Andrei Goga - US grants

[unreadable] DESCRIPTION (provided by applicant): Understanding how selective inhibition of cyclin-dependent kinases (Cdks) affects normal and tumor cell growth may lead to novel treatment strategies for cancer. Cdks are a conserved family of serine/threonine kinases that serve a central role in regulating the eukaryotic cell cycle. Cdk2 and Cdk1 are generally thought to control the S and M phases of the vertebrate cell cycle, respectively. Their exact function in vertebrate cells has been difficult to define because there are no truly specific inhibitors that distinguish between Cdk1 and Cdk2. The essential role of Cdks in cell cycle progression makes them a target of great interest for the development of specific inhibitors as anti-cancer agents. Tumor cells develop a deregulated cell cycle that may render their growth especially sensitive to Cdk inhibition. This raises the possibility that Cdk inhibition may result in the predominant killing of tumor cells while sparing normal cells. Currently no murine model exists to validate the role of specific Cdk inhibition in developing tumors. The long-term goal of the candidate, and this proposal, is to understand how normal and tumor cells respond to Cdk inhibition and to apply this knowledge to develop new therapeutic strategies against cancer. First, we will examine the effect of Cdk inhibition in the context of cells transformed by a variety of oncogenes. Transgenic mouse tumor model systems will be bred to strains harboring oncogenes that render cells sensitive to Cdk inhibition. We will test if Cdk inhibition can mediate significant regression of endogenous tumor, or prevent de novo tumor development. Second, mutant forms of Cdks that can be selectively inhibited in vivo by soluble small molecules will be generated. Third, we will determine the precise cell cycle arrest points and cellular consequences associated with Cdk1 and Cdk2 inhibition. The candidate is a physician-scientist currently pursuing post-doctoral training who is proposing a 5-year mentored research experience with J. Michael Bishop and David Morgan at the University of California, San Francisco. The proposed training program is designed with the goal of preparing this applicant to establish an independent laboratory in an academic oncology department. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): The promise of molecular targeted therapy for cancer is to provide selective killing of tumor cells while sparing normal cells. Targeted therapy, however, requires that the oncogenic pathways activated in tumor cells can be defined, and that selective inhibitors can be found to abrogate these pathways. One major limitation to targeted therapeutic approaches is that many oncogenic pathways, especially those involving transcription factors, cannot be directly inhibited with small molecule compounds. An alternative approach is to use small molecule inhibitors that target basic cellular processes, such as the cell cycle, which merely arrest normal cells, but which in combination with activation of particular oncogenic pathways result in synthetic-lethal combinations. Cyclin-dependent kinases (CDKs) are a conserved family of protein kinases that play a central role in regulating the eukaryotic cell cycle. CDK1 and CDK2 are thought to be particularly important for driving the major cell cycle events in normal and neoplastic mammalian cells and these kinases might therefore be important targets for cancer therapy. The overall hypothesis that is being tested is whether inhibition of different CDKs can result in selective killing of tumor versus normal cells. (1) We seek to determine the genetic context in which cells are rendered especially sensitive to CDK inhibitors, resulting in cell death or another abortive cell cycle program. (2) We seek to determine how MYC oncogene over- expression sensitizes to cell death following CDK1 inhibition. (3) We seek to understand the molecular basis for cell death induced by CDK inhibition. To accomplish our goals we will utilize two complementary approaches to address this question. Both conventional small-molecule CDK inhibitors as well as a chemical-genetic approach will be employed to identify the genetic context in which CDK inhibitors may prove to be useful therapeutics. Our hypothesis, if confirmed, will significantly improve our understanding of how CDK inhibitors may be useful to target specific oncogenic pathways and should lead to novel therapeutics for cancer.

