Martin McMahon - US grants

DESCRIPTION (provided by applicant): Mutationally activated BRAF is expressed in a wide variety of human cancers with a strikingly high prevalence in melanoma. Mutated BRAF promotes sustained activation of the RAF->MEK->ERK MAP kinase signaling pathway that in turn contributes to many of the aberrant behaviors of the melanoma cell. Using conditionally active forms of BRAF, it has been demonstrated that BRAF->MEK->ERK signaling can suppress apoptosis in a variety of mammalian cells. These data are consistent with the embryonic lethal phenotype of 5/WFnullizygous mice that display a variety of apoptotic defects. BCL-2 family proteins, comprising both pro- and anti-apoptotic members, play an essential role in the regulation of apoptosis. Hence, we find it highly provocative that we have identified a direct biochemical connection between activation of BRAF and the phosphorylation of the BH3 domain only protein BIM, a pro-apoptotic member of the BCL-2 family that plays an essential role in melanocyte apoptosis. Here we propose experiments to test the hypothesis that BRAF activation can influence apoptosis in normal melanocytes and in melanoma cells through alterations in BIM phosphorylation. To test this hypothesis we will utilize a system for the conditional activation of BRAF signaling in melanocytes. This system will be used to explore the global effects of BRAF activation in melanocytes as well as the specific effects of BRAF on apoptosis. To complement these studies, we will use genetic (RNAi) and pharmacological inhibitors of BRAF->MEK->ERK signaling or BIM expression to assess the importance of this pathway in melanocyte/melanoma cell apoptosis in response to trophic factor deprivation, matrix detachment or DNA damage. Although there is strong genetic and biochemical evidence for an important role for BRAF signaling in the initiation and progression of human melanoma, and for BIM in the regulation of melanocyte survival, we will test the hypothesis that these proteins are linked in a direct biochemical pathway. We shall elucidate the biochemical mechanism(s) by which BRAF regulates the proapoptotic effects of BIM in mammalian cells with an emphasis on the role of phosphorylation in regulating BIM expression/activity. We will characterize the extent of BRAF-induced BIM phosphorylation, identify the specific sites and analyse the heterogeneity of BIM phosphorylation in response to BRAF activation. We shall confirm these sites by expression of normal and mutated BIM and by the generation of phospho-specific antisera. Finally, we will use biochemical and genetic techniques to explore the mechanism by which BRAF-induced phosphorylation inhibits BIM's pro-apoptotic activity. We believe that the proposed experiments will provide a general system for the analysis of the earliest effects of BRAF in the conversion of normal melanocytes into metastatic melanoma cells. Moreover, these studies will lead to a better understanding of the role(s) of BRAF and BIM in controlling apoptosis in mammalian cells. Finally, we anticipate that these studies will ultimately lead to the design and implementation of new therapeutic strategies to target metastatic melanoma cells for apoptosis leading to remission of this dread disease.

The Pancreas Cancer Program comprises a multi-disciplinary group of investigators focused on pancreatic cancer of either endocrine or ductal origin (PDAC). These investigators include a multidisciplinary team of clinicians and clinical scientists in endocrinology, gastroenterology, medical oncology, radiation oncology and surgical oncology who are poised to address critical issues in pancreatic cancer, with full integration of faculty in basic and population sciences. The scientific goals of the program are to: Understand the genetic and environmental factors that influence risk for PDAC and pancreatic neuroendocrine tumor development. Understand the genetic and biologic events occurring early in pancreas cancer, with a view to developing better diagnostic tools. Understand the factors that promote progression of pancreas cancer, with a special emphasis on PDAC stroma and the biology of invasion and metastasis. Understand the basis of chemotherapy resistance and sensitivity in PDAC and pancreatic neuroendocrine tumors. Develop new paradigms and model systems to evaluate the role of specific genes in the initiation and progression of these cancers. Use such model systems to evaluate new targeted therapeutics, either alone or in combination with conventional cytotoxic agents, in the treatment of PDAC and pancreatic neuroendocrine tumors. The program includes 15 members from 8 departments in the School of Medicine. The Program has $2,444,680 Total peer reviewed support for the last budget year. The Program has 8% intra-programmatic and 34% inter-programmatic publications.

