Abstract 3157: Novel GD2-specific chimeric antigen receptor-modified T cells targeting osteosarcoma
Authors: Chulanetra Monrat , Sayour Elias , Eldjerou Lamis , Lagmay Joanne , Milner Rowan , Slayton William , Chang Lung-Ji 
Published: August 2015

Abstract
Despite intensive multi-modality therapies employed in the treatment of osteosarcoma (OS) the prognosis for children with high-risk disease remains suboptimal. Since standard systemic therapy for OS has remained unchanged in the past thirty years, with no new agents identified to complement conventional regimens, there has been a significant plateau in the survival of OS patients thus necessitating the development of more targeted and efficacious therapeutics. Chimeric antigen receptor (CAR) modified T cells can meet this need by utilizing the immune system's surveillance capacity and potent cytotoxic mechanisms against tumor specific antigen targets with exquisite specificity. Since OS highly expresses the GD2 antigen, a viable immunotherapeutic target, we sought to assess if a novel 4th generation CAR modified T cell could target GD2 and induce cytotoxicity against OS.
We have engineered a newer generation of CAR T cells that harness the vital co-stimulatory domains imperative for T cell activation while embedding the safety features of suicide induction. The latter, an advanced safety design iCasp9, is based on AP1903-induced caspase 9 dimer formation to trigger apoptosis, which has been incorporated in our CAR design. The 4th generation lentiviral CAR (4S-CAR) encodes a combination of CD28/CD137/CD27 T cell co-stimulatory domains and CD3zeta domain fused with a mutated FKBP12 (F36V) and caspase 9. To illustrate the self-destructive function, 4S-CAR T cells were treated with AP1903 to induce iCasp9 dimerization. High efficiency of apoptosis was observed within 30 min, and in 2 hours, more than 80% CAR T cells died. After demonstrated the ability to screen patient OS samples for GD2 expression, we found that OS cells expressing high levels of GD2 were effectively targeted and killed by 4S-GD2-CAR-T cells. 4S-GD2 CAR modified T cells were shown to overexpress the exhaustion marker PD-1 when in contact with target tumor cells, and multiple OS expressing cell lines were shown to overexpress PD-L1, and the PD-L1 level increased after cocultured with 4S-GD2 CAR modified T cells. The combination effects of immune checkpoint inhibitors such as anti-PD-L1 and anti-PD-1 antibodies with CAR T cells will be further investigated.
Anti-GD2 expressing CAR T cells provide a novel targeted approach to eliminate residual disease with consummate specificity. As cellular therapeutics using CAR T cells become further entrenched in the forefront against refractory malignancies, OS provides a new and compulsory niche to further test the feasibility, safety and potential efficacy of this promising new technology.
Citation Format: Monrat Chulanetra, Elias Sayour, Lamis Eldjerou, Joanne Lagmay, Rowan Milner, William Slayton, Lung-Ji Chang. Novel GD2-specific chimeric antigen receptor-modified T cells targeting osteosarcoma. [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 3157. doi:10.1158/1538-7445.AM2015-3157