Background: Epigenetic alterations affect virtually all cellular pathways associated with tumorigenesis and cancer progression. Importantly, the multifaceted immunomodulatory activity of DHA has been shown to improve the immunogenicity and immune recognition of neoplastic cells; thus, we predicted DHA could be part of new and potentially more effective immunotherapeutic combinations in cancer (Maio et al., Clin Can Res, 2015). Targeting immune check-point(s) with immunomodulatory monoclonal antibodies (mAb) is a novel and rapidly evolving strategy to treat cancer. The prototype approach of this therapeutic modality relies on the inhibition of negative signals delivered by CTLA-4 expressed on activated T lymphocytes. CTLA-4 blockade has changed the therapeutic landscape of metastatic melanoma (MM) by significantly improving the long-term survival of mm patients; however, objective clinical responses are limited, thus opening the path to combination regimens to improve its efficacy. Based on the immunomodulatory activity of the second-generation DHA guadecitabine (Covre et al., Semin Oncol, 2015) we designed the NIBIT-M4 study. This trial will sequence guadecitabine and ipilimumab in mm patients to provide proof-of-concept to the immunologic and clinical efficacy of DHA combined with CTLA-4 blockade. Methods: This is a Phase 1b, dose-escalation study in treatment naïve or pretreated unresectable Stage III or Stage IV melanoma patients, amenable to serial tumor biopsies. Primary objective will assess MTD and safety of guadecitabine combined with ipilimumab. Secondary objectives will include immune-related (ir) -DCR, -ORR, -PFS, median OS, and survival rate at 1 and 2-years. Immune-biologic correlates will be exploratory objectives. The dose escalation of guadecitabine will follow a 3+3 design. Cohorts of 3-6 patients will receive ipilimumab i.v. 3 mg/kg on W1, 4, 7 and 10 day 1 q21d and guadecitabine s.c. on W0, 3, 6, 9, days 1-5 q21d at the one of following doses: Dose Level (DL) -1: 15 mg/m² day; DL 0: 30 mg/m² day; DL +1: 45 mg/m² day. Sample size will range from 6 to 19 patients. Four patients have been enrolled to date. Clinical trial information: NCT02608437