions, or indeed any set of 1-4 can be selected and inserted into the four question slots shown in Figure 1, Panel D. Once in the panel, the questions can be edited by the researcher, usually to change the question from a ‘yes/no’ or a ‘list’ to one which is more conversational, requiring a more elaborate, evocative answers.

The researcher may return to the Idea Coach, rerun the Idea Coach with the same ‘squib’, or even edit the squib to produce a better question. Idea Coach has been developed to allow the researcher a great deal of latitude in exploring different types of questions. Often the researcher uses Idea Coach to create a number of such sets of questions, because of the additional information and ‘analyses’ provided by Idea Coach when it summarizes each set of 15 questions. In other words, the material provided in Table 1 is provided for the next set of 15 questions. Finally, the questions in subsequent runs of Idea Coach with the same ‘squib’ may generate some repeating questions. Each run of Idea Coach is complete unto itself.

The Idea Coach returns with a set of analyses, as follows:

  • Topic Questions. These are the questions themselves, in the form of questions
  • Key Ideas: These the are questions, but in the form of an idea, rather than a question
  • Themes: The AI embedded in Idea Coach attempts to summarize the 15 questions or ideas into a limited group of more general ideas.
  • Perspectives: The AI now attempts to move from themes to positive versus negative points of view about the themes
  • What is missing: The AI now considers the topic once again, looking for issues that may have been ignored in the body of the 15 questions. Note that each of the subsequent reruns of the Idea Coach to create questions may come up with these missing ideas, but there is no causal connection. AI does not ‘learn’ from one creation of 15 questions to the next creation of 15 questions. Every effort is expended to keep the AI efforts independent from one iteration to the next.
  • Alternative viewpoints: AI now attempts to move to the other spectrum, to the opposite point of view, and explore those.
  • Interested audiences to the ideas created by the Idea Coach
  • Opposing audiences to the ideas created by the Idea Coach
  • Innovations: New ideas suggested for products and services from the questions and ideas presented in this run of 15 questions.
  • Creating Answers to Questions Using AI

    Once the researcher has created the set of four questions, with or without the help of AI-driven Idea Coach, it is time to create the answers or ‘elements’, four for each question. It will be combinations of these answers which will constitute the test stimuli, the so-called vignettes or combinations of answers. In the actual evaluation done by the respondents, the respondent will see only unconnected combinations of answers. Figure 2 shows the second section of the study, where the question is presented at the top, and the research or Idea Coach is requested to present four answers. Figure 2 Panel A shows the request for four answers. Figure 2 Panel B shows the answers that were selected by the research, and perhaps edited manually to make the answer easier when the answer in embedded into the test vignette.

    FIG 2

    Figure 2: Panel A: The first question, and the request for Idea Coach. The question has been modified so that Idea Coach will return with a short answer. Panel B: The four answers returned by Idea Coach and modified slightly by the researcher.

    It is important, therefore, that the answers be both relevant, as well as paint a word picture through a phrase. For this reason, the Idea Coach one again provides the question, then 15 answers. The researcher may repeat the Idea Coach, indeed as many times as desired. Each repetition generates 15 answers. The researcher must select a total of four answers, and in free to edit the answers to make the answers seem less like a list, and more like a stand-alone description.

    The Idea Coach once again stores each iteration, allows the researcher to edit the answer, and even allows the researcher to go back and edit the question to instruct the Idea Coach to focus more on expanding the answer. In the end, however be, it is the researcher who chooses the answers, edits the answers, even edits the question to be more general or to move to a different ‘angle’. Idea Coach returns with one page for each iteration of answers for each question. Thus, Idea Coach generates a minimum of four pages of results when Idea Coach is used once for each question but could generate 20 pages of answer when Idea Coach is applied five times for each question.

    Finally, as was the case with the questions, Idea Coach provides additional AI summarization, synthesis, and exploration for each page. At the end of the creation of the answers, when all have been selected, and the researcher moves to the next step, the BimiLeap program returns the now-complete Idea Book to the research in the form of an Excel file. Each page or each tab of the Idea Book has the squib or the specific question on topic, followed by the AI analysis. The book can be quite large. For example, if the researcher were to use Idea Coach five times for the squib and each of the four questions, Idea Coach would with return with 25 pages of results, one per page or tab, with page capturing the output from AI as well as the AI summarization of the suggested questions or answers.

    Once the researcher has selected the elements for the study, the BimiLeap program moves to the orientation and the rating scale. Figure 3 shows the orientation (Panel A) and the rating scale (Panel). The rating scale shown below is a minimal scale, with very little information presented to the respondent about the purpose of the study. This minimalist approach is adopted to allow the elements, the specific phrases, to convey the information. Any additional information presented in the respondent orientation ends up weakening the contribution of the actual elements.

    FIG 3

    Figure 3: Respondent orientation (Panel A), and anchored rating scale (Panel B)

    The five scale points are labelled. Close inspection of the questions show that there are two dimensions. The first dimension of the scale is the respondent’s own feeling, whether that be like it or don’t like it, respectively. The second dimension of the scale is what the researcher thinks the rating will be as assigned by the respondent. Again, there are two options, like it or don’t like. The actual scale seems daunting at first, and indeed professionals often complain that they cannot make sense of the scale. Even worse, often the respondent feels that the two dimensions interfere with each other, and in frustration these professionals simply stop participating. Although disappointing, the response of professionals is understandable. They are trying to ‘assign the right answer’ and find rating scale hard to ‘game’.

    Rating question: Please select how you feel

    1=I don’t like it and the patient does not like it

    2=I don’t like it but the patient is ok with it

    3=I can’t answer

    4=I am ok with it but the patient is not ok

    5=We both are ok with it

    Table 2 shows the actual information about the study, taken from the information used when the researcher set up the study. The study set up enables the researcher to obtain a great deal of additional information about the respondent by means of the preliminary questions, also known as the self-profiling classification questions. The BimiLeap program automatically collects information on the respondent’s age and gender (not shown in Table 2). In addition, the program allows the researcher to ask up to eight additional questions, and for each question allow up to eight answers. The preliminary questions in Table 2 show the types of information that can be obtained from the study.

    Table 2: Study information obtained from the set-up, as well as the number of respondents who completed the study

    TAB 2

    Launching the Study and the Respondent Experience

    Once the researcher has finished setting up the study and previewing it, the researcher launches the study. The actual Mind Genomics study is completed on the internet. The participants receive an invitation to the site, log in to the site. The BimiLeap program provides a number of ways to acquire the respondents, as shown in Figure 4. These range from having the BimiLeap program allow the researcher to tailor a panel from various sets of qualifications, or to have the BimiLeap representative create the panel. Other ways include working with a different on-line panel provider instead of Luc.id, Inc., or sourcing the respondents oneself. It is this last option that was selected by the researchers. The respondents were young medical students, interns, and residents associated with the clinic of Dr. Riswan Hameed in Brooklyn, NY, USA, as well as fellow medical professionals of the respondents scattered around the world. All respondents participated voluntarily. The BimiLeap program is set up to preserve respondent confidentiality. No personal identifying information is kept as part of the study. When private information is obtained, it is usually in the form of general questions in the self-profiling classification, but that information does not suffice to reveal respondent identity.

    FIG 4

    Figure 4: Sourcing options for respondents

    Once the respondent agrees to participate, the study begins. The respondent reads the very short introduction, and then proceeds to the self-profiling classification. To keep the appearance spare and ‘clean’, the self-profiling classification questionnaire comprises a pull-down menu, shown in Figure 5. The respondent pressed the check button, and the appropriate question drops down, along with the answer.

    FIG 5

    Figure 5: The self-profiling classification, completed by the respondent at the start of the study. The classification comprises a set of pull-down questions.

    Rather than giving each respondent a set of 16 phrases, the four sets of answers to the four questions, Mind Genomics creates small, easy-to read s/. The basic experimental design used by Mind Genomics is a simple 4 variable x 4 level main effects design. The design comprising 16 elements generates 24 combinations. Each combination comprises a minimum of two elements or answers from (different) questions, and a maximum of four elements or answer from (different) questions. No question ever contributes more than one answer or element to a vignette. However, the underlying design ensures that quite a number of the vignettes are absent from one element or answer form a question, and some vignettes are absent two elements or answers from two questions.

    The experimental design ensures that each of the 16 elements or answers appears in a statistically independent fashion. This will become important when the data are submitted to OLS (ordinary least-squares) regression analysis to link together the elements and the responses. Furthermore, the combinations are not all complete, allowing the researcher to use the regression analysis to estimate the absolute level of contribution of the elements to the ratings [13]. This will be discussed below.

    A continuing issue in consumer researcher is the implicit belief that the test is being conducted with the stimuli that are believed to be important. The reality is that when the researcher tests combinations of answers, it is not clear that these are the appropriate combinations to test. The researcher may end up testing these combinations in order to reach closure, and get the study done. Such a requirement means that the test stimuli, the vignettes, end up being the ‘best guesses’ about what to test.

    The Mind Genomics system permuted the combination of 24 vignettes creating new combinations with the same mathematical properties. All that has changed is that the combinations change, but the property of statistical independence is maintained, along with the property that each vignette has at most one answer from a question. This approach, the permuted experimental design [14] ends up allowing the researcher to have each respondent test a different set of vignettes, but at the same tie a set of vignettes precisely designed for OLS regression at the individual respondent level.

    Figure 6 shows a test combination that was evaluated by the respondent. The question and rating scale are at the top. Below, and indented are the elements as prescribed by the underlying experimental design. The elements are simply presented in unconnected form. No effort is made to present a felicitous, well-crafted combination. Rather, the objective is to present the respondent with the necessary information in a simplistic manner. Respondents move through these vignettes quite quickly, ‘grazing’ for information in the manner called by economic Daniel Kahneman as System 1. The word ‘graze’ is particularly appropriate here. Rather than forcing the respondent to adopt an intellectualized, judgment approach, the Mind Genomics system seemingly ‘throws’ the information at the respondent in a manner reminiscent of daily life. It is left to the mind of the respondent to process the information and assign a rating.

    FIG 6

    Figure 6: Example of a test vignette evaluated by the respondent

    It is worth noting here that all too often professionals attempt to be analytical in the evaluation of these vignettes, a behavior which ends up being frustrating both for them and for the researcher. The professionals often try to ‘answer correctly,’ looking for underlying patterns to guide. All too often, the respondents get angry, irritated, refuse to participate, or simple complain at the end of the evaluation. Although the data ends up being quite information, especially after the respondents are clustered or segments by the pattern of their responses, many of the critics of Mind Genomics simply reject the system as being, in the words of Harvard’s legendary psychologist, William James, a ‘blooming, buzzing confusion.’ Nothing could be further from the truth, as will be shown below, especially once the clustering is done.

    Uncovering Patterns – How Elements Drive Responses

    The objective of Mind Genomics is to uncover patterns in daily life. The test stimuli are the different vignettes, 24×50 or 1200 mostly different vignettes across the 50 respondents, with each respondent evaluating 24 combinations, and most combinations different from each other as a result of the deliberate permutation strategy. The rating scale, however, is not simple, but rather asks two questions, namely how does the medical specialist feel about the vignette in relation to diabetes management, and how does the medical specialist think the patient will feel about the vignette in relation to diabetes management. There are two answers, OK and not OK. (Note the use of the colloquial, which in fact is the lingua franca of the everyday).

    The choice of a ‘two-faceted’ scale is deliberate. The single scale allows the researcher to probe how the medical specialists feels. There are really two scales here, a scale for how the medical professional feels (ok vs. not ok), and for how the medical professional thinks the patient will react (ok vs. not ok).

    The easiest way to deal with this data is to create five scales by simple transformation:

    Professional OK Ratings 5 and 4 transformed to 100, ratings 3, 2 and 1 transformed to 0

    Professional Not OK Ratings 1,2 transformed to 100, ratings 3,4, and 5 transformed to 0

    Patient OK Ratings 5 and 2 transformed to 100, ratings 4,3 and 1 transformed to 0

    Patient Not OK Ratings 1 and 4 transformed to 100, ratings 2,3,and 5 transformed to 0

    I can’t answer Rating 3 transformed to 100, ratings 1,2,4 and 5 transformed to 0

    To all the transformed variables a vanishingly small random umber is added. This is a prophylactic step to ensure that all of the newly transformed binary variables exhibit some minimal variability, allowing the ordinary least-squares regression to work.

    Once the transformation is complete, the five newly created variables can be related to the presence/absence of the 16 elements by means of a simple linear equation:

    Binary Dependent Variable = k1(A1) + k2(A2).. k16(D4)

    The database comprises columns which ‘code’ the structure of the vignette. Each of the 16 columns is reserved for one of the 16 elements. When the element or message appears in the vignette, dictated by the underlying experimental design, the value is ‘1’ for that element and that vignette. When the element or messages does not appear in the vignette, again dictated by the underlying experimental design, the value is ‘0’. The coding method is called ‘dummy variable’ coding because all we know about the variable is that it either appears in the vignette or does not appear in the vignette [15].

    The regression analysis is run five times, using the equation above. The regression model is estimated without an additive constant, so that all of the variation can be traced to the contribution of the 16 elements. The experimental design ensures that all 16 elements are statistically independent of each other, so that one can compute ratios of coefficients in a meaningful manner.

    The interpretation of the coefficients is straightforward, and easiest to explain by a simple example. A coefficient of +20 for an element means that when the element is inserted into a vignette, there is an increase of 20% in the likelihood that the vignette will reach the value of 100.

    Before looking at the results for the total panel for the newly created binary scales, it is helpful to anchor the performance. A coefficient of +4 or lower is simply not shown. It is irrelevant as a driver of the specific binary variable. A coefficient of approximately +15 to +16 approaches statistical significance. Important coefficients, however, move beyond simple statistical significance to much higher values, here +21 or higher.

    Tables 3 and 4 show the coefficients for the total panel. The five columns show the newly created binary variables. The numbers in the body of Table 4 are the coefficients. Coefficients less than 5 are simply not shown. Coefficients 21 or higher are shown in shaded cells. The most important outcome from Table 4 are that no elements perform strongly for any newly created dependent variable. There are some which are close, but none reach the imposed threshold of a coefficient of 21 or higher.

    Table 3: Coefficients for models relating the presence/absence of elements to the newly created binary variables

    TAB 3

    Table 4: Coefficients for models relating the presence/absence of elements to the Binary dependent variable=R54 (Medical professional says OK, medical professional believes patient would say ok). The table shows the total panel, and then the coefficients divided by the types of patients the respondent dislikes versus the types of patients the respondent likes.

    TAB 4

    The analysis now moves to dividing the respondents by the type of patients they dislike strongly, and by the type of patient they like stray. The coefficients appear in Table 5. Once again, the elements with coefficients lower than 5 are shown with blank cells, and the elements with strong coefficients greater than or equal to 21 are shown with shaded cells. There are five elements which perform strongly in the different breakouts of respondents. Although these are strong performing elements, the reality is that there is no clear pattern. Were we not to ‘know’ the meaning of the elements, we would say that knowing what the medical professional likes and dislikes gives us a few stronger opinions, but that is all. The reason we see that there is no pattern comes from the reality that the elements have cognitive meaning, and thus we can see similarities when they exist, or at least superficial similarities. The use of test stimuli with deep cognitive meaning, our elements. Allows us this ability to reject random strong performing elements because there is no apparent pattern emerging, despite that strong performance.

    Table 5: Coefficients for models relating the presence/absence of elements to the Binary dependent variable=R54 (Medical professional says OK, medical professional believes patient would say ok). The table shows the total panel, and then the coefficients divided by two and then three emergent mind-sets.

    TAB 5

    Uncovering New-to-the-World ‘Mind-Sets’

    Mind Genomics adds value to our understanding by dividing the respondents into groups based upon the pattern of their coefficients. Rather than assuming that respondents who seem to think alike when they describe themselves (Table 5), Mind Genomics looks for strong performing, interpretable groups of people in the population, these groups emerging from how the people respondent to a specific set of granular messages. These are the so-called ‘mind-sets’. Mind-sets are defined as homogeneous groups of individuals, the homogeneity limited to a specific and concrete situation. Mind-sets emerge as statistically coherent groups for the situation, but people in the same mind-set for one situation may be in different mind-sets for another situation.

    Creating the mind-sets is straightforward, to a great extent the result of creating the vignettes according to an underlying experimental design, and then permuting the design. Each respondent thus ends up evaluating an ‘appropriate’ but random portion of the design space. The word ‘appropriate’ is used in view of the subsequent analysis, which creates an individual-level model for each respondent. Although the respondents each evaluated different sets of combinations, their individual sets of ratings can be submitted to the regression analysis described above. The result for this study is 50 rows of coefficients, one for each respondent. Those 50 rows can be clustered into a small number of groups, such as two or three groups, not based upon who the respondent is, but rather on a measure of similarity between the rows, or more correctly, a measure of dissimilarity between pairs of rows. Rows whose set of 16 coefficients move in ‘opposite directions’ suggest that the respondents see this particular world of diabetics n different ways. Rows who set of 16 coefficients move in the same way suggest the respondents in this particular world of diabetics see the world in the same way.

    The computational approach is known as k-means clustering [16]. The clustering ends up dividing the group of 50 respondents into two groups, and then into three groups. These groups are ‘mind-sets.’ Table 6 shows the coefficients for the Total Panel, then for the two miond0-sets (viz., two clusters), and then for the three mind-sets (viz., three clusters).

    Table 6: The underlying ‘stories’ (viz., interpretations) of the strong performing elements for each mind-set

    TAB 6(1)

    TAB 6(2)

    TAB 6(3)

    TAB 6(4)

    Jumping out of the table is the far greater number of strong performing elements for the three-cluster solution. Not only are there’re more strong performing elements of coefficient 21 a d higher, but a story begins to emerge. The story emerges from the data and is not imposed. It is the meanings of the ‘cognitively rich’ elements which tell the stories:

    Mind-set 1: The medical professional focuses on eating

    The patient feels overwhelmed by the idea of changing her eating habits.

    Our patient’s current diet plan focuses on balanced meals with appropriate portion sizes.

    She believes her current diet is sufficient and sees no need for modifications.

    Mind-Set 2: The medical professional wants to educate the patient about herbal remedies.

    An online course or video was provided to educate the patient about herbal remedies and their effects on diabetes.

    Infographics or visual aids were utilized to convey the potential risks and benefits of herbal remedies.

    The patient attended a workshop or seminar dedicated to informing them about the risks and benefits of herbal remedies.

    A healthcare professional explained the potential risks and benefits of herbal remedies in diabetes management.

    Mind-Set 3: The medical professional is sensitive to the patient’s fear of missing out on favorite foods.

    Emotional attachment to comfort foods.

    AI can be used to summarize the data from the different groups. Table 7 presents the AI analysis for each of the three mind-sets, from the three-mind-set solution. Once again the researcher should winnow down the large amount of data produced by Mind Genomics. Rather than trying to synthesize a story using all the elements, a story based upon the coefficients, the Mind Genomics program uses only those elements with coefficients of 21 or higher. There are far fewer elements to generate the pattern. The AI creates its own story from the data, but at least that story is grounded in the data from strong performers only. Table 8 shows the answers to the queries put to AI, in the effort to discern the ‘story’ in the pattern generated by the strong performing elements.

    Table 7: Calculation of the IDT=Index of Divergent Thought, which measures the strength of the coefficients, and thus the strength of the study.

    TAB 7

    Gamifying the Process –‘How We Did We Do, and How Much Better Can We Do?’

    Mind Genomics is a science emerging only in recent years. With the ease, rapidly, and low cost of doing these experiments, the natural question is to ask, ‘how good are the data?’ After all, when a study can be designed, executed and analyzed in a matter of hours, with very low cost, by virtually anyone, how does one differentiate between good science and poor science? One cannot see the researcher. It might be possible for a researcher to offer an ‘oeuvre’ of work across several years, so that the overall quality of this oeuvre can be judged. But what about one-off studies which may be brilliant? Or, equally likely, maybe a simple waste. One way to assess quality is to look at the coefficients generated by the study. The vignettes are evaluated by people who cannot ‘guess’. When the coefficients are low, the coefficients are essentially ‘noise’. When the coefficients are consistently high, however, we must conclude that a meaningful pattern is emerging. Table 8 shows a way of estimating the performance of the elements. The calculations are simple. One divides the data into the six groups (total, two groups from the two-mind-set solution, three groups from the three-mind-set solution). Each has a relative base size, as Table 7 shows. The calculations show the progression to a single number, 50, which is the average squared coefficient. It is called the IDT, index of divergent thought. There are no norms for the IDT, at least not yet. When thousands of studies are done, there will be sufficient values for the IDT to be considered across topics, ages of researchers, and so forth. With this ability to ‘gamify’ the process, developing the IDT, one can imagine the competition among researchers. One potential of gamification is the ability to offer prizes to the highest-scoring studies in a topic presented as a challenge in a prize competition. The study here on diabetes is only one topic in diabetes. What might be the outcome if a ‘meaningful prize’ were to be offered for all studies having IDT values of 70 or higher, with additional incentive for studies with IDT values of 80 or higher. Such an approach done world-wide might well produce a plethora of new ideas.

    Assigning a New Person to a Mind-Set

    The final step in the analysis of these data creates a tool, a mind-set assigner, which enables the medical professional to assign a new person to one of the three mind-sets for this granular, relatively minor issue. A key benefit of the Mind Genomics project is the ability to do the research quickly and inexpensively. Yet, it is simply not realistic for a new person to have to go through the research protocol, evaluate 24 vignettes, and be assigned to a mind-set. The process would simply not work because there is no way to incorporate the data of this new respondent into the original data, rerun the data one again, this time with the 51st person and then re-discover the mind-sets anew.

    Figure 7 shows the system developed to assign a new respondent to a mind-set. This is called simply the mind-set assigner. The approach works with six questions, taken from the original study, and runs a Monte-Carlo c with the patterns of these six questions. The outcome is the selection of the proper six questions from the original study, and the assignment of each of the 64 possible response patterns to one of the three mid-sets.

    FIG 7

    Figure 7: The Mind-Set Assigner tool. Panel A shows the bookkeeping information, including the request for participation. Panel B shows additional ‘background questions’ requested of the respondent. Panel C show the actual se of six questions, and the two possible answers.

