Abstract: Modified vaccinia Ankara-Bavarian Nordic (MVA-BN)-Brachyury followed by fowlpox virus-BN-Brachyury was well tolerated upon administration to patients with advanced cancer. Sixty-three percent of patients developed CD4+ and/or CD8+ T-cell responses to brachyury after vaccination. BN-Brachyury vaccine also induced T-cell responses against CEA and MUC1, which are cascade antigens, that is, antigens not encoded in the vaccines. Background Brachyury, a transcription factor, plays an integral role in the epithelial–mesenchymal transition, metastasis, and tumor resistance to chemotherapy. It is expressed in many tumor types, and rarely in normal tissues, making it an ideal immunologic target. Bavarian Nordic (BN)-Brachyury consists of vaccination with modified vaccinia Ankara (MVA) priming followed by fowlpox virus (FPV) boosting, each encoding transgenes for brachyury and costimulatory molecules. Methods Patients with metastatic solid tumors were treated with two monthly doses of MVA-brachyury s.c., 8 × 108 infectious units (IU), followed by FPV-brachyury s.c., 1 × 109 IU, for six monthly doses and then every 3 months for up to 2 years. The primary objective was to determine safety and tolerability. Results Eleven patients were enrolled from March 2018 to July 2018 (one patient was nonevaluable). No dose-limiting toxicities were observed. The most common treatment-related adverse event was grade 1/2 injection-site reaction observed in all patients. Best overall response was stable disease in six patients, and the 6-month progression-free survival rate was 50%. T cells against brachyury and cascade antigens CEA and MUC1 were detected in the majority of patients. Conclusion BN-Brachyury vaccine is well tolerated and induces immune responses to brachyury and cascade antigens and demonstrates some evidence of clinical benefit.

Abstract: Purpose: The transcription factor brachyury has been shown in preclinical studies to be a driver of the epithelial-to-mesenchymal transition (EMT) and resistance to therapy of human tumor cells. This study describes the characterization of a Modified Vaccinia Ankara (MVA) vector–based vaccine expressing the transgenes for brachyury and three human costimulatory molecules (B7.1, ICAM-1, and LFA-3, designated TRICOM) and a phase I study with this vaccine. Experimental Design: Human dendritic cells (DC) were infected with MVA-brachyury-TRICOM to define their ability to activate brachyury-specific T cells. A dose-escalation phase I study (NCT02179515) was conducted in advanced cancer patients (n = 38) to define safety and to identify brachyury-specific T-cell responses. Results: MVA-brachyury-TRICOM-infected human DCs activated CD8+ and CD4+ T cells specific against the self-antigen brachyury in vitro. No dose-limiting toxicities were observed due to vaccine in cancer patients at any of the three dose levels. One transient grade 3 adverse event (AE) possibly related to vaccine (diarrhea) resolved without intervention and did not recur with subsequent vaccine. All other AEs related to vaccine were transient and ≤grade 2. Brachyury-specific T-cell responses were observed at all dose levels and in most patients. Conclusions: The MVA-brachyury-TRICOM vaccine directed against a transcription factor known to mediate EMT can be administered safely in patients with advanced cancer and can activate brachyury-specific T cells in vitro and in patients. Further studies of this vaccine in combination therapies are warranted and planned. Clin Cancer Res; 23(22); 6833–45. ©2017 AACR.

