Abstract 1767: Biodistribution of zoledronate and effects on gd PSCA-CAR T cells in a model of bone metastatic prostate cancer
Authors: Tordesillas Leticia , Cianne Junior , Frieling Jeremy S. , Bustos Xiomar , Lynch Conor C. , Abate-Daga Daniel 
Published: April 2023

Abstract
Background: Metastatic castrate resistant prostate cancer (mCRPC) is frequently manifested in the bone, leading to increased morbidity and mortality. We have previously demonstrated that γδ Chimeric Antigen Receptor (CAR) T cells targeting prostate stem cell antigen (PSCA) led to significant regression of established prostate cancer cells in the bone. Regression was further increased by combination with the bisphosphonate zoledronate (ZOL), routinely administered to mCRPC patients to prevent bone loss. To further optimize the use of γδ CAR T cells for mCRPC, we aimed to determine the kinetics of γδ CAR T cell in a mouse model of bone metastatic prostate cancer, either as single treatment or in combination with ZOL. In addition, we identified the sites of systemic ZOL accumulation in an immune competent model that could potentially induce off-target effects of γδ CAR T cells.
Methods: Male NSG mice were injected with C4-2B prostate cancer cells expressing PSCA and luciferase in the left tibia, while the right tibia received PBS. ZOL (25 μg/kg) was injected every other day subcutaneously in half of the mice and was discontinued one day prior to administering T cells. When tumors were established, mice received γδ PSCA-CAR T cells, γδ untransduced (UT) T cells or were left untreated. Bone marrow from tumor-bearing tibia, tumor-naive tibia, femur, spleen and blood were recovered at multiple time points, and the number of γδ T cells was analyzed by flow cytometry. To determine systemic ZOL uptake, C57BL/6 mice were injected with PTE-82 prostate cell line in both tibias. One week after injection, mice received ZOL-AF647 every other day for 2 weeks. Bone marrow, liver, kidney, peritoneal lavage, skin-draining lymph nodes, Peyer’s patches and spleen were recovered and analyzed by flow cytometry.
Results: γδ CAR T cells showed a rapid accumulation in the bone marrow recovered from tumor-bearing tibias, with 3 times more cells than those from mice treated with γδ UT cells (p=0.0002). The number of γδ T cells peaked at 5 days post infusion and were still detected after 21 days. Increased γδ CAR T cell accumulation was not observed in tumor-free bones or in spleen or blood, suggesting a preferential localization of γδ CAR T cells at tumor sites. Treatment with ZOL did not significantly affect the number or phenotype of γδ T cells accumulated in bone. Analysis of ZOL distribution showed significant uptake by macrophages, specially from liver and peritoneal lavage (p&lt;0.0001), with an average of 30% and 70% of macrophages showing positive staining for ZOL-AF647, respectively (p&lt;0.0001).
Conclusion: γδ PSCA-CAR T cells accumulate and get activated at tumor sites with limited distribution outside the bone, while their kinetics is not affected by ZOL treatment in the NSG xenograft model. Additional studies will be necessary to determine the impact of ZOL uptake by myeloid cells on γδ CAR T cells migration.
Citation Format: Leticia Tordesillas, Junior Cianne, Jeremy S. Frieling, Xiomar Bustos, Conor C. Lynch, Daniel Abate-Daga. Biodistribution of zoledronate and effects on gd PSCA-CAR T cells in a model of bone metastatic prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1767.