    The Mind Set assigner comprises three panels:

    Panel 7A shows the introduction as the respondent sees it. The introductions requests permission to acquire the data and presents the respondent with a variety of questions about background. All, some, or even just the permission itself can be asked. The researcher sets up the specific information desired.

    Panel 7B shows specific background questions that can be asked of the respondent. These questions are additional to the actual assigner tool. Often when one wishes to ‘type’ the larger population, the objective ay be to add in additional information about the individuals in the different mind-sets. Panel 7B allows the researcher to ask up to 16 additional questions, with up to four possible answers for each.

    Panel 7C shows the six questions in the mind-set assigner, these questions taken from the actual study. The questions can be slightly modified by the researcher, but good practice dictates that the questions be the same as the elements of the study. The order of the six questions is randomized from one respondent to the next. It is the pattern of responses which is used to assign the new person to one of the three mid-sets.

    It is worth noting that the researcher can do several studies, create mind-set assigning tools for each, and then combine these in a simple system allowing the research to ‘sequence the mind’ of the respondent.

    Discussion and Conclusions

    The Mind Genomics way of thinking, from conceptualizing a problem to providing a rapid and affordable experiment, offers science a new way to understand how people think. Researchers are accustomed to expensive, well-thought out, often laborious and time-consuming studies. One of the great problems today is the head on collision of the need to understand the person but the cost and time necessary to do so. So very often much of the research effort is spent getting the funding to do the research, and then cutting the problem down to a size where it can be solved. As a consequence, much of today’s research is done by committee, piecemeal, after inordinate wait, and only when the funding comes through. These are factors which act as a drag on our understanding of people, and our use of the data to improve the lives of people.

    The study reported here represents one simple project, readily done by one or a few researchers, at low cost. Beyond the benefits of speed and cost lie the potential to create a corpus of knowledge about people in their daily lives, how they react, and how they interact. Up to now there has been a dearth of knowledge about people from the point of view of their everyday lives, what they think, what they say, and how they should say it. The Mind Genomics project provides this corpus of knowledge in a scalable form, anywhere in the world, and at any time. One could imagine one of these studies for each country, for each topic area. The opportunity to create this database of the mind may now be a reality. This paper shows the tesserae, the pieces of that reality, for a specific, granular, almost minor topic. One need not ‘triage’ the research, at least for these types of topics. All the topics can be addressed. The vision of science driving a personalization of the medical experience, with better interactions, healthier populations, and improved economics, lies within reach [17].

    Acknowledgments

    The authors would like to thank Dr. Rizwan Hameed for the ongoing inspiration to pursue this work.

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Communication Styles Regarding Child Obesity: Investigation of a Health and Communication Issue by a High School Student Researcher, Using Mind Genomics and Artificial Intelligence

DOI: 10.31038/MGSPE.2023313

Abstract

One hundred low-income respondents in New York City each evaluated 24 unique vignettes, combinations of 2-4 elements (messages). The vignettes were created according to a permuted experimental design, with each respondent evaluating full set of vignettes, allowing regression analysis to reveal the motivating power of each element, and k-means clustering to divide the 100 respondents into different groups, mind-sets, based upon the specific group of elements that were most motivating. The three emergent mind-sets were: MS1=Work on self and health; MS2=Set goals and structured reporting; MS3=Get family involved to focus on eating habits. To assign new individuals to a mind-set, the paper shows the construction and use of a PVI (personal viewpoint identifier) typing tool. The paper finishes with the use of artificial intelligence (AI) for three automated post-study applications; identify recurrent-themes in the data, suggest new areas of knowledge to pursue, and suggest new products and services for each mind-set.

Introduction

The world of 2023 is experiencing a rapidly growing epidemic, that of obesity, with innumerable consequences, diabetes the main source of worry, but many other diseases following behind [1,2]. One need not go through the litany of diseases, nor look for causes. There are many causes, a great number having to do with lifestyle alone with genetic proclivity [3,4]. The world of energy saving devices has made physical exercise into something often pursued specifically by regimented exercise, either alone, in a gym or with a personal trainer. To further complicate matters, the foods eaten are often simply not as healthy as they could be, perhaps because our lifestyle allows us to snack more than we should, on food, which is more delicious, but not healthful.

The literature is vast. One can find causes, report incidence of disease, create registries, attempt modifications. Much of the knowledge of what to do comes from the medical and the nutrition professionals. What happens, however, when we move from professionals to young people who explore obesity and the complex it comprehends. What more or what in general can we learn when we empower a young person to do the research, a person who is not yet jaded and hammered into conformity? What types of insights do we obtain? The Mind Genomics approach is a continuation of a stream of work using young minds to explore topics. What is presented here has already been used a number of times by elementary and middle school students, with significant new learning emerging from the process [5].

The approach of this paper is based on Mind Genomics, an emerging science of the ‘everyday’. The goal of Mind Genomics is to find out how people think about different aspects of ordinary life. The Mind Genomics studies works by presenting respondents with combinations of elements (messages about a topic), instructing the respondent to rate the combination, and deconstruct the ratings into the contributions of the different messages. The objective is to let the respondent evaluate almost realistic combinations of messages, a system which cannot be gamed, and which quickly reveals how the respondent thinks about the different aspects of a situation. Mind Genomics has a long history, beginning with conjoint measurement [6], moving to functional measurement [7] and finally to the form presented here [8,9]. Mind Genomics has already been used in many settings, ranging from marketing to social issues to health issues, to the law and beyond [10-14].

Part 1 – Searching for Questions, Answers, and Insights at the Initial Stage of Setting Up the Study

The Mind Genomics process is templated, with the website guiding the researcher through the different steps. The website itself is at www.BimiLeap.com. The first objective is to create the raw materials. This effort occurs only after the researcher creates the framework for the study, gives the study a name, selects the language for the prompts to the researcher, and finally agrees agree not to collect personal information unless knowingly provided by the respondent (viz. the survey-taker).

It is now time to create the raw materials. The raw materials for Mind Genomics are questions which tell a story, and then answers to those questions. The template for today’s Mind Genomics studies comprises four questions created by the researcher, and then four answers to each question. The questions will never be shown to the respondent, but the answers will be shown in combinations called vignettes.

When confronted with the Mind Genomics process, many researchers freeze, becoming nervous at the prospect of asking four questions which tell a story. Once the questions are asked, it becomes reasonably simple to provide answers. Despite the ability to provide answers, the nervousness of the researcher ends up being a major stumbling block hindering the adoption of Mind Genomics. The reality is that people are simply not educated in a way which encourages them to create questions and answers in the form of a narrative that can be used by researchers.

The idea Coach was invented in 2022 to circumvent the problem of the ‘stymied researcher’ the researcher who simply experiences too much fear and cannot move through that fear to produce the requisite questions and answers. Often after a few attempts the researcher become comfortable with the process of one question → multiple answers. The Idea Coach was created to help the novice advance to that stage.

Figure 1 shows the screens from the study template requesting the four questions (A), the Idea Coach selected by the researcher (B), the computer screen showing the first few of the of 15 questions emerging from one request from the Idea Coach program within BimiLeap (C), and finally the four questions dropped into the template. The researcher can edit the questions to make it simpler for the Idea Coach. Table 1 shows the full text of each of the 15 questions generated by AI through Idea Coach.

FIG 1

Figure 1: The request for four questions, the Mind Genomics Idea Coach, and the four questions which emerge

Table 1: The 15 questions generated by Idea Coach in its first pass. The 15 questions were generated in response to the paragraph written for Idea Coach in Figure 1, Panel B.

TAB 1
 

The Mind Genomics process continues with the creation of answers to each question, again by the AI-driven Idea Coach. Each iteration returns with 15 answers to a question, with an estimate of 15-30 seconds for the 15 answers to emerge. The answers are presented to the researcher, who can choose either to incorporate the answers into the template, or to request a re-run.

Note also that after the questions and answers have been presented to the respondent by Idea Coach, the BimiLeap program returns with the ‘Idea Book’, containing one page for each request for 15 questions, and one page for each request for 15 answers. Below the record of the questions and answers, the Idea Coach inserts the analysis by AI of additional understanding that the research can learn from those 15 questions or answers, the analysis done by having AI use a set of queries to summarize the questions or answers. Appendix 1 shows an example of this AI summarization and expansion of the information for the questions. The same work is done on each page, viz., for each request for questions, and for each request for answers.

Appendix 1 shows the set of questions returned by Idea Coach.

Appendix 2 shows the four sets of different answers

Figure 2 shows screenshots of the process, proceeding from the template with no answers to the question (Panel A), the Idea Coach Request (Panel B), the partial output of Idea Coach for that first iteration (Panel C), and the four final answers selected (Panel D). Although the researchers ran only one iteration and obtained the answers, as well as editing them before finalizing the answers, Idea Coach makes it easily to do iteration after iteration, enabling the researcher once again to have an AI-driven tutor become essentially ‘Socrates as a service.’

FIG 2

Figure 2: The template showing the request for four answers to the first question (Panel A), the Idea Coach which is either selected to skipped (Panel B), a set of prospective answers to question 1 returned by the Idea Coach (Panel C), and the four selected answers ‘dropped into’ the template.

It is important to keep in mind that this first step with Idea Coach becomes a teaching tool, viz., almost a Socratic tool. The researcher need not create the questions or the answers. The AI embedded in the BimiLeap program ends up providing questions and answers, each of which can be edited by the researcher. Table 2 presents the four questions and the four answers. Note that the answers have been edited for clarity by the researchers. The Mind Genomics template enables the researcher to edit or even entirely replace any question or answer that has been written by the researcher or by Idea Coach.

With the introduction of AI into the BimiLeap program in the form of Idea Coach, the AI has been given a new task, summarizing the different questions created in a single pass, or summarizing the 15 answers created in one pass to answer one question. Appendix 1 shows the AI summarization and expansion/interpretation of the 15 suggested questions for the first pass through Idea Coach, where the effort was to create four questions. (Note, the material returns in a separate book called the Idea Book. A researcher can run the effort many times, with each run creating a separate page of analysis and interpretation for the 15 questions generated by that pass.)

Running the Mind Genomics Study

Figure 3 (Panel A) shows one of eight possible self-profiling questions chosen by the researcher. For each question the researcher may provide up to eight different answers but must have at least two. The respondent selects the most appropriate answer to each question. Not shown here are the questions of gender and age.

Figure 3 (Panel B) shows the very brief orientation screen shown to the respondent at the start of the study. The rationale for having a very short introduction, virtually a single sentence, is the desire to have the individual elements or messages generate the differences in the response. Where a specific frame of mind is desired, one with deeper knowledge of the topic, such as the background of a law or medical case, the orientation page can extend to half a page or more.

Figure 3 (Panel C) shows the rating question and the scale points. The scale is set up to be 5, 7 or 9 points. The researcher selects the anchors. The researcher is required to anchor the top and bottom of the scale but has the option to provide anchors for the other scale points.

Figure 3 (Panel D) shows the open-ended question, allowing the respondent to add additional reactions.

FIG 3

Figure 3: Additional study set-up screens. Panel A: Example of a self-profiling classification question. Panel B: Respondent orientation. Panel C: The rating scale and anchors. Panel D: Open-ended question.

Figure 4 shows two final screens in the set-up. Figure 4, Panel A shows the researcher’s final thoughts and key words. Mind Genomics studies are easy and quick to set-up. It is important to capture the thinking underlying the specific study, especially since a project might involve as many as 5-10 parallel studies, run at the same time, with different aspects covered by the various studies. It is always a good idea to record the momentary thinking underlying the specific study. The key words are also requested, with a minimum of one word required. The key words allow for database searches.

Figure 4, Panel B shows the request for panel sourcing. The researcher can choose from different sources, giving the researcher flexibility. When the researcher opts for the default provider, to provide the respondents, the researcher is led to the API for Luc.id Inc., a panel aggregator, and from there to the selection of respondents according to specific criteria. When the researcher wants to work with other panel providers or even source the respondents from a pool of specific individuals, the researcher chooses another option. The objective is to make respondent selection easier, driving the researcher to options which will greatly increase the likelihood of a successfully completed study, with the desired number of appropriate respondents.

FIG 4

Figure 4: Final thoughts (Panel A), and sourcing the respondents (Panel B)

The Respondent Experience

Once the study has been set up, and the financial aspects agreed upon (viz., platform charge and panelist recruitment fees) the respondents are invited to participate, usually by email. The respondents who agree to participate are led to the website which introduces the study through the short orientation (Figure 3, Panel B), and then proceed to the self-profiling classification (Figure 5). The self-profiling classification is presented as a pull-down menu, uncluttered and easy to complete.

FIG 5

Figure 5: The screen showing the self-profiling classification for the study

The final steps are the presentation of the test vignettes (Figure 6), and the open-ended question (Figure 7). The test vignettes appear to be random combinations of elements, viz., answers to the question. Figure 6 shows a test vignette comprising the rating scale at the top, and the vignette in the middle. The respondent evaluates 24 unique vignettes, some having two elements, some having three elements, some having four elements. To the untutored eye, the 24 vignettes appear as a hodge-podge, a collection of screens comprising seemingly random mixtures of elements. The elements are the answers in Table 2. The questions never appear, and indeed their sole reason for being is that they motivate the different answers. The questions are not relevant for the respondents, whereas the answers are. Most respondents confronted with this seeming randomness end up looking at the combination on the screen and assigning a rating. Exit interviews with respondents, and especially with academics, end up with the same observation, viz., that everything seemed so ‘random’. Many respondents confess that they attempted to assign the ‘appropriate rating’ to the vignette, but had a hard time, and so they felt they just ‘guessed.’ This state of ‘indifference’, of lack of involvement, allows the respondent to answer honestly. The actual structure of the vignettes are anything but random. The basic design, a main effects experimental design, creates 24 combinations, the aforementioned vignettes. Each vignette has a minimum of two elements (answers in the terminology of Table 2), and a maximum of four elements. The experimental design ensures that each vignette has at most one element or answer from each question, but in those with three elements one question does not contribute an element, and in those with two elements two questions do not contribute elements. The underlying experimental design, known informally as a 4×4, ensures that the elements will appear five times in 24 vignettes, and be absent from 19 vignettes. Furthermore, the design ensures that the 16 elements will be statistically independent of each other, allowing for the analysis of the data by regression modeling, either at the group level or at the individual respondent level. Finally, the underlying experimental design is only a template. The design stays constant, but the elements can be permuted so that the elements are still associated with questions, but the combinations change. This strategy is called permuting the experimental design and ensures that the combinations or vignettes cover a lot of the possible combinations. At the level of science, this permutation means that the researcher does not have to limit the focus to pre-specified combinations that are thought to be most promising. Rather, he permuted design enables the researcher to quickly explore a large number of combinations of elements, and, at after-the-fact develop a detailed picture of the different elements. A good analogy is what the MRI ends up doing, taking pictures of the same tissue from different angles, and then combining these pictures after the fact [15]. When the respondent finishes reading the vignette the respondent simply presses the appropriate key on the computer or smartphone, the rating is registered along with the response time. The response time is defined as the number of thousandths of a second between the appearance of the vignette on the screen and the rating assigned by the respondent. After the respondent has assigned the rating the BimiLeap program automatically advances to assemble the next vignette at the local site viz., at the respondent’s computer or smartphone. Over a period of 20+ years the Internet-based surveys or experiments have gotten shorter. When the efforts were first launched in a major way around 1999, it was fairly easy and inexpensive to obtain respondents. The Internet was fairly new, respondents were interested in participating and the attention span seemed to be much longer. It was not unusual to be able to get hundreds of respondents for a study, and for the web-based interaction with the respondent to last 15-20 minutes. Today, however, people are time-starved, and have substantially shorter attention spans. The 4×4 experimental designs used here require about 3-4 minutes on the internet. Even with that short time, it is vital to work with a panel provider, unless one has a captive audience. The panel providers have access to millions of respondents. It becomes cost effective and time saving to work with these providers, an effort which ends up virtually guaranteeing that the study will be completed, often in an hour or sooner.

FIG 6

Figure 6: Example of a vignette

Table 2: The four questions and the four answers to each question provided by the Idea Coach, viz., AI embedded in the BimiLeap program.

TAB 2

The Mind Genomics Study about Low-income Parent’s Response to What Do for an Obese Child

This study was run by the senior author (BK) following his interest in nutrition education. The focus of the study was on what are the aspects, concerns, and messages regarding a low-income parent thinking about a child who is obese. The study was created on the BimiLeap platform, and sent for participation to parents in New York City, with self-declared income less than $45,000, and with self-declared parent of at least one child under 12 years old.

The study required about 30 minutes to set up with the assistance of the Idea Coach. The study was run by Luc.id, a panel provider, on July 13, 2021. Table 3 presents the set-up information taken from the set-up screens.

Analyzing the Data by Transforming the Ratings, and Using OLS (Ordinary Least-squares) Regression

The study comprised the responses of 100 individuals, each of whom evaluated 24 unique vignettes. Across all of the 2400 vignettes the vast majority of the vignettes were unique. This uniqueness means that the ordinary analyses of averaging ratings for a common stimulus won’t work, since the stimuli tested by the respondents are vignettes. What is important, however, is the response to the individual elements, the phrases which give the message. There were only 16 elements, each element appearing five times in 24 vignettes, so each element appearing 500 times the full set of 2400 vignettes.

The analysis first transforms the ratings. The rationale for transforming the ratings comes from the reality that most users of data do not know what to do with the actual ratings. For example, what does a 4.5 mean on a five-point scale? The scientists who do the research have a difficult time explaining the meaning of the intermediate scale point. This difficulty is glossed over because the research conclusions end up with ‘statistically speaking, these two ‘items’ are same, different, lower, higher.’ The pervasive use of inferential statistics, of same vs different, higher vs lower, ends up making the rating value a simple way by which to compare two or more groups on these simple ‘same/different/lower/higher’ categories of reporting. The reality of the scientific project is to find effects, find differences, using the rating as key indicator, with the discussion moving away from the focus on the rating value and on to the hypothesis.

The objective of Mind Genomics is to measure the mind or at least to put numbers onto messages so that one can create a metric of thoughts, of attitudes. The use of responses to vignettes is a good way to do that but the analysis has to extract the important information. That information must be a number, comparable across studies, amenable to being databases, to being used as a key performance indicator (KPI). To create these KPI-level numbers requires a few simple steps, which end up creating an easy-to-master system.

Step 1: Transform the Ratings to a Binary Scale

Most managers who use the data focus on simple numbers by which they understand and by which they take action. It is easy to communicate the percent of respondents saying yes vs percent saying no. To achieve this binary scale, the Mind Genomics convention is to pick some point on the scale which divides the Top from the Bottom. For the five-point scale, this cut-point is 4. Ratings of 5 and 4 are transformed into 100, ratings 1,2 and 3 are transformed into 0. To the newly created transformed variable is added a vanishingly small random number (<10-5). The rationale for the random number is that in the case that all of the ratings assigned by a respondent were either 1-3 or 4-5, there would be some randomness, preventing the regression from crashing.

The transformation is really a mapping of the anchored rating scale to a binary scale. For our analysis in this paper the ratings 5 and 4 have in common that the vignette is ‘motivating’ whereas the ratings 1, 2, and 3 have in common that the message is not ‘motivating’. The messages may not be demotivating, but rather simply are not motivating.

Finally, to finish the discussion of mapping, one could also create another binary variable ‘Work For Us’. This variable would take on the value 100 for ratings of 5 and 2, and then take on the value of 0 for ratings of 4, 3, and 1. This paper does not deal with the variable ‘Work For Us’, although everything that has been done for ‘Motivating’ can be done for ‘Work For Us’.

Step 2: Estimate the Parameters of the Equation Relating the Presence/Absence of the 16 Elements to the Dependent Variable

The equation is expressed as: DV (dependent variable)=k1(A1) + k2(A2)…k16(D4)

The equation has no additive constant. This is deliberate in order to make all coefficients comparable, both within an individual, across individuals within a single study, and across studies with different sets of elements.

The underlying experimental design ensures that each group of respondents will generate data that can be subject to OLS regression, ranging from data generated by all respondents together (Total Panel) and down to definable subset (e.g., based upon the answers of the respondent in the self-profiling classification), and down to the level of the individual.

The coefficients for the total panel are shown in screen shot in Figure 7. These are ‘flash results’ appearing on the researcher’s screen. The BimiLeap program updates the data every minute or so, providing a report of the coefficients for each defined group. The numbers are the coefficients of the model, shown for the Total Panel. We interpret the coefficient as the percent of responses achieving the rating 5 or 4 (viz., motivates) when the element is inserted into the vignette. Most of the coefficients are positive. Statistically ‘significant’ coefficients have values around 15-16.

FIG 7

Figure 7: Screen shot of the visual report of the data. The visual report is updated when the researcher refreshes the screen. The screen shows the number of respondents who started the study, the number who completed, the elements and the coefficients estimated for the Total Panel.

Step 3: Create the Models for the Total Panel and for the Self-defined Groups

Once the data have been incorporated into the database, the OLS regression rapidly reveals the strong performing as well as the weak performing elements. Table 3 shows coefficients for the 16 elements for key self-defined subgroups. Only those subgroups with 10 or more respondents are shown. Fewer than 10 respondents generate readable data, but the base size makes the coefficients less robust.

Table 3 shows many blank cells. These correspond to coefficients which do not reach the cutoff point of 16 or higher. The coefficient value of 16 approaches statistical significance in terms of a t-test of coefficients. The strong performing elements are operationally defined as having coefficients of 25 or higher. These strong performers are shown as shaded cells.

Table 3: Study summary, provided when the data report is issued at the end of the field work and analysis.

TAB 3

A cursory look at the performance of the elements in Table 3 suggests that there may be patterns among strong performing elements, but these patterns are hard to discern. The same finding occurs in study after the study. Simply looking at the respondents by WHO they are, what they say they THINK, and what they say they DO produces data, but interpretable patterns usually fail to not emerge. The reason for the pattern not emerging is that there are 16 elements, not one or two. With 16 elements, the patterns which emerge should tie together a reasonable number of elements. With one or two elements, the temptation is to create a plausible ‘hypothesis.’ With its abundance of elements, patterns will emerge readily when present, letting the researcher focus on data, not on hypotheses which fit sparse data.

