Research Grants Awarded
Validating Myc As A Therapeutic Target In Breast Cancer
Investigator Initiated Research
Breast cancers harbor mutations in many molecular pathways. Hitherto, most attention has focused on targeting molecules that are overtly mutated in breast cancers, assuming that they are essential drivers of the malignancy. Most notable are the HER and EGFR receptor tyrosine kinases, which present tractable therapeutic targets. Unfortunately, recent studies indicate that oncogenic signaling through RTKs is both redundant and adaptive, perhaps explaining why RTK inhibitors have failed to demonstrate the hoped for efficacy in most solid tumors. At the distal end of oncogenic signaling lie a multitude of biological outputs necessary for breast tumor maintenance - tumor cell proliferation, survival, de-differentiation, angiogenesis, invasion and metastasis. These processes are both mechanistically diverse and highly redundant, further complicating and confounding identification of effective therapeutic targets.
The c-, N- and L-Myc proteins encode isofunctional transcription factors of remarkable pleiotropy. Over-expression of c-Myc is directly implicated in ~70% of breast cancers. Substantial work indicates that Myc proteins are essential integrators of cell expansion: Myc-deficient normal or transformed cells exhibit profound deficits in cell proliferation and replicate only slowly and inefficiently. More recently, in vivo studies using switchable forms of the Myc oncoprotein to drive tumors in many distinct tissues in mice have shown that Myc, in addition to tumor cell proliferation, instructs and maintains many other aspects of the tumor phenotype ? both cell-intrinsic (e.g. cell growth, de-differentiation, loss of cell adhesion) and cell-extrinsic (angiogenesis, invasion, inflammation). Consequently, subsequent de-activation of Myc triggers tumor regression through multiple mechanisms - both cell-intrinsic (e.g. senescence, re-differentiation and apoptosis) and extrinsic (e.g. collapse of tumor vasculature and microenvironment). Nonetheless, Myc itself appears not to be activated by mutation in the majority of breast cancers. Instead, over-expression of endogenous Myc appears driven by "upstream" oncogenic signals that arise, typically, in mutated HER receptor tyrosine kinases or the Wnt/APC/beta-catenin axis. The extent to which such endogenous Myc acts as a non-redundant integrator of the tumor phenotype remains unknown.
Taken together, the above suggest that Myc function ? non-redundant and underpinning multiple oncogenic programs required for maintenance of established tumors - would be an excellent therapeutic target. However, there are two perceived problems. First, current dogma deem Myc proteins as "undruggable" by small molecules. Their most obvious and specific Achilles' Heel is the dimerization interface they all share with their obligate partner Max. This interface is a particularly attractive drug target since it is common to all Myc family members and, moreover, required for Myc oncogenic function: hence, mutations in the Myc dimerization domain that render it refractory to drug binding will likely also disable Myc's requisite interaction with its partner Max ? limiting the capacity of breast tumor cells to acquire drug resistance Myc mutants. However, the Myc:Max dimerization interface is an extended coiled-coil interaction that has hitherto proven an elusive target. Nevertheless, a variety of novel chemistries (e.g. macrolide and stapled alpha-helices) are changing these preconceptions. Indeed, some early stage compounds have shown promise, albeit in vitro. Second, Myc is deemed essential for normal cell proliferation, raising concerns that inhibiting Myc would elicit catastrophic side effects in continuously or episodically proliferating tissues such as intestine, bone marrow, skin or mammary epithelium.
We have constructed a unique transgenic mouse in which expression of a dominant-negative Myc dimerization mutant can be reversibly induced in all tissues of the live animal. Hence, this model allows us to inhibit Myc function systemically and reversibly by targeting the shared Myc dimerization interface ? essentially a genetic model for a Myc inhibitory drug that at once indicates both the therapeutic efficacy of Myc inhibition in tumors and the collateral side effects for normal tissues. Using this model we have shown that systemic inhibition of Myc triggers dramatic phenotypes in skin and intestinal epithelium, involving slowing of cell cycle in epidermal follicles and intestinal crypts. However, mice show no ill effects and, moreover, all phenotypes rapidly and completely reverse when the Myc-inhibitor is shut down. By contrast, the same blockade of Myc in orthotopic lung tumors driven by sporadic activation of endogenous L-Ras triggers rapid and complete regression. We will extend these studies to ascertain the role played by Myc in genesis, progression and maintenance of breast cancer, initially using two well-described mouse models in which there is no overt activation of Myc itself: the MMTV-HER2 model of lobular and alveolar breast cancer and the MMTV-Wnt-1 model of metastatic disease. Using these models we will establish the acute consequences of systemic Myc inhibition for early, late stage and (where feasible) metastatic breast cancers and ascertain whether, and if so how, resistance to Myc inhibition can emerge. In this proposal we will test the hypothesis that the Myc transcription factors serve as non-redundant, core conduit of the diverse signaling pathways that are oncogenically activated in breast cancers ? coupling oncogenic signaling to the many programs involved in tumor maintenance, expansion and invasion.
