Laura Soucek

Modelling Myc inhibition as a cancer therapy

Myc is a pleiotropic basic helix–loop–helix leucine zipper transcription factor that coordinates expression of the diverse intracellular and extracellular programs that together are necessary for growth and expansion of somatic cells. In principle, this makes inhibition of Myc an attractive pharmacological approach for treating diverse types of cancer. However, enthusiasm has been muted by lack of direct evidence that Myc inhibition would be therapeutically efficacious, concerns that it would induce serious side effects by inhibiting proliferation of normal tissues, and practical difficulties in designing Myc inhibitory drugs. We have modelled genetically both the therapeutic impact and the side effects of systemic Myc inhibition in a preclinical mouse model of Ras-induced lung adenocarcinoma by reversible, systemic expression of a dominant-interfering Myc mutant. We show that Myc inhibition triggers rapid regression of incipient and established lung tumours, defining an unexpected role for endogenous Myc function in the maintenance of Ras-dependent tumours in vivo. Systemic Myc inhibition also exerts profound effects on normal regenerating tissues. However, these effects are well tolerated over extended periods and rapidly and completely reversible. Our data demonstrate the feasibility of targeting Myc, a common downstream conduit for many oncogenic signals, as an effective, efficient and tumour-specific cancer therapy.

Mast cells are required for angiogenesis and macroscopic expansion of Myc-induced pancreatic islet tumors

An association between inflammation and cancer has long been recognized, but the cause and effect relationship linking the two remains unclear. Myc is a pleiotropic transcription factor that is overexpressed in many human cancers and instructs many extracellular aspects of the tumor tissue phenotype, including remodeling of tumor stroma and angiogenesis. Here we show in a β-cell tumor model that activation of Myc in vivo triggers rapid recruitment of mast cells to the tumor site—a recruitment that is absolutely required for macroscopic tumor expansion. In addition, treatment of established β-cell tumors with a mast cell inhibitor rapidly triggers hypoxia and cell death of tumor and endothelial cells. Inhibitors of mast cell function may therefore prove therapeutically useful in restraining expansion and survival of pancreatic and other cancers.

Temporal dissection of p53 function in vitro and in vivo

To investigate the functions of the p53 tumor suppressor, we created a new knock-in gene replacement mouse model in which the endogenous Trp53 gene is substituted by one encoding p53ERTAM, a p53 fusion protein whose function is completely dependent on ectopic provision of 4-hydroxytamoxifen. We show here that both tissues in vivo and cells in vitro derived from such mice can be rapidly toggled between wild-type and p53 knockout states. Using this rapid perturbation model, we define the kinetics, dependence, persistence and reversibility of p53-mediated responses to DNA damage in tissues in vivo and to activation of the Ras oncoprotein and stress in vitro. This is the first example to our knowledge of a new class of genetic model that allows the specific, rapid and reversible perturbation of the function of a single endogenous gene in vivo.

c-Myc recruits P-TEFb for transcription, cellular proliferation and apoptosis.

c-Myc promotes cellular proliferation, sensitizes cells to apoptosis and prevents differentiation. It binds cyclin T1 structurally and functionally from the positive transcription elongation factor b (P-TEFb). The cyclin-dependent kinase 9 (Cdk9) in P-TEFb then phosporylates the C-terminal domain of RNA polymerase II, which is required for the transition from initiation to elongation of eukaryotic transcription. Inhibiting P-TEFb blocks the transcription of its target genes as well as cellular proliferation and apoptosis induced by c-Myc.

Inhibition of Myc family proteins eradicates KRas-driven lung cancer in mice.

The principal reason for failure of targeted cancer therapies is the emergence of resistant clones that regenerate the tumor. Therapeutic efficacy therefore depends on not only how effectively a drug inhibits its target, but also the innate or adaptive functional redundancy of that target and its attendant pathway. In this regard, the Myc transcription factors are intriguing therapeutic targets because they serve the unique and irreplaceable role of coordinating expression of the many diverse genes that, together, are required for somatic cell proliferation. Furthermore, Myc expression is deregulated in most-perhaps all-cancers, underscoring its irreplaceable role in proliferation. We previously showed in a preclinical mouse model of non-small-cell lung cancer that systemic Myc inhibition using the dominant-negative Myc mutant Omomyc exerts a dramatic therapeutic impact, triggering rapid regression of tumors with only mild and fully reversible side effects. Using protracted episodic expression of Omomyc, we now demonstrate that metronomic Myc inhibition not only contains Ras-driven lung tumors indefinitely, but also leads to their progressive eradication. Hence, Myc does indeed serve a unique and nondegenerate role in lung tumor maintenance that cannot be complemented by any adaptive mechanism, even in the most aggressive p53-deficient tumors. These data endorse Myc as a compelling cancer drug target.