Abstract: BN-CV301 is a poxviral-based vaccine comprised of recombinant (rec.) modified vaccinia Ankara (MVA-BN-CV301; prime) and rec. fowlpox (FPV-CV301; boost). Like its predecessor PANVAC, BN-CV301 contains transgenes encoding tumor-associated antigens MUC1 and CEA as well as costimulatory molecules (B7.1, ICAM-1, and LFA-3). PANVAC was reengineered to make it safer and more antigenic. Patients and Methods: This open-label, 3+3 design, dose-escalation trial evaluated three dose levels (DL) of MVA-BN-CV301: one, two, or four subcutaneous injections of 4 × 108 infectious units (Inf.U)/0.5 mL on weeks 0 and 4. All patients received FPV-CV301 subcutaneously at 1 × 109 Inf.U/0.5 mL every 2 weeks for 4 doses, then every 4 weeks. Clinical and immune responses were evaluated. Results: There were no dose-limiting toxicities. Twelve patients enrolled on trial [dose level (DL) 1 = 3, DL2 = 3, DL3 = 6). Most side effects were seen with the prime doses and lessened with subsequent boosters. All treatment-related adverse events were temporary, self-limiting, grade 1/2, and included injection-site reactions and flu-like symptoms. Antigen-specific T cells to MUC1 and CEA, as well as to a cascade antigen, brachyury, were generated in most patients. Single-agent BN-CV301 produced a confirmed partial response (PR) in 1 patient and prolonged stable disease (SD) in multiple patients, most notably in KRAS-mutant gastrointestinal tumors. Furthermore, 2 patients with KRAS-mutant colorectal cancer had prolonged SD when treated with an anti-PD-L1 antibody following BN-CV301. Conclusions: The BN-CV301 vaccine can be safely administered to patients with advanced cancer. Further studies of the vaccine in combination with other agents are planned. See related commentary by Repáraz et al., p. 4871

Abstract: Concurrent ETBX-011, ETBX-051, and ETBX-061 can be safely administered to patients with advanced cancer. All patients developed CD4+ and/or CD8+ T-cell responses after vaccination to at least one tumor-associated antigen (TAA) encoded by the vaccine; 5/6 patients (83%) developed MUC1-specific T cells, 4/6 (67%) developed CEA-specific T cells, and 3/6 (50%) developed brachyury-specific T cells. The presence of adenovirus 5-neutralizing antibodies did not prevent the generation of TAA-specific T cells. Background A novel adenovirus-based vaccine targeting three human tumor-associated antigens—CEA, MUC1, and brachyury—has demonstrated antitumor cytolytic T-cell responses in preclinical animal models of cancer. Methods This open-label, phase I trial evaluated concurrent administration of three therapeutic vaccines (ETBX-011 = CEA, ETBX-061 = MUC1 and ETBX-051 = brachyury). All three vaccines used the same modified adenovirus 5 (Ad5) vector backbone and were administered at a single dose level (DL) of 5 × 1011 viral particles (VP) per vector. The vaccine regimen consisting of all three vaccines was given every 3 weeks for three doses then every 8 weeks for up to 1 year. Clinical and immune responses were evaluated. Results Ten patients enrolled on trial (DL1 = 6 with 4 in the DL1 expansion cohort). All treatment-related adverse events were temporary, self-limiting, grade 1/2 and included injection site reactions and flu-like symptoms. Antigen-specific T cells to MUC1, CEA, and/or brachyury were generated in all patients. There was no evidence of antigenic competition. The administration of the vaccine regimen produced stable disease as the best clinical response. Conclusion Concurrent ETBX-011, ETBX-051, and ETBX-061 can be safely administered to patients with advanced cancer. Further studies of the vaccine regimen in combination with other agents, including immune checkpoint blockade, are planned.

Abstract: There are several situations for which vaccines can be employed in the prevention and treatment of human carcinomas. True prevention of cervical cancer has been demonstrated in the use of the HPV vaccine (Gardasil). The Sipuleucel-T “dendritic cell”-based vaccine (Dendreon) has recently been approved by the FDA for the treatment of metastatic prostate cancer. The low levels of toxicity seen with many cancer therapeutic vaccines has led to interest in the potential use of these vaccines in patients with high-risk preneoplastic conditions such as familial adenomatous polyposis (FAP), high-grade prostatic epithelial neoplasia (PIN), and in patients with preneoplastic breast lesions. Clinical studies are ongoing and planned in the use of vaccines in adjuvant settings in patients rendered disease free radiographically, but with a high risk of developing metastatic disease. Preclinical Studies: We have developed a recombinant poxviral vaccine platform (recombinant vaccinia prime and multiple avixpox booster vaccination) with each vector containing the transgenes for one or more tumor-associated antigens and the transgenes for a triad of T-cell costimulatory molecules (TRICOM). These are “off-the-shelf” vaccines that can and have been widely distributed for clinical studies. In preclinical studies, these vaccines have demonstrated the ability to induce increased levels of high-avidity antigen-specific T cells and induce antitumor effects in several animal models. The use of a CEA-TRICOM vaccine with Celebrex was shown to enhance survival in a preneoplastic model for FAP at far greater levels than either agent alone. Preclinical