Mind Sets

A key tenet of Mind Genomics is that at the granular level of behavior, the everyday, people differ from each other in the way they make choices, in the way they value the information they receive. Furthermore, these person-to-person variations exist as basic, explainable differences among people. It is the job of the Mind Genomics researcher to uncover these different groups, these mind-sets.

The way to uncover these basic mind-sets at the level of granular and everyday topics is by clustering together people who show similar patterns of coefficients. The patterns of coefficients for our study on obesity show what is important to various people. Mind Genomics enables the researcher to discover these mind-sets using empirical methods.

The actual mechanics for discovering mind-sets are simple. The researcher creates an individual level for each respondent, and then clusters the set of individual coefficients using one or another method for clustering. The method used here is k-means clustering [16]. The metric for ‘distance’ between people is the value (1-Pearson Correlation), with the Pearson Correlation computed across the 16 coefficients for two people. Each respondent ends up being assigned to one of the segments or mind-sets.

Segmentation is a heuristic, aiming to simplify data by putting the items (viz., respondents) into non-overlapping groups. Depending upon the specific algorithm for doing the clustering, the result may end up as few clusters versus many clusters, and as clusters which may be easy-to-interpret or hard-to-interpret. The researcher makes the judgment considering the criteria of interpretability (do the mind-sets or segments make intuitive sense), and parsimony (the fewer the number of segments or min-sets the better, as long as the mind-sets tell a coherent story).

Table 4 shows the three mind-sets emerging from these data. The mind-sets all focus on obesity, medical interventions, and lifestyle interventions. What is important, however, is the fact that even with a fairly constrained topic such as obesity, there are clear and meaningful differences in the way people respond to the messaging. The nuanced differences among the groups emerge clearly, along with key messages for that group. In effect, the clustering provides a springboard for better thinking, better understanding, and hopefully improved communications.

Appendix 3 shows AI interpretation of the strong performing elements from Table 4. The opportunity to use AI to interpret the results becomes more important when the clustering reveals interpretable clusters. Once again the interpretability of these clusters, these mind-sets, come from the commonality of elements which motivate the different groups of respondents. It is the sheer simplicity of similar ideas, important for clustering, which also boosts the ability of AI to create meaningful interpretations, enhancing patterns which are already obvious from the similar ‘meanings’ of the strong elements.

Table 4: Performance of elements across different self-defined groups of respondents. Only elements of +16 or higher are shown.

TAB 4

Appendix 4 shows AI suggestions about new products and services for the three mind-sets. These suggestions also emerge from the AI summarization of each set of answers.

Moving Beyond Knowledge to Actionable Communication – The Personal Viewpoint Identifier (PVI)

The ‘project’ of Mind Genomics does not stop at the experiments and at the identification of relevant interpersonal messaging and actions. Rather, having been nurtured in the business environment from the pioneering efforts of Wharton Business School professors Paul Green and Yoram Wind [17], Mind Genomics carries in its DNA the opportunity for application. The specific application is a bank of knowledge about interpersonal communications in the world of professional and client. The objective is not to prescribe clinical specifics but rather to tailor the style of communication to that which is empirically most appropriate for the patient. The data in this granular level study of the general nature of what to say to parents of children regarding obesity is a good example. What are the types of words to which the three different mind-sets will attend? The topic of childhood obesity seems so well defined that it’s quite likely the health professional might not even realize that more success could be had by knowing how to frame one’s messages It is to the end of assigning a person to a mind-set for purposes of general understanding ‘how to communicate’ that we now turn. During the past several years, the notion of developing a personal viewpoint identifier has come to the fore for a variety of issues [18,19]. The rationale for the of a personal viewpoint identifier, or indeed, for any typing tool is that once the science is established, those who need to know what to communicate are given actionable suggestions. The experienced person might not need that PVI typing tool, but the inexperienced person does, the person with decades of trial and error which makes the person an expert. The objective of the PVI is to present an individual with a simple to complete questionnaire, shown by Figure 8. Panel 8A shows the consent form, and background material about the respondent. Panel 8B shows the six-question typing tool. Panel C shows the assignment of the respondent to one of the three mind-sets, as well as feedback for the three mind-sets. The researcher creates the titles for the mind-sets, and the messages. The underlying algorithm sorts create the assignment method. There are many available statistical methods to create typing tools, such as discriminant function analysis or CHAID. It will be the large set of these easy-to-create databases and typing tools that will allow a new vista to emerge. This opportunity with be deeper, quickly and easily obtained knowledge of specifics in style, in language, to help professionals understand those who seek their advice. In other words, a system to understand the way the patient wants to interact as a person with the medical professional. The approach is not to diagnose the patient, not to suggest anything other than revealing the most likely ‘best style’ of communication.

FIG 8

Figure 8: The PVI (personal viewpoint identifier) for the obesity topics covered in this study

Table 5: How the 16 elements perform among the Total Sample vs among three emergent mind-sets

TAB 5

Setting up the PVI is straightforward (see www.pvi360.com). The setup is formatted to accept Excel-type data, meaning that either the entries are entered by hand, or a complete excel matrix can be copied and pasted. Figure 9 shows the formatted sheet, which needs only be completed, with data easy to copy from the results file.

FIG 9

Figure 9: Set-up form for the PVI (personal viewpoint identifier), coded by colors to make the set up easy even for beginners.

Discussion and Conclusions

The foregoing study represents just one effort to generate deep knowledge about messaging to individuals regarding a health and wellness condition. The contribution of this paper is to present an approach which can generate large amounts of data about how people think, the data coming from different topics, or the same topic with different messages, or even the same topic with the same message across the world. The potential now exists for the industrial-level acceleration of curated primary information about how one should communicate with people, either within a topic area such as obesity, across different areas such as better living, and across different populations [20]. The challenge for today is straightforward. The challenge is to create this depth of knowledge on a daily basis, for all topics where people need to speak with those who are tasked with helping them on the arc of wellness and health to the point where they need intervention. Can these studies of effective language be automated around the world so that the precious, diminishing time of health professionals can be spent communicating in the best, most effective, kindest way possible, for every individual who shows up requesting health. It is that vision, of truly industrial-scale knowledge of the ‘how to effectively communicate’ which might well return a modicum of interpersonal intimacy, trust and communication effectiveness to a system thought by many to be slowly breaking down.

Acknowledgments

The author acknowledges the help provided by Yehoshua Deitel of Sifra Digital Inc., in Israel, both in the development of the Mind Genomics system and the Idea Book. The authors also acknowledge Professor Attila Gere of Hungarian University of Agriculture and Life Sciences and Mr. Robert Sherman of Robertsoft for their efforts in creating the PVI.

References

  1. Han JC, Lawlor DA, Kimm, SY (2010) Childhood obesity. The Lancet. [crossref]
  2. Lakshman R, Elks CE, Ong, KK (2012) Childhood obesity. Circulation. [crossref]
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Epidemiological and Anatomopathological Profile of Prostate Cancer in Thiès

DOI: 10.31038/CST.2023831

Abstract

Prostate cancer is a cancer of the elderly: it rarely appears before the age of 50, and the incidence increases very rapidly with age. Age is the main risk factor identified for prostate cancer. This risk is 1% to 7% between 50 and 64 years of age, and rises from 14% to 26% between 65 and 74 years of age. These risks increase by 40% between the ages of 75 and 79 and reach 50% at the age of 80. The most common problems are: prostatitis or inflammation of the prostate, urinary urgency, urinary frequency, dysuria, acute retention of urine, or more rarely, initial hematuria. The objective of this study was to describe the epidemiological, clinical and histological aspects of prostate cancer in the Thiès region. Our study was conducted between January 2020 and December 2022 with a population of 165 patients with prostate cancer in the urology department of the regional hospital of Thiès and the Saint Jean de Dieu hospital of Thiès. The variables studied were age, PSA levels, Gleason score and histological grades. The mean age of the patients was 71.18 years with extremes ranging from 47 years to 90 years, and the mean PSA level was 965.33 ng/ml with extremes from 5 ng/ml to 5000 ng/ml. Our results show that 87% of our patients were older than 65 years. Gleason score 7 (4+3) was more represented with a rate of 37% corresponding to grade III according to WHO-ISUP 2016. The incidence is in average decline between 2020 and 2022.

Keywords

Prostate, Cancer, Epidemiology, Senegal

Introduction

In 2020, according to the World Health Organization (WHO), prostate cancer is the third most common diagnosed malignancy. With 1,414,259 cases (7.3% of the total), prostate cancer is preceded only by lung and colorectal cancer with 2,206,771 and 1,148,515 cases (11.4 and 10.0%), respectively [1]. It is the most commonly diagnosed cancer in more than 50% of the world’s countries (112 out of 185) and its incidence varies considerably between high and low human development index (HDI) countries, 37.5 versus 11.3 per 100,000 population respectively. Mortality rates are less variable (8.1 vs. 5.9 per 100,000 people). respectively. Mortality rates are less variable (8.1 vs. 5.9 per 100,000 people). Despite the significant burden of prostate cancer, established risk factors for this malignancy are limited to age, ethnicity, and a positive family history of the disease [2,3]. In fact, cancer incidence and mortality rates are strongly associated with increasing age, with a mean age at diagnosis of 66 years [4]. They also vary considerably by region and population, with higher rates among men of African descent and lower rates among those of Asian descent [3,5,6]. Epidemiological studies have also found that first-degree relatives of a patient with prostate cancer have two to three-fold increased risk of developing the disease compared to the general population, and the risk further increases with the number of affected relatives [7]. The aim of this prospective study was to investigate the clinical and histological characteristics of prostate cancer in the Thies region.

Material and Methods

This is a prospective descriptive study including 165 patients with prostate cancer. These patients were recruited from the urology department of the Thiès regional hospital and the Saint Jean de Dieu hospital in Thiès between January 2020 and December 2022. Inclusion criteria were a suspicious digital rectal exam with a PSA level above 4 ng/ml, then biopsies were performed for histopathological diagnosis. All histologically confirmed malignant prostate tumors were included in this study. Data were collected by consulting the hospitalization records, which are kept in the archives on a pre-established sheet, we collected in current family records the demographic data (name, surname, age, ethnicity, reason for consultation) and medical history. Data entry and analyses were performed using Microsoft Office Excel.

Results

Our epidemiological survey was conducted in a regional population of one hundred and sixtyfive (165) prostate cancer patients from January 2020 to December 2022. The mean age was 71.18 years with extremes ranging from 47 years to 90 years, and the mean PSA level was 965.33 ng/ml with extremes from 5 ng/ml to 5000 ng/ml. Only 16.36% of patients were less than or equal to 65 years of age (27/165). 82% of the patients were between 60 and 80 years of age and 11.5% were older than 80 years (Figure 1). In this study, the most common reasons for consultation were: urinary urgency, urinary frequency, dysuria, acute retention of urine, or more rarely, initial hematuria. The incidence of prostate cancer decreased with the years with 67 cases in 2020, 55 in 2021 and 47 cases in 2022 (Figure 2).

fig 1

Figure 1: Distribution of patients by age group

fig 2

Figure 2: Incidence of prostate cancer from 2020 to 2022

When examining histopathologic differentiation, patients with Gleason score 7 (4+3) prostate adenocarcinoma were more represented with a total of 61 individuals out of 165. According to WHO-ISUP 2016 grade, grade III was more frequent with 37% of cases, followed by grade IV with a frequency of 27%; only 2% of cases were grade V (Figure 3).

fig 3

Figure 3: Distribution of patients by WHO-ISUP Grade 2016

Discussion

We recruited 165 patients newly diagnosed with prostate cancer. These cases of prostate cancer, which made it possible to study the epidemiological and histopathological aspects of these cancers in Thiès, could not be considered as the totality of prostate cancers at the regional level because only anatomopathological specimens received in the laboratory were considered. The mean age of the patients was 71.18 years, comparable to those reported in the African literature, particularly that observed in Senegal by Gueye et al. and in Congo-Brazzaville by Peko et al. which is 69 years [8,9]. In the statistical studies of Amégbor et al. and Brawley et al. the average age of onset of prostate cancer is respectively 70 and 71 years [10,11]. In our study, the age range between 60 and 80 years was more represented with a frequency of 82%, this age range is found in Amegbor et al. and Ndoye et al. [12]. All these data confirm that prostate cancer is a disease of the elderly. According to Boyle, it is the most frequent cancer in men over 50 years old [13].

Moreover, the frequency of prostate cancer increases with age [14]. In our study 84% of the patients were older than 65 years. In addition, the low incidence rate in men under 50 years of age (0.06%) provided further evidence that the prevalence of prostate cancer is closely related to the increasing age of the patients [15,16]. The average age at diagnosis was high, hence the presence of these very advanced forms beyond any therapeutic resources. The delay in diagnosis is related to the natural history of prostate cancer but also to the apprehension that men had to come to the urologist. In addition, there is a lack of information and awarenessraising policy for the population about this condition, and difficulties in accessing health services. Contrary to the work carried out in the West where the average PSA level oscillates between 15 and 25 ng/ml [17], we note an elevation of the PSA level with an average PSA of 965.33 ng/ml, which is in line with the results of Nzamba et al. in Ivory Coast [18]. This PSA profile is consistent with the litterature, as several studies have reported a higher PSA level in African Americans than in Caucasian Americans [19-21]. This high PSA value could indicate early metastatic extension. In this study, Gleason score 7 (4+3) was the most represented with 61 out of 165 cases or 37%. This result is different from that of Jalloh et al. [22] and Amégbor et al. who found score 6 with a rate of 52% and 60% of cases. The patients in our study present a high proportion of advanced stage tumors (stage III and stage IV), i.e. 64% of cases; these results are matched with those of Gueye et al. and Benseba et al. in Algeria [23]. Only 2% of patients are stage V, these results confirm the progress of screening We note in this study population a relatively small decrease in the incidence of diagnosed cancers over the years, from 67 cases in 2020 to 43 cases of cancer in 2022. This relative decrease could be explained by a more targeted screening practice and improved diagnostic tools. Mass screening is not recommended. Early individual diagnosis is based on an annual PSA test associated with digital rectal examination in men between 50 and 75 years of age with a life expectancy of more than 10 years.

Conclusion

Prostate cancer in this study population is characterized by an advanced age at diagnosis that can lead to an advanced tumor grade correlated with a high PSA level and a huge frequency of metastasis.

The results of this study showed a relatively small decrease in the incidence of prostate cancer in the Thiès population for the period from 2020 to 2022. However, the relatively declining incidence in the Thies region indicates the need to optimize methods of timely diagnosis of prostate cancer by focusing on high-risk incidence groups.

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In Search of a Better Doctor Visit: A Mind Genomics Exploration

DOI: 10.31038/IMROJ.2023821

Abstract

This paper explores the messaging about a doctor visit from the perspective of survey takers (respondents) acting as prospective patients. Respondents evaluated systematically varied vignettes, combinations of messages about a visit to the doctor. The elements were developed using artificial intelligence embedded in the BimiLeap program. Respondents each evaluated 24 unique vignettes comprising systematically varied combinations of the messages. Each vignette presented 2-4 messages, combined according to a main- effects permuted experimental design. Based upon the response to the vignettes about a ‘visit to the doctor’s office,’ regression and cluster analyses revealed three different-sets of prospective and current patients (Patient Mind-Set 1: Focus on connection with the doctor after the visit; Patient Mind-Set 2 – Doctor is attentive to my needs and involves me; Patient Mind-Set 3 – Visit ends with what specifically to do..) The Mind Genomics approach presented in this paper can be used to educate medical professionals about what they believe patients want and what patients actually want regarding the visit to their health care office.

Introduction

A search through the academic literature reveals what many practitioners and patients already know, namely that the patient-facing system today is ‘broken’, or at least can be optimized. One need only read the titles of papers to get a sense of the massive dissatisfaction, both from the viewpoint of the patient books detailing the issues, but perhaps a more productive solution is to find out what patients need and want from the point of view of human interaction. The clinical issues are best left to the experts, but what about the issues of patient experience? [1-4].

In today’s climate of hyper measurement, of everything, the patient experience is measured with an almost religious fervor. Following most visits, one receives the now-expected follow-up survey of the experience (e.g., Press-Ganey, Siegrist, 2013) [5]. No patient visit is left unexamined, as the patient is warned to expect a follow-up satisfaction survey and requested to be sure to uprate the scores for the experience if at all possible. Survey after survey, whether from a survey professional or from the office of the practitioner show remarkably similar structure, namely rating general statements about the experience on some type of point scale, along with questions about recommendations. These general surveys continue to show a decreasing satisfaction with the interaction with the doctor in the office, a trend that people may talk about in casual conversation but has now become a topic in the world of professional medicine [6]. Some of the issues may be individual differences, with doctors and nurses varying in their so-called bedside manner. Those differences are built into the system. People behave the way they behave. They may be taught some ways around their behavioral shortcomings once these shortcomings are identified, but the people may take years to really improve their behaviors. The other issue is the change in the economics, with insurance companies and venture capital using the medical system to optimize financial yield by treating the visit with the patient as a product whose ‘financials’ are to be optimized as if optimizing the production of any item to be sold to customers.

Exploring Granular Thinking about the Visit to the Doctor through Mind Genomics

Over the past forty years an alternative way of thinking about measuring experience has emerged, the origins of which go back to the pioneering work of functional measurement, and the foundations of mathematical psychology embodied in conjoint measurement. The common property here is to present respondents with combinations of alternatives and get their ratings of these alternatives or their choices. The subsequent mathematical analysis relating the choices or ratings to the composition of the test combinations reveals the underlying strength of the individual options or elements. The rationale for this approach comes from the realization that people react to stories, to vignettes of experience, not to single statements. It is more ecologically valid to present ‘small stories’ to be judged, and in turn identifying ‘what’ in the stories drives the reaction of the person judging the stories. It is from this worldview that Mind Genomics evolved, as an attempt to explore the granularity of experience in a way that does not allow the respondent to ‘game’ the system [7,8].

Mind Genomics evolved from this pioneering work, focusing on simple, DIY (do it y 0urself) templates, and automated analyses Underlying the DIY template is a carefully structured path which ensures that the data emerging from the Mind Genomics study will be statistically correct, even to the level of an individual respondent. The Mind Genomics approach has been used for many different types of problems, ranging from medical to legal, ethics, consumer products, social issues and education. The approach is similar for virtually all of the studies, with the exception of the specific elements or messages, and the underlying experimental design. The specific steps for the Mind Genomics studies have been presented in various papers. The approach followed in those papers will be one used here, with slight variations, such as the use of regression modeling without an additive constant, a change which makes the data easier to understand [9-11].

This study focuses on the application of the Mind Genomics approach to the issue of what do patients want in the sessions with their doctors, and in turn what do doctors, nurses, and senior medical students feel they want in their interaction with the patient.

The Mind Genomics World View

When one thinks about the mind of the patient regarding the session with a doctor, or vice versa, the mind of a doctor or other health professional thinking about the session with a patient, the topic at first seems easy, almost self-evident. There is an almost instinctive positive or negative reaction when patients describe their experiences with doctors, a reaction that can range from rapturous to disillusioned. A lot of it is emotion, with the catch-all phrase of bed-side manner summarizing a great deal of the feelings about the experience, even when the meeting occur at an appointment rather than at the bedside of a sick person [12]. The important thing to note is the reality that for most patients the doctor or nurse is the professional with whom they will interact, from whom they may get good news or less fortunately, bad news. Whether the doctor is proficient or not may be relevant and can be determined from reviews and from talking to one’s friends, but the immediate situation is one of emotion. The emotional tension may be mild, such as the visit to the doctor’s office for a routine physical, or the emotional tension may be significant as in the case of the doctor calling the patient to come in talk about some issues which have just surfaced for the patient.

At the end of the visit, the patient is often asked to complete a survey about responses to the visit. Press-Ganey surveys are well known in this regard [13], although patients may be asked to fill out any of many different ‘home-grown’ surveys, devised by the staff of the specific medical practice. T the typical survey might end up having the patient rate the doctor’s behavior at the visit, perhaps doing so with one overall rating or perhaps dimensionalizing the visit into such direct issues as the rating of promptness, explanation of the situation, the amount of time spent with the doctor, and so forth. These numbers are tabulated and produced into a report profiling the visit as a series of scaled responses, like a report card, albeit one with more emotion but perhaps absence of soul anyway.

The foregoing approach allows the researcher to cover many topics, but in superficial terms only. In the effort to capture as many aspects as possible about the visit of patient and health professional, the researcher ends up giving each topic short shrift, usually covering the topic by one general question or perhaps two or three general questions. There is usually none of the richness of language to capture the experience, and the feeling about such experience.

It is at this point that Mind Genomics departs from the conventional methods. Mind Genomics presents vignettes, combinations of messages, to the respondent, and instructs the respondent to rate the feeling about the vignette. The respondent is not asked to be analytical, but simply to rate the feeling on a scale. The respondent ends up rating a set of these vignettes, combinations of messages, with each vignette comprising a minimum of two and a maximum of four elements. The vignettes are simple to read, convey detailed information, and simply require the respondent to scan them and assign a rating. From the evaluation of 24 such vignettes, each rated by a respondent, the researcher can assemble a profile of how each of test phrases (16 of them) drive the respondent’s feelings. The respondent feels that she or he is guessing, but nothing can be further from the truth. Each set of 24 vignettes evaluated by a respondent differs in combination from every other set of 24 vignettes. Underneath the combinations is a carefully designed layout, the experimental design, which puts these combinations together in a structured manner, allowing the respondent to react to a compound description, but with the ability to tease out the contribution of each element of the vignette. Often the survey-taker’s response is that the interview seemed jumbled, the elements seemed randomly combined, and instead of trying to answer honestly (another way for saying ‘giving the correct response’), survey-taker confesses that she or he simply guessed.

The analysis of the results provides a deep snapshot of how the respondent feels about the elements. The system cannot be gamed. The ability to probe a topic deeply rather than superficially means that it is now possible to deeply understand the topic. The data make a great deal of sense as will be shown below. Almost always, faced with 24 seemingly random combinations of messages about a topic, the respondent feels she or he gives up trying to guess and simply assigns a rating which seems ‘correct.’ It is that level of focus, the same level that economic psychologists Daniel Kahneman calls ‘System 1’ [14].

