Engineering CAR-T cells for solid tumors: bispecific antigen targeting, tumor microenvironment modulation, and...

Chimeric antigen receptor T (CAR-T) cell therapy has revolutionized the treatment of hematologic malignancies, yet its efficacy in solid tumors remains limited due to antigen heterogeneity, immunosuppressive tumor microenvironments, and therapy-associated toxicities. This review highlights advances across CAR-T generations, emphasizing co-stimulatory domains and cytokine-armed TRUCKs to enhance persistence and function. Viral (lentiviral, gamma-retroviral) and non-viral (CRISPR, transposons, mRNA electroporation) delivery systems are compared for efficiency, safety, and scalability, with CRISPR enabling multiplex edits for improved specificity. Dual-targeting CARs counter antigen heterogeneity, while hypoxia-inducible and SynNotch CARs restrict activity to tumor sites. Chemokine receptor engineering enhances infiltration, and armored CARs secreting IL-12 or checkpoint inhibitors remodel the TME. Nanobody-based CAR-T cells further expand design versatility, offering improved stability, tumor penetration, and reduced immunogenicity compared with single-chain variable fragment constructs. Safety innovations include iCasp9 Suicide switches, dasatinib-controlled activation, and cytokine blockade. Clinical trials of bispecific CAR-Ts show promise, yet challenges Like manufacturing complexity and off-target effects persist. Integrating AI-driven design and Personalized neoantigen targeting may unlock CAR-T 2.0 for solid tumors, pending scalable production and regulatory harmonization.