Specificity of Molecular Responses to Erk1/2 and Mkk1/2 Inhibitors in Melanoma Cells

Skin cancer is the most commonly diagnosed cancer in the U.S. and 75% of skin cancer related deaths are due to malignant melanoma, a cancer originating in melanin producing melanocytes. The RAF/MKK/ERK signaling cascade is constitutively activated in over 90% of melanomas and 52% of tumors contain BRAF V600E/K oncogenic driver mutations. Although small molecule inhibitors specifically targeting mutant BRAF V600E/K and the downstream kinases MKK1/2 have been successful in clinical settings, resistance invariably develops. In preclinical studies, inhibitors of ERK1/2 can overcome resistance to BRAF V600E/K and MKK1/2 inhibitors, making them promising alternative pathway inhibitors for the treatment of melanoma. However, the specificity of molecular responses to ERK1/2 inhibitors remains unknown. In this thesis, I use SILAC-based phosphoproteomics to quantify molecular responses to the clinically available MKK1/2 inhibitor, trametinib, and the ERK1/2 inhibitors, SCH772984, GDC0994 and Vertex-11e in WM239a human metastatic melanoma cells. I observed significant responses in approximately 5% of all phosphosites identified. Significantly regulated phosphosites showed a high degree of overlap between all inhibitors, suggesting that the pathway functions linearly with relatively little evidence for branchpoints that lead to bifurcation upstream of ERK1/2. I also observe phosphosites responsive to only one of four MKK1/2 or ERK1/2 inhibitors. For example, trametinib shows an ability to block activating phosphorylation sites on p38α MAPK, which are not shared by ERK1/2 inhibitors SCH772984 and GDC0994, the MKK1/2 inhibitor selumetinib, or the BRAF V600E/K inhibitor, vemurafenib. Trametinib directly inhibits MKK6 in vitro, although with a IC50 10-fold higher than its inhibition of p38α MAPK in cells. This suggests the potential that the direct target of trametinib in cells is upstream of MKK6. Further analyses of phosphoproteomics responses to two MKK1/2 inhibitors and two ERK1/2 inhibitors identifies phosphorylation sites that can be classified as (i) known or novel targets of BRAF-MKK-ERK signaling, (ii) potential branchpoints at MKK1/2 upstream of ERK1/2, and (iii) off-targets of different inhibitors. My results show how information from phosphoproteomics comparisons of multiple MKK1/2 and ERK1/2 inhibitors can be combined to provide a deeper understanding of pathway specificity

Identification of research priorities in CHD:Empowering patients and families through participation in the development of formal research agendas

Background: Conquering CHD, formerly known as the Pediatric Congenital Heart Association (PCHA), is the leading congenital heart disease (CHD) patient advocacy organisation in the United States of America, and places high priority on patient engagement in the research process. Participatory design is an approach to problem-solving that utilises the knowledge and opinions of groups of people to generate plans and new ideas. Utilising this mode of patient engagement, patients and families engaged with Conquering CHD assisted in developing a list of research priorities which was then distributed to the larger membership with instructions to rank the priorities in order of importance. Upon completion, these items were compared to the current scientific literature to assess correlation with current publications. This cross-sectional study and literature review aimed to assess the priorities of patients and families in CHD research and to determine the reflection of these areas in the current body of scientific literature. Methods: This cross-sectional study utilised a survey asking participants to rank the importance of research items within categories including Technology Advances, Genetic and Cellular Research, Broad Understanding of CHD, and Psychosocial Outcomes which was distributed through social media and email to 43,168 accounts across all platforms. Respondents were asked to place each item in a ranked order in each category, with the value 1 representing the most preferred for each participant. Anyone engaged with Conquering CHD was eligible to complete the study, including patients and families. Subsequently, a literature review of the largest medical databases including PubMed, Scopus, and ScienceDirect was undertaken to determine the number of articles published per each topic which was then assessed to determine if there is a correlation between patient-ranked priorities and the current body of literature. Results: The study generated a total response of 527 participants. Regarding Technology Advances, valve replacement was the preferred topic (mean rank 2.07, IQR 2). Stem cell research was the favoured topic in Genetic and Cellular Research (mean rank 2.53, IQR 2). Access to care was the priority in the Broadening Understanding of CHD (mean rank 1.24, IQR 1). Pertaining to Psychosocial Outcomes, psychological/emotional effects was the highest ranked topic (mean rank 1.46, IQR 1). The literature review returned a total of 135,672 articles in the areas of interest. For Valve Replacement, 8361 articles resulted reflecting a proportion of 0.097 of total articles. For Stem Cell Research, 9921 articles resulted reflecting a proportion of 0.115 of total articles. For Access to Care, 7845 articles resulted reflecting a proportion of 0.091 of total articles. For Psychological/Emotional Effects, 6422 articles resulted reflecting a proportion of 0.074 of total articles. A Spearman's correlation demonstrated no correlation between the preferred domain of CHD research and the number of articles published for that domain (rs = 0.02, p = 0.94). Conclusions: This process demonstrates the effectiveness of participatory design, using a patient and family network to determine the research items of concern to those affected by CHD. The cross-sectional survey was effective in assessing patient and family priorities but was limited by access to reliable internet and delivery only in English. Though the study had a large response rate, it was limited to patients already engaged with Conquering CHD. For these reasons, it may not completely reflect the opinions of the total population affected by CHD. However, this offers valuable insight into patient-determined priorities and reveals that the current scientific literature does not correlate with these items. These data serve to inform individual and institutional research agendas to better reflect the needs and desires of this population.</p

