CAR-NK vs CAR-T: Which Cell-Based Therapy Works Better?

Cell-based immunotherapies have transformed cancer treatment, with chimeric antigen receptor (CAR) engineering enhancing immune cells’ ability to target tumors. CAR-T therapy has been widely adopted for certain blood cancers, while CAR-NK cells offer a promising alternative with distinct advantages. Understanding how these approaches compare is key to optimizing future treatments.

T Cells And NK Cells In Immunotherapy

T cells and natural killer (NK) cells play distinct roles in immunotherapy. Cytotoxic CD8+ T cells recognize malignant cells through the major histocompatibility complex (MHC), allowing for highly targeted responses but also making them susceptible to tumor immune evasion via MHC downregulation. NK cells, in contrast, function independently of MHC, using a balance of activating and inhibitory receptors to detect abnormal cells, making them effective against tumors that escape T cell surveillance.

CAR-T cells can proliferate extensively in vivo, leading to durable remissions in conditions like B-cell acute lymphoblastic leukemia (B-ALL) and diffuse large B-cell lymphoma (DLBCL). However, this prolonged activity can also cause severe side effects, such as cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS), requiring careful management. NK cells have a shorter lifespan in circulation, reducing the risk of prolonged toxicity but potentially necessitating repeated dosing or modifications to enhance persistence.

The source of these cells also impacts therapeutic applications. Autologous T cell therapies, derived from a patient’s immune system, require extensive ex vivo expansion and genetic modification, leading to high costs and long preparation times. NK cells, particularly those from umbilical cord blood or induced pluripotent stem cells (iPSCs), provide an off-the-shelf alternative with lower manufacturing complexity, enabling broader accessibility and faster deployment.

Genetic Modification Techniques For T Cells

Engineering T cells for therapy involves precise genetic modifications to improve efficacy, persistence, and safety. Viral vector-mediated gene transfer, particularly using lentiviral and retroviral vectors, is a common method for introducing CARs. Lentiviral vectors are preferred for their ability to transduce both dividing and non-dividing cells, ensuring stable CAR expression. However, concerns about insertional mutagenesis and oncogenic risks have led to alternative gene delivery methods.

Non-viral approaches, such as transposon-based systems like Sleeping Beauty and PiggyBac, offer a cost-effective alternative by enabling stable genomic integration without the biosafety concerns of viral production. Electroporation-based techniques, including CRISPR-Cas9 gene editing, allow precise modifications, such as knocking out inhibitory receptors like PD-1 to enhance CAR-T cell persistence and resistance to tumor-induced immunosuppression.

CAR design optimization has significantly influenced therapeutic outcomes. First-generation CARs contained only an antigen-binding domain linked to a CD3ζ signaling motif, providing limited activation. Second-generation CARs added costimulatory domains like CD28 or 4-1BB, improving proliferation and durability. Third-generation CARs combined multiple costimulatory signals, while fourth-generation CARs, known as TRUCKs, incorporate inducible cytokine expression to modulate the tumor microenvironment. These advancements have expanded CAR-T therapy beyond hematologic malignancies, with ongoing efforts to improve efficacy against solid tumors.

Genetic Modification Techniques For NK Cells

Modifying NK cells for therapy presents unique challenges due to their resistance to genetic manipulation and limited proliferative capacity ex vivo. Unlike T cells, NK cells exhibit lower transduction efficiencies with viral vectors, necessitating alternative strategies. While lentiviral and retroviral vectors have been used with some success, non-viral methods like electroporation and transposon systems offer scalable and safer approaches for CAR-NK engineering.

CRISPR-Cas9 gene editing through electroporation allows precise modifications without permanent genomic integration, minimizing the risk of insertional mutagenesis. However, transient gene expression requires combination with transposon-based systems like Sleeping Beauty or PiggyBac for stable integration. mRNA electroporation provides a short-lived but potent anti