A hallmark of human cancer is global DNA hypomethylation (GDHO), but the mechanisms accounting for this defect and its pathological consequences have not been defined in human epithelial ovarian cancer (EOC). In EOC, GDHO was associated with advanced disease and reduced overall and disease-free survival. GDHO(+) EOC was enriched for a proliferative gene expression signature, including CCNE1 and FOXM1 overexpression. DNA hypomethylation preferentially occurred within genomic blocks (hypomethylated blocks) overlapping late-replicating, lamina-associated domains, PRC2 binding, and H3K27me3. Increased proliferation coupled with hypomethylated block formation at late replicating regions suggested passive hypomethylation, which was further supported by the observation that cytosine DNA methyltransferases (DNMTs) and UHRF1 showed significantly reduced expression in GDHO(+) EOC, after normalization to proliferation markers. Importantly, GDHO(+) EOC showed elevated chromosomal instability (CIN), and copy number alterations (CNA) were enriched at hypomethylated blocks. Together, these findings implicate a passive demethylation mechanism for GDHO that promotes genomic instability and poor prognosis in EOC.

The cancer-testis/cancer-germline antigen NY-ESO-1 is a vaccine target in epithelial ovarian cancer (EOC), but its limited expression is a barrier to vaccine efficacy. As NY-ESO-1 is regulated by DNA methylation, we hypothesized that DNA methyltransferase (DNMT) inhibitors may augment NY-ESO-1 vaccine therapy. In agreement, global DNA hypomethylation in EOC was associated with the presence of circulating antibodies to NY-ESO-1. Pre-clinical studies using EOC cell lines showed that decitabine treatment enhanced both NY-ESO-1 expression and NY-ESO-1-specific CTL-mediated responses. Based on these observations, we performed a phase I dose-escalation trial of decitabine, as an addition to NY-ESO-1 vaccine and doxorubicin liposome (doxorubicin) chemotherapy, in 12 patients with relapsed EOC. The regimen was safe, with limited and clinically manageable toxicities. Both global and promoter-specific DNA hypomethylation occurred in blood and circulating DNAs, the latter of which may reflect tumor cell responses. Increased NY-ESO-1 serum antibodies and T cell responses were observed in the majority of patients, and antibody spreading to additional tumor antigens was also observed. Finally, disease stabilization or partial clinical response occurred in 6/10 evaluable patients. Based on these encouraging results, evaluation of similar combinatorial chemo-immunotherapy regimens in EOC and other tumor types is warranted.