Identification of a Serum-Induced Transcriptional Signature Associated With Type 1 Diabetes in the BioBreeding Rat

OBJECTIVE - Inflammatory mediators associated with type 1 diabetes are dilute and difficult to measure in the periphery, necessitating development of more sensitive and informative biomarkers for studying diabetogenic mechanisms, assessing preonset risk, and monitoring therapeutic interventions. RESEARCH DESIGN AND METHODS - We previously utilized a novel bioassay in which human type 1 diabetes sera were used to induce a disease-specific transcriptional signature in unrelated, healthy peripheral blood mononuclear cells (PBMCs). Here, we apply this strategy to investigate the inflammatory state associated with type 1 diabetes in biobreeding (BB) rats. RESULTS - Consistent with their common susceptibility, sera of both spontaneously diabetic BB DRlyp/lyp and diabetes inducible BB DR+/+ rats induced transcription of cytokines, immune receptors, and signaling molecules in PBMCs of healthy donor rats compared with control sera. Like the human type 1 diabetes signature, the DRlyp/lyp signature, which is associated with progression to diabetes, was differentiated from that of the DR+/+ by induction of many interleukin (IL)-1-regulated genes. Supplementing cultures with an IL-1 receptor antagonist (IL-1Ra) modulated the DRlyp/lyp signature (P < 10-6), while administration of IL-1Ra to DRlyp/lyp rats delayed onset (P = 0.007), and sera of treated animals did not induce the characteristic signature. Consistent with the presence of immunoregulatory cells in DR+/+ rats was induction of a signature possessing negative regulators of transcription and inflammation. CONCLUSIONS - Paralleling our human studies, serum signatures in BB rats reflect processes associated with progression to type 1 diabetes. Furthermore, these studies support the potential utility of this approach to detect changes in the inflammatory state during therapeutic intervention

Integrating Teaching and Research in Undergraduate Biology Laboratory Education

A course recently designed and implemented at Stanford University applies practical suggestions for creating research-based undergraduate courses that benefit both teaching and research

Advocacy at the Eighth World Congress of Pediatric Cardiology and Cardiac Surgery

