SUMMARY DNA methylation at the 5-position of cytosine (5-mC) is a key epigenetic mark critical for various biological and pathological processes. 5-mC can be converted to 5-hydroxymethylcytosine (5-hmC) by the Ten-Eleven Translocation (TET) family of DNA hydroxylases. Here we report that “loss of 5-hmC” is an epigenetic hallmark of melanoma with diagnostic and prognostic implications. Genome-wide mapping of 5-hmC reveals loss of 5-hmC landscape in the melanoma epigenome. We show that down-regulation of Isocitrate Dehydrogenase 2 (IDH2) and TET family enzymes is likely one of the mechanisms underlying 5-hmC loss in melanoma. Rebuilding the 5-hmC landscape in melanoma cells by reintroducing active TET2 or IDH2 suppresses melanoma growth and increases tumor-free survival in animal models. Thus, our study reveals a critical function of 5-hmC in melanoma development and directly links the IDH and TET activity-dependent epigenetic pathway to 5-hmC-mediated suppression of melanoma progression, suggesting a new strategy for epigenetic cancer therapy.

SUMMARY Therapeutic antibodies targeting programmed cell death-1 (PD-1) activate tumor-specific immunity and have shown remarkable efficacy in the treatment of melanoma. Yet, little is known about tumor cell-intrinsic PD-1 pathway effects. Here we show that murine and human melanomas contain PD-1-expressing cancer subpopulations and demonstrate that melanoma cell-intrinsic PD-1 promotes tumorigenesis, even in mice lacking adaptive immunity. PD-1 inhibition on melanoma cells by RNA interference, blocking antibodies, or mutagenesis of melanoma-PD-1 signaling motifs suppresses tumor growth in immunocompetent, immunocompromised and PD-1-deficient tumor graft recipient mice. Conversely, melanoma-specific PD-1 overexpression enhances tumorigenicity, as does engagement of melanoma-PD-1 by its ligand, PD-L1, whereas melanoma-PD-L1 inhibition or knockout of host-PD-L1 attenuate growth of PD-1-positive melanomas. Mechanistically, the melanoma-PD-1 receptor modulates downstream effectors of mTOR signaling. Our results identify melanoma cell-intrinsic functions of the PD-1:PD-L1 axis in tumor growth and suggest that blocking melanoma-PD-1 might contribute to the striking clinical efficacy of anti-PD-1 therapy.

Increasing evidence suggests that tumor dormancy represents an important mechanism underlying the observed failure of existing therapeutic modalities to fully eradicate cancers. In addition to its more established role in maintaining minimal residual disease after treatment, dormancy might also critically contribute to early stages of tumor development and the formation of clinically undetectable micrometastatic foci. There are striking parallels between the concept of tumor dormancy and the cancer stem cell (CSC) theory of tumor propagation. For instance, the CSC hypothesis similarly predicts that a subset of self-renewing cancer cells-that is CSCs-is responsible for tumor initiation, bears the preferential ability to survive tumor therapy, and persists long term to ultimately cause delayed cancer recurrence and metastatic progression. Additionally, many of the biological mechanisms involved in controlling the dormant state of a tumor can also govern CSC behavior, including cell cycle modifications, alteration of angiogenic processes, and modulation of antitumor immune responses. In fact, quiescence and immune escape are emerging hallmark features of at least some CSCs, indicating significant overlap between dormant cancer populations and CSCs. Herein, we crucially dissect whether CSCs occupy specific roles in orchestrating the switch between dormancy and exuberant tumor growth. We elucidate how recently uncovered CSC biological features could enable these cells to evade immunologic clearance and regulate cancer expansion, relapse, and progression. We propose that the study of CSC immunobiological pathways holds the promise to critically advance our understanding of the processes mediating tumor dormancy. Ultimately, such research endeavors could unravel novel therapeutic avenues that efficiently target both proliferating and dormant CSCs to minimize the risk of tumor recurrence in cancer patients.