The last decade has ushered in remarkable advances in our understanding of the molecular architecture of breast cancer. In particular, new analytical techniques have uncovered a bewildering diversity and number of genetic mutations that affect many different cellular and tissue processes. Cataloging and sequencing these mutations is now a major focus of breast cancer research, with particular emphasis on identifying recurrent lesions based on the notion that these are likely bottleneck event during mammary carcinogenesis. Unfortunately, cataloging mutations in tumors provides us with no direct information as to which of them is especially critical for maintenance of the established tumor and, hence, a candidate for targeted therapy. Tumors, like all evolving systems, pass through restrictive bottlenecks and bottlenecks select for new traits. However, once through that selective chicane, there is no surety that such traits will remain relevant in the tumor?s later guise as an invasive, metastatic malignancy. The problem is compounded by the fact that breast cancers, like other solid human tumors, pass through episodes of genomic instability, generating a vast amount of mutational noise that obscures the underlying mutations that drive the disease 1. Nonetheless, the evident genetic complexity of breast cancers has fostered the common belief that they are irreducibly complex, adaptable and able to evolve resistance to almost any targeted therapy.
In this proposal, we confront this pessimism by taking a completely different approach to identifying and validating therapeutic targets in breast cancer. We reason that substantial evidence in vivo and in vitro supports the notion that oncogenic mutations signal their pathological effects through a core cell machinery which then serves to couple such oncogenic signals to the various processes that drive and maintain breast tumors. We hypothesize that this core machinery is shared by many, perhaps all breast tumor types and is both essential and non-redundant ? and so has many of the desired features of an ideal therapeutic target. A pivotal component of this core machinery is Myc, a transcription factor that controls and coordinates the diverse gene expression programs necessary for tumor growth, spread and blood supply. Myc is currently deemed an intractable therapeutic target by the pharmaceutical industry ? in the main because it exerts its protean influences through direct interactions with other protens, a class of biological process that has proved difficult to disrupt. However, a variety of novel chemistries are now changing our views on this and it is time to revisit the previously "undruggable." The other major concern is that Myc, aside from its obligate role in tumor cells, is a core component of normal cell proliferation/regeneration too. Hence, pharmacological inhibition of Myc might trigger unacceptably severe side effects akin to those elicited by conventional radio and chemotherapy. Whether there exists an exploitable therapeutic window for an anti-Myc drug depends upon the relative importance of Myc function to normal versus tumor tissues. In this regard, studies suggest that tumor cells may have a far more urgent and continuous need for Myc activity than normal tissues: however, this has never been confirmed by experiment.
In this application, we propose to model the efficacy of anti-Myc therapy in mammary cancers in vivo ? both the therapeutic impact of Myc inhibition on established breast cancers and any side effects of Myc inhibition on normal breast epithelium and other tissues. To do this, we have constructed a unique mouse model in which we can reversibly inhibit Myc function, systemically in all tissues including tumors, essentially modeling genetically what an anti-Myc drug of the future would do. Our preliminary data in lung tumors are most encouraging and indicate that transient Myc inhibition triggers rapid and dramatic regression of established tumors with mild, well-tolerated and completely reversible side effects on normal tissues. We will use our model to determine short and long term effects of transient Myc inhibition on normal mouse mammary epithelium during puberty and pregnancy, and to ascertain its therapeutic impact in two, initial well-validated mouse models of breast cancer. Should, as we predict, Myc inhibition prove therapeutically efficacious in treating mammary tumors in the mouse, our over-arching goal will be to present the pharmaceutical industry with the strongest possible case for re-investigating Myc as a therapeutic target in the treatment of this disease.
1. Greenman, C. et al., Patterns of somatic mutation in human cancer genomes. Nature 446, 153-8 (2007).