The Eighth World Congress of Pediatric Cardiology and Cardiac Surgery (WCPCCS) will be held in Washington DC, USA, from Saturday, 26 August, 2023 to Friday, 1 September, 2023, inclusive. The Eighth World Congress of Pediatric Cardiology and Cardiac Surgery will be the largest and most comprehensive scientific meeting dedicated to paediatric and congenital cardiac care ever held. At the time of the writing of this manuscript, The Eighth World Congress of Pediatric Cardiology and Cardiac Surgery has 5,037 registered attendees (and rising) from 117 countries, a truly diverse and international faculty of over 925 individuals from 89 countries, over 2,000 individual abstracts and poster presenters from 101 countries, and a Best Abstract Competition featuring 153 oral abstracts from 34 countries. For information about the Eighth World Congress of Pediatric Cardiology and Cardiac Surgery, please visit the following website: [www.WCPCCS2023.org]. The purpose of this manuscript is to review the activities related to global health and advocacy that will occur at the Eighth World Congress of Pediatric Cardiology and Cardiac Surgery. Acknowledging the need for urgent change, we wanted to take the opportunity to bring a common voice to the global community and issue the Washington DC WCPCCS Call to Action on Addressing the Global Burden of Pediatric and Congenital Heart Diseases. A copy of this Washington DC WCPCCS Call to Action is provided in the Appendix of this manuscript. This Washington DC WCPCCS Call to Action is an initiative aimed at increasing awareness of the global burden, promoting the development of sustainable care systems, and improving access to high quality and equitable healthcare for children with heart disease as well as adults with congenital heart disease worldwide

Academic misconduct, misrepresentation and gaming: a reassessment

The motivation for this Special Issue is increasing concern not only with academic misconduct but also with less easily defined forms of misrepresentation and gaming. In an era of intense emphasis on measuring academic performance, there has been a proliferation of scandals, questionable behaviors and devious stratagems involving not just individuals but also organizations, including universities, editors and reviewers, journal publishers, and conference organizers. This introduction first reviews the literature on the prevalence of academic misconduct, misrepresentation and gaming (MMG). The core of the article is organized around a life-cycle model of the production and dissemination of research results. We synthesize the findings in the MMG literature at the level of the investigator or research team, emphasizing that misbehavior extends well beyond fabrication and falsification to include behaviors designed to exaggerate or to mislead readers as to the significance of research findings. MMG is next explored in the post-research review, publication, and post-publication realms. Moving from the individual researcher to the organizational level, we examine how MMG can be engaged in by either journals or organizations employing or funding the researchers. The changing institutional environment including the growth of research assessment exercises, increased quantitative output measurement and greater pressure to publish may all encourage MMG. In the final section, we summarize the main conclusions and offer suggestions both on how we might best address the problems and on topics for future research

Impact Metrics

Virtually every evaluative task in the academy involves some sort of metric (Elkana et al. 1978; Espeland & Sauder 2016; Gingras 2016; Hix 2004; Jensenius et al. 2018; Muller 2018; Osterloh and Frey 2015; Todeschini & Baccini 2016; Van Noorden 2010; Wilsdon et al. 2015). One can decry this development, and inveigh against its abuses and its over-use (as many of the foregoing studies do). Yet, without metrics, we would be at pains to render judgments about scholars, published papers, applications (for grants, fellowships, and conferences), journals, academic presses, departments, universities, or subfields. Of course, we also undertake to judge these issues ourselves through a deliberative process that involves reading the work under evaluation. This is the traditional approach of peer review. No one would advocate a system of evaluation that is entirely metric-driven. Even so, reading is time-consuming and inherently subjective; it is, after all, the opinion of one reader (or several readers, if there is a panel of reviewers). It is also impossible to systematically compare these judgments. To be sure, one might also read, and assess, the work of other scholars, but this does not provide a systematic basis for comparison – unless, that is, a standard metric(s) of comparison is employed. Finally, judging scholars through peer review becomes logistically intractable when the task shifts from a single scholar to a large group of scholars or a large body of work, e.g., a journal, a department, a university, a subfield, or a discipline. It is impossible to read, and assess, a library of work

Making Research Data Accessible

This chapter argues that these benefits will accrue more quickly, and will be more significant and more enduring, if researchers make their data “meaningfully accessible.” Data are meaningfully accessible when they can be interpreted and analyzed by scholars far beyond those who generated them. Making data meaningfully accessible requires that scholars take the appropriate steps to prepare their data for sharing, and avail themselves of the increasingly sophisticated infrastructure for publishing and preserving research data. The better other researchers can understand shared data and the more researchers who can access them, the more those data will be re-used for secondary analysis, producing knowledge. Likewise, the richer an understanding an instructor and her students can gain of the shared data being used to teach and learn a particular research method, the more useful those data are for that pedagogical purpose. And the more a scholar who is evaluating the work of another can learn about the evidence that underpins its claims and conclusions, the better their ability to identify problems and biases in data generation and analysis, and the better informed and thus stronger an endorsement of the work they can offer