Mind Genomics data are deconstructed into the contribution of the different messages. The story is in the pattern of coefficients from models or equations relating the presence/absence of elements the messages, the response. These coefficients emerge from regression. The pattern of coefficients often points to different groups of people, the differences now coming from who they are but from how they think about the specific topic. These are the mind-sets, the desired information emerging from the study of the granular experience.

Setting Up the Study on the Mind Genomics Platform

To illustrate the approach of Mind Genomics we present a study on what low-income respondents feel they want from a visit with the doctor. Thus, the Mind Genomics projects here are done in the spirit of patient satisfaction studies (e.g., like Press Ganey survey), or the very many after-the-fact customer satisfaction surveys which try to dimensionalized the experience with the professional, the sales representative, or the help desk..

The study was suggested by a constant topic surfaced in the daily online, world-wide meeting among clinicians and allied parties, the Global Population Health Management Forum. A continuing theme of the FORUM is the recognition that people feel shortchanged by the current medical care system, especially in the United States, but increasing in other countries. These feelings about ‘shortchange’ actually came from the doctors themselves and were supported by both medical literature [15], and by popular literature and advertisement.

Figures 1-4 show the steps of the process represented by screen shots. Figure 1, Panel A show the first screen, requiring the respondent to assign the study a name, to select the language of the prompts (e.g., English, Chinese, etc.), and to agree not to request personal information unless specifically agreed to by the survey taker before the start of the study.

Figure 1 Panel B shows introduction to the AI-powered Idea Coach. Often the researcher is unable to formulate questions. This inability to formulate a string of questions is increasingly common because it requires critical and structured thinking. During the years 20222-2023 the emergence of easily available AI in the form of Chat GPT allowed for the Mind Genomics program to incorporate a system to suggest questions, based upon the input of the researcher. These questions are suggestions for discussion, and not meant to be informational. They teach about the topic by presenting different questions that the researcher can ask. The Idea Coach can be accessed dozens of times until the researcher has discovered the four questions that are of greatest promise. Each use of the Idea Coach generates 15 questions. With many uses of Idea Coach for the same ‘squib’ or problem description, Idea Coach will produce a number of different questions, but some questions will repeat.

Figure 1 Panel C shows a set of questions produced by AI through Idea Coach. To reinforce the spirit of experimentation and inquiry and to reduce the fear of asking question, the Idea Coach can be re-interrogated as many times as desired. After a while the same questions will appear. The different suggestions for questions from Idea Coach will be stored for subsequent analysis and returned to the researcher in a comprehensive package called the ‘Idea Book’. The Idea Book is separate from the study, set up as a document to help learning.

Figure 1 Panel D shows the final four questions selected by the researchers with the aid of AI (Idea Coach), but with the language edited by the researcher to make the question easier to understand. Figure 2, Panel A shows the output of one run of Idea Coach to select answers for question A (How do you want to spend your visit with the doctor). Each iteration of the Idea Coach to provide answers to the questions will generate 15 answers. As in the case of generating questions, generating answers will produce both new answers and repeats.

fig 1

Figure 1: The first part of the set-up for the study, using a templated system and AI (Idea Coach) to suggest questions.

Figure 2, Panel B shows the four answers that were accepted by the researcher. As was the case for the generation of questions, the idea Coach returns with a mix of previously used answers and new answers. Once again the researcher can modify the answer and may return to the question section to modify the language of the question.

Figure 2, Panel C shows the self-profiling question, set up so that the researcher can find out more detailed information about who the survey taker is, what the survey taker thinks, and what the survey taker does. The language in Panel C for eight questions is left to the researcher. The two remaining questions are age and gender.

Figure 2, Panel D shows the rating question that will be used by the respondent, who in the course of the study will evaluate 24 vignettes, created by combining answers together. For right now, it is only important to keep in mind that the scale has a minimum number of points (five), and that the scale has two dimensions, first For Me vs Not For Me, and then Positive Gut Feel vs Negative Gut Feel. In this way the study generates a deeper picture of how the survey taker feels. Figure 3 shows final thoughts and the open-ended questions.

fig 2

Figure 2: Creating the answers, the self-profiling classification question(s), and the rating scale

Figure 3 (Panel A) shows the box where the respondent. Figure 3B shows the box where the researcher can record the purpose of the study. The researcher is required to write something in this box. The rationale is that the BimiLeap system is used for teaching as well as for exploring real situations. As such, it is a good idea for the person designing the study to record the rationale. The study is also meant to be searchable on a big database, requiring that the researcher select key worlds.

fig 3

Figure 3: The opened-ended question and the final thoughts about the study as written by the researcher

The respondent experience begins with the greeting to the respondent, and then the self-profiling questionnaire as shown in Figure 4. The questions were created at the set-up time (see Figure 2, Panel C). To make the introduction less daunting, the BimiLeap program presents the questions in one page, but the answers in pull-down form. The respondent provides the necessary information, including the agreement not to provide any information that would identify the respondent. For those cases where it is necessary to know who the respondent actually is, the study must be augmented by permission forms. Otherwise, the default is total privacy.

fig 4

Figure 4: The self-profiling questionnaire page

Figure 5 shows the vignette as it looks on a PC. The vignette presented to the respondent is a stark collection of phrases, put into the different groups as answers to questions. The vignette shows only the answers, not the questions. The layout of the vignette throws information at the respondent in what must seem like a ‘blooming, buzzing confusion’ in the words of Harvard psychologist William James when asked to describe the perceptual world of the newborn baby. Despite the stark appearance, the vignette is effective as a means to throw information at the respondent in a way which allows them to ‘graze’, to pick up information quickly, rate the vignette, and move on to the next. After 24 vignettes the respondent does not feel ‘drained’ by having had to read an enormous amount of prose. The sheer starkness of the layout allows the researcher to move quickly through the vignettes, rather than being caught in the quicksand of too much verbiage.

fig 5

Figure 5: Example of a vignette as it looks on a PC. Each respondent evaluated a unique set of 24 such vignettes.

The actual combinations of elements (vignettes) are prescribed by an underlying experimental design. The experimental design was developed in a fashion which allows each respondent to evaluate a unique set of 24 vignettes. Each vignette has a minimum of two elements, and a maximum of four elements. The elements are answers to the questions. A vignette never contains more than one answer from a question, but many vignettes are absent from one question or absent from two questions, respectively. Finally, the experimental design is created so that each respondent ends up evaluating an isomorphic experimental design, viz., the same mathematical design but with different combinations. This is called an isomorphic permuted design [16].

How Low-income Respondents in New York Design Their Visit to the Doctor

This study focused on the design of a visit to the doctor by the patient. The respondents were chosen to be low-income individuals. The respondents were provided by a Mind Genomics vendor specializing in on-line survey-takers. The vendor, Luc.id Inc., provides totally anonymized respondents who fit the above-mentioned criteria (Table 1).

Table 1: Specifics for study 1 (Low-income respondents design visit to doctor)

tab 1

Each respondent evaluated a full set of vignettes, as structured by the underlying experimental design. To reinforce the point made above, each respondent evaluated a totally different combination of vignettes. The ratings on the 5-point scale were transformed to a binary scale. Ratings of 5 and 4 (For Me) were transformed to 100, ratings of 3, 2 and 1 were transformed to 0. The conversion of a Likert scale to a simple binary scale makes the results easier to communicate.

After the transformation, the data from each self-defined group was subject to an OLS (ordinary lease-squares) regression. The regression is expressed by the statement: Top2 = k1(A1) + k2(A2)…K16(D4). The coefficients tell us the additive percent of respondents who will rate the vignettes 5 or 4 (viz., ‘Me’) when the vignette contains the specific element.

Often researchers and respondents feel that the evaluation of vignettes complicates an otherwise easy task. Table 2 shows the strong performing coefficients across the 16 elements, and all of the subgroups. There are no clear patterns across groups, a situation which typically appears in Mind Genomics studies when the focus is on clearly different groups, but when there is no method for understanding the deep differences in the way of thinking. The clear patterns will emerge from mind-set segmentation, shown in the next section.

Table 2: High scoring elements for the rating of ‘Fits Me’. Coefficients of 21 and above are shaded

tab 2(1)

tab 2(2)

Mind-Sets in the Population

Mind Genomics was developed as a response to the psychophysics of the 1950’s and 1960’s, which searched for invariance, for the ‘one’ or ‘correct’ relation between physical stimulus level and subjective response. Psychophysicists typically work with well-defined physical stimuli, such as tones of varying sound pressure levels, weights of varying mass, circles of different areas, or money of various amounts. The standard approach espoused by Harvard psychophysicist, S Smith Stevens was to present unpracticed respondents with stimuli of various magnitudes, instruct the respondents to rate the perceived intensity, and then plot the relation between the number assigned (so-called magnitude estimate) and the physical magnitude [17]. The relation conformed to a power equation pf the form Rating = k(Physical Magnitude)n. The exponent n becomes the slope when the foregoing power equation or power function was linearized by being plotted in log-log coordinates, viz. log Rating = log k + n(Log Physical Magnitude). Note that it was within this tradition the author HRM received his PhD with Professor Stevens, in 1969.

The linearizable power function breaks down when the rating is degree of liking. In that case the relation is a curve perhaps like a parabola. There is an optimal level of liking somewhere in the middle stimulus range [18]. Just as important, the optimum point varies across people. The optimal level of liking may be of low intensity, medium intensity or high intensity. One need only think of the addition of sweetener or whitener to coffee/ some people like sweet dark coffee, others like light but non sweet coffee, and so forth.

With the differences in optimal points, one needs to cluster the respondents to identify meaningful, although operationally defined groups, called taste segments. The same thing can be done for the different messages in a Mind Genomics study to identify mid-sets. The thinking is the same; create a measure for each individual showing the pattern of elements which drive interest, and then cluster the respondents based upon similarities these patterns.

The process to develop these segments, not of taste but of thinking, follows a straightforward path, one which does not make any assumptions but rather combines statistical analysis by k-means clustering [19], followed by regression analysis to create the ‘mind-set’ equations, and then interpretation. The interpretation of the clusters is left to the researcher, with the suggestion that there be as few clusters or ‘mind-sets’ as possible (parsimony), but with the mind-sets interpretable.

Table 3 shows the coefficients for the mind-sets. The data could have been limited to two mind-sets, but the clustering solution for two mind-sets was unclear. When three mind-sets were extracted the results made more sense. Table 3 shows the strongest performing elements for each mind-set. From time to time an element might perform well in two of the three mind-sets, almost never in three of the three mind-sets.

Table 3: Performance of the 16 coefficients among respondents assigned to the Total Panel and then to one of three mutually exclusive and exhaustive mind-sets. Strong performing elements, coefficients of 21 or higher, are shown in shaded cells.

tab 3

The three mind-sets are not mutually exclusive, but rather reflect the existence of individuals who stress different aspects of the visit with the doctor or other medical professional.

Patient Mind-Set 1: Focus on connection with the doctor after the visit

Patient Mind-Set 2 – Doctor is attentive to my needs and involves me

Patient Mind-Set 3 – Visit ends with what specifically to do.

During the past several years the emergence of AI, artificial intelligence, has become of increasing interest to researchers. The Mind Genomics program in BimiLeap now incorporates a set of queries for the strong elements of each key subgroup. Table 4 presents the AI ‘summarization’ of the three mind-sets. The summarization is not meant to replace the human interpretation but rather to highlight some possible patterns that would not have been suspected.

Table 4: AI summarization of the strong performing elements for each mind-set by using Chat GPT to identify commonalities among these elements.

tab 4(1)

tab 4(2)

tab 4(3)

During the past several years the emergence of AI, artificial intelligence, has become of increasing interest to researchers. The Mind Genomics program in BimiLeap now incorporates a set of queries for the strong elements of each key subgroup. Table 4 presents the AI ‘summarization’ of the three mind-sets. The summarization is not meant to replace the human interpretation but rather to highlight some possible patterns that would not have been suspected.

From Knowledge to Application: Creating ‘Service-Based Products’

As part of the AI ‘summarization’ by fixed queries about strong performing elements (Table 4), the notion emerged that perhaps armed with the strong performing ideas the AI might be able suggest new innovative products, services, experiences, or policies. Table 5 shows these AI-driven suggestions. It is important to keep in mind that the raw materials for these suggestions are the elements that were found to be most appealing by the mind-sets of actual people, the respondents or survey-takers participating in the study. Whether the suggestions are good or poor, meaningful or meaningless, is not the issue here. Rather, the ease with which the researcher can work with ordinary people to understand in the particulars of the wellness-illness continuum means that one can now use AI to suggest possible solutions to the problem. With a Mind Genomics study taking less than one hour to set up with the Idea Coach, about one-to-three hours to ‘field’ with a paid panel of survey takers, and about 30 minutes for complete analysis, the potential is here to systematize the array of problems and arrive at prospective solutions that can be tested in the subsequent iterations of the Mind Genomics process, perhaps a day later.

Table 5: AI driven suggestions for new or innovative products, services, experiences or policies, based upon the analysis of the strong performing elements in a Mind Genomics study.

tab 5

Table 6: The form used to create the PVI (personal viewpoint identifier). The format is a drag-and-drop powered by Microsoft Excel®.

tab 6

The PVI (Personal Viewpoint Identifier): Understanding New People through a Short Interview

The final topic of this paper is the creation of a tool to assign people to one of the three mind-sets. The notion of mind-set as a way of looking at the world is clear. What has become increasingly obvious is that people differ from each other in the style that they find most comfortable, whether the situation is buying food, interacting with friends, or even dealing with medical professionals during a visit. The differences are not in the substance of what is discussed, but rather the general style, the types of words, the types of feelings that are conveyed during the interaction. In the world of commerce this is known as the nature of the interaction such as the interaction between a sales prospect and a salesperson [20]. The knowledgeable salesperson adjusts the language and behavior to what is deemed most likely to make the sales prospect be interested in listening and perhaps even buying. In the medical world this sensitivity to how a patient likes to interact with the medical professional is also important. Often in part it is referred to as the doctor’s ‘bedside manner.’

The next question to apply this knowledge is to recognize how a person wants to be treated in the meeting with the medical professional, e.g., in the doctor’s office, in the hospital, even on the phone with telehealth. Is there a way to discover the person’s desired ‘style of interaction’ in a rushed, crowded environment, with say a new and inexperienced, young medical professional, perhaps doing a rotation in a foreign country? In other words, can the Mind Genomics results be incorporated into an easy-to-use tool, administered in less than a minute, to tell the medical professional the type of interaction that the person might find to be most comfortable. The questions are simply those asked by any consumer researcher, on the web. The analysis of the answers puts the individual into one of the three groups, with the new benefit that the medical professional has a sense of how to interact with the patient because of some new, codified knowledge [21].

During the past three years a great deal of effort has gone into creating a system which allows a person to develop a typing tool, based upon the summary data from the study, data which parallels the numerical results of Table 3, along with the option to provide feedback and recommendations to the user of the tool, and the ability to show a video, as well as obtain additional information from up to four new questions. Table 5 shows the input structure for the PVI, in three sections; names/feedback/rating questions, additional questions to be answered (chosen by the researcher), and the summary data from the three mind-sets used to create the PVI. In turn, Figure 6 shows the PVI as the respondent see it. The left panel shows database questions about the respondent. The right panel shows the six questions. The output ends up being information to the clinician about the style that the respondent finds best, viz., the style preferred by one of the three mind-sets. Thus, the clinician understands the mental ‘WHO’ in terms of what is relevant at the level of interpersonal behavior, perhaps allowing the clinician to fine tune the interaction to make it smoother [22-26].

fig 6

Figure 6: The PVI (personal viewpoint identifier), as shown to the respondent who completed the questionnaire. The results are immediately databased, and returned to the clinician and, when desired, to the patient.

Discussion and Conclusions

As the medical system continues to ‘break down’, at least in the minds of many medical professions as well as the rank-and-file individuals who are the patients, opportunities exist to improve the system, even without improvement in clinical aspects. The improvements presented in this paper are simple to discover with the Mind Genomics technology and with Idea Coach. The decisions about which improvements are most promising emerge from treating the effort as a conventional market research study. The output of the effort ends up being suggestions for behavior from the Idea Coach, and initial suggestions of promise from work with consumer survey-takers, the respondents in the study. These individuals can be stratified by who the people are (viz. geo-demographics), what the people do, what the people believe. The Mind Genomics technology through the BimiLeap platform works with those divisions of people but adds to those divisions the ability to identify new-to-the-world groups of individuals, not based on general behaviors, but rather base on their responses to granular level descriptions of the situation. It is the compilation of such data which promise the ability to know what to do, at least at the level of person-to-person interaction to create a better medical experience, here specifically a better visit to the doctor.

References

  1. Anderson R, Barbara A, Feldman S (2007) What patients want: a content analysis of key qualities that influence patient satisfaction. Journal of Medical Practice Management 22: 255-261. [crossref]
  2. Barsky AJ (1981) Hidden reasons some patients visit Annals of Internal Medicine 94: 492-498.
  3. Berry DC, Gillie T, Banbury S (1995) What do patients want to know: an empirical approach to explanation generation and validation. Expert Systems with Applications 8: 419-428.
  4. Topol E (2015) The Patient Will See You Now: The Future of Medicine Is In Your Basic Books.
  5. Siegrist Jr RB (2013) Patient satisfaction: history, myths, and misperceptions. AMA Journal of Ethics 15: 982-987. [crossref]
  6. Sia B (2016) Physician burnout: a global Lancet 388: 2272-2281. [crossref]
  7. Anderson NH (1976) How functional measurement can yield validated interval scales of mental quantities. Journal of Applied Psychology 61: 677-692.
  8. Green PE, Krieger AM, Wind Y (2004) Thirty years of conjoint analysis: Reflections and prospects. Springer US 117-139.
  9. Moskowitz HR, Gofman A (2007) Selling Blue Elephants: How to Make Great Products That People Want Before They Even Know They Want Them. Pearson Education.
  10. Davidov S, al Humaidan M, Gere A, Cooper T, Moskowitz H (2021) Sequencing the ‘Dairy Mind’ using Mind Genomics to create an “MRI of Consumer Decisions”. New Advances in the Dairy Industry. Intech Open.
  11. Losavio J, Gollub E (2022) Application of mindsets to Health education and behavior change. Programs. Health 14: 407-417.
  12. Epstein RM (2006) communication research matter: what do patients notice, what do patients want, and what do patients need? Patient Education and Counseling 60: 272-278.
  13. Newgard CD, Fu R, Heilman J, Tanski M, Ma OJ, et al. (2017) Using Press Ganey provider feedback to improve patient satisfaction: a pilot randomized controlled trial. Academic Emergency Medicine 24: 1051-1059.
  14. Kahneman D, Frederick S (2007) Frames and brains: Elicitation and control of response tendencies. Trends in Cognitive Sciences 11: 45-46. [crossref]
  15. Masege D (2022) Managed Healthcare: Ethical implications on the doctor-patient relationship (Doctoral dissertation, School of Clinical Medicine, University of the Witwatersrand, Johannesburg)
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  17. Stevens SS (1975) Psychophysics: An Introduction to its Perceptual, Neural and Social Prospects, New York, John Wiley.
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  20. Moon Y (2002) Personalization and personality: Some effects of customizing message style based on consumer personality. Journal of Consumer Psychology 12: 313-326.
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Raman Spectroscopic Determination of Water in Glasses and Melt Inclusions: 25 Years after the Beginning

DOI: 10.31038/GEMS.2023554

Abstract

The method of water determination in glasses and melt inclusions with Raman spectroscopy is shown. Furthermore, it is demonstrated that by the linearity of the calibration curve in a concentration range from 50 ppm to nearly 50%, the practicability of the method can be significantly simplified.

Keywords

Raman spectroscopy, H2O and D2O determination, Reference samples, Calibration

Introduction

Water is the most essential magmatic and pegmatitic volatile. Exact knowledge about the amount of water during magmatic and pegmatitic crystallization processes is critical in understanding the behavior of volatiles in silicate melts. Therefore a simple method for determining total water is necessary. The technique must be simple, fast, and cover a large concentration range. Because most information about the water in magmatic-pegmatitic systems comes from melt inclusions, the method must also have a high spatial resolution and sensitivity.

The author used micro-Raman spectroscopy from the beginning (1998) for this challenge. The author published the first data produced between 1998 and 1999, together with Webster and Heinrich [1]. The most significant problem was the acquisition of reference glasses with testified and independently determined water concentrations. The demand for a simple Raman spectroscopic method was in the air [2]. However, they are standing in this time only at the beginning.

Reference Samples

The prerequisite for developing such a method is the availability of good standards. In the first step, the author used synthetic glasses (of albite, granitic, or pegmatitic composition) with water concentrations determined with the Karl-Fischer method. These glasses come from different authors quoted in Thomas [3]. Water concentrations higher than 12% in silicate glasses are unstable over time. Therefore the author used for higher concentrations water-rich melt inclusions in pegmatite quartz. To obtain the necessary homogenous glass, the melt inclusions must be homogenized under pressure and subsequently fast-cooled using rapid quenching methods (see Thomas et al. [4,5] and the Electronic supplementary material – ESM). Generally, homogeneous, water-rich melt inclusions (>25%) are metastable and disintegrate after the first measurement into a water-bearing stable glass and a water-rich volatile sub-phase. Then the bulk-water concentration results from the water in the partial volumes. We used pure glasses as references with very low water concentrations below the ppm level. Thomas [6] determined the water content of these glasses with the self-calibrating proton-proton (pp) scattering method [7].

