Abstract 2814: Anti-CAIX Immune Restoring (IR) CAR-T cells display superior antitumor activity and reverse immunosuppressive TME in a humanized ccRCC orthotopic mouse model
Authors: Wang Yufei , Buck Alicia , Grimaud Marion , Culhane Aedin , Braun David , Kodangattil Sreekumar , Razimbaud Cecile , Chang Matthew , Fayed Atef , Apollon Audrey , Kastrunes Gabriella , Zerefa Luann , Piel Brandon , Ivanova Elena , Bonal Dennis , Jones Kristen , Nguyen Quang-De , Zhu Zhu , Wei Kevin , Hayden Nicholas , O'Donnell Madison , Huang Ying , Jenning Rebecca , Ficial Miriam , Sticco-Ivins Maura Aliezah , Signoretti Sabina , Wu Catherine , Choueiri Toni , Wee Jon , Paweletz Cloud , Barbie David A. , Freeman Gordon , Marasco Wayne A. 
Published: June 2022

Abstract
Chimeric Antigen Receptor (CAR) T cell therapy is a new type of “living drug” that has proven to be a powerful, clinically translatable immunotherapy for hematologic malignancies. To date, there are five CAR-T products approved by FDA, four CD19 targeted CAR-T cells, and one targeting B-cell maturation antigen (BCMA). However, this success has not yet been transferred to solid tumors. A major challenge is the immunosuppressive tumor microenvironment (TME). Development of immunotherapies has traditionally been hampered by discrepancies observed between in vitro and in vivo studies and actual clinical trial outcomes. The lack of clinically relevant mouse models for human immunotherapy testing is often seen as the primary cause. Existing clear cell renal cell carcinoma (ccRCC) in vivo models poorly recapitulate the tumor microenvironment (TME). Here we report a ccRCC orthotopic humanized NSG-SGM3 mouse model (ccRCC-hNSG-SGM3) with reconstituted human lymphocytes and bearing human ccRCC skrc-59 cells under the kidney capsule. Human leukocyte antigen (HLA) matched CD34+ human stem cells were used for the humanization to reduce T cell alloreactivity against skrc-59 human ccRCC cells. Tumors were harvested, sorted for CD45+ tumor infiltrating leukocytes (TILs) and single cell RNA sequencing (scRNAseq) was performed to profile the TME in ccRCC-hNSG-SGM3. By comparing to patient data from prospective clinical trials of the anti-PD-1 monoclonal antibody (mAb) nivolumab in advanced ccRCC, the results demonstrated that CD45+ TILs from ccRCC-hNSG-SGM3 reconstitute most CD45+ cell types, including dendritic cells, exhausted CD8 T cells, and regulatory T cells (Tregs), that are observed in advanced ccRCC patient TME. Furthermore, we generated HLA matched Immune Restoring (IR) CAR-T cells which can secrete anti-PD-L1 monoclonal antibody (mAb) locally to restore active antitumor immunity, and we assessed the efficacy and safety of IR CAR-T cells in this model. Anti-CAIX CAR-T cells armored with anti-PD-L1 mAb showed superior efficacy in tumor regression and significantly decreased TIL exhaustion compared to irrelevant CAR or irrelevant payload in this model. In addition, in situ hybridization (ISH) results showed CAR-T cells infiltration in tumor but no CAR-T cells were observed in normal tissues. In summary, the ccRCC-hNSG-SGM3 system is able to model the advanced ccRCC TME and provides a powerful tool for ccRCC TME study and immunotherapy assessment. IR CAR-T cells exhibited superior tumor regression and reversed immunosuppressive TME in humanized mice.
Citation Format: Yufei Wang, Alicia Buck, Marion Grimaud, Aedin Culhane, David Braun, Sreekumar Kodangattil, Cecile Razimbaud, Matthew Chang, Atef Fayed, Audrey Apollon, Gabriella Kastrunes, Luann Zerefa, Brandon Piel, Elena Ivanova, Dennis Bonal, Kristen Jones, Quang-De Nguyen, Zhu Zhu, Kevin Wei, Nicholas Hayden, Madison O'Donnell, Ying Huang, Rebecca Jenning, Miriam Ficial, Maura Aliezah Sticco-Ivins, Sabina Signoretti, Catherine Wu, Toni Choueiri, Jon Wee, Cloud Paweletz, David A. Barbie, Gordon Freeman, Wayne A. Marasco. Anti-CAIX Immune Restoring (IR) CAR-T cells display superior antitumor activity and reverse immunosuppressive TME in a humanized ccRCC orthotopic mouse model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2814.