Dual-action nanotherapy: Temozolomide-loaded, anti-PD-L1 scFv-functionalized lipid nanocarriers for targeted glioblastoma therapy

• A novel dual-action nanostructured lipid carriers combining chemotherapy and immunotherapy is developed for glioblastoma treatment • Anti-PD-L1 scFv functionalization ensures active targeting of tumor cells overexpressing PD-L1, enhancing therapeutic specificity • Conjugated scFv successfully binds to PD-L1 expressions without any binding efficiency loss • These NLCs demonstrate effective blood-brain barrier (BBB) penetration Glioblastoma (GBM) is a highly malignant brain tumor with limited treatment options and poor prognosis. GBM exhibits resistance to conventional therapies, including temozolomide (TMZ), radiotherapy, and immunotherapy, partly due to immunosuppressive mechanisms such as programmed death-ligand 1 (PD-L1) overexpression. To address these challenges, we developed TMZ-loaded nanostructured lipid carriers (NLCs) conjugated with anti-PD-L1 single-chain variable fragments (scFv) for dual chemo-immunotherapy. The anti-PD-L1 scFv enabled active targeting of PD-L1-expressing tumor cells while mitigating immune evasion, and the NLCs efficiently crossed the blood-brain barrier (BBB), delivering TMZ to the tumor site. In vitro studies confirmed nanoparticle internalization by GL261 glioma cells and specific binding of the conjugated scFv to PD-L1. In vivo studies using the GL261/C57BL/6 mouse model demonstrated that TMZ-NLCs conjugated with anti-PD-L1 scFv (TMZ-NP-scFv) stabilized tumor growth by the third week, unlike other treatment groups. While all therapeutic groups (TMZ solution, TMZ-NP, and anti-PD-L1 scFv alone) significantly improved survival compared to controls, the TMZ-NP-scFv group exhibited the most pronounced survival benefit. These results highlight the potential of TMZ-loaded, PD-L1-targeted NLCs as a synergistic strategy to enhance GBM treatment by combining chemotherapy and immune checkpoint blockade.