300 - A Novel Therapeutic Approach Using citVIM-specific CAR-T cells and Radiotherapy to Target NSCLC and Pancreatic Cancer Cells

02:35pm - 02:45pm ET

Room 160

Presenter(s)

Yorleny Vicioso Mora, MD, PhD - Duke University, Durham, NC

Y. M. Vicioso Mora¹, D. Carrion Estrada², L. Lopez Bailon², R. Chaurio Gonzalez², G. hernandez Galicia², N. Plappert², C. Hsiang Lai², J. O’Brien³, J. Mine², J. Kowalczyk³, S. R. Floyd¹, S. J. Antonia³, and J. R. Conejo-Garcia²; ¹Department of Radiation Oncology, Duke University School of Medicine, Durham, NC, ²Department of Integrative Immunobiology, Duke University School of Medicine, Durham, NC, ³Department of Medicine, Duke University School of Medicine, Durham, NC

Purpose/Objective(s): We identified citrullinated vimentin (citVIM) as the target of a tumor-reactive antibody (Ab) produced in response to pembrolizumab in a patient with Non-Small Cell Lung Cancer (NSCLC) (NCT02818920). Using the variable region of this Ab, we generated a new Chimeric Antigen Receptor-T (CAR-T) cell redirected against citVIM. We found that citVIM is expressed on the surface of tumors (epithelial origin), tumor-promoting myeloid cells, and fibroblasts, but not in healthy tissues. Therefore, citVIM-directed CAR-T cells represent a potential new therapy for multiple malignancies. However, despite progress in CAR-T cell therapy for patients with refractory hematological cancers, its application in solid tumors has been limited by hypoxic, immunosuppressive TME, poor antigen expression, and limited CAR-T cell infiltration. Radiotherapy (RT) has been shown to enhance CAR-T cell therapy by improving the hypoxic TME and promoting CAR-T cells tumor infiltration. Because RT induces cellular stress and citVIM is expressed under such conditions, here we present a novel approach using RT to sensitize NSCLC and pancreatic cancer cells to citVIM-specific CAR-T cells by inducing citVIM expression on the tumor cells' surface.

Materials/Methods: We irradiated NSCLC and pancreatic cancer cell lines with conventional, hypo-fractionated, and ablative doses of RT and assessed the expression of citVIM on the surface of tumor cells post RT compared to untreated controls. We further developed citVIM-directed CARs using the scFv cloned from our patient-derived Ab. We then assess the antitumor efficacy of citVIM CAR-T cells against NSCLC and pancreatic cancer cells, both in vitro and using orthotopic tumor-bearing syngeneic mice in combination with RT.

Results: We found that surface expression of citVIM was upregulated in NSCLC and pancreatic cancer cells after doses of RT = 5Gy. We observed upregulation in the expression of the enzyme that catalyzes the posttranslational modification converting arginine to citrulline, leading to the expression of citVIM on tumor cells (Peptydil deaminase enzyme 2 or PADI2). citVIM-directed CAR-T cells showed target-specific cytotoxicity compared to mock control CAR-T cells. CRISPR/Cas9-mediated knockout of PADI2 in target cells confirmed the specific killing ability of citVIM CAR-T cells. RT (10 Gy) significantly increased the cytotoxicity of CitVIM-directed CAR-T cells against NSCLC and pancreatic cancer cell lines in vitro and enhanced the therapeutic response of citVIM-specific CAR-T cells in vivo, resulting in superior survival time compared with the RT-only group in our pancreatic tumor-bearing mice.

Conclusion: Our findings suggest that RT induces citVIM expression on cancer cells surface, leading to improved citVIM-directed CAR-T cells cytotoxicity against NSCLC and pancreatic cancer cells. This is the first study to establish RT as a key regulator of citVIM expression on tumor cells, providing a rationale for combining RT and cellular therapy.