DESCRIPTION (provided by applicant): Despite our rapidly growing understanding of how oncogenes signal, relatively little is known about the underlying mechanisms that cause abrupt cell cycle arrest, cell death and tumor regression upon acutely inactivating an oncogene. This application seeks to address NIH's Provocative Question #22: Why do many cancer cells die when suddenly deprived of a protein encoded by an oncogene? We propose that acute inactivation of different oncogenes in diverse tissue types results in a common 'metabolic catastrophe.' Acute inhibition of driver oncogenes results in widespread collapse of tumor-associated metabolic reprograming. The resulting metabolic state of tumor cells can neither supply them with sufficient energy nor metabolic intermediates for anabolism resulting in a state of 'metabolic catastrophe' resulting in tumor cell death and regression of cancers. The aims of the application seek to: 1) Use innovative hyperpolarized 13C-pyruvate imaging to visualize the earliest metabolic events associate with tumor regression. 2) We will perform global gene expression and metabolomic profiling of liver cancers driven by MYC, RAS or MYC and RAS together to define metabolic pathways altered as a consequence of acute oncogene inactivation. 3) We will compare the metabolic consequences of oncogene inactivation in diverse tissue types, including breast, lung and liver tumors to define which oncogene-regulated metabolic pathways are common across different tissue types and driver oncogenes. The overarching goal of these studies is to identify metabolic pathways critical for tumor survival, against which novel therapeutics can be developed. PUBLIC HEALTH RELEVANCE: A major unanswered question in cancer biology is why and how tumors regress when the initiating oncogene is acutely inhibited. This application seeks to answer this question by examining diverse metabolic changes which occur when different oncogenes are acutely inactivated. We will test the effects of inactivating two canonical oncogenes, MYC and RAS and both combined, in breast lung and liver tumor tissues. An innovative approach to study tumor formation and regression will be employed; including novel imaging technology as well as genetic and metabolic profiling of diverse tumor types. We anticipate that knowledge gained from these studies will be rapidly translated to the development of novel therapeutics to target human cancer.

Project Summary The MYC oncogene is overexpressed in some of the most aggressive and difficult to treat human cancers, including receptor triple-negative breast cancers (TNBCs), as well as high-grade lymphomas and aggressive subtypes of liver cancers. MYC overexpression induces a highly malignant state by driving proliferation and altering various metabolic programs within tumor cells. We recently discovered that MYC-driven transgenic models of breast cancer have reprogramed cellular metabolism that favor fatty acid oxidation (FAO) as an energy source. This finding was also observed in MYC-high TNBCs and has recently been confirmed by an independent research group. Targeting FAO in human cell lines, MYC-driven transgenic animal models and patient-derived xenograft (PDX) models of breast cancer results in diminished tumor growth and increased cell death. What remains unknown is what are the molecular mechanisms through which MYC reprograms tumor metabolism to favor FAO. Furthermore, we seek to understand why MYC-high tumors are dependent on the FAO pathway for their growth and survival. Finally, we seek to improve upon current therapies for TNBCs by performing advance preclinical studies in PDX models to determine if blocking FAO alone or in combination with other metabolic pathways in vivo will provide improved therapeutic response. Our studies will improve our understanding of how MYC reprograms metabolism in vivo and will lead to novel therapeutics for difficult to treat MYC overexpressing cancers.

