Enhanced melanoma-specific cytotoxic T-cell expansion and the clinical outcomes with simlukafusp alfa and pembrolizumab following checkpoint inhibitor progression

Background Despite advances in checkpoint inhibitor (CPI) therapy, a substantial proportion of patients with advanced melanoma experience disease progression, and the capacity of CD8 + cytotoxic T lymphocytes (CTLs) specific for wild-type melanoma-associated antigens to persist and remain functionally inducible after CPI failure is poorly understood. Here we report exploratory translational analyses from the BP41054 Phase I trial investigating the combination of simlukafusp alfa (FAP-IL2v), a tumor stroma-targeted IL-2 variant, with pembrolizumab in CPI-experienced patients with advanced melanoma.MethodsPeripheral blood mononuclear cells from a subset of 33 CPI-experienced patients were profiled at baseline (C1D1_pre) and on treatment (C2D8) using quantitative immunophenotyping and a panel of 28 melanoma-associated peptide–HLA tetramers spanning 14 antigens and multiple HLA alleles. Antigen-specific CTLs were further characterized functionally by ex vivo peptide challenge and interferon (IFN)-γ intracellular staining. Associations with progression-free survival (PFS) were assessed as exploratory endpoints.Results Treatment was associated with increased circulating natural killer cells and CD8 + T cells, including effector memory and terminally differentiated effector subsets, as assessed in peripheral blood. CTLs specific for 11 unique peptide-major histocompatibility complexes were identified, with increased frequencies detected during treatment in a subset of patients. These antigen-specific CTLs demonstrated functional activity as indicated by IFN-γ production on peptide challenge, though responses were variable across peptide specificities and patients, and direct cytotoxic activity was not assessed. Signs of incomplete functional rescue, including persistent co-expression of exhaustion markers, were observed. In exploratory descriptive analyses, patients whose antigen-specific CTL diversity increased on treatment had a longer median PFS than those in whom diversity was reduced; with three patients per group, these outcomes are reported descriptively, without formal statistical comparison. Baseline melanoma-associated immunity did not predict clinical outcome.Conclusions These hypothesis-generating findings indicate that melanoma-specific CD8 + T-cell immunity persists and remains inducible in CPI-experienced patients, supporting the rationale for strategies that re-engage residual antitumor immunity. Validation in larger cohorts with paired tumor and blood sampling will be required to establish clinical relevance.