Abstract 111: Establishing orthotopic renal cell carcinoma xenograft in PBMC-humanized immunodeficient NSG-MHC I/II double knock-out mice
Authors: Kang Wonyoung , Wang Yufei , Laimon Yasmin N. , Pham Oanh , Matran Ben , Bottoms Lauren , Khang Sheng , Yuan Hsien-Chi , Murugan Nithyassree , Sheshdeh Aseman B. , Yao Li-chin , Yang Jiwon , Cheng Mingshan , Signoretti Sabina , Marasco Wayne A. , Keck James G. 
Published: March 2024

Abstract
Clear cell renal cell carcinoma (ccRCC), the predominant histological subtype of RCC, is highly correlated with immune cells, an association that is linked to a worse prognosis. Our study aimed to establish an orthotopic ccRCC mouse model reconstituted with human peripheral blood mononuclear cells (PBMCs) and implanted with human ccRCC SKRC-59 cells under the kidney capsule in immunodeficient NSG-MHC I/II double knock-out (DKO) mice. The study was conducted using two experimental designs to optimize humanization and tumor formation. In the first experimental design, the mice were engrafted with SKRC-59 cells under the left kidney capsule 24 hours post-irradiation on study day 0. The PBMCs were then injected intravenously on study day 9. The second experimental design involved irradiating mice on study day -1, followed by engraftment with PBMCs either perfectly (6/6) or partially (3/6) matched human leukocyte antigens (HLA) with the tumors. Subsequently, SKRC-59 cells were placed under the kidney capsule on study day 0. Daily post-operative observations were conducted for six days following surgery to ensure the well-being of the mice. Tumor burden was assessed twice weekly using an in vivo imaging system, along with monitoring body weights and clinical signs. The humanization rates were evaluated 7, 14, and 21 days post-PBMC engraftment, as well as at the end of the study, using peripheral blood. Approximately 5 to 6 weeks post-tumor implantation, the mice were euthanized, and both left and right kidneys were collected and weighed for further analysis. The humanization rates were significantly higher when the PBMCs were engrafted on the same day as irradiation compared to 10 days post-irradiation. The PBMC humanization, and HLA-type PBMC donors did not affect tumor burden. Additionally, we found that distant metastasis in the lungs developed in several mice. Using flow cytometry and immunohistochemistry (IHC) to analyze tumor infiltrating leukocytes (TILs), a predominant number of CD3+ T cells were found in human CD45+ TILs, indicating that this model could be useful for evaluating T cell-mediated immunotherapies. Moreover, lympho-myeloid aggression (LMA) was observed in this peripheral blood leukocytes (PBL) mouse model, showing early stage tertiary lymphoid structures (TLS), which potentially provides a powerful tool to recapitulate TLS formation and study associated mechanisms and therapies. Our study demonstrated the growth of ccRCC SKRC-59 tumors under the renal capsule and observed TILs and metastatic patterns in PBMC humanized DKO mice. This PBMC-humanized orthotopic ccRCC xenograft model can recapitulate tumor microenvironment (TME), and be potentially used to assess the safety and efficacy of various immunotherapeutics.
Citation Format: Wonyoung Kang, Yufei Wang, Yasmin N. Laimon, Oanh Pham, Ben Matran, Lauren Bottoms, Sheng Khang, Hsien-Chi Yuan, Nithyassree Murugan, Aseman B. Sheshdeh, Li-chin Yao, Jiwon Yang, Mingshan Cheng, Sabina Signoretti, Wayne A. Marasco, James G. Keck. Establishing orthotopic renal cell carcinoma xenograft in PBMC-humanized immunodeficient NSG-MHC I/II double knock-out mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 111.