Methodology

For the measurements, we used primarily a Dilor XY Laser Raman Triple 800 mm spectrometer equipped with Olympus optical microscope and long-distance 80x and 100x objectives. The spectra were collected with a Peltier-cooled CCD detector [8]. We used the 514 and 488 nm lines of a Coherent Ar+ Laser Model Innova 70-3 and a power of 150 mW for the sample excitation. We obtained effectively interference-free Raman spectra of tiny melt inclusions embedded in the transparent quartz matrix with the confocal technique. The integral intensity in the 3100-3750 cm-1 frequency range was used for all measurements. Starting in 2006, we used a LabRam HR800 UV-VIS spectrometer for all further measurements. This exchange gave no problems because we could, using well-studied reference samples, transform the calibration data for the new device. Furthermore, starting in 2006 [6,9], we used the so-called “Comparator Technique,” justified by the fact that the calibration curve is strongly linear and goes practically through the zero point. However, note here that the exact zero point is for the integral intensity not defined. With this work in 2006, we could also determine the water content in homogenized melt inclusions deep in the quartz matrix by extrapolation to a deep of zero because the integral intensity increases linearly with the decrease of the surface layer (for example, by polishing). Another way is the use of different deep inclusions with the same composition. If on both sides, the polished sample is not too thick (~200 µm) and the inclusion is not precisely between both surfaces, then can, from two measurements, the water content adequate be determined or estimated. In 2006 we also showed that integral intensity between 2250 and 2900 cm-1 for D2OT forms a linear relationship with the concentration and can be used to determine D2O beside the water. We have made such determination (H2O and D2O) for water-rich melt inclusions in the Shaw meteorite [10]. Regarding the rare appearance of fluorescence in the frequency range around 3500 cm-1, we have used the weaker band at about 1630 cm-1 (see McMillan) [11] to quantify water.

Results

The primary aim was to complete a general calibration curve for an extensive concentration range. Also, the simplicity of the method was always a request. That was only possible because the research on melt inclusion was an essential target over a long time. Forty-eight different glasses or melt inclusions were used for the calibration curve plot. Each point represents at least 10 determinations [8]. For simplification, the ± 1s standard deviation is not shown (however. can be seen in Thomas) [8] (Figure 1).

The proof of a general calibration curve was the prerequisite for applying the Raman spectroscopy in the simplified form of the “Comparator Technique.” That means only one certified reference sample is necessary to determine water in glasses and melt inclusions.

fig 1

Figure 1: Generalized calibration curve for water in silicate glasses and melt inclusions in the concentrations range from about 50 ppm to nearly 50% – that is five orders of magnitude. The upper smaller diagram shows the concentration range from 50 ppm (determined with pp-scattering – see Reichard et al. (2004) and Thomas et al. (2008) to 1%.

Conclusions

The outlined Raman method for determining H2O and D2O can also be used in analogy for other components. Thomas (2002) [3] has shown that for boric acid [H3BO3] in fluid and melt inclusions and later also for sulfate [12], carbonate/hydrogen carbonate (see, e.g., Thomas et al. 2020) [13].

Acknowledgment

I thank many colleagues and coauthors who accompanied me for over 50 years in my research, mostly on melt inclusions. Furthermore, I think here first to Jim D. Webster (1955-2019), who was very interested in my work on melt inclusions in granites and pegmatites of the Variscan Erzgebirge. I am, above all, grateful to H. Behrens and F. Holtz (University of Hannover, Germany), Robert (Bob) Bodnar (Virginia Tech, Blacksburg), H.R. Westrich (Sandia National Laboratories, Albuquerque, New Mexico), and M. Leschik (University Clausthal, Germany) for providing glass standards with defined H2O and D2O concentrations. Bob Bodnar encouraged me at the ECROFI XV in 1999 to publish the first results on water determination very fast.

References

  1. Thomas R, Webster JD, Heinrich W (1999) Melt inclusions in pegmatite quartz: Complete miscibility between silicate melts and hydrous fluids. ECROFI Abstracts, Terra Nostra 99/6, 305-307.
  2. Chabiron A, Pfeiffert C, Pironon J, Cuney M (1999) Determination of water content in melt inclusions by Raman spectroscopy. ECROFI Abstracts, Terra Nostra 99/6, 68-69.
  3. Thomas R (2002) Determination of the H3BO3 concentration in fluid and melt inclusions in granite pegmatites by laser Raman microprobe spectroscopy. American Mineralogist 87: 56-68.
  4. Thomas R, Davidson P, Rhede D, Leh M (2009) The miarolitic pegmatites from the Königshain: a contribution to understanding the genesis of pegmatites. Comtrib. Mineral Petrol 157: 505-523.
  5. Thomas R, Davidson P (2016) Origin of miarolitic pegmatites in the Königshain granite/Lusatia. Lithos 260: 225-241.
  6. Thomas SM, Thomas R, Davidson P, Reichart P, Koch-Müller M, et al. (2008) Application of Raman spectroscopy to quantify trace water concentrations in glasses and garnets. American Mineralogist 93: 1550-1557.
  7. Reichart P, Datzmann G, Hauptner A, Hertenberger R, Wild C, et al. (2004) Three-dimensional hydrogen microscopy in diamond. Science 306: 1537-1540.
  8. Thomas R (2000) Determination of water contents of granite melt inclusions by confocal laser Raman microprobe spectroscopy. American Mineralogist 85: 868-872.
  9. Thomas R, Kamenetsky VS, Davidson P (2006) Laser Raman spectroscopic measurements of water in unexposed glass inclusions. American Mineralogist 91: 467-470.
  10. Thomas R, Davidson P (2019) Shaw meteorite: water-poor and water-rich melt inclusions in olivine and enstatite. Mineralogy and Petrology 113: 1-5.
  11. McMillan PF (1994) Water solubility and speciation models. In: Mineralogical Society of America. Reviews in Mineralogy 30: 131-156.
  12. Thomas R, Davidson P (2017) Hingganite-(Y) from a small aplite vein in granodiorite from Oppach, Lusatia Mts., E-Germany. Miner Petrol 111: 821-826.
  13. Thomas R, Davidson P, Rericha A (2020) Emerald from the Habachtal: new observations. Mineralogy and Petrology 114: 161-173.

Macrophage Activation Syndrome in a Patient with Systemic Lupus Erythematosus Undergoing Cyclophosphamide Treatment: A Case Report

DOI: 10.31038/JCRM.2023631

Summary

Macrophage Activation Syndrome (MAS) is a disorder related to hemophagocytic lymphohistiocytosis and is a life-threatening complication of rheumatic diseases. The diagnosis is challenging because MAS symptoms are quite similar to those of many active autoimmune diseases or severe sepsis. We describe the case of a female patient with systemic lupus erythematosus that presented with symptoms suggesting acute decompensation of autoimmune disease and sepsis. She was later diagnosed with MAS. Despite an aggressive immunosuppressive treatment, she developed a fatal outcome.

Keywords

Macrophage activation syndrome, Systemic lupus erythematosus, Hemophagocytic lymphohistiocytosis, Infection

Introduction

Macrophage Activation Syndrome (MAS) is a disorder related to hemophagocytic lymphohistiocytosis (HLH) and is a life-threatening complication of rheumatic diseases [1]. HLH is divided into primary and secondary. While primary (or familial) HLH is an inherited disease, secondary HLH is triggered by other diseases, including infections, malignancy, and autoimmune diseases [2]. MAS is a secondary HLH associated with autoimmune diseases. The most common is Systemic Juvenile Idiopathic Arthritis (SJIA), followed by Systemic Lupus Erythematosus (SLE) [3].

The diagnosis is challenging because MAS symptoms are quite similar to those of many active autoimmune diseases or severe sepsis. The typical signs and symptoms are persistent fever, hepatosplenomegaly, lymphadenopathy, and hemorrhagic manifestations. Abnormal results include cytopenia, coagulopathy, and hyperferritinemia. Since the mortality rate is up to 50% in adults, early recognition of MAS is important to improve prognosis [4].

We describe a case report of a SLE female patient that presented with symptoms suggesting acute decompensation of the auto-immune disease and sepsis. She was later diagnosed with MAS. Despite aggressive immunosuppressive treatment, she developed a fatal outcome.

Case Report

A 29-year-old female patient with a known diagnosis of SLE since 2013 was admitted to our Department with a 38.5°C fever, diffuse edema, and dyspnea with one-week duration. Her medical history was characterized by severe SLE with recent exacerbation of the disease while on maintenance treatment with azathioprine and oral low-dose corticosteroids. Therefore, she was started on treatment with cyclophosphamide. At admission, laboratory examination revealed anemia (hemoglobin of 4.3 mg/dL), leucocytes of 8,980/mm3 (35% immature forms), and low platelet count (83,000 per microliter of blood). There were signs of renal impairment function (creatinine of 1.97 mg/dl) and proteinuria (urinary protein-to-creatinine ratio of 1.8). The albumin concentration was low (0.8 g/dL). The complement levels were reduced (C3 of 70 mg/dL and C4 of 30 mg/dL). An abdominal ultrasound revealed splenomegaly.

At this point, cyclophosphamide was withdrawn due to the possibility of medication toxicity. She was kept on a high dose of prednisone. Given the possibility of systemic infection, the patient was started on large-spectrum antibiotics (piperacillin plus tazobactam). An upper Gastrointestinal (GI) endoscopy revealed signs of esophageal candidiasis, but no ulcer or gastritis. Therefore, oral fluconazole was also started. Blood and urine cultures posteriorly showed negative results.

Despite the treatment with antibiotics and antifungals and the suspension of cyclophosphamide, the patient presented an unfavorable evolution. After 3 days of blood and filtered platelet transfusion, she recurred with anemia (hemoglobin of 5.2 mg/dL) and low platelets (13,000 per microliter of blood). Additional blood exams also revealed leucopenia (5,120/mm³), normal fibrinogen (216 mg/dL), and high ferritin levels (10,508 ng/mL – reference of 13-150 ng/mL). The aspartate aminotransferase was elevated (65 units/L), as well as triglycerides (236 mg/dL). After a few days, the renal function continued to deteriorate (creatinine level of 2.86 mg/dL) with metabolic acidosis.

A bone marrow biopsy was performed after hematologic consultation, showing evidence of hemophagocytosis (Figure 1). At this point, treatment was switched to high dose of methylprednisolone (1000 mg per day) associated with intravenous immunoglobulin (30 g per day for 5 days). Despite this approach, on the third day, the patient presented with massive GI bleeding. A new endoscopy did not reveal any ulcer or severe gastritis. In a few days, respiratory failure treated with mechanical ventilation occurred, with the patient dying due to multiple organ failures.

fig 1

Figure 1: Bone marrow biopsy showing focal signs of hemophagocytosis (arrows). Hematoxylin and eosin stained, 1000× magnification.

Discussion

MAS is a secondary HLH related to rheumatic diseases, and it was first described as a complication of SJIA in 1985 by Hadchouel et al. [5]. It is a life-threatening condition characterized by cytopenia, high fever, liver insufficiency, andl coagulopathy.

Excessive activation and proliferation of T lymphocytes and macrophages or histiocytes lead to extensive hemophagocytosis in the bone marrow and cytokine storm [1]. The exact incidence of MAS in rheumatic diseases is still unknown. Although MAS is by far the most common disease in the pediatric population with SJIA, there have been increased reports of MAS related to SLE. The occurrence of MAS-associated SLE in adults is uncommon, ranging from 0.9 to 4.6%. It usually occurs in young female patients [3]. The clinical characteristics of MAS-associated SLE and active SLE are very similar. Both entities share clinical and laboratory features, which include fever, cytopenia, and splenomegaly. This makes the differential diagnosis very difficult. Hyperferritinemia is considered the best parameter to distinguish between MAS and SLE, with a sensitivity and specificity of about 100% favoring MAS [6]. Following multiple organ failures, if MAS is left untreated or even undetected, the mortality rate can rise to 42% in adults [4]. The index of suspicion for MAS is higher when an infection is ruled out or inflammation persists and does not respond to treatment of an underlying infection. In the present case, it could be useful to start an immunomodulatory therapy even in the face of infection. Systemic antibiotics were started as soon as the patient was admitted, even in the absence of a clear infection [7]. Although an identifiable precipitating factor is often not clearly identified, MAS has been related to numerous triggers, including among others a flare of the underlying disease, the toxicity of nonsteroidal antiinflammatory drugs, and viral infections [2]. Most cases are triggered by high activities of autoimmune diseases or infectious agents, resulting in a prolonged immune activation, predominantly by the cytotoxic T cells and the macrophages [3]. In a systematic review of the literature, Aziz et al. described the major risk factors that led to the development of MAS in SLE [8]. According to these authors, the most important precipitating factors were the lupus flare itself, its time of onset, and a high systemic lupus erythematosus disease activity index. Other factors identified were infections, drugs, underlying malignancy, and pregnancy [3,8]. Although there is no obvious cause of MAS occurrence in the present case, we considered the presence of systemic infection as the most probable cause. However, one should consider SLE decompensation or cyclophosphamide toxicity as potential triggers.

In 2014, Fardet et al. developed the HScore, a clinical tool that may be used to determine the probability of getting secondary HLH in adults [9]. This score encompasses 9 variables (known underlying immunosuppression, high temperature, organomegaly, triglyceride, ferritin, serum glutamic oxaloacetic transaminase, fibrinogen, cytopenia, and hemophagocitosys features on bone marrow aspirate). Each variable has a distinct weight, as reported by Fardet et al. [9] (Table 1).

Table 1: The HScore. Reproduced from Fardet et al. (2014) with permission of John Wiley and Sons.

Parameter

No. of points (criteria for scoring)

Known underlying immunosuppression 0 (no) or 18 (yes)
Temperature (°C) 0 (<38.4)

33 (38.4-39.4)

49 (>39.4)

Organomegaly 0 (no)

23 (hepatomegaly or splenomegaly) 38 (hepatomegaly and splenomegaly)

Number of cytopenias 0 (one lineage)

24 (two lineages)

34 (three lineages)

Ferritin (ng/mL) 0 (<2000)

35 (2000-6000)

50 (>6000)

Triglyceride (mmol/L) 0 (<1.5)

44 (1.5-4)

64 (>4)

Fibrinogen (g/L) 0 (>2.5)

30 (<2.5)

Aspartate Aminotransferase (U/L) 0 (<30)

19 (>30)

Hemophagocytosis on bone marrow 0 (no)

35 (yes)

According to the authors, the probability of having hemophagocytic syndrome ranged from <1% with an HScore of ≤90 to >99% with an HScore of ≥250 [9]. In the present case, the HScore was 263, suggesting a greater than 99% probability of having secondary HLH/MAS.

To date, several therapeutic options are available, including non-biologic and biologic treatments. The mainstay of MAS treatment is glucocorticoid therapy, usually with intravenous methylprednisolone 30 mg/Kg/dose (maximum 1 g) for 1 to 3 days. For the nonresponders, additional therapy with cyclosporin is recommended. For patients who are refractory to the high dosages of the above medications, alternative options like etoposide, cyclophosphamide, and plasma exchange can be useful, although no randomized trials are showing consistent results of these medications [3]. Recently, reports with biological agents in refractory cases have shown promising results, including infliximab, rituximab, and intravenous immunoglobulin [7].

In the present case, some factors may have contributed to the unfavorable outcome. Firstly, we must acknowledge that there was some delay in the proper diagnosis of MAS. Secondly, despite the recent use of cyclophosphamide and oral corticosteroid, none of these therapies were the first-line treatment for MAS; therefore, the institution of high-dose methylprednisolone contributed to an additional delay in correct treatment. Finally, our patient was treated in a public tertiary Brazilian institution that lacks the more recent options for the treatment of MAS refractory cases, like immunobiological agents. Consequently, we could only opt for intravenous immunoglobulin in the present case.

Conclusions

MAS is a challenging and life-threatening disorder related to HLH. The occurrence of MAS-associated SLE in adults is relatively uncommon. The clinical characteristics of MAS-associated SLE and active SLE are very similar. Both entities share clinical and laboratory features, which include fever, cytopenia, and splenomegaly. This makes the diagnosis very difficult. A high index of suspicion, associated with immediate treatment is essential for the achievement of better outcomes.

References

  1. Dhote R, Simon J, Papo T, Detournay B, Sailler L, et al. (2003) Reactive hemophagocytic syndrome in adult systemic disease: report of twenty-six cases and literature review. Arthritis Rheum 49: 633-639. [crossref]
  2. Ravelli A (2002) Macrophage activation syndrome. Curr Opin Rheumatol 14: 548-552.
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  5. Hadchouel M, Prieur AM, Griscelli C (1985) Acute hemorrhagic, hepatic, and neurologic manifestations in juvenile rheumatoid arthritis: possible relationship to drugs or infection. J Pediatr 106: 561-566. [crossref]
  6. Egües Dubuc CA, Uriarte Ecenarro M, Meneses Villalba C, Aldasoro Cáceres V, Hernando Rubio I, et al. (2014) Hemophagocytic syndrome as the initial manifestation of systemic lupus Reumatol Clin 10: 321-324. [crossref]
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  8. Aziz A, Castaneda EE, Ahmad N, Veerapalli H, Rockferry AG, et al. (2021) Exploring macrophage activation syndrome secondary to systemic lupus erythematosus in adults: a systematic review of the literature. Cureus 13: e18822. [crossref]
  9. Fardet L, Galicier L, Lambotte O, Marzac C, Aumont C, et (2014) Development and validation of the HScore, a score for the diagnosis of reactive hemophagocytic syndrome. Arthritis Rheumatol 66: 2613-2620. [crossref]

An Intriguing Half Leaf from the Middle Jurassic of China

DOI: 10.31038/GEMS.2023553

Abstract

Most leaves in angiosperms have reticulate venation, but not all leaves with reticulate venation belong to angiosperms. Although angiosperms flowers have been reported in the Jurassic, leaves similar to angiosperms are lacking in the Jurassic. Here we report an unnamed fossil leaf from the Middle Jurassic Yan’an Formation of Ningxia, China. Although its partial preservation does not allow us to determine the affinity of the fossil, its occurrence underscores the probablity that future digging may uncover angiosperm leaves in the Jurassic.

Keywords

Fossil, Middle Jurassic, China, Angiosperms, Leaf

The origin and early evolution of angiosperms, which have more than 300,000 species and account for more than 90% species diversity of land plants, have been foci of botanical debates for long time. Some palaeobotanists thought that angiosperms did not occur on the Earth until the Cretaceous. But recent years witnessed increasing reports of angiosperms in the pre-Cretaceous age. Among the reports, Schmeissneria [1,2] and Nanjinganthus [3,4], both from the Early Jurassic, are based on tens even hundreds of specimens, making a strong case that angiosperms have long existed in the pre-Cretaceous. Theoretically, angiosperm leaves have more potential to be preserved as fossils. But the fact is that angiosperm leaves are almost completely lacking in the Jurassic. This situation makes any trace of angiosperm-like leaves in the Jurassic especially badly wanted. Here we report a partially preserved leaf from the Yan’an Formation (the Middle Jurassic) of Lingwu, Ningxia, China (37°43’N, 106°26’E, Figure 1). The character assemblage of reticulate venation and intramarginal vein of this leaf are only seen in angiosperms hitherto, making it unique in the fossil record. This discovery makes future discovery of angiosperm leaves in the Jurassic more likely.

FIG 1

Figure 1: Geographical information of the fossil locality in Lingwu, Ningxia, China (37˚43’N, 106˚26’E). a. Fossil locality (black Square) in northwestern China. b. Detailed position of fossil locality (black square) in Lingwu, Ningxia, China.

The specimen (No. SGY007-16, deposited in Ningxia Geological Museum) was preserved as a compression with some coaly residue, uncovered from the Yan’an Formation (the Middle Jurassic), which is widely distributed in Northeastern China and has yielded various fossil plants [5-8]. The specimen is a grey siltstone slab 36 mm x 26 mm. The details were imaged using a Nikon SMZ1500 stereomicroscope equipped with a Nikon DS-Fi1 digital camera. All figures are organized using a Photoshop 7.0.

The leaf is incomplete, at least 15 mm long, 4 mm wide (Figure 2a). The leaf is smooth-margined, with an intramarginal vein (Figure 2a, 2b and 2d). Lateral veins parallel each other, branching from the midvein at an angle between 40° and 50° (Figure 2a and 2b). Lateral veins and transverse veins in between form angular meshes, which are 0.44-1.22 mm long and 0.2-0.45 mm wide (Figure 2a-2c). Lateral and transverse veins form right angles or acute angles, about 88 μm wide, with no obvious differentiation between lateral and transverse veins (Figure 2a-2c). Rarely, there is a freely-ending veinlet in an areole (Figure 2c).

FIG 2

Figure 2: The partially preserved leaf and its details. All scale bar=1 mm. a. The general view of the specimen, showing the partial lamina and partially preserved midvein (black arrow) and the freely ending veinlets (white arrows). b. Detailed view of the basal portion of the leaf, showing a branch in the background (white arrow) and vein (black arrow) after skeletonization. c. Vein meshes after skeletonization, showing the transverse veins (black arrows) between lateral veins. Note the freely ending veinlet (white arrow). d. Detailed view showing smooth margin with intramarginal vein (black arrow) and parallel lateral veins (white arrows).

Other than in angiosperms, reticulate venation has been in several fossil taxa, including ferns (Clathropteridaceae, Dipteridaceae, Polypodiaceae), uncertain group (Gigantopteridales), seed plants (Glossopteridales, Caytoniales, Ginkgoales, Cycadales, Gnetales, angiosperms, uncertain groups) (Table 1). Therefore the occurrence of reticulate venation does not ensure that a taxon with reticulate venation is an angiosperms [9-12].

Table 1: Comparison of our leaf and previously reported taxa with reticulate venation. Note that intramarginal vein is restricted to angiosperms.