Making decisions through Myc

c‐Myc is a transcriptional regulator involved in carcinogenesis through its role in growth control and cell cycle progression. Here we attempt to relate its role in stimulating the G1–S transition to the ability to affect functioning of key cell cycle regulators, and we focus on how its property of modulating transcription of a wide range of target genes could explain its capacity to affect multiple pathways leading to proliferation, apoptosis, growth, and transformation.

THE UPS AND DOWNS OF MYC BIOLOGY

The basic helix-loop-helix protein Myc is a renowned transcription factor controlling disparate aspects of cell physiology that, together, allow efficient proliferation of somatic cells. This ability, together with the observation that its deregulated expression occurs in the majority of human cancers, suggests that Myc could be a good therapeutic target. However, several aspects of Myc biology remain elusive: what is the major difference between oncogenic and physiological Myc? How does oncogenic Myc evade the intrinsic tumor surveillance pathways provided by evolution? If Myc inhibition were even possible, what would be the consequences for the homeostasis of normal proliferating tissues versus the fate of cancer cells? Here we summarize the latest works addressing these issues.

Endogenous Myc maintains the tumor microenvironment

The ubiquitous deregulation of Myc in human cancers makes it an intriguing therapeutic target, a notion supported by recent studies in Ras-driven lung tumors showing that inhibiting endogenous Myc triggers ubiquitous tumor regression. However, neither the therapeutic mechanism nor the applicability of Myc inhibition to other tumor types driven by other oncogenic mechanisms is established. Here, we show that inhibition of endogenous Myc also triggers ubiquitous regression of tumors in a simian virus 40 (SV40)-driven pancreatic islet tumor model. Such regression is presaged by collapse of the tumor microenvironment and involution of tumor vasculature. Hence, in addition to its diverse intracellular roles, endogenous Myc serves an essential and nonredundant role in coupling diverse intracellular oncogenic pathways to the tumor microenvironment, further bolstering its credentials as a pharmacological target.

Interrogating open issues in cancer precision medicine with patient-derived xenografts

Patient-derived xenografts (PDXs) have emerged as an important platform to elucidate new treatments and biomarkers in oncology. PDX models are used to address clinically relevant questions, including the contribution of tumour heterogeneity to therapeutic responsiveness, the patterns of cancer evolutionary dynamics during tumour progression and under drug pressure, and the mechanisms of resistance to treatment. The ability of PDX models to predict clinical outcomes is being improved through mouse humanization strategies and the implementation of co-clinical trials, within which patients and PDXs reciprocally inform therapeutic decisions. This Opinion article discusses aspects of PDX modelling that are relevant to these questions and highlights the merits of shared PDX resources to advance cancer medicine from the perspective of EurOPDX, an international initiative devoted to PDX-based research.

Design and properties of a Myc derivative that efficiently homodimerizes

bHLH and bHLHZip are highly conserved structural domains mediating DNA binding and specific protein-protein interactions. They are present in a family of transcription factors, acting as dimers, and their selective dimerization is utilized to switch on and off cell proliferation, differentiation or apoptosis. Myc is a bHLHZip protein involved in growth control and cancer, which operates in a network with the structurally related proteins Max, Mad and Mnt. It does not form homodimers, working as a heterodimer with Max; Max, instead, forms homodimers and heterodimers with Mad and Mnt. Myc/Max dimers activate gene transcription, while Mad/Max and Mnt/Max complexes are Myc/Max antagonists and act as repressors. Modifying the molecular recognition of dimers may provide a tool for interfering with Myc function and, in general, for directing the molecular switches operated via bHLH(Zip) proteins. By molecular modelling and mutagenesis, we analysed the contribution of single amino acids to the molecular recognition of Myc, creating bHLHZip domains with altered dimerization specificity. We report that Myc recognition specificity is encoded in a short region within the leucine zipper; mutation of four amino acids generates a protein, Omomyc, that homodimerizes efficiently and can still heterodimerize with wild type Myc and Max. Omomyc sequestered Myc in complexes with low DNA binding efficiency, preventing binding to Max and inhibiting Myc transcriptional activator function. Consistently with these results, Omomyc produced a proliferation arrest in NIH3T3 cells. These data demonstrate the feasibility of interfering with fundamental biological processes, such as proliferation, by modifying the dimerization selectivity of a bHLHZip protein; this may facilitate the design of peptides of potential pharmacological interest.