? DESCRIPTION (provided by applicant): Lung cancer is a global scourge responsible for 1.4 million deaths worldwide and ~160,000 deaths in the USA this year alone. Despite the grim clinical picture, cancer genetics has revealed that lung cancers can be divided into genetically-defined subsets based on driver oncogene mutations that, in turn, serve as predictive biomarkers for the clinical deployment of new first-line pathway-targeted therapies in lung cancer patients. However, to date, only a minority of such patients have benefitted from these advances such that treatment options for most patients remain poor and are limited to conventional approaches including surgery, radiation, and/or conventional chemotherapy that are ineffective against tumor cells and toxic to normal cells. The overarching, long-term goal of this research is to aid the deployment of rational, evidence-based therapeutic strategies to treat lung cancer patients. However, the immediate objectives of this proposal are to mechanistically define how parallel pathways, such as WNT?ß-catenin?c-MYC or PI3'-kinase?PDK?AKT signaling, cooperate with oncogenic KRASG12D or BRAFV600E in the genesis and maintenance of lung cancer. To do so we will use: State-of-the-art genetically engineered mouse models of lung cancer; Lung cancer- derived cell lines whose aberrant behavior is driven by relevant genetic abnormalities; Pathway-targeted therapeutics, many of which are in cancer clinical trials and one of which is specifically being tested in lung cancer and; An innovative transposon mutagenesis system to simultaneously elicit lung cancer progression and identify its underlying genetic cause. Building on a solid foundation of studies published in the previous cycle of this grant, in Aim 1 we will identify the tumor cell autonomous mechanisms by which WNT?ß-catenin signaling cooperates with oncogenic KRASG12D or BRAFV600E in the genesis and maintenance of lung cancer. In Aim 2 we will explore the mechanisms by which PI3'-kinase signaling promotes the proliferative expansion and maintenance of both early- and late-stage BRAFV600E-induced lung tumors. BRAFV600E-induced lung tumors remain uniformly benign unless cooperating genomic events unleash malignant progression. In Aim 3 document the use of Sleeping Beauty (SB) transposon mutagenesis to promote progression of BRAFV600E- driven lung cancers. Genetic progression factors identified will be validated using lentivirus-mediated cDNA expression or by in vivo CRISPR/Cas9-mediated tumor suppressor gene silencing in the mouse lung. Moreover, we will mine the TCGA lung adenocarcinoma genome, RNA sequencing and RPPA databases to credential genes implicated in our SB screen in mice as being directly relevant to bona fide human lung cancer. Finally, although this proposal is focused primarily on studies of genetic cooperation in the genesis of lung cancer, we will test the anti-tumor effects of novel pathway-targeted inhibitors of BRAFV600E, WNT or PI3'- kinase signaling that are likely to have important translational implications for the design and evaluation of new pathway-targeted strategies to treat patients with this ubiquitous, devastating and poorly understood disease.

DESCRIPTION (provided by applicant): Melanoma is noted for its alarming increase in incidence, especially amongst the young, aggressive clinical behavior and propensity for lethal metastasis, illustrating an urgent need for new treatment strategies for this disease. Despite the bleak clinical and epidemiological picture, genetic analysis has uncovered key driver oncogenes in melanoma such as mutationally activated BRAF. Indeed, when the BRAFV600E oncoprotein is pharmacologically inhibited with vemurafenib, BRAF mutated melanoma patients, even those with widely disseminated, metastatic disease, have enjoyed dramatic tumor regression coupled with significant health improvement. However, since not all BRAF mutated melanoma patients respond to vemurafenib and, those that do, generally relapse with lethal drug resistant disease, the paradigm of melanoma therapy is evolving towards rationally-targeted combinations of pathway-targeted inhibitors acting against BRAFV600E and cooperating signaling modules such as the PI3'-kinase->AKT pathway. Indeed, despite vemurafenib's clinical success and our growing scientific knowledge of the melanoma cell's inner workings, the challenge remains how best to employ BRAF inhibitors for melanoma therapy to: 1. Maximize therapeutic benefit; 2. Minimize emergence of lethal drug resistant disease; and; 3. Mitigate potentially harmful side effects. Consequently, the long-term, overarching goal of this research is to design BRAF inhibitor-based combination therapies that enhance the overall response rate and the depth of each patient's primary anti-tumor response and extend indefinitely the duration of remission of patients with BRAF mutated melanoma in collaboration with colleagues in the UCSF Melanoma Clinic. Towards these goals, the short-term aims of this proposal are to employ state-of-the-art genetically engineered mouse (GEM) models, bona fide human melanoma cells and a portfolio of clinically relevant pathway-targeted inhibitors to elucidate the molecular mechanism(s) of BRAFV600E/PI3'- kinase pathway cooperation in melanomagenesis and the consequences of combined pathway-targeted blockade in the pre-clinical setting. In Aim 1, GEM models will be used to probe the importance of PI3'- kinase->AKT signaling in cooperating with oncogenic BRAFV600E in converting normal melanocytes to metastatic melanoma cells. In Aim 2 the superiority of combined versus single agent inhibition of BRAFV600E or PI3'-kinase in promoting tumor regression will be evaluated using GEM models of BRAFV600E-initiated melanoma. To complement analysis of mouse melanoma specimens in Aim 2, a panel of bona fide human BRAFV600E expressing melanoma cell lines will be employed in Aim 3 to elucidate molecular mechanism(s) by which BRAFV600E cooperates with PI3'-kinase->AKT signaling in regulating the melanoma cell division cycle and/or apoptosis through effects on protein synthesis. When completed, these studies will provide the mechanistic foundation for the development of robust and rational, pathway-targeted combination therapeutic strategies to increase both the quantity and quality of life of patients with BRAF mutated melanoma.