Affinity

Margin

Reticulate venation

Intramarginal vein

Vein order

Midrib

Secondary vein

Freely ending veinlet

Age

Ref

Clathropteris Clathropteridaceae toothed frequent absent 4+ present unbranched absent Mesozoic
Hausmannia Dipteridaceae toothed frequent absent 3 absent dichotomous absent Mesozoic 9
Polypodium Polypodiaceae toothed frequent absent 3 present unbranched absent extant 10
Woodwardia Polypodiaceae smooth frequent absent 3? present dichotomous absent extant 10
Onoclea Polypodiaceae smooth frequent absent 2 present ? absent extant 10
Linopteris Seed plants smooth frequent absent 1 present? dichotomous absent Palaeozoic 10
Reticulopteris Seed plants smooth frequent absent 2 present dichotomous absent Palaeozoic 10
Lonchopteris Seed plants smooth frequent absent 2 present dichotomous absent Palaeozoic 10
Ginkgo biloba Ginkgoales infrequent 1 absent absent extant 10
Stangeria Cycadales frequent 2 multi-strand dichotomous absent extant 10
Ctenis Cycadales infrequent 1 absent absent Mesozic 10
Dictyozamites Bennettitales frequent 1 absent absent Mesozoic 10
Drewria Gnetales frequent 2 absent unbranched absent Cretaceous 10
Welwitschia Gnetales frequent 2 absent unbranched absent extant 10
Gnetum Gnetales frequent 4 multi-strand dichotomous simple-branched extant 10
Glossopteris Glossopteridales smooth frequent 2 present dichotomous absent Permian 10
Gangamopteris Glossopteridales smooth frequent 1 multistrand dichotomous absent Permian 10
Gigantonoclea Gigantopteridales frequent 4 multi-strand unbranched absent, branched Permian 10
Delnortea Gigantopteridales frequent 4 multi-strand unbranched absent Permian 10
Sagenopteris Caytoniales frequent 2 present dichotomous absent Jurassic 10
Sanmiguelia Uncertain variable 4 absent variable absent Triassic 10
Marcouia Uncertain frequent 2 present dichotomous absent Triassic 10
Furcula Uncertain frequent 3–4 present excurrent/dichotomous present Triassic 10
Pannaulika Uncertain frequent 4 present excurrent present Triassic 10
Myrtophyllum geinitzii Angiosperms smooth frequent present 4 present dichotomous Cretaceous 11,12
Myrtophyllum angustum Angiosperms present 11
Grevilleophyllum constans Angiosperms present 11
Eucalyptophyllum oblongifolium Angiosperms present 11
Eucalyptolaurus depreii Angiosperms smooth frequent present 3 present brochidodromous absent Cretaceous 11
Eucalyptolaurus Angiosperms smooth frequent present 3 present brochidodromous absent Cretaceous
Callianthus Angiosperms smooth frequent present 1 absent dichotomous absent Cretaceous
Our leaf smooth frequent present 2? present unbranched rare Jurassic

Our survey of fossil and extant taxa with reticulate venation indicates that, besides the implication given by reticulate venation, the occurrence of intramarginal vein appears to be restricted to angiosperms (Table 1). Therefore the occurrence of intramarginal vein in our new leaf seems to underscore its possibility of an angiosperm. This inference is further strengthened by the occurrence of freely ending veinlet in areole, which, although not strictly restricted to angiosperms, is only seen in angiosperms, Gnetales, and fossil taxa of uncertain affinity. Taking all together, despite its Jurassic age and partial preservation, our new leaf with reticulate venation, intramarginal vein, and freely ending veinlet suggests that, unlike widely-believed, angiosperms are more likely to be a truthful existence in the Jurassic, in line with previous reports of Jurassic flowers  [1-4, 13] and implication given by molecular clock studies [14].

Acknowledgement

This research was supported by the National Natural Science Foundation of China (42288201, 41688103, 91514302), Strategic Priority Research Program (B) of Chinese Academy of Sciences (XDB26000000), and Natural Science Foundation of Ningxia (2021AAC03471).

References

  1. Wang X (2010) Schmeissneria: An angiosperm from the Early Jurassic. Journal of Systematics and Evolution 48: 326-335. [crossref]
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  6. Wang X (1995) Study on the middle Jurassic flora of Tongchuan, Shaanxi Province. Chinese Journal of Botany 37: 81-88.
  7. Deng S, Fang L, Lu Y, Fan R, Yuan X (2010) The Mesozoic stratigraphy of the Ordos Basin. (Shehong, Sichuan, China).
  8. Deng S, et al. (2003) Stratum introduction. Petroleum Industry Press, Beijing.
  9. Golovneva LB, Grabovskiy AA (2019) The genus Hausmannia (Dipteridaceae) in the Cretaceous of the North-East of Russia and its paleobiogeographic implications. Cretaceous Research 93: 22-32.
  10. Trivett ML, Pigg KB (1996) A survey of reticulate venation among fossil and living land plants, in ” Flowering plant origin, evolution and phylogeny ” (eds) Taylor DW, Hickey LJ, Ch. 2: 8-31 (Chapman and Hall).
  11. Coiffard C, Gomez B, Thiébaut M, Kvacek J, Thévenard F, et al. (2009) Intramarginal veined Lauracease leaves from the Albian-Cenomanian of Charente-Maritime (western France). Palaeontology 52: 323-336.
  12. Gall L (2022) Paleobotany Division, Yale Peabody Museum. Yale University Peabody Museum. Occurrence dataset.
  13. Han L, Zhao Y, Zhao M, Sun J, Sun B, Wang, X (2023) New fossil evidence suggests that angiosperms flourished in the Middle Jurassic. Life 13.
  14. Li H, et al. (2019) Origin of angiosperms and the puzzle of the Jurassic gap. Nature Plants 5: 461-470.

To the Geochemistry of Beryllium: The Other Side of the Coin

DOI: 10.31038/GEMS.2023552

Abstract

Two very different examples of Be mineralizations show that Be can be enriched to very high values. This short paper tries to find the origin of such unusual behavior. Possibilities are discussed.

Keywords

Water-rich melt inclusions, Be-enrichment, Gaussian and Lorentzian element distribution, Supercritical phases, Mantle-crust connection

Introduction

There are a couple of comprehensive contributions to the chemistry, mineralogy, petrology, and geochemistry of beryllium: Gmelin [1] and the following issues up to 1997, Beus [2], Everest [3], Grew ES and coauthors (2002) [4], London D [5]. Considering such substantial work, it should be said all essential facts. However, careful microscopical and Raman spectroscopic studies of beryl mineralization tell us a different, maybe second, story. That is related to the supercritical melt and fluid state. While studying minerals from pegmatites related to the Variscan tin mineralization of Ehrenfriedersdorf in central Erzgebirge/Germany, we often found high concentrations of beryllium, which does not fit the expected ideas. In one melt inclusion (No. 61), Webster et al. [6] determined 1130 ppm Be with ion microprobe. This runaway value was the start-point for an intense search for higher Be values. And we were successful. In 2011 Thomas et al. [7] could, for the first time, present a more significant number of Be data for the Ehrenfriedersdorf pegmatite. Conspicuously, the data showed a strong dependence on the melt inclusions’ homogenization temperature and water concentration. As daughter minerals, we found in Ehrenfriedersdorf pegmatite beryllonite [NaBePO4] and hambergite [Be2BO3(OH, F)]. Beryllonite is related chiefly to water-rich melt inclusions in beryl of the beryl-quartz veins. In other beryl mineralizations, for example, Orlovka, Eastern Transbaikalia/Russia (Thomas et al. 2009 and unpublished data together with Badanina [8], and the Habachtal emerald deposit Thomas et al. 2020) [9], we found Be-carbonates as the main Be daughter phase. Many data and thoughts are presented in the concise papers by Thomas et al. [10,11]. Taleb’s book, “The Black Swan. The Impact of the Highly Improbable” [12], significantly influenced our thinking. Particularly the finding of water-rich melt inclusions, which depict a pseudobinary melt-water solvus in emerald, has stimulated our understanding of processes that are not so obvious. In this short contribution, we want to show that beryllium can enrich to very high concentrations – far away from all expectations. The first indications came from melt and fluid inclusions produced synthetically in morganite crystals from the Muiane pegmatite (Thomas et al., 2010) [13] during heating at 700°C, 1.0 kbar for 20 hours.

Samples

For this study, we used two different samples: well-transparent morganite crystals from the Muiane pegmatite [13] and beryl from a small beryl-quartz vein related to the Variscan tin deposit Ehrenfriedersdorf in the central Erzgebirge, Germany [14] – see Figure 1.

Other photos of such beryl-quartz samples from the Sauberg mine are in Thomas et al. [15,16].

FIG 1

Figure 1: Specimen from a beryl-rich quartz (Qtz) vein from the Sauberg mine near Ehrenfriedersdorf. The sample is from a vertically directed vein – is rotated by 90°. The dark minerals on the roof and the bottom are cassiterite, molybdenite, and other minor ore minerals.

Methods

Samples used in this study have been prepared over a long time, starting in 1996. For this, we used different high-pressure devices and analytical methods. A concise description of the methods used is summarized in Thomas et al. [17,18] and the ESMs. This study used a petrographic polarization microscope with a rotating stage coupled with the RamMics R532 Raman spectrometer working in the spectral range of 0-4000 cm-1 using a 60mW single mode 532nm laser. For Raman spectroscopic routine measurements, we used an Olympus long-distance LMPLN100x objective. Details are given in Thomas et al. [15,16]. In a paper published last year [19] used for the homogenization of melt inclusions the HDAC (Hydrothermal Diamond Anvil Cell) technique because water-rich melt inclusions show the tendency to leak or decrepitate during heating on the microscopic heating stage due to the growing pressure inside the inclusions. However, the HDAC method has the disadvantage that only a single or only a small couple of inclusions can be studied. Therefore we used different high-pressure devices coupled with rapid quenching because the obtained samples are larger and contain many inclusions which can be analyzed with other methods over the years. Of course, we also used the HDAC technique to see that our interpretations obtained from rapid quench experiments were realistic.

Results

Morganite from the Muiane Pegmatite, Mozambique

Morganite is the pale pink variety of beryl colored by Mn2+. Crystals from the Muiane pegmatite contain a lot of melt and daughter mineral-rich fluid inclusions. From first homogenizations at 700°C and 1.0 kbar for 20 hours, some melt inclusions homogenized totally, and during fast cooling, these inclusions heterogenized into a silicate melt and a fluid subphase containing hambergite [Be2BO3(OH,F)] and small bromelite [BeO] daughter crystals. According to Raman measurements, the silicate glass phase beside the water-rich phase has 7.0 ± 0.2% (n=36) H2O. After heating and quenching, a microscopic study of the morganite ships showed newly formed, very plane melt and fluid inclusions in newly formed halos around large melt inclusions produced during partial decrepitation under pressure. And to our surprise, these inclusions are rich in bromelite (Figure 2). These inclusions were not present before the heating procedure under constant CO2 pressure.

FIG 2

Figure 2: Bromelite [BeO]-rich inclusions in morganite, produced by heating to 700°C at a constant pressure of 1.0 kbar (20 hours) and fast quenching after the run.

That means that bromelite in water or water-rich melts are at 700°C highly mobile. This test tube-type experiment under pressure showed us Be’s high solubility and mobility. We obtained 14 to 17% Be in the H2O solution from well-formed inclusions. That is far from all ideas, for the natural examples given in Grew [4]. Another point is essential. There are two newly formed inclusion types: Melt- or glass-rich inclusions (Type-A melt inclusions) and water-rich inclusions with only a tiny part of the glass or melt (Type-B melt inclusions). Both inclusion types formed at the same time at 700°C. Furthermore, the observations show that the Be-content in the silicate glass with 7% water is very low and in the water-rich glass very high, higher than in the extreme water-rich inclusions.

Beryl-quartz Vein from the Sauberg Mine Near Ehrenfriedersdorf, Erzgebirge, Germany

This beryl contains a lot of fluid and melt inclusions. Many melt inclusions have beryllonite daughter crystals, which can be used to estimate the inclusions’ natural Be content [7]. Note here the inclusions are samples of the specific mineral-forming phases [20]. New studies [14] show that the beryl-quartz vein has a complicated history. This mineralization shows clear hints of the participation of supercritical melts or fluids – showing an interaction between mantle and crust [21]. The beryllium distribution of water-rich melt inclusions (Figure 3) around the solvus crest of a pseudobinary melt-water solvus shows this exceptionally well.

FIG 3

Figure 3: Gaussian distribution of Be (in ppm) in water-rich melt inclusions in beryl from the Sauberg mine near Ehrenfriedersdorf. The abscissa represents the measured inclusions’ determined bulk water content (H2O). All points are the mean of at least five determinations. The center of the peak is at 26.4% H2O, the width is 9.5% H2O, the height is 12075 ppm Be, and the offset is 214 ppm Be. The offset represents the general enrichment level of the mineralization in question.

Note, however, that the shown Gaussian curve represents average values produced by the limited number of analyses. The highest up-to-now measured Be value is 71500 ppm (from the volume of a well-formed beryllonite daughter crystal in a melt inclusion). By such values, the Gaussian curve degenerates to a Lorentzian curve type – typical for supercritical conditions. In normal beryl mineralization [5], such extreme enrichment is entirely out of the question. We must find an acceptable answer because the measured data are correct and always checked (starting with Webster et al.) – [6]. In the case of the beryl-quartz veins from the Sauberg mine near Ehrenfriedersdorf, enrichment in a miarolitic cavity is here not in question. The steady presence of HP and HT minerals (nanodiamond, moissanite, beryl-II, kumdykolite, and others) in the beryl-quartz mineralization makes it probable that supercritical phases from mantle depths participate in this mineralization. The finding of high-pressure and high-temperature minerals related to the Variscan granites and mineralizations supported this idea. To such phases belong spherical nanodiamond crystals, moissanite, stishovite, coesite, cristobalite-X-I, and beryl-II intergrown with moissanite and kumdykolite [15,16,21]. These minerals are all foreign crystals not in natural equilibrium with present surrounding mineralizations and rocks. Fundamental questions arise from the direct paragenetic relationship of beryl and moissanite at around 700°C and a pressure ≤ 2 kbar: (i) is the supercritical phase (melt, fluid?) primarily rich in beryllium? (ii) which role acts the spherical beryl-moissanite intergrow? (iii) what is the mechanism for the simultaneous growth of beryl and moissanite? (iv) Can such a mechanism be used for the technological crystallization of moissanite at significantly lower temperatures?.

Discussion

In cooperation with my coauthors, I found many element distributions showing such extreme enrichment, and such enrichment shows precise Gaussian or Lorentzian distribution curves [10,11]. For the Ehrenfriedersdorf deposit, we found Gaussian or Lorentzian distributed plots for the elements Li, Be, B, P, Cl, Zn, As, Sn, Cs, Ta, and W. A prerequisite for this analytical approach was developing a simple, destructions-free and fast analytical method for determining water in homogenized melt inclusions [22] on the base of the Raman spectroscopy. Different techniques were developed and used to determine the elements in question. Noteworthy was the Raman spectroscopic and electron microscopic determination of B, Be, and other elements [23,24], as well as the synchrotron radiation XRF (SXRF) method [25,26], the femtosecond LA-ICP-QMS microanalysis [27]. Why would up to now not similar distribution found? Is it a question of the direct availability of necessary experimental and analytical techniques? Or are such distributions related to the supercritical phases? Further studies on melt inclusions, for example, performed on mineralizations related to the Lusatian Block, made the last possibility highly plausible. More work is necessary!

Acknowledgment

I have written these lines in “we-form” because progress in the inclusion research would not be possible without the advisors and coauthors. I thank many colleagues who accompanied me for over 50 years in my research. Thanks go to Edwin (Ed) W. Roedder (1919-2006), which brought me on the right path. Furthermore, I think here first to Jim D. Webster (1955-2019), who was very interested in my work on melt inclusions in granites and pegmatites of the Variscan Erzgebirge, and I-Ming Chou and William (Bill) A. Bassett, who introduced me to work with the HDAC device and enabled me to judge this technique. Unforgotten is also the longstanding cooperation with Paul Davidson.

References

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The Role of the Nucleotide-Excision Repair (NER) Pathway in Soft Tissue Sarcomas: A Review and a Focus on Its Potential as a Therapeutic Target

DOI: 10.31038/CST.2023824

Simple Summary

The nucleotide excision repair (NER) pathway involves more than thirty protein-protein interactions and removes DNA adducts caused by chemotherapy drugs. The key genes of NER are often over-expressed in cancer cells and alterations of this pathway are responsible for increased or decreased sensitivity to specific therapeutic agents. This is particularly relevant in soft tissue sarcomas (STS), rare mesenchymal-originated tumors whose underlying mechanisms still lack understanding. Altogether the NER pathway components can be potential therapeutic targets in STS. The subtle regulation of NER activity may be clinically relevant as a surrogate prognostic marker or to predict sensitivity to chemotherapy agents. Further prospective evaluation of NER should be performed to address this question.

Abstract

Soft tissue sarcomas (STS) are low-incidence, mesenchymal-derived tumors represented by more than 50 his to types. Despite the latest developments, the rates of patients developing recurrent and metastatic disease are high. Many of the mechanisms underlying STS are still unknown, but there is evidence of the possible role of DNA damage response (DDR) pathways. DDR pathways include a variety of pathways used by cells to repair DNA damage of various kinds; they also have roles in protecting cancer cells from exogenous agents that target DNA, for these reasons are one of the main targets of potential anticancer therapeutic strategies. Nucleotide excision repair (NER) is one of the key repair pathways that can remove various bulky DNA lesions, often given by UV light, and is the main repair mechanism of DNA damage caused by carcinogens and chemotherapeutic drugs. Defects in NER are often the cause of several autosomal recessive genetic diseases. Variations in NER pathway actors can lead to a NER proficiency or NER deficiency condition, and this can be a risk, prognostic, and treatment response factor in cancer. This review focuses on the association between variations in the NER pathway in STS and is intended to point to NER as a pathway to focus on in the next future to optimize the treatments in use and improve the possibilities of personalizing therapies in STS patients in clinical practice.

Keywords

DNA damage response (DDR) mechanism, Nucleotide excision repair (NER), Soft tissue sarcoma (STS), Chemosensitivity

Introduction

Soft tissue sarcomas (STS) are rare tumors with more than 100 different histological subtypes. The scientific community has focused over the years on the search for biomarkers in STS [1] and the identification of variations at the genomic [2], expression [3], and protein [4] level.

Maintaining the integrity of genetic material is critical for the survival of all cell lines, yet various factors, both endogenous and exogenous, can compromise DNA stability. DNA damage repair (DDR) mechanisms are necessary for the maintenance of genome integrity. This is particularly important in cancer cells, in which mechanisms of resistance and sensitivity to radio- and chemotherapeutic cytotoxic agents are directly controlled by DDR pathways. For these reasons, DDR pathways are one of the main targets of potential anticancer therapeutic strategies [5]. The main DDR pathways are direct repair (DR), base excision repair (BER), mismatch repair (MMR), nucleotide excision repair (NER), non-homologous end joining (NHEJ), and homologous recombination repair (HRR) [6]. The absence or deficiency of a specific DDR mechanism can result in genomic instability and tumor progression. Certain modifications of specific genes of a DDR pathway are typical, with some frequency, of some specific cancers [7]. Transcriptomic profiling of tumor tissues suggested codependences between DDR pathways, indicating a potential benefit of combination therapies, which were confirmed by in vitro studies. Somatic alterations in the NER pathway, especially in ERCC genes, are common in various types of cancer. In a 2007 study, Castro et al showed that in cancer cells, NER and apoptosis pathways are the most impaired, with a high diversity of gene expression profiles in comparison to normal cells [8]. Other studies have shown how a deficiency in the NER pathway correlates with increased sensitivity to irofulven and cisplatin [9,10] and decreased sensitivity to trabectedin [11]. Moreover, inhibiting NER has been shown to increase sensitivity to alkylating agents in multiple myeloma cases [12]. DDR pathway alterations are present in numerous histologic subtypes of sarcoma. In a study conducted on STS specimens, at least one pathogenic mutation of the DDR pathway was detected in 15.9% of the patients, the most altered gene was ATRX (10%) and furthermore mutations were observed in 25 sarcoma subtypes. [13]. Recent studies have analyzed the genotype and expression profile of NER genes in STS patients, showing a correlation between ERCC1 and ERCC2 specific single nucleotide polymorphisms (SNPs) and a higher expression of both genes [14].

Nucleotide Excision Repair (NER) Pathway

NER mechanism recognizes and repairs various types of DNA damage caused by UV irradiation, cisplatin, and other damaging agents. NER pathways can be classified as either global genome repair (GGR), which repairs DNA damage anywhere in the genome, or transcription-coupled repair (TCR), which specifically restores DNA strands that are being transcribed (Figure 1). NER mechanisms rely on a series of reactions: recognition of DNA damage, unwinding double-strand DNA in the neighborhood of the damage, excision of the damaged nucleotides, and filling of the single-stranded gap by DNA synthesis. In GGR, DNA damage is recognized by XPC/Rad23 (xeroderma pigmentosum, C/Rad23 complementation group) or UV-DDB (UV-damaged DNA binding protein) [15] while in TC-NER, DNA damage blocks RNA polymerase II (RNAPII) interacting with CSB (ERCC excision repair 6, chromatin remodeling factor) and CSA (ERCC excision repair 8, subunit of CSA ubiquitin ligase complex)-CSB. After damage recognition, in both pathways, RNA polymerase II H transcription initiation factor (TFIIH) is recruited. It is subsequently recruited to the XPG (ERCC excision repair 5, endonuclease) complex, a single-stranded DNA-specific endonuclease. TFIIH unwinds DNA in the vicinity of damage, XPD (ERCC excision repair 2, helicase subunit of the TFIIH core complex), XPB (ERCC excision repair 3, helicase subunit of the TFIIH core complex) and XPA (DNA damage recognition and repair factor) are in charge of recognizing and verifying the damage. XPA binds to the chemically altered nucleotides in a single strand of DNA and recruits the XPF (ERCC excision repair 4, endonuclease catalytic subunit)-ERCC1 (ERCC excision repair 1, endonuclease non-catalytic subunit) catalytic subunit, which makes a cut on the damaged strand of 5′ to extract the damage. Next, XPG makes a 3′ cut that leads to the excision of a single-stranded DNA fragment containing the damage. Then thanks to PCNA (proliferating cell nuclear antigen) and DNA polymerase δ or ε new DNA is synthesized, finally DNA ligase 1 or 3 seals the DNA [16]. Defects in the NER pathway can be attributed to several inherited human diseases, including xeroderma pigmentosum (XP), an autosomal recessive genetic disease characterized by increased sensitivity to UV radiation [17] (Figure 1).

fig 1

Figure 1: NER proficient and NER deficient tumoral cells. Effects of variations in the NER pathway leading to a NER proficient condition (left) with correct repair of DNA damage or NER deficient (right) with DNA damage persisting. The NER deficient condition can be reversible, resulting in DNA repair, or irreversible, resulting in permanent DNA damage that can lead to cellular damage.

