A bio-orthogonally engineered probiotic-T cell chimera amplifies adoptive cell therapy through spatial DC-T cell immune licensing.

Abstract

Adoptive cell transfer (ACT) has achieved durable clinical responses in hematological malignancies, yet remains limited in solid tumors owing to poor T cell infiltration, inadequate persistence, and tumor-driven immunosuppression. Here, we report a bio-orthogonally engineered probiotic-T cell chimera (T-FOLactis) that creates a mutually reinforcing cellular-microbial system, in which FOLactis, an engineered Lactococcus lactis expressing a Flt3L-OX40L fusion protein, provides localized immunostimulatory cues to enhance T cell activation and effector function, while adoptively transferred T cells serve as cytotoxic effectors and mobile carriers that facilitate delivery of the bacterial payload to the tumor microenvironment. Using strain-promoted azide-alkyne cycloaddition chemistry, FOLactis bacteria were covalently anchored onto the surface of T cells, generating a cell-bound platform for spatially restricted immune modulation. In syngeneic colorectal cancer models, T-FOLactis reduced tumor burden by 78.1% compared with saline-treated controls and by 65.0% relative to conventional ACT, while prolonging median survival from 22 to 46 d without overt systemic toxicity. Mechanistically, T-FOLactis promoted dendritic cell (DC)-T cell proximity and established a spatially organized immune-licensing niche that amplified the functional impact of FOLactis-induced inflammatory cues, including IL-18. This spatial configuration was associated with enhanced coupling of cytokine availability, DC maturation, local co-stimulation and cytotoxic CD8+ T cell effector programming. IL-18 blockade impaired this licensing program and reduced therapeutic benefit, supporting IL-18 as a key functional mediator within the DC-T cell licensing niche. Together, these findings establish a modular cell-surface engineering strategy for augmenting ACT in colorectal cancer through coordinated immune network engagement.