PROJECT SUMMARY ! The objective of this training program is to provide post-doctoral fellows with didactic and research experience in cellular and molecular aspects of cancer to prepare them for independent investigative careers in basic and translational cancer research. The program forms the core of cancer biology training in the Helen Diller Family Comprehensive Cancer Center (HDCCC) at the University of California, San Francisco (UCSF). The faculty, who are all members of the HDCCC, consists of basic researchers, laboratory-based physician-scientists, and more applied clinician-investigators who share common interests in the multifaceted fields of cellular, molecular and structural biology applied to the understanding of mechanisms of cancer initiation, progression, diagnosis and therapy. The areas of didactic and research training will expose trainees to a spectrum of approaches, concepts and opportunities from altered gene and protein structure and expression, cancer microenvironment and immunity, cell cycling and signaling to differentiation and development. The goal of this approach is to further the understanding of cancer incidence and progression so that the trainees will have an appropriate perspective to approaching basic cancer research as well as to address, prevention, biomarkers and translation to patients. Post-doctoral trainees will join one of 37 research groups involved in studying these basic mechanisms. To broaden their experience, the trainees will have secondary mentors and will be encouraged to seek out collaborations with other research groups at UCSF or outside. Because of the significance and need for training in Bioinformatics and Computational Biology for modern cancer research, we have initiated a workshop for our trainees, taught by the Program faculty. Trainees will have access to all the academic resources available at UCSF. In this way, trainees will be provided with an in-depth research experience in an environment that covers the broad forefront of molecular and cellular dysregulation in cancer. Seminar programs, research-in-progress discussions and journal clubs complement the research training. Trainees must have a Ph.D. or equivalent degree in cell or molecular biology, genetics, biochemistry or an applicable discipline, or an M.D. or M.D., Ph.D. The trainees will be selected on the basis of past accomplishments and promise, course work, grades achieved, suitability for the research projects and a commitment to a research career. Trainees will receive a stipend for an average of 2 years, but will be part of the program throughout their training period of at least 3 years. The program will consist of 8 trainees, complemented by the larger group of other trainees in the host laboratories to make a significant critical mass of basic cancer researchers in the CCC. Upon completion of the program, it is anticipated that the trainees will continue careers in basic and translational cancer research in academic institutions, governmental agencies or the biotechnology industry.

ABSTRACT A progressive breakdown in the bilayered structure of the mammary gland is the hallmark of all breast cancers, but the structural change that occurs between ductal carcinoma in situ (DCIS) and invasive ductal carcinoma (IDC) is of particular importance because it represents a major inflection point in risk for patients. Breast cancers originate in the inner luminal layer of the mammary epithelium, where transformed luminal epithelial cells (LEP) proliferate to fill the ducts and lobules in DCIS. Surprisingly, LEP in DCIS have acquired all the necessary genetic aberrations to invade, but remain constrained within the tissue by an intact outer myoepithelial (MEP) layer?a group of cells that forms a dynamic barrier blocking access of the in situ tumor to the basement membrane (BM, the specialized extracellular matrix (ECM) that surrounds the mammary epithelium). Thus, we propose that translocation of transformed LEP past the MEP layer, and not genetic mutations, is a key rate- limiting step in progression to IDC. Here, we aim to identify the physical and molecular changes that must occur in LEP to facilitate this structural transition. We approach this challenge through the lens of mammary epithelial self-organization. We previously demonstrated that normal human LEP and MEP can self-organize in vitro, and that the capacity of MEP to exclude LEP from the BM is determined by hard-wired and lineage-specific interfacial tensions at each cell-cell and cell-ECM interface. We showed using experiments and mathematical modeling that the LEP-ECM interface is highly unfavorable energetically compared to the MEP-ECM interface, which prevents LEP from positioning themselves next to the BM. We hypothesize the existence of a rate-limiting and high-energy structural intermediate during the progression of DCIS to IDC, where LEP translocate into the MEP layer, next to the BM. We propose a statistical mechanical framework for understanding how perturbations to the interfacial properties and dynamics of tumor cells facilitate the formation of this intermediate. Specifically, we predict that changes to the LEP-ECM interfacial energy are a critical physical change necessary to promote basal translocation of transformed LEP. Preliminary studies support this hypothesis: we found that a frequently dysregulated gene?PIK3CA?disrupts self-organization when activated in LEP by rendering the LEP-ECM interface more energetically favorable. In this proposal, we will determine whether this and other physical changes to LEP are necessary for their basal translocation, and identify the molecular changes downstream of PIK3CA that give rise to these physical changes. We will test our hypothesis using complementary in vitro and in vivo experimental systems: using organoids reconstituted from human reduction mammoplasty tissues and genetically engineered mouse models. Our long-term goal is to reveal the changes that promote and inhibit progression from DCIS to IDC. Better physical and molecular predictors of progression would benefit DCIS patients who would otherwise be over-treated, as only a third of DCIS cases progress to IDC. Further, blocking LEP translocation would represent a therapeutic strategy to prevent breast cancer progression.