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BackgroundA vaccine containing 6 melanoma-associated peptides to stimulate helper T cells (6MHP) is safe, immunogenic, and clinically active. A phase I/II trial was designed to evaluate safety and immunogenicity of 6MHP vaccines plus programmed death 1 (PD-1) blockade.Participants and methodsParticipants with advanced melanoma received 6MHP vaccines in an incomplete Freund’s adjuvant (6 vaccines over 12 weeks). Pembrolizumab was administered intravenously every 3 weeks. Tumor biopsies at baseline and day 22 were analyzed by multiplex immunohistochemistry. Primary end points were safety (Common Terminology Criteria for Adverse Events V.4.03) and immunogenicity (ex vivo interferon-γ ELISpot assay). Additional end points included changes in the tumor microenvironment (TME) and clinical outcomes.ResultsTwenty-two eligible participants were treated: 6 naïve to PD-1 antibody (Ab) and 16 PD-1 Ab-experienced. Median follow-up was 24.4 months. Most common treatment-related adverse events (any grade) included injection site reactions, fatigue, anemia, lymphopenia, fever, elevated aspartate aminotransferase, pruritus, and rash. Treatment-related dose-limiting toxicities were observed in 3 (14%) participants, which did not cross the study safety bound. A high durable T cell response (Rsp) to 6MHP was detected in only one participant, but twofold T cell Rsps to 6MHP were detected in 7/22 (32%; 90% CI (16% to 52%)) by week 13. Objective clinical responses were observed in 23% (1 complete response, 4 partial responses), including 4/6 PD-1 Ab-naïve (67%) and 1/16 PD-1 Ab-experienced (6%). Overall survival (OS) was longer for PD-1 Ab-naïve than Ab-experienced participants (HR 6.3 (90% CI (2.1 to 28.7)). In landmark analyses at 13 weeks, OS was also longer for those with T cell Rsps (HR 6.5 (90% CI (2.1 to 29.2)) and for those with objective clinical responses. TME evaluation revealed increased densities of CD8+T cells, CD20+B cells, and Tbet+cells by day 22.ConclusionsTreatment with the 6MHP vaccine plus pembrolizumab was safe, increased intratumoral lymphocytes, and induced T cell Rsps associated with prolonged OS. The low T cell Rsp rate in PD-1 Ab-experienced participants corroborates prior murine studies that caution against delaying cancer vaccines until after PD-1 blockade. The promising objective response rate and OS in PD-1 Ab-naïve participants support consideration of a larger study in that setting.

BackgroundPeptide vaccines designed to stimulate melanoma-reactive CD4+ T cells can induce T cell and antibody (Ab) responses, associated with enhanced overall survival. We hypothesized that adding toll-like receptor 3 agonist polyICLC to an incomplete Freund’s adjuvant (IFA) would be safe and would support strong, durable CD4+ T cell and Ab responses. We also hypothesized that oral low-dose metronomic cyclophosphamide (mCy) would be safe, would reduce circulating regulatory T cells (T-regs) and would further enhance immunogenicity.Participants and methodsAn adaptive design based on toxicity and durable CD4+ T cell immune response (dRsp) was used to assign participants with resected stage IIA-IV melanoma to one of four study regimens. The regimens included a vaccine comprising six melanoma peptides restricted by Class II MHC (6MHP) in an emulsion with IFA alone (Arm A), with IFA plus systemic mCy (Arm B), with IFA+ local polyICLC (Arm C), or with IFA+ polyICLC+ mCy (Arm D). Toxicities were recorded (CTCAE V.4.03). T cell responses were measured by interferon γ ELIspot assay ex vivo. Serum Ab responses to 6MHP were measured by ELISA. Circulating T-regs were assessed by flow cytometry.ResultsForty-eight eligible participants were enrolled and treated. Early data on safety and dRsp favored enrollment on arm D. Total enrollment on Arms A-D were 3, 7, 6, and 32, respectively. Treatment-related dose-limiting toxicities (DLTs) were observed in 1/7 (14%) participants on arm B and 2/32 (6%) on arm D. None exceeded the 25% DLT threshold for early closure to enrollment for any arm. Strong durable T cell responses to 6MHP were detected ex vivo in 0%, 29%, 67%, and 47% of participants on arms A-D, respectively. IgG Ab responses were greatest for arms C and D. Circulating T-regs frequencies were not altered by mCy.Conclusions6MHP vaccines administered with IFA, polyICLC, and mCy were well tolerated. The dRsp rate for arm D of 47% (90% CI 32 to 63) exceeded the 18% (90% CI 11 to 26) rate previously observed with 6MHP in IFA alone. Vaccination with IFA+ polyICLC (arm C) also showed promise for enhancing T cell and Ab responses.