studies also demonstrated the synergy in the use of TRICOM vaccines with local radiation of tumor or certain chemotherapeutic agents (both modalities have been shown to alter the phenotype of tumor cells and rendered them more susceptible to T-cell killing). Anti-CTLA4 MAb was also shown to synergize with TRICOM vectors to enhance T-cell avidity. Several of the above findings have led to completed and ongoing clinical studies. Clinical Studies: A 43-center placebo (empty vector)-controlled randomized trial employing PSA-TRICOM vaccine (PROSTVAC) in patients with metastatic prostate cancer demonstrated that PROSTVAC vaccinated patients had a superior overall survival at 3 years post-study (30% vs. 17% alive) and a longer median overall survival by 8.5 months (25.1 vs. 16.6 months, p=0.0061, HR 0.56). As with the Sipuleucel-T vaccine trial, this survival advantage was seen in the absence of any advantage in time to progression. Toxicity in this and other TRICOM trials was minimal. A second single-arm trial with PROSTVAC demonstrated a similar survival benefit. Patients who survived longer on vaccine had higher ratios of effector-to-regulatory T cells. A phase III trial with PROSTVAC is scheduled to begin in 2011. Preliminary evidence of patient benefit is being observed in the use of PROSTVAC vaccine in ongoing randomized trials employing (a) Samarium 153 chelate (Quadramet) plus or minus vaccine in prostate cancer patients with metastatic bone lesions, (b) hormone therapy plus or minus vaccine in hormone refractory “nonmetastatic” prostate cancer patients with rising PSA, and (c) in a multicenter trial in hormone naive prostate cancer patients following treatment of their primary tumor but with rising PSA. A nonrandomized study has also provided evidence of clinical benefit for the use of PROSTVAC with anti-CTLA4 MAb in patients with metastatic prostate cancer. Clinical trials are also ongoing in the use of CEA-MUC-1-TRICOM vaccine (PANVAC). A multicenter trial (M. Morse, H.K. Lyerly, PIs, Duke Cancer Center) in patients receiving PANVAC following surgery for metastatic colorectal cancer also provided preliminary evidence of increased patient survival in the absence of time to progression. An ongoing randomized trial at NCI in patients with metastatic breast carcinoma employing Docetaxel plus or minus PANVAC is also providing preliminary evidence of patient benefit in the vaccine arm. Immune Trageting of Cells with Enhanced Metastatic Potential: The use of surgery, radiation and/or adjuvant chemotherapy will control most primary carcinomas. It is believed, however, that a subpopulation of cells with higher invasive and metastatic potential, and greater resistance to conventional therapy is responsible for eventual metastatic spread. The goal is to identify molecules involved in these processes of tumor progression and to target them immunologically. We have recently identified the T-box transcription factor Brachyury as a master driver of the epithelial-to-mesenchymal transition (EMT) phenotype process for a range of human carcinoma cells. The EMT process has been implicated by others as a major factor in tumor invasion and metastases. We have now demonstrated in preclinical studies that enhanced expression of Brachyury in a range of human carcinoma cells leads to a switch in phenotype from epithelial to mesenchymal, increased migration and invasion in vitro, and increased metastatic spread in vivo; silencing Brachyury RNA in carcinoma cells has shown reciprocal results. Unlike several other EMT drivers examined, Brachyury is selectively expressed in a range of human carcinomas vs. normal adult human tissues with the exception of testes. We have now demonstrated the ability to generate Brachyury-specific human T cells, which in turn have the ability to lyse human carcinoma cells expressing Brachyury in an MHC-restricted manner. We are in the process of generating clinical grade recombinant Brachyury vaccine. References: 1. Kantoff P, Schuetz TJ, Blumenstein BA, Glode LM, Bilhartz DL, Wyand M, Manson K, Panicali DL, Laus R, Schlom J, Dahut W, Arlen PM, Gulley JL, Godfrey WR. Overall survival (OS) analysis of a Phase II randomized controlled trial (RCT) of a poxviral-based PSA targeted immunotherapy in metastatic castration-resistant prostate cancer (mCRPC). J Clin Oncol. 2010;28:1099-1105. 2. Gulley JL, Arlen PM, Madan RA, Tsang K-Y, Pazdur MP, Skarupa L, Jones JL, Poole DJ, Higgins JP, Hodge JW, Cereda V, Vergati M, Steinberg SM, Halabi S, Jones E, Chen C, Parnes H, Wright JJ, Dahut WL, Schlom J. Immunologic and prognostic factors associated with overall survival employing a poxviral-based PSA vaccine in metastatic castrate-resistant prostate cancer. Cancer Immunol Immunother. 2010;59:663-674. 3. Palena C, Polev DE, Tsang KY, Fernando RI, Litzinger M, Krukovskaya LL, Baranova AV, Kozlov AP, and Schlom J. The human T-box mesodermal transcription factor Brachyury is a candidate target for T-cell mediated cancer immunotherapy. Clin Cancer Res. 13:2471-2478, 2007. 4. Fernando RI, Fernando RI, Litzinger M, Trono P, Hamilton DH, Schlom J, and Palena C. The T-box transcription factor Brachyury promotes epithelial-to-mesenchymal transition in human tumor cells. J Clin Invest.120:533-544, 2010. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr SY24-02. doi:10.1158/1538-7445.AM2011-SY24-02