Immune Cell Stimulation Primes Tumor Cells for Radiation Effect

Abstract

<h3>Purpose/Objective(s)</h3> Only fifteen years ago, renal cell carcinoma (RCC) was considered chemotherapy and radiation resistant, with surgery as the exclusive modality of treatment. Even today, with the advances of immunotherapy extending treatment options for RCC, adjunctive radiation is used sparingly. In the metastatic treatment setting there is a growing interest in adding stereotactic radiation to potentially enhance the effects of immunotherapy via the abscopal effect. The activity of radiation to be applied for both local and systemic effects is underdeveloped in the RCC treatment strategies. In this study we developed patient derived organoid (PDO) cultures from renal cell carcinoma tumors (RCC) to test the hypothesis that previously radiation resistant cultures can be sensitized with immune cell activation. <h3>Materials/Methods</h3> We analyzed PDO cultures generated from fresh nephrectomy samples and grown in 3D Matrigel culture. These PDO cultures contain a tumor and stromal cells as well as immune cells that are representative of the kidney tumors they are derived from. PDO cultures were immune stimulated with anti-CD3 and anti CD-28 antibodies for 3-5 days to target T-cells for activation and irradiated at peak stimulation on day 4-5. Applying a range or radiation dosages, following irradiation, PDO are incubated in culture for an additional 72hrs before further analysis. Using flow cytometry and a luminescent cell viability assay, we analyzed stimulated and unstimulated immune cell phenotype and characterized the remaining tumor cell populations. <h3>Results</h3> We observed that ccRCC tumor PDO cultures are highly resistant to radiation. At all tested dose levels, without stimulation both immune and tumor cell populations are minimally affected by radiation. There is a small decrease in intra-tumor CD45+ cells that is not reflected in subsequent CD3+, CD4+, and CD8+ populations, as these cells are unchanged. However, when antibodies are used to induce immune cell activation, both tumor and immune cell populations display increased sensitivity to radiation. In our experiments we observed ∼ 25% reduction in tumor cell populations (CD45-) and ∼65% reduction in immune cells (CD45+). Subsequent phenotyping of CD3+, CD4+, and CD8+ populations showed similar reductions. <h3>Conclusion</h3> Using a localized multicellular primary tumor model system, we showed that immune stimulation promoted the sensitivity of nearby tumor cells to the effects of ionizing radiation. These results have important implications for the combined use of immune-based therapies and radiation modalities.