ERCC1

The product of this gene is required for the repair of DNA lesions such as those induced by UV light or formed by electrophilic compounds including cisplatin. The encoded protein forms a heterodimer with the XPF endonuclease, and the heterodimeric endonuclease catalyzes the 5′ incision in the process of excising the DNA lesion. The heterodimeric endonuclease is also involved in HRR and in the repair of inter-strand crosslinks [18]. Overexpression of ERCC1 is correlated with better progression-free survival (PFS) in patients treated with doxorubicin plus trabectedin [19] and favorable overall survival (OS) [19]. Additionally, high expression of ERCC1, and BRCA1 haplotype were associated with the improved progression-free rate (PFR), PFS, and OS in STS [20]. Increased ERCC1 and XPF expression were associated with improved disease-free survival (DFS) and distant disease-free survival (DDFS) in STS [21]. A study carried out on STS patients showed that regarding the SNP rs11615, the alternative allele has a higher germline frequency than the general population and ERCC1 is overexpressed in 75% of STS samples analyzed compared to healthy corresponding tissue and its expression varies according to the genotype [14].

ERCC2 (XPD)

ERCC2 is part of the BTF2/TFIIH complex, which is essential in TCR. The translated protein has ATP-dependent DNA helicase activity and belongs to the RAD3/XPD subfamily of helicases. Defects in this gene are related to Cockayne syndrome, XP cancer-prone complementation group D syndrome, and trichothiodystrophy. [22]. ERCC2 gene is overexpressed in STS and its expression varies according to the genotyping of rs13181 and rs1799793, in addition, these SNPs have a higher frequency than the general population [14]. Additionally, ERCC2 is mutated in 3% of epithelioid sarcoma and 6.5% in perivascular epithelioid cell tumors and is significantly associated with increased homologous recombination deficiency (HRD) scores [13].

ERCC3 (XPB)

This gene encodes an ATP-dependent DNA helicase that is a subunit of basal transcription factor 2 (TFIIH) and, therefore, also functions in class II transcription. Mutations in this gene are associated with XP B, Cockayne’s syndrome, and trichothiodystrophy [23] ERCC3 overexpression is associated with disease progression in STS patients treated with trabectedin [24]. A study analyzing the genetic background, by whole-exome analysis, of a family with a 4-year-old child who has a Li-Fraumeni tumor (often associated with STS) hypothesized that ERCC3 may be a potential TP53-related modifier candidate responsible for accelerated tumor onset by the proband compared with the mother, who carries the same TP53 mutation [25].

ERCC4 (XPF)

XPF forms a complex with ERCC1 by playing a role in the 5′ incision made during NER. This complex is a DNA repair-specific endonuclease that interacts with meiotic structure-specific essential endonuclease 1 (EME1). Variations in this gene can underlie xeroderma pigmentosum complementation group F (XP-F), or xeroderma pigmentosum VI (XP6) [26]. In a study that performed targeted genomic sequencing within an Asian cohort of sarcoma patients, a truncating mutation in ERCC4 (p.Cys723*) was found in two patients with sarcoma diagnosed under 25 years of age [27]. In a retrospective study on angiosarcoma, ERCC4 was found mutated in 6% of patients [28].

ERCC5 (XPG)

This gene encodes a single-stranded DNA-specific endonuclease that makes the 3′ incision in DNA excision repair. XPG also plays a role in RNAPII transcription. Variations in this gene can cause xeroderma pigmentosum complementation group G (XP-G) and Cockayne syndrome [29]. A study of 113 STS samples showed a correlation between high expression of the common allele (aspartic acid at codon 1104) and better PFR, PFS, and OS [20]. A translational study showed that an overexpression of ERCC5 correlates with trabectedin activity and is associated with longer PFS in advanced STS treated with trabectidine [30]. Furthermore, in a cohort of STS, the frequency of SNP rs1047768 is the same as that of the general population, while the frequency of SNP rs2296147 is lower than that of the general population; the gene is overexpressed in 42% of the STSs analyzed and its expression correlates with that of the ERCC2 gene. Finally, the effect of SNP rs1047768 in protein structure was hypothesized for the first time in this study, suggesting a possible effect in ssDNA binding [14]. A meta-analysis associated variations on the ERCC5 gene with an increased risk of STS [31].

ERCC6 (CSB)

The encoded protein has ATP-stimulated ATPase activity, interacts with several transcription and excision repair proteins, and may promote complex formation at DNA repair sites. CSB interacts with RNAPII at the damaged site, and by direct interaction it recruits CSA [32], forming a complex responsible for the association and stabilization of UV-stimulated scaffold protein A (UVSSA), which stimulates TC-NER [33]. Mutations in this gene are associated with Cockayne syndrome type B and cerebro-oculo-facio-skeletal syndrome (COFS) [34]. On dbSNP are reported 68 clinically significant pathogenic variants of ERCC6 [35]. A retrospective translational study on STS showed that ERCC6 was underexpressed in L-sarcomas, compared with other STS subtypes [24].

ERCC8 (CSA)

CSA, encoded by ERCC8 (chr 10), is part of an E3-ubiquitin-ligase complex. CSA, in TC-NER, is required for recovery of DNA synthesis after repair is responsible for ubiquitination and proteasomal degradation of CSB, is required for recovery of DNA synthesis after repair [36] and interacts with CSB and with p44, a subunit of TFIIH. Mutations in this gene have been identified in patients with the hereditary disease of Cockayne syndrome (CS). [34], however genetic polymorphisms are shown to increase breast [37], gastric [38] and oral [39] cancer risk. On dbSNP are reported 32 clinically significant pathogenic variants of ERCC8 [35]. There are no data at present on the correlation of ERCC8 and STS.

XPA

The XPA protein is a zinc finger protein that plays a central role in NER by interacting with DNA and other proteins, forming the structure required to assemble the NER etching complex [40]. A retrospective translational study on STS showed that high levels of XPA expression correlated with better efficacy of trabectedin. [24].

XPC

It is a key component of the XPC complex, which plays an important role in the early stages of GG-NER. It has higher affinity for single-stranded DNA, and is important for damage detection [41]. At present, there are no experimental data on the role of XPC in STS.

TFIIH

TFIIH is a 10-subunit protein complex involved in both transcription and DNA repair, highly conserved in the entire eukaryotic domain. It can be divided in a 7-subunit CORE complex, consisting of XPB, XPD, p62, p44, p34, p52 and p8, and a CAK module (Cyclin Activated Kinase), comprised of CDK7, cyclin H and MAT1 [42]. XPB and XPD are both ATP-dependent DNA helicase and they catalyze the ATP-dependent opening of the DNA at the transcription starting site or at the damaged site. XPB, encoded by ERCC3 in chromosome 2, unwinds the DNA helix in the 3′-5′ direction and can also function as a 5′-3′ DNA traslocase [43], while XPD, encoded by ERCC2 in chromosome 19, acts in 5′-3′ direction and it’s responsible for recruiting the CAK complex [44]. The remaining 5 subunits (p62, p44, p34, p52 and p8) are encoded respectively by GTF2H1 in chromosome 11, GTF2H2 in chromosome 5, GTF2H3 in chromosome 12, GTF2H4 and GTF2H5 in chromosome 6; they carry out structural and ATPase regulation roles [45]. The three-subunit detachable CAK module has a fundamental role as a regulator of both transcription and damage repair pathways; in particular it is necessary for transcriptional activation, but its presence inhibits damage-repair functions [46]. Of the seven genes encoding the CORE components of TFIIH, mutations in ERCC3 and ERCC2 affect both RNA transcription and DNA repair pathway, causing severe disorders such as Xeroderma Pigmentosum, Cockayne Syndrome and Trichothiodystrophy [47]. NCBI dbSNP reports 14 clinically significant pathogenic or likely-pathogenic variants of ERCC3 [35]; in addition, various ERCC3 polymorphisms have been linked to increased risk of lung cancer [48] and osteosarcoma [49]. Regarding ERCC2, dbSNP reports 41 pathogenic or likely-pathogenic variants and polymorphisms in this gene have been associated with a higher risk of lung [50] and colorectal cancer [51]. Furthermore, a significant link has been reported between specific ERCC2 and ERCC3 SNPs and their predisposition to specific types of sarcomas [52]. The other 5 genes of the CORE complex are less affected by clinically significant polymorphisms, although it has been reported that mutations in p52, p8, and p44 are associated with developmental disorders [45]. Regarding the CAK module, high expression of cyclin H has been associated with trabectedin sensitivity in STS [24].

Ubiquitylation in NER Pathway

Ubiquitin is a 76-amino acid protein used for labeling targeted proteins, regulating their stability and function. Ubiquitylation is a sequential process that involves the action of E1 ubiquitin activating enzyme, E2 ubiquitin-conjugating enzyme and E3 ubiquitin ligating enzyme [53]. There are 2 E1, 40 E2 and about 600 E3 enzymes in the human genome [54], which emphasizes the specificity of the ligation process [55]. The initial ubiquitin is attached to the target protein in a lysine (K) residue in the C-terminal portion of the target. Subsequent ubiquitin molecules are sequentially attached to lysine residues of the previous molecule. Poly-ubiquitin chains can have different functions depending on the lysine residue to which the molecules link. The K48-linked chains signal proteasomal degradation of the target protein, whereas K63-linked chains regulate target protein function [56]. Ubiquitylation is reversible by the action of deubiquitinating enzymes (DUBs) [57]. Complex processes such as NER require similarly complex regulation through easily inducible and reversible post-translational modifications. Ubiquitylation has been shown to play a key role in this pathway [58].

GG-NER Regulation by Ubiquitylation

In the presence of UV damage to DNA, the COP9 signalosome is released from the CRL4DDB2 complex, an E3 ubiquitin ligase comprising CUL4, ROC1, and DDB2. In normal conditions the COP9 signalosome inhibits the CRL4DDB2 complex activity, in the absence of this inhibition CUL4 can be neddylated by NEDD8, leading to the activation of the E3 complex [59]. At this stage, recognition of DNA damage by XPC and DDB2 occurs [60]. The CRL4DDB2 complex carries out the action of E3 ubiquitin-ligase on histones H2A, H2B, H3, and H4, weakening the histones-DNA interactions in the damaged area [61]. The complex auto-ubiquitinates DDB2, decreasing its affinity to damaged DNA [62]. This process competes with the presence of XPC at the damaged site, which stabilizes DDB2 [63], and with PARylation of DDB2 by PARP1, which inhibits its ubiquitination [64]. Deubiquitinase BAP1 also appears to be involved in this regulation process [65]. DDB2 is extracted from the complex by VCP/p97 and targeted to the proteasome. The removal of DDB2 from DNA increases the binding affinity of XPC to TFIIH, which is recruited at the damaged site. TFIIH promotes, through its p62 subunit, DDB2 extraction from the complex. XPC-TFIIH-XPA complex formation allows the initiation of the DNA damage verification process [66]. Simultaneously with DDB2 ubiquitylation, the CRL4DDB2 complex also ubiquitinates XPC, not resulting in degradative signaling, but increasing its affinity to DNA [67]. XPC then undergoes SUMOylation, induced by UV damage to DNA, which results in the recruitment of RNF111/Arkadia (SUMO-targeted ubiquitin ligase), responsible for XPC ubiquitylation that leads to its removal from damaged DNA [68]. Extraction of XPC by VCP/p97 allows the other factors of GG-NER to be recruited.

TC-NER Regulation by Ubiquitylation

In the presence of DNA damage, RNA-Pol II is interrupted, recruiting CSA and CSB, which in turn recruit UVSSA. CSA is part of CRL4CSA E3 ubiquitin-ligase complex, comprising CUL4, ROC1 and CSA. As in the GG-NER regulation, this complex is regulated by the COP9 signalosome, which detaches in the presence of DNA damage and allows the neddylation of CUL4, thereby activating the E3 complex [59]. CSB undergoes modification by multiple factors, as it is ubiquitylated by CRL4CSA complex and BRCA1-BARD [69] and deubiquitylated by the deubiquitylating enzyme USP7 [70], which is recruited by UVSSA [71]. This fine-tuned regulation controls the stability of CSB before its extraction by VCP/p97, allowing the recruitment of the other damage repair factors. CRL4CSA complex also mono-ubiquitylates UVSSA, allowing recruitment of TFIIH, which is then linked to RBP1 [60]. If the damage is not repaired, as in the case of mutations in CSA or CSB, the RBP1 subunit is ubiquitylated by NEDD4 [72] and the Elongin A ubiquitin ligase complex [73], inducing its extraction by VCP/p97 and subsequent proteasome degradation.

Role of Chromatin in NER Pathway

Activation of the NER pathway requires DNA not wrapped around histones, as various proteins need to access the double helix. UV damage to DNA provides the signal that leads to post-translational modifications of histones or ATP-dependent chromatin remodeling. These mechanisms allow the relaxation of chromatin around histones and increase the efficiency of the NER pathway [74,75].

Histone Modifications

Acetylation is mediated by histone acetyltransferases (HATs) and the reverse process, deacetylation, is catalyzed by deacetylases (HDACs). Histone acetylation promotes chromatin relaxation and activation, promoting DNA transcription [76]. Histone acetylation stimulates NER after UV damage [65]. Recent studies report that DDB2 interacts with HBO1 (HAT) in a UV-dependent manner leading to acetylation of H3 and H4, which increases chromatin accessibility [77]. DDB2 appears to be responsible for deacetylation of H3 and H4 through HDACs, leading to the stimulation of XPC recruitment [78]. Methylation acts differently on chromatin state depending on the number of methyl groups and the residues on which they are added [79]. Recent work has shown that DDB2 recruits the methyltransferase ASH1L to damaged DNA regions, leading to tri-methylation of histone H3K4. This process induces XPC binding to nucleosomes [80]. UV irradiation also stimulates tri-methylation of H3K79 by DOT1L, which also correlates with XPC recruitment. Deletion of DOT1L is present in many cases of melanoma [81]. Finally, another histone modification is phosphorylation which leads to a more relaxed state of chromatin and serves as a checkpoint in several processes, including DNA repair [82]. Phosphorylation of specific histones does not appear to affect the NER pathway; rather, it is the NER pathway that induces phosphorylation of histone H2AX via single-strand DNA production [83].

ATP-dependent Chromatin Remodeling

Chromatin remodelers are enzymes with an ATPase domain, which use ATP energy to modify the structure of nucleosomes. They are divided into 4 families: SWI/SNF, CHD, INO80, ISWI [84]. During NER, the SWI/SNF complex catalyzes the relaxation of chromatin, making it more accessible. CSB, which plays a key role in the NER pathway, belongs to this group [85]. An additional role of SWI/SNF is its association with XPC, which promotes the recruitment of subsequent repair factors [86]. The remodeler INO80 interacts at the level of damaged sites with DDB1, suggesting a role in XPC recruitment [87]. CHD is recruited following UV damage and mediates XPC binding to TFIIH [88].

Histone Chaperones

Proteins are involved in the transport and mobilization of histones at the chromatin level [89]. CAF-1 and HIRA are associated with NER, as they are recruited in the late stages of the pathway and are involved in the deposition of neo-synthesis histones after damage repair [90,91].

Conclusions

DDR pathways, including NER, play a key role in the formation of various tumor types and their sensitivity or resistance to treatment. The NER pathway is composed of a variety of processes that are finely regulated by each other and integrated with many other cellular pathways and alterations in this pathway play an important role in many tumor types, including STS. Recent studies contribute to the notion that NER pathway deficiencies constitute potential cancer therapeutic targets. It has been found that putative damaging germline and somatic alterations in NER genes were present in STS [14]. Moreover, recent findings provide novel insights into a synthetic lethal relationship between clinically observed NER gene deficiencies and sensitivity to irofulven and its potential synergistic combination with other drugs [9]. There is still little evidence on the association between cancer risk and variations in NER genes; in fact, there are no FDA-approved targeted therapies that target germline or somatic mutations in NER pathway genes. However, mutations in NER genes can have multiple roles as biomarkers: they can act as predictive biomarkers, indicating an increased risk of developing cancer, being useful for early cancer detection by subjecting the population to higher levels of screening; at the same time, they can also be used as prognostic biomarkers, giving precise indications to physicians about the degree of sensitivity to drugs targeting DNA repair deficiencies, if variations in NER genes are present. Recent studies focus instead on the therapeutic role of NER inhibitors, such as spironolactone [92] well as triptolide, which inhibits NER by affecting XPB and transcription. NER inhibition has been shown to reverse acquired resistance to alkylating agents in multiple myeloma cells [12]. Hence, it may be another adjunct target to be considered in combination therapies. Their further investigation in these tumor types is necessary for the identification of new biomarkers or therapeutic targets (Figures 1 and 2).

fig 2

Figure 2: Nucleotide excision repair pathway (NER). NER pathways can be classified as either global genome repair (GGR) and transcription-coupled repair (TCR). The NER pathway consists of a series of reactions: recognition of DNA damage, unwinding double-stranded DNA in the neighborhood of the damage, excision of the damaged nucleotides, and filling the gap by DNA synthesis and ligation.

Author Contributions

Conceptualization, S.P.; writing—original draft preparation, F.S., A.P., A.B., E.C, S.F., O.C., E.G. and S.P; writing—review and editing, I.P, A.B. and G.S.; supervision, A.L. and S.P.; funding acquisition, I.P., D.A.C, S.P.; figures preparation: S.F. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by Regione Toscana Bando Salute 2018, (Research project CUP n. D78D20000870002), grant number D78D20000870002.

Acknowledgments

Our special memory goes to Alessio Cerasola, an unforgettable guy who fights the disease with his ever-present smile.

Conflicts of Interest

The authors declare no conflict of interest

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Alpha-Fetoprotein-derived Segments as Integrin Peptidomimetics for Potential Cancer Cell Targeting and Therapy: A Review and Commentary

DOI: 10.31038/CST.2023823

Abstract

Integrins constitute a group of dimeric polypeptide chains that function as natural agonists of cell surface receptor-dependent cell activities. The integrins themselves comprise a superfamily of hetero-dimeric (alpha and beta chains) transmembrane cell surface receptors whose functions include cell adhesion, growth, migration, and angiogenesis. In comparison, the integrin-like peptides (ILP) comprise groups of protein derived segments, namely, short peptides derived from naturally occurring proteins from intrinsic subdomain fragments or short motifs present on larger proteins or enzymes. Certain ILPs can bind or compete for amino acid sequence sites located on integrin beta-1 and beta-3 chains of heterocomplex receptors. Binding at major sites or allosteric minor sites can inhibit or block cell migration, angiogenesis, metastasis, and platelet aggregation. Recently, a small integrin-like peptide derived from naturally occurring alpha-fetoprotein (AFP), similar to a disintegrin, has been reported to inhibit growth and adhesion functions associated with integrin-dependent cell activities. The present report describes an example of an AFP integrin-like peptide and lends credence to support its proposed use in adjunct cancer therapies.

Keywords

Alpha-fetoprotein, Integrins, Cell adhesion, Migration, Cell-to-cell contact, Breast cancer

Introduction

A) General

Integrins comprise a superfamily of hetero-dimeric (alpha and beta chain) transmembrane receptors present on multiple cell types including tumor cells [1]. The numerous functions that integrins mediate include cell-to-cell and cell-to-extracellular matrix (ECM) adhesion, cell growth, migration and spreading, metastases, angiogenesis, cytoskeletal-induced locomotion, and platelet aggregation [2,3]. Naturally occurring antagonists of the integrin receptor are termed disintegrins (DTs) which block or inhibit integrin cell functions [4,5]. While the integrins comprise 23 or more different alpha and beta chain combinations, the DTs constitute families of only two types of molecules [6,7]. Integrin-like peptides function in a similar manner to the disintegrins which are derived from metalloproteinases. It has been demonstrated in previous publications, that small peptides, derived from naturally occurring serum-related proteins, can mimic portions of the integrin polypeptide chains [8,9]. Such an action could interfere, compete, interrupt, or block signal transduction in the integrin receptors. Thus, small integrin-like peptides (ILP) are being proposed that can inhibit or compete with adhesion functions associated with metastasis, cell migration, cell-to-cell contact, and cell spreading. Since integrins show promise as potential molecular targets for cancer, the integrin-like peptides could possibly serve as formidable anticancer therapeutic agents for cell migration and metastatic targets.

B) Objectives and Aims

The objectives in the present report comprise several in number. First, the integrin biologic functions and activities are described. Second, the types and family members of the heterodimeric cell-adhesion integrin molecules are discussed on an overview fashion. Third, naturally occurring peptides derived from “mother” proteins will be addressed as integrin-like peptidomimetics. Finally, a prime example of an integrin-like non-toxic peptide mimetic is described which displays activities such as inhibition of platelet aggregation, suppression of cell-to-matrix adhesion, cell migration/spreading, and cell-to-cell contact activities.

C) The Integrin Cell Surface Receptors

The integrin superfamily of cell surface receptors consists of hetero-dimeric (alpha and beta chains) transmembrane glycoproteins that mediate cell-to-extracellular matrix (ECM), adhesion, and cell-to-cell contact interactions [10,11]. The integrins are integral cell surface single pass-transmembrane receptors consisting of two paired chains of non-covalently linked alpha and beta polypeptide chains. Both integrins and the ECM molecules play important roles in ontogenetic development, maintenance of adult cell physiology, tissue repair, hyperplastic growth, hemostasis, and tumor oncogenesis [12-14]. The dimeric hetero-complexed integrins further serve as cell membrane receptors capable of forming focal adhesion contact linkages to the cytoskeleton; such links are located on the inner layer of cell membranes. Integrins can further bind to multiple ECM ligand proteins such as: fibronectin, laminin, vitronectin, collagen, thrombospondin, entactin, fibrinogen, talin, the intracellular adhesion molecule (ICAM), and the vascular cell adhesion molecular (VCAM) [15-17]. Studies have further linked integrin signaling to cytoplasmic cytoskeletal filament-associated proteins such as vinculin, talin, α-actinin, paxillin, and divalent cation-dependent proteins such as calreticulin [18,19]. The integrins further play a major role in cell adhesion activities in the immune system [20-22].

