Beyond CAR-T: Multi-layered engineering strategies shaping the next generation of cellular cancer immunotherapy

Abstract

Adoptive cellular immunotherapy has transformed the treatment landscape for hematologic malignancies; however, its broader application to solid tumors remains limited by antigen heterogeneity, inadequate persistence, restricted tumor infiltration, and the profoundly immunosuppressive tumor microenvironment (TME). While conventional CAR-T cell therapy has demonstrated remarkable clinical success, no single immune-cell platform is likely to overcome all biological barriers encountered in complex solid tumors. Emerging evidence instead supports a shift toward an integrated cellular immunotherapy ecosystem that combines the complementary strengths of multiple engineered immune cell populations. In this review, we present a unifying framework for the multi-layered engineering of next-generation immune cell therapies, encompassing CAR-natural killer (CAR-NK) cells, TCR-engineered αβ T cells, γδ T cells, CAR-natural killer T (CAR-NKT) cells, and CAR-macrophages. Rather than considering these platforms as isolated technologies, we examine how shared engineering strategies, including cytokine armoring, CRISPR-based genome editing, protein-level receptor optimization, tumor-responsive gene circuits, metabolic reprogramming, and induced pluripotent stem cell (iPSC)-based manufacturing, can be integrated to enhance persistence, functionality, safety, and resistance to TME-mediated suppression. We further discuss the biological rationale, preclinical advances, and emerging clinical evidence supporting each platform, highlighting their distinct and complementary mechanisms of tumor recognition and immune activation. Finally, we propose that the future of cancer immunotherapy lies not in the optimization of a single cellular modality, but in the development of synergistic, scalable, and off-the-shelf engineered immune-cell networks capable of delivering durable and effective responses against heterogeneous solid tumors. This graphical abstract illustrates an integrated framework for the rational engineering of next-generation immune cell therapies aimed at overcoming the major barriers of solid tumor treatment. At the center, the immunosuppressive tumor microenvironment (TME)—characterized by antigen heterogeneity, hypoxia, metabolic stress, and inhibitory cellular and molecular networks (e.g., PD-L1, TGF-β, Tregs, MDSCs, and CAFs)—represents the primary obstacle to durable therapeutic responses. Surrounding this core are diverse immune effector platforms, including CAR-NK, TCR-T, γδ T, CAR-NKT, and CAR-macrophage cells, each contributing complementary mechanisms such as antigen-specific cytotoxicity, MHC-independent recognition, phagocytosis, immune bridging, and TME remodeling. The outer layer highlights advanced engineering strategies that enhance therapeutic performance, including cytokine armoring, CRISPR-based genome editing, signaling optimization, tumor-responsive promoters, iPSC-derived manufacturing platforms, and metabolic reprogramming. Together, these innovations aim to improve persistence, resistance to immunosuppression, tumor infiltration, and scalability for off-the-shelf production. The proposed multi-layered design strategy envisions a transition from conventional single-platform CAR-T therapies toward a coordinated cellular immunotherapy ecosystem capable of achieving safer, more effective, and durable responses against heterogeneous solid tumors while addressing the remaining challenges of persistence, manufacturing standardization, and TME resistance.