54Allogeneic HSPC-engineered CD70-directed CAR-NKT cells for renal cell carcinoma targeting tumor, microenvironment, and alloreactive T cells

Abstract Background Renal cell carcinoma (RCC), originating from renal epithelium, is the most prevalent type of kidney cancer, accounting for over 90% of all cases. Although targeted therapies such as vascular endothelial growth factor (VEGF) and mammalian target of rapamycin (mTOR) inhibitors have improved clinical outcomes, approximately 33% of patients still progress to metastatic disease, with a 5-year survival of only 12%. These limitations highlight the urgent need for more effective and Innovative treatment options. Chimeric antigen receptor (CAR)-engineered T (CAR-T) cell therapy targeting CD70 is emerging as an attractive approach for RCC. Clinical trials investigating CD70-directed CAR-T (CAR70-T) cell therapies in RCC are currently underway. Despite their therapeutic potential, current CAR70-T cell therapies face several key limitations. Clinical responses have been modest, likely due to the inherent challenges posed by solid tumors, including antigen heterogeneity and a highly immunosuppressive tumor microenvironment (TME). To overcome these limitations, the development of potent, off-the-shelf CAR70-based cell therapies that can address RCC tumor immune evasion and TME-associated suppression is critically needed. Methods To address these challenges, we employed our previously established hematopoietic stem and progenitor cell (HSPC) gene engineering technology and a clinically guided culture method to generate allogeneic CD70-directed CAR-engineered invariant natural killer T (AlloCAR70-NKT) cells for the treatment of RCC. Through the use of a comprehensive array of experimental models, including primary RCC patient samples, patient-derived tumor cell lines, in vitro functional assays, and both orthotopic and metastatic in vivo xenograft models, we comprehensively evaluated the AlloCAR70-NKT cells, including their manufacturing, in vitro and in vivo antitumor efficacy, mechanism of action, pharmacodynamics and pharmacokinetics, safety, and immunogenicity. Results In this study, we characterize primary RCC patient samples and identify a distinct opportunity to leverage CAR-NKT cells for therapeutic intervention. Utilizing a clinically guided culture method, we successfully generated AlloCAR70-NKT cells from hematopoietic stem and progenitor cells, with high purity, robust expansion, and no fratricide risk. These cells demonstrated multimodal targeting capabilities, including potent cytotoxicity against orthotopic and metastatic RCCs via both CAR- and NK receptor-mediated mechanisms, as well as selective engagement of the immunosuppressive TME through TCR recognition. Notably, host alloreactive T cells express elevated levels of CD70 and can be efficiently targeted by AlloCAR70-NKT cells, leading to enhanced in vivo persistence of therapeutic cells. Conclusions Taken together, our findings support the therapeutic potential of AlloCAR70-NKT cells as a next-generation, off-the-shelf immunotherapy with dual tumor- and TME-targeting functionality, and the added advantage of alloreactive T cell elimination, offering a compelling strategy for treating RCC.