PRaG 5.0: Personalized thymosin alpha-1 and radioimmunotherapy for advanced refractory solid tumors—Data from an expanded cohort.

2516 Background: Treatment-related lymphopenia and inadequate anti-tumor immune activation remains major barriers to improving outcomes of combination immunoradiotherapy, especially for advanced refractory solid tumors. PRaG 5.0 innovatively integrates personalized thymosin alpha-1 (Tα-1) into the PRaG regimen (PD-1 inhibitor + radiotherapy + GM-CSF) to preserve lymphocytes and enhance anti-tumor immunity. We report updated efficacy and safety data of PRaG 5.0 in advanced refractory solid tumors. Methods: This is a prospective, single-armed, phase II study enrolled 43 heavily pretreated patients. Eligible patients were stratified by baseline T-lymphocyte counts to receive personalized Tα-1 dosing: 7-day loading dose for low baseline T-lymphocyte counts, and maintenance Ta-1 (1.6 mg, thrice weekly) for those with normal counts. Tα-1 was combined with the PRaG regimen for at least 2 cycles. After PRaG cycles, patients continued PD-1 inhibitor plus Tα-1 until disease progression or intolerable toxicity. The primary endpoint was objective response rate (ORR) per RECIST v1.1. Secondary endpoints included median progression-free survival (mPFS), disease control rate (DCR), safety, and immune cell dynamics (assessed by flow cytometry, single-cell sequencing, and TCR sequencing). Results: Among the 43 screened subjects, 39 received treatment, of whom 30 required loading dose thymosin α1 (Tα1) therapy, 36 were treated with PRaG regimen and 17 entered the maintenance phase with PD-1 inhibitor combined with Tα1. The objective response rate (ORR) of the 39 patients was 30.8% (95%CI 17.0%, 47.6%), including 4 cases of complete response (CR) and 8 cases of partial response (PR); the median progression-free survival (PFS) was 4.2 months (95%CI 2.3, 6.9). No Grade 3 or above TRAE was reported. Immune analysis revealed significant increases in CD8+ T cells, NK cells, and CD4+TEM cells post-Tα-1 loading, with a notable decrease in Tregs (p = 0.001). Single-cell sequencing demonstrated higher clonality in GZMK-CD8+T in responders, while TRDV2-CD8+T cells expanded in non-responders. Conclusions: Integrating of Tα-1 into the PRaG regimen is a promising therapeutic strategy, offering enhanced efficacy and a favorable safety profile. Detailed immune profiling identified distinct T-cell dynamics associated with treatment response, highlighting potential biomarkers for clinical application. Further validation in larger cohorts is warranted. Clinical trial information: NCT05790447 .