? DESCRIPTION (provided by applicant): Lung cancer is a global scourge responsible for 1.4 million deaths worldwide and ~160,000 deaths in the USA this year alone. Despite the grim clinical picture, cancer genetics has revealed that lung cancers can be divided into genetically-defined subsets based on driver oncogene mutations that, in turn, serve as predictive biomarkers for the clinical deployment of new first-line pathway-targeted therapies in lung cancer patients. However, to date, only a minority of such patients have benefitted from these advances such that treatment options for most patients remain poor and are limited to conventional approaches including surgery, radiation, and/or conventional chemotherapy that are ineffective against tumor cells and toxic to normal cells. The overarching, long-term goal of this research is to aid the deployment of rational, evidence-based therapeutic strategies to treat lung cancer patients. However, the immediate objectives of this proposal are to mechanistically define how parallel pathways, such as WNT?ß-catenin?c-MYC or PI3'-kinase?PDK?AKT signaling, cooperate with oncogenic KRASG12D or BRAFV600E in the genesis and maintenance of lung cancer. To do so we will use: State-of-the-art genetically engineered mouse models of lung cancer; Lung cancer- derived cell lines whose aberrant behavior is driven by relevant genetic abnormalities; Pathway-targeted therapeutics, many of which are in cancer clinical trials and one of which is specifically being tested in lung cancer and; An innovative transposon mutagenesis system to simultaneously elicit lung cancer progression and identify its underlying genetic cause. Building on a solid foundation of studies published in the previous cycle of this grant, in Aim 1 we will identify the tumor cell autonomous mechanisms by which WNT?ß-catenin signaling cooperates with oncogenic KRASG12D or BRAFV600E in the genesis and maintenance of lung cancer. In Aim 2 we will explore the mechanisms by which PI3'-kinase signaling promotes the proliferative expansion and maintenance of both early- and late-stage BRAFV600E-induced lung tumors. BRAFV600E-induced lung tumors remain uniformly benign unless cooperating genomic events unleash malignant progression. In Aim 3 document the use of Sleeping Beauty (SB) transposon mutagenesis to promote progression of BRAFV600E- driven lung cancers. Genetic progression factors identified will be validated using lentivirus-mediated cDNA expression or by in vivo CRISPR/Cas9-mediated tumor suppressor gene silencing in the mouse lung. Moreover, we will mine the TCGA lung adenocarcinoma genome, RNA sequencing and RPPA databases to credential genes implicated in our SB screen in mice as being directly relevant to bona fide human lung cancer. Finally, although this proposal is focused primarily on studies of genetic cooperation in the genesis of lung cancer, we will test the anti-tumor effects of novel pathway-targeted inhibitors of BRAFV600E, WNT or PI3'- kinase signaling that are likely to have important translational implications for the design and evaluation of new pathway-targeted strategies to treat patients with this ubiquitous, devastating and poorly understood disease.

