Immune-cytokine signature predicts survival in patients with advanced melanoma treated with oncolytic adenovirus TILT-123 and chemotherapy and IL-2-free adoptive TIL therapy.

Metastatic melanoma resistant to immune checkpoint inhibitors remains difficult to treat, and while adoptive tumor-infiltrating lymphocyte (TIL) therapy has shown durable responses, its reliance on lymphodepleting chemotherapy and high-dose interleukin (IL)-2 causes toxicity that may limit patient eligibility. The dual-cytokine-armed oncolytic adenovirus igrelimogene litadenorepvec (TILT-123) was administered with TILs in the TUNINTIL trial (trial registration: NCT04217473) in patients with metastatic melanoma resistant to immune checkpoint inhibitors, without lymphodepleting chemotherapy or IL-2 post-conditioning. This study presents a correlative immunological analysis of the phase I TUNINTIL trial evaluating TILT-123 in combination with TIL therapy. The TUNINTIL trial was a first-in-human, open-label, dose-escalation, multicenter, multinational phase I trial. 17 patients with checkpoint-inhibitor-resistant metastatic melanoma received up to six intratumoral TILT-123 injections followed by TIL infusion, without lymphodepleting chemotherapy or IL-2 post-conditioning. Systemic immune profiling (serum proteomics, flow cytometry, interferon-γ ELISpot assay), intratumoral immune cell-cell profiling (multiplex immunofluorescence, H&E, adenovirus E1a immunohistochemistry), quantitative PCR, and neutralizing antibody responses were assessed at defined time points through the trial, with survival follow-up updated to March 2026. Response criteria were evaluated using Response Evaluation Criteria in Solid Tumors V.1.1 and positron emission tomography-based criteria. Statistical analyses included Kaplan-Meier survival with log-rank tests, Mann-Whitney U tests, Pearson correlation, and receiver operating characteristic/area under the curve analysis for biomarker cut-off determination. Tumor biopsy analyses revealed an early innate immune activation marked by natural killer-cell expansion and cytotoxic gene upregulation, followed by increased intratumoral T-cell infiltration. This occurred without lymphodepleting chemotherapy or post-conditioning IL-2. Intratumoral viral DNA was detectable in a subset of patients. The enrichment of CD27+CD28+ memory-precursor CD8+ T cell was associated with favorable clinical outcomes. Elevated monocytic myeloid-derived suppressor cells and angiogenic/inflammatory cytokines following combination treatment were associated with disease progression, highlighting the role of immunosuppressive myeloid subsets as potential mediators of therapeutic resistance. Additionally, correlative analysis in pooled TILT-123 cohorts identified serum epidermal growth factor as a candidate biomarker for stratifying and monitoring patients. These findings provide mechanistic insights into TILT-123 combined with TIL therapy and propose future directions for biomarker-guided clinical studies. NCT04217473.