Abstract 2508: Multi-modal treatment with peptide vaccine, metronomic cyclophosphamide and anti-PD1 monoclonal antibody provides effective control of tumors in multiple models
Authors: Weir Genevieve , Hrytsenko Olga , Stanford Marianne , Karkada Mohan , Berinstein Neil , Mansour Marc 
Published: August 2015

Abstract
Future cancer immunotherapies will combine multiple treatments to improve immune responses to cancer through synergistic, multi-modal mechanisms. In a phase I clinical trial, we found that metronomic cyclophosphamide (mCPA; 50 mg BID) enhanced the immunogenicity of a DepoVaxTM (DPX) based cancer vaccine (DPX-Survivac) in ovarian cancer patients. We emulated these results in preclinical transplantable tumor models which allowed us to study the underlying mechanisms of cyclophosphamide induced immune modulation and explore additional combinations to enhance the therapeutic effect. In syngeneic murine models for ovarian cancer (MOSE), melanoma (B16-F10) and HPV-induced cancer (C3), metronomic low dose cyclophosphamide (mCPA) on alternating weeks (20 mg/kg/day PO) in combination with a DPX peptide vaccine provided long-term control of established tumors. The addition of mCPA caused selective enrichment of antigen-specific CD8+ T cells resulting in increased immune responses detected by IFN-γ ELISPOT and in vivo cytotoxicity assay, as well as improved protection from tumors. Efficacy of the vaccine and mCPA combination was limited in mice bearing advanced tumors, yet antigen-specific CD8+ T cells could be detected infiltrating the tumor by flow cytometry along with increased expression of PD-1 by qPCR, suggesting that the combination therapy was able to generate strong cytotoxic T cell response but was still subject to tumor induced suppression within the tumor microenvironment. A triple combination therapy of mCPA, vaccine and anti-PD1 enabled the control and/or regression of established later-treated tumors which could not be achieved with a combination of mCPA and anti-PD1. To better understand the mechanism of the enhanced protection observed with the tri-therapy, mice were implanted with a mixture of C3 and B16 tumor cells and then treated with a vaccine against the C3 tumors alone, in combination with mCPA and anti-PD1. Splenocytes of mice treated with the tri-therapy produced IFN-γ ELISPOT responses towards both cell lines, suggesting epitope spreading. This effect could not be detected in tumor bearing mice treated with mCPA anti-PD1 without vaccine. We conclude that enhanced tumor control mediated by the tri-therapy is mediated by highly active, tumor-specific cytotoxic T lymphocytes produced by a strongly immunogenic vaccine, resulting in epitope spreading that can further facilitate tumor rejection or control. These results provide a rationale for clinical testing of checkpoint blockade therapy in combination with our highly immunogenic combination of mCPA and DPX-Survivac targeting the tumor associated antigen survivin.
Citation Format: Genevieve Weir, Olga Hrytsenko, Marianne Stanford, Mohan Karkada, Neil Berinstein, Marc Mansour. Multi-modal treatment with peptide vaccine, metronomic cyclophosphamide and anti-PD1 monoclonal antibody provides effective control of tumors in multiple models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2508. doi:10.1158/1538-7445.AM2015-2508