EXPERIMENTAL THERAPEUTICS PROGRAM ABSTRACT The Experimental Therapeutics (ET) Program of Huntsman Cancer Institute (HCI) is organized to facilitate transdisciplinary collaboration and to accelerate the progress of patient-oriented research to clinical translation and care of patients with cancer. The Program is organized into two main research themes: Drug Discovery/Drug Delivery and Clinical Research. Program members work to discover new agents that can be developed into drugs used to target cancer cells, develop second-generation polymeric carriers of anticancer drugs, design macromolecular targeted therapeutics with selective inhibitory effects on various types of malignancy, and develop drug-free macromolecular therapeutic constructs to increase efficacy of currently used therapeutics. Program members also ensure the execution of clinical research at HCI and the translation of important Cancer Center discoveries to human applications. Program members are also active in the leadership and conduct of clinical trials, including investigator-initiated trials. This clinical research focus aligns with the goals of NCI to develop and conduct state-of-the-art cancer treatment and advanced imaging multi- institutional clinical trials to evaluate new cancer therapies and related clinical approaches across a broad range of populations and cancer types. Program members are engaged in the effort to transform the previous NCI Clinical Trials Cooperative Group Program into a consolidated and integrated National Clinical Trials Network (NCTN), by providing clinical trials leadership and access to vitally important accruals among patients in the Intermountain West. The goals of the program are to: 1) provide new approaches to individualized cancer treatment as well as the most up-to-date clinical treatments via investigator-initiated therapeutic, imaging, and imaging-guided clinical trials; 2) foster bidirectional translation of discoveries between the laboratory and the clinic through collaborations and scientific synergy among ET program members, other program members, and HCI practicing clinicians; 3) develop novel therapies to meet unmet clinical needs in cancer through collaboration and interaction with the HCI multidisciplinary disease groups and disease-oriented research teams. The 39 ET Program members are drawn from 11 different University of Utah departments in the School of Medicine and Colleges of Pharmacy and Science. The Program includes 24 professors, seven associate professors, seven assistant professors, and one instructor, with 11 PhDs, 22 MDs, and six MD, PhDs. Funding includes $14.8M in 2013 total costs (NCI: 28%, other NIH: 20%, other peer-reviewed sources: 15%). From 2009?2013, ET members published 550 peer-reviewed papers, of which 28% involved collaboration with other Cancer Center members and 66% involved external partnerships.

EXPERIMENTAL THERAPEUTICS PROGRAM ABSTRACT The Experimental Therapeutics (ET) Program at Huntsman Cancer Institute (HCI) is the hub that connects and coordinates discoveries made in the basic sciences with translational research and clinical trials to improve diagnostics, disease management, and treatment outcomes for cancer patients. This broad-reaching program coordinates faculty from numerous and diverse departments conducting research on mechanisms underlying normal and abnormal cellular biochemistry, physiology, pharmacology, and molecular and cell biology with physician-scientists and clinical researchers. The overarching goal is to improve patient outcomes by identifying and validating novel treatment approaches, fostering translational research to adapt laboratory observations into novel interventions, and conducting clinical research to assess the effectiveness of new treatment approaches in humans. ET Program Aims encompass discovery and translation of HCI research into clinical practice through development of novel treatment interventions, original investigator-initiated trials (IITs), industry trials (including first-in-human trials), and trials in cooperation with the National Cancer Trials Network (NCTN), all with special emphasis on cancers of particular relevance in HCI?s catchment area?the State of Utah. The Specific Aims are: 1) to develop new cancer therapies based on discoveries from HCI science, and 2) to develop innovative treatment interventions and conduct translational and clinical research, with special emphasis on cancers relevant to HCI?s catchment area. The ET Program?s 59 members belong to 15 departments within three University of Utah schools/colleges. Peer-reviewed cancer-related funding was $4.9M at the end of 2018, with $2.6M (53%) of that from the National Cancer Institute. Program members also hold $9.8M in industry funding. During the previous cycle, ET members published 970 peer-reviewed cancer research papers. Of these, 20% resulted from intra- programmatic collaborations, while 23% represent inter-programmatic collaborations. Further, the Program houses 288 interventional treatment trials that accrued 676 patients in 2018. The ET Program is instrumental to the success of HCI?s overall mission to improve the outcomes of cancer patients. In particular, we: 1) translate deep basic mechanistic understandings and pioneering preclinical translational research into original clinical trials, and 2) collect and conduct iterative analysis of specimens from patients on clinical trials to improve our understanding of disease mechanisms and therapeutic sensitivity or resistance in human beings. Going forward, the ET Program will strive to bring research full circle to fuel a virtuous cycle of discovery, preclinical translation, clinical testing, and bedside-to-bench analyses.