BackgroundA multipeptide vaccine containing 6 melanoma-associated peptides to stimulate helper T cells (6MHP) is safe and immunogenic and has clinical activity. A phase I/II trial was designed to evaluate safety and immunogenicity of 6MHP vaccines plus PD1 blockade.MethodsParticipants with measurable advanced melanoma, age ≥ 18 years, and ECOG performance status 0–1 were administered 6MHP vaccine intradermally and subcutaneously in an incomplete Freund’s adjuvant on days (D) 1, 8, 15, 43, 64, and 85. Pembrolizumab 200 mg was administered intravenously every 3 weeks for up to two years. Biopsies of accessible tumors at baseline and D22 were analyzed by multiparameter immunofluorescence histology. Primary endpoints were safety (CTCAE 4.03) and immunogenicity (ex vivo IFNγ ELIspot assay). Secondary and exploratory endpoints included changes in the tumor microenvironment (TME), and clinical outcomes.ResultsTwenty-two eligible participants were enrolled and treated, including 6 naïve to PD-1 Ab and 16 anti-PD-1 Ab-experienced. Median follow-up was 20 months. Treatment-related adverse events (any grade) experienced by >20% were injection site reactions, fatigue, anemia, nausea, fever, bruising, and rash. Treatment-related dose limiting toxicities (grade 3 elevated AST, skin ulcer, or uveitis) were observed in 3 (14%), which did not cross the study safety bound. Objective clinical responses were observed in 23% (1 CR, 4 PR), including 4/6 anti-PD-1 Ab-naive (67%) and one 1/16 anti-PD1 Ab-experienced (6%). Four participants (18%) had SD as best radiographic response (18%), all in the Ab-experienced cohort. T cell responses to 6MHP were detected in seven participants (32%) by week 13 and were associated with clinical response (CR/PR 80% vs. SD/PD 18%; p = 0.01). Overall survival was prolonged for anti-PD-1 Ab naïve vs experienced (p = 0.0048), for those with T cell response (p = 0.045, landmark analysis after week 13), and for those with objective response (p = 0.0148). TME evaluation in 12 participants revealed significant increases by D22 in the densities (per mm2) of CD8+ T cells (p = 0.0186), CD20+ B cells (P = 0.002), and Tbet+ cells (p = 0.034).ConclusionsIn patients with advanced melanoma, combined treatment with the 6MHP vaccine plus pembrolizumab was safe, increased intratumoral T and B cells, as well as Th1 (Tbet+) cells, and induced T cell responses that were associated with objective response and with overall survival. The promising objective response rate and overall survival in patients naive to PD1 blockade supports consideration of a larger study to assess definitive benefit in that clinical setting.AcknowledgementsWe acknowledge the support of Merck for providing pembrolizumab at no charge, and the University of Virginia Cancer Center Support grant P30 CA044579 for support of shared resource facilities.Trial RegistrationThe clinical trial Mel64 (PATHVACS) is registered with Clinicaltrials.gov (NCT02515227).Ethics ApprovalThe clinical trial Mel64 (PATHVACS) was performed with IRB (#18174) and FDA approval (IND #10825) and is registered with Clinicaltrials.gov (NCT02515227). Written informed consent was obtained from each participant prior to participation in the study.