Each integrin subfamily is characterized by a combination of a small number of β-chains associated with a large number of α-chains. To date, eight different β-chains and 14 different α-chains have been described, accounting for at least 20 combinatorial variations of the two heterodimeric receptors [9]. Both the α- and β- subunits are integral membrane glycoproteins containing variable long-lengths of extracellular domain chains linked to short intracellular chains [10]. The α-chains exhibit four repeat amino acid segments which bind calcium (Ca++) and other divalent cations such as Mg++ and Mn++ [18,19]. The β-subunits display at least four cysteine-rich repeats in linear juxtaposition; these repeats stabilize the chains of the extracellular amino terminal loops [9,20]. In overview, both chains contribute to the formation of an interface which forms the ligand binding pocket. In contrast to their extracellular domains, the intracellular domains of both the α- and β- chain constitute short amino acid segments capable of binding to cytoskeletal-associated proteins that can link the integrins to G-proteins, actin, and calreticulin, a Ca++ influx regulator involved in cell migration [12,13].

D) The Integrins-ECM Interaction and Signal Transduction

Studies of ECM interaction with cells via their integrin receptors have shown that integrins function as bidirectional transducers of extra- and intracellular signals. The two-way (bidirectional) signaling can occur from “outside-to-inside” and from “inside-to-outside” the cell [21,22]. The regulation of cell proliferation, differentiation, survival, and immediate gene expression is influenced by integrin mediation of cell interaction associated with the ECM. The disruption of epithelial and endothelial cell interactions with the ECM can induce programmed cell death, while fibroblast-integrin adhesion can affect cell cycle activities by influencing cyclin-A and D expressions [23,24]. In addition to signal transduction with the actin cytoskeleton, the cytoplasmic domains of the integrins interact in a cascade fashion with protein kinases, calcium-binding proteins, focal adhesion kinases, Na+/H+ antiporters, tyrosine MAP kinases, and transcription nuclear factors such as NFkB and AP1 [25-27].

The integrins must be activated in order to undergo adhesion and binding to the ECM. Activation of integrins occurs through local soluble mediators such as hormones, cytokines, growth factors, or by interfaces with the ECM. Thus, cell activation is known to involve adhesion to clusters of stimulated integrins which culminate in signals triggered by local events in the cellular environment, such as thrombogenic agonists, antigen stimulation/processing, and T-cell activities [20,28,29]. In contrast, integrin activation can be blocked by ILP and/or disintegrins to disallow cell adhesion and ECM binding at inopportune times and locations. Untimely adhesions can lead to unwanted thrombosis and inflammation, while already adhered cells may need to detach in order to undergo mitosis and cell migration [21,27]. As previously reported, both the disintegrins and ILPs can effectively contribute to blocking, inhibiting, reducing, and dysregulation of integrin function.

Protein-Encrypted Peptides; Growth Inhibitory Peptide (GIP)

The inclusion of a class of growth regulatory factors, extracellular ligands, and angiogenic peptide fragments encrypted within a polypeptide chain of a full-length protein is known but is not widely recognized [30]. However, some of the most potent growth inhibitors are derived from short peptide fragments (segments) already existent in naturally occurring mammalian full length proteins. Such intrinsic segments themselves can affect cell growth and proliferation in an opposite function from that of the “mother” protein [31,32]. This less recognized concept of a protein-derived body reserve containing peptide growth inhibitor fragments is becoming a recurring theme in the field of growth regulation, intracellular signaling, and crosstalk among and between signal transduction pathways. Classical examples of such occult (cryptic) peptides derived from proteins include the following examples,

  1. Tenacin binding peptide derived from fibronectin;
  2. Angiostatin from plasmin;
  3. Endostatin from type XVIII collagen;
  4. Vasostatin from calreticulin; and
  5. Constatin from type IV collagen.

Such cryptic hidden peptide sites can be exposed following a conformational change on a protein or can be revealed following proteolytic cleavage from a larger protein [33,34]. Such peptides can also be chemically synthesized as single fragments of 20-45 amino acids. A well-published example of a peptide site revealed following a conformational transition change on a full-length protein is an encrypted “growth inhibitory” site on the alpha-fetoprotein (AFP) molecule [35]. The AFP protein is normally a growth promoting molecule, but can be temporarily converted to a growth inhibitory molecule.

The encrypted peptide segment on AFP, termed the growth inhibitory peptide (GIP), is a 34 amino acid segment concealed in a hydrophobic cleft of the tertiary folded AFP molecule. The GIP-34 site is revealed following protein unfolding in chemical environments consisting of high ligand concentrations of estrogens, fatty acids, and growth factors [32,35]. The exposed transitory GIP site converts the usually growth-enhancing AFP molecule into a temporary growth-inhibiting molecule. This conversion occurs via protein unfolding via a conformational change resulting in a denatured intermediate state that reflects a molten globular form (MGF) of the AFP protein [36]. Since the MGF of AFP is a transitory intermediate form, AFP can refold back to its native tertiary fold following removal of excess ligands (agents) in the microenvironment [37]. Because the AFP-MGF form is unstable, the GIP-34 amino acid segment alone has been synthesized, purified, and characterized as a free and distinct 34-mer synthetic peptide segment [33-35]. Thus, 34-mer GIP fragment can inhibit growth factor, fatty acid, and estrogen-induced growth in a concentration-dependent manner in addition to blocking metastatic and cell migration-associated activities.

GIP-34 Physicochemical Properties

GIP-34 has been synthesized by classical F-MOC (9-fluronylemethoxy-carbonxyl)- protected solid phase synthesis, as previously described [38]. Following peptide syntheses, the lyophilized peptide was purified by reverse-phase high-performance liquid chromatography (HPLC), producing a peptide whose major peak displayed a molecular mass of 3573 (34-mer) as determined by electrospray ionization mass spectroscopy. Cyclization of GIP-34-mer can be accomplished by addition of reducing agents to form a disulfide bridge construct at the time of the linear peptide synthesis. Circular dichroism (CD) analyzed in the UV wavelength for GIP-34 displayed a negative maximum at approximately 201nm. Computer modeling and analysis of the GIP-34 CD spectrum revealed a secondary structure comprising 45% β-sheets and turns, 45% random coli (disordered), and 10% α-helix structure [35].

Amino Acid Sequence Matches

The GIP-34 AA sequence was subjected to a FASTA search in the Genbank (GCG Wisconsin Program) database, as described [32,33,35]. The GCG search found identity/similarity sequence matches to receptor-binding proteins, such as the fibroblast growth factor (FGF) receptor, insulin growth factor II receptor (IGFIIR), transforming growth factor-β (TGF-β), and the dopamine (DOPA) receptor [39]. Other Genbank matches revealed transcription-associated proteins, including homeodomain proteins and FTZ-F1 (the AFP transcription factor), which have been previously reported [40-42]. These AA matches provide evidence that the GIP fragments contain short recognition cassettes for multiple and varied receptor involvement and interactions. Matches with cell-adhesion related proteins were also found; these included collagen XIII, collagen IV, laminin, fibrinogen, and fibronectin [41,42], (Table 1). Finally, identities/similarities were identified with transcription-associated factors, such as Hox, c-myc, forkhead, and Pax. GIP-34 matches were further found with integrin-associated proteins, the ECM proteins, cell mitosis proteins, and other adhesion proteins (Tables 1 and 3). Further identities were found with the integrin α/β chain proteins such as α11bβ3, α1β3, and αvβ1. Such integrins can serve as receptors for ECM proteins and are known to participate in cell-to-cell activities such as cell adhesion and migration (spreading) activities. Finally, matches were also made with ECM-associated proteins, such as the Von-Willebrand Factor, VLA-1, and PG-IIIa proteins, which are involved in cell adhesion, aggregation, and the action of metalloproteinases (i.e., the Adams Family) (Table 3). Thus, GIP-34 shows an identity/similarity matches to integrins, basement membrane proteins, and ECM proteins, all of which are involved in cell-to-cell and cell-to-ECM interactions. A comparison of the properties and traits of integrins versus GIP are displayed on Table 2.

Table 1: Growth Inhibitory Peptide (GIP) Amino Acid Sequences * were matched in the Genbank to Various Integrin Alpha/Beta Chain Complexes and the Compared to their Extracellular Matrix (ECM) Adhesion Inhibition by GIP. Note that many of the Integrins are expressed on a variety of tumor cells.

Integrin Subunits

*GIP Amino Acid Sequence

AA Identity%

ECM Binding Ligand

Tumor to ECM Adhesion% Inhibition

Cell/Tissue and Tumor Distribution

α

β

αVβ3A LSEDKLLACGEGAAD,

SEDKLLACG

100(9)

47(15)

FIB, VTN, FBN, TSP

40-50

Melanomas and angiogenic cell
αMβ2 (Mac) SEDKLLACG,

LACGEGAADI

66.7(9)

43(10)

FBN, C3bi, I CAM

50

Immune, Inflammatory cells
αVβ6 SEDKLLA

100(7)

50(12)

FBN

50

Carcinoma cells virus associated fusion
α6β1 GEGAADIII

78(9)

75(8)

LAM-1

10-45

NSCL carcinoma
αVβ1 SEDKLLA-CGEG

100(7)

75(4)

VTN, FBN

40-50

Analytic tumors
α1β1 CGEGAADIIIGH

43(12)

75(8)

LAM COLL

10-45

Breast carcinoma
αLβ2 (LFA-1) CGEGAADIIIG

80(11)

43(10)

FBN, C3i

50

Myeloid cells, Leucocytes
α4β7 GEGAADIII

MTPVNPGV

78(9)

56(9)

FBN, VCAM MADCAM

50

Endothelial mucosal cells
α3β1 DKLLACGEGAADIIICGEG

43(14)

75(4)

FBN, COLL LAM

30-55

Many tumor cells
αVβ8 IRHEMTPVNPG

67(12)

50(12)

Not reported

not done

Reproductive tissues
αVβ5 CGEGAADIIIGHLCIRHEM-TPBNPGVGQ

67(12)

80(25)

VTN, FBN

45-50

Epithelium carcinoma cells
α6β4 IRHEMTPVPVNPGV

78(8)

50(12)

LAM-1, LAM-2

10-45

Keratinocyte malignancy
α2β1 IIGHLCIRHE

MTPVNPGV

53(17)

75(8)

COLL, LAM

10-55

Epithelium, endothelium leucocytes

Table 2: Comparison of properties shared by integrin-related components and the AFP-derived Growth Inhibitory Peptide (GIP).

Activity and/or Property

Integrin-related Properties

GIP Peptide Related Properties

Cell Toxicity Non-toxic Non-toxic (cytostatic)
Working Range Nanogram concentrations Nanogram concentrations
Platelet Physiology Activate platelets for aggregation Inhibits platelet aggregation
Cell Type Localization Most body cells, platelets, uterus, breast cancer cells Platelet, uterus, breast cancer cells
Ligand Binding Extra-cellular matrix proteins. (fibromectin, virtomectin, etc.) Extra-cellular matrix protein interaction
Protein Homology C3b complement & C2 component, Factor B Von Willebrand factor, Mac-1 Von Willebrand factor, fibronectin precursor
Aggregation Form dimers, receptor aggregation (clustering) Forms dimers, trimers & oligomers
Adhesion Cell-to-cell, cell-to-ECM Cell-to-cell &cell-to ECM
Cellular Internalization Soluble ligand/integrin internalization Apparent cellular internalization
Secondary Structure Beta sheets & turns in extracellular subunits Mainly beta sheets & turns in soluble peptide
Distinctive Amino Acid Presence Cysteine relative to aspartic acid spacing Display 2 cysteine with aspartic acid spacing
Ligand Binding Region N-terminal half of α and β subunits Short sequence homologies to α chain component
Cellular Localization Cell surface transmembrane peptides extending into cytoplasm Fluorescence localization at cell surface and intercytoplasmic sites
Ligand Recognition Specificity  Controlled by the α subunit AFP-peptide more homologous to α chain subunit
Influence of Estradial Estradiol suppresses integrin ligand regulation of α2 subunit Peptide suppresses estrogen-sensitive growth
Integrin α (I-domain) Homology Similar to collagen binding domain of Von Willebrand Factor Similar to collagen binding domain of Von Willebrand Factor

Table 3: Integrin-associated Protein (IAP) amino acid sequences (left column) are matched to Growth Inhibitory Peptide (GIP) amino acid sequence stretches (middle column). Numbers to the left of the single letter amino acid code of GIP signify the amino acid number located on the full-length alpha-fetoprotein polypeptide.

I. Mitosis-associated Proteins

Protein (IAP) Name

Growth Inhibitory Peptide Amino Acid Sequences

Biological Activity or Function Affected by GIP

Contactin-associated Proteins 481 IGHLCIRH Cell adhesion
Neurotropic Tyrosine Kinase Receptor-3 461 CCQLSEDK Cell migration and invasion
Matrix metalloproteinase-13 497 ADIIIGHL

485 CIRHEMTP

Collagenases (ADAM-13)
ADAM-22, Integrin α2β1 481 IGHLCIRH Cell-to-Cell contact, cell migration, cell adhesion
Integrin α6 (IGAG) linked to Beta chain (VLA-6) 485 CIRHEMTP Cell-to-cell contact, cell migration, cell adhesion

II. Extracellular Matrix Proteins

Protein Name

Alpha-fetoprotein Growth Inhibitory Peptide Sequence Matches

Biological Analysis or Function Affected by GIP

Receptor for Peptin-54 (G-coupled receptor) 481 IGHCIRH G-coupled receptor for signal transduction
Fibroblast Growth Factor receptor-4 497 ADIIIGHL Regulates growth and proliferation, blood vessel angiogenesis
Ephrin Receptor 2B 481 IGHCIRH Regulates bidirectional signaling related to tumor growth/metastasis
Met Oncogene Hepatocyte Factor Receptor (C-Met) 481 IGHCIRH Tyrosine Kinase Receptor, axon guidance, cell segmentation, angiogeneis

III. Growth Factor Associated Proteins

Protein Name

Alpha-fetoprotein Growth Inhibitory Peptide Sequence Matches

Biological Activity or Function Affected by GIP

Vascular Endothelial Growth Factor 477 ADIIIGHL Stimulates vascular permeability
P53 Protein Cell Tumor Antigen 477 ADIIIGHL Prevents cancer growth, a tumor suppressor
Tyrosine Phosphate Non-Receptor-7 477 ADIIIGHL Tyrosine kinase related
Cell Growth Regulator 477 ADIIIGHL Enzyme that regulates cell growth/proliferation
NF-KB Signal Factor 477 ADIIIGHL Signal transduction factor regulating phosphorylation

Cell Adhesion Assays with the AFP-derived Peptide

AFP-derived GIP has been subjected to cell adhesion studies involving many of the ECM ligand proteins known in the literature and discussed herein [32,33]. Various ECM proteins were coated on microtiter plates to serve as solid attachment surfaces for two breast cancer cell types: the human MCF-7 and the murine mammary 6WI-1 cell culture lines (Table 1). The adhesion of MCF-1 and 6WI-1 tumor cells either in the presence of AFP peptide or in peptide-free medium were assayed on ECM-coated microtiter plates with soluble GIP used as a competitive inhibitor. GIP-34 was capable of inhibiting cell adhesion of the ECM ligand proteins in both tumor cell lines which spanned inhibition of 30-50%. Inhibition of the mouse and human tumor cell adhesion was roughly equivalent on microtiter plates coated with either collagen IV, fibrinogen, fibronectin, or thrombospondin and slightly less for laminin, collagen-I, and vitronectin in the two cell types. Human MCF-7 breast cancer cells, in the presence of GIP-34, further displayed substantial inhibition of vitronectin-induced adhesion, while mouse 6WI-1 cells demonstrated similar peptide inhibition of laminin coated adhesion [34,35]. Overall, the AFP peptide was found to competitively inhibit both MCF-7 and 6WI-1 cell-to-ligand attachments which ranged from 40-60%. Finally, it was found that rabbit anti-GIP antibodies could also block the ligand adhesion inhibition effects, similar to the GIP fragment itself.

Inhibition of Cell Migration Spreading and Metastasis by GIP

Cell adhesion receptors and their ligands (i.e., ECM proteins), provide the traction and stimulus for the migration and spreading of tumor cells [28,41,43] (Table 3). In general, most cells including tumor cells, use adhesion molecules to execute cell migration, which is termed cell spreading in cell culture. The integrins initiate migration of adherent cells such as fibroblasts, epithelial cells, and tumor cells upon the ECM surfaces. Cell migration requires multivalent binding of integrins to matrix bound ligands such as collagen, laminin, and fibronectin [15,16,27]. Analysis of coverslip cell migration assays revealed that the GIP inhibited more than 60% of the MCF-7 cancer cells’ spreading and migration on the surface of coverslips [34,35]. The MCF-7 cells that exhibited migration displayed distorted morphology such as star-shaped configurations, cytoplasmic spiking, surface spiny spheres, membrane ruffling, and extensions of cytoplasmic processes, all coupled with low cell viability. In cancer movements, it is noteworthy that cell migration and spreading constitute the initial steps in the metastatic process; furthermore, GIP has been reported to inhibit metastases in vivo in animal models [32,33,36].

Tumor Cell Adhesion to the Extracellular Matrix

Tumor cell adhesion to the ECM is an essential step in the tumor cell migration and metastases process, providing a means for migrating cancer cells to transiently attach to the connective tissue substratum while spreading [41]. A tumor cell adhesion ECM assay was utilized to assess whether the AFP derived GIP-34 could influence or modify tumor cell attachment to a protein substratum or matrix [38]. Various ECM proteins were absorbed to the walls of microtiter plates and screen for their ability to serve as a substratum for enhanced tumor cell adhesion, as compared to non-ECM protein-coated microtiter plates [32,33]. Using 6WI-1 mouse mammary tumor cells, substantial cell attachment was observed with vitronectin, laminin, fibrinogen, fibronectin, and collagens I and IV after 2.0 hours of incubation at 37°C. GIP-34 was then tested for its ability to compete with tumor cell adhesion to the ECM substratum. GIP-34 was capable of inhibiting many of the ECM proteins spanning from 40% to 60% [32,33,43,45].

Cross-talk signaling between the ECM and the tumor cell membrane is known to occur. Overall, GIP-34 was capable of inhibiting both the attachment of tumor cells to the substratum and the subsequent growth of remaining tumor cells on that particular ECM substratum. Based on the ECM adhesion data, tables of integrin-association inhibition with GIP fragments are presently presented, which exemplify integrin α- and β- chain to ECM interactions (Tables 1 and 3, and above references).

Additional Activities of Integrin-like Peptides (GIP)

It is germane to this discourse that additional insight and perspectives be addressed regarding the use of integrin-like peptides (ILPs) in cancer therapies. For example, short ILPs can be structurally altered and modified to produce more potent forms of such inhibitors. Recombinant and chimeric forms of ILPs and AFP subdomains have been synthesized for use in studies of integrin inhibition/competition of tumor growth, proliferation, adhesion, migration, and angiogenesis of cancers such as liver, breast, lung, melanoma, and others [32,33,46,47]. In addition, ILPs such as GIP have been reported to induce apoptosis in radio-sensitized cultured lymphocytes [33]. Moreover, it has been reported that ADAM-22, a disintegrin-like metalloproteinase, is an active participant in the development of breast cancer resistance during endocrine hormone therapy in women [48-50]. With regard to this report, GIP administered to cultured MCF-7 human breast cancer cells was shown to down-regulate the expression of ADAM-22 by 30-fold as determined by a global RNA microarray analysis [44]. These data would suggest that GIP treatment not only could down-regulate the expression of ADAM-22, but could also block the development of hormone-resistance in breast cancer. In a further study, GIP was reported to further suppress the growth of MCF-7 human breast cancer cells in vitro and in vivo [51].

Concluding Remarks

It now seems plausible that interference with integrin signaling could provide a rational basis for the development of aids in the therapeutic treatments for cancer growth, progression, and metastases. Anti-integrin antibodies, disintegrins and ILPs all predict promise in future anti-cancer therapy studies. Integrin interruption of the adhesive interaction of tumor-to-tumor cells and platelets to tumor cells should be capable of serving to arrest or impede cancer cell migration and metastasis [41,45]. The observations that different integrins are expressed on various tumor types and are differentially expressed during tumor transformation, progression, and metastasis suggest that integrins might also serve as prognostic biomarkers [10]. Integrin-like mimetic agents that block or interfere with the initial attachment of integrins to ECM components, can also blunt signal transduction events thus inhibiting proliferation, cell migration/invasion, and platelet aggregation. Such agents could constitute a formidable armamentarium of non-toxic anti-cancer agents. Such anti-adhesive agents might further find potential application in the treatment of the five major classes of human disorders, namely; neoplasia, inflammation, trauma, wound healing, and infection.

Since integrin dysfunction frequently results in cancer pathology, integrins represent an appealing array of targets for anti-tumor therapy. Because ILPs specifically bind or compete with integrins, they serve to interfere with and/or block functions such as cancer cell growth and proliferation, and the migration activities described herein. All such activities described above suggest that integrins might have the potential to serve as prominent candidates for molecular cancer targets and as such, make integrin-like peptides promising non-toxic therapeutic adjunct agents to treat cancers.

Acknowledgment

The author extends his thanks and gratitude to Ms. Sarah Andres for her commitment and time expenditure in the skilled typing and processing of the manuscript, references, tables of this report.

Abbreviations

Coll: Collagen; FBG: Fibrinogen; FIB: Fibrin; LAM: Laminin; TSP: Thrombospondin; VTN: Vitronectin; VWF: Von Willebrand Factor; *: Amino Acid Single Letter Code; C3i: Complement Factor-3 inhibited. Integrin data obtained from References 34 and 35.
ECM: Extracellular Matrix; C: Complement Protein; C3b: Complement Subunit; AFP: Alpha-Fetoprotein; GIP: Growth Inhibitory Peptide-34.

Disclosures

Financial

None; no U.S. federal grants were used in the preparation of this paper.

Interest

The author declares that there are no known conflicts of interest in the preparation of this manuscript.

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