Expansion and characterization of tumor-infiltrating lymphocytes from primary and metastatic immune checkpoint inhibitor-exposed renal cell carcinoma tumors.

Abstract

548 Background: Adoptive cell therapy (ACT) with tumor-infiltrating lymphocytes (TILs) has demonstrated efficacy in patients with immune checkpoint inhibitor (ICI)-refractory melanoma and non-small cell lung cancer. TIL expansion from treatment-naïve and ICI-exposed melanoma tumors has shown comparable feasibility; however, expansion feasibility from ICI-exposed renal cell carcinoma (RCC) tumors has not been established. Additionally, RCC metastases remain underexplored as a tissue source for viable TILs. We investigated TIL expansion from primary and metastatic ICI-exposed RCC tumors and characterized the phenotypic and functional properties to explore the potential for TIL-based ACT in ICI-refractory RCC. Methods: Following IRB approval, surgically resected specimens from 17 patients were collected and cultured with high-dose IL-2 (6000 IU/mL) for four weeks. All patients were treated with at least one cycle of immune checkpoint inhibitor therapy. TIL expansion success was defined as at least one fragment expanding to a minimum of 2 wells. TILs from 5 randomly selected metastatic tumor fragments from 3 different patients (2 adrenal tumor fragments and 3 lung tumor fragments) underwent rapid expansion protocol (REP) to assess the feasibility of therapeutic-scale manufacturing. Pre- and post-REP TIL phenotypes and reactivity to autologous tumor were analyzed with flow cytometry and co-culture assays. Results: 11 primary and 6 metastatic RCC tumor specimens (lung n = 4, adrenal n = 1, bone n = 1) were collected. TIL expansion was successful in 10/11 (90.9%) primary tumor specimens and 6/6 (100%) metastatic specimens. Primary TILs from both the primary and metastatic RCC tumors contained significantly higher proportions of T-cells versus NK cells (p = 0.002 and p < 0.0001, respectively), with no significant difference observed in CD4+ versus CD8+ T-cell populations in either group. In co-culture assays, Pre-REP TILs from 2/5 (40%) primary and 3/5 (60%) metastatic specimens demonstrated reactivity to autologous tumor. The mean fold expansion of TILs from metastatic samples following REP was 576.3-fold. The average post-REP TIL population consisted of 59.3% CD4⁺ and 29.3% CD8⁺ T cells. 3/5 (60%) post-REP TIL samples produced TNF-a and Granzyme B, and 2/5 (40%) produced IFN-g when co-cultured with autologous tumor. Conclusions: Tumor-reactive TILs can be successfully expanded from primary and metastatic ICI-exposed RCC tumors, with lymphocyte populations comprised predominantly of T-cells. Post-REP TILs from metastatic ICI-exposed RCC specimens exhibited retention of tumor-specific reactivity via cytokine production.