Abstract 4575: Chimeric antigen receptor macrophages (CARMA) for adoptive cellular immunotherapy of solid tumors
Authors: Klichinsky Michael , Ruella Marco , Shestova Olga , Kenderian Saad S. , Kim Miriam Y. , O'Connor Roddy , Scholler John , June Carl , Gill Saar 
Published: July 2017

Abstract
Chimeric antigen receptor (CAR) T cell immunotherapy has demonstrated profound results in hematologic malignancies but clinical efficacy in the solid tumor setting has not been observed. Barriers to T cell entry and function may partially explain this observation. As solid tumors actively recruit myeloid cells, we hypothesized that macrophages have the potential to be a powerful cellular immunotherapeutic agent in this setting if properly activated and redirected. We here describe the development of CAR macrophages (CARMA), demonstrating the feasibility, mechanism, and efficacy of this platform. To examine the function of CARs in macrophages, first generation anti-CD19, anti-mesothelin, or anti-HER2 CARs with a CD3ζ intracellular domain were introduced into the THP1 macrophage model. In vitro function was assessed via quantitative phagocytosis and luciferase-based specific killing assays. CARMA selectively phagocytosed and cleared cognate antigen-bearing tumor cells. To demonstrate the requirement for CAR-mediated intracellular signaling for activity, a CD3ζ-null CAR construct was tested in vitro. The deletion of CD3ζ significantly reduced the phagocytic and killing capacity (p&lt;0.01) of CARMA. We identified Ad5f35, a chimeric adenovirus, as a novel and highly efficient viral vector for the transduction of normal donor and cancer patient macrophages (&gt;70% CAR expression). Ad5f35 transduction polarized human macrophages toward a durable immunostimulatory M1 phenotype and rendered CARMA resistant to subversion toward the immunosuppressive M2 phenotype, as defined by surface markers and metabolomics. CARMA enhanced the proliferative capacity of CD8+ T cells in phytohemagglutinin activation assays and secreted factors that activated by-stander macrophages. Primary human anti-HER2 CARMA demonstrated targeted phagocytosis and killing of HER2 expressing ovarian and breast cancer cell lines, and exhibited a six-fold higher luciferase-based killing capacity of SKOV3 cells compared to trastuzumab in vitro (p=0.002). Anti-HER2 CARMA was evaluated in vivo in an intraperitoneal (IP) SKOV3 ovarian cancer xenograft model. Mice that received IP CARMA had a decrease in tumor burden of approximately two orders of magnitude and had a 30-day survival benefit relative to untreated or control macrophage treated mice (p=0.018). In a systemically disseminated SKOV3 model, a single dose of IV CARMA led to a durable anti-tumor response (38-fold reduction relative to control on day 31 post-treatment; p=0.016) Lastly, we demonstrated that the blockade of the anti-phagocytic CD47/SIRPα axis enhanced the phagocytic capacity of CARMA. In summary, we here demonstrate that human macrophages engineered with a CAR exhibit targeted anti-tumor function in both in vitro and in vivo preclinical models. This novel cellular immunotherapeutic approach has a clear translational potential for the treatment of solid tumors.
Citation Format: Michael Klichinsky, Marco Ruella, Olga Shestova, Saad S. Kenderian, Miriam Y. Kim, Roddy O'Connor, John Scholler, Carl June, Saar Gill. Chimeric antigen receptor macrophages (CARMA) for adoptive cellular immunotherapy of solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4575. doi:10.1158/1538-7445.AM2017-4575