Immunologic determinants in procured melanoma tissue as drivers of durable response to tumor-infiltrating lymphocyte therapy.

Abstract

9511 Background: Predictive biomarkers that identify melanoma tumors capable of generating therapeutically effective tumor-infiltrating lymphocytes (TIL) remain inadequately defined. We conducted an integrated spatial and histopathologic evaluation of resected melanoma specimens to characterize tumor microenvironmental features associated with durable clinical benefit (≥12 months) and improved survival following TIL therapy. Methods: Data were extracted from early-phase clinical trials conducted at Moffitt Cancer Center enrolling patients with advanced melanoma treated with TIL. Tumor specimens from 45 of 50 patients undergoing TIL manufacturing were evaluable based on tissue availability. Histopathology, multiplex immunofluorescence, and NanoString GeoMx spatial transcriptomics were performed on procured tumors. Tertiary lymphoid structures (TLS) were defined as organized aggregates of CD3⁺ T cells and CD20⁺ B cells and classified as immature or mature based on CD21/CD23 expression. Associations with ex vivo TIL expansion, infused product composition, clinical response, progression-free survival (PFS), and overall survival (OS) were assessed. Results: Procurement sites included subcutaneous/soft tissue (78%), lymph node (18%), and lung (4%). Viable tumor content and necrosis were not associated with ex vivo TIL expansion or response. TLS were present in 56% (25/45) of tumors and were strongly associated with clinical response (18/45 responders overall; response rate 56% [14/25] in TLS-positive tumors vs 20% [4/20] in TLS-negative tumors; p=0.014), improved PFS after adjusting for procurement site (HR=0.32; p=0.005) and improved OS (p<0.0001). Evaluation of ex vivo TIL expansion demonstrated that TLS presence did not affect the total number of expanded TIL (p = 0.3) but was significantly associated with a higher proportion of CD8⁺ T cells within the infusion product (83% vs 56%, p=0.027). Spatial transcriptomics of 132 immune-enriched regions of interest demonstrated that responders exhibited marked upregulation of antigen-presentation and interferon-response programs, including MHC I/II genes, STAT1/2, IFI6, and CXCL9 . A patient-level immune activation score derived from the top 25 responder-associated transcripts correlated with improved PFS (p<0.0001). To independently validate the significance of this immune gene set score, we analyzed an external dataset from the Lauss et al., 2017 cohort. Consistent with our findings, the gene set score was significantly higher in TIL responders compared with non-responders (p=0.048). Conclusions: TLS and spatial immune activation signatures in procured melanoma tumors are strongly associated with response and survival following TIL therapy. These findings support the integration of tumor microenvironment biomarkers into patient selection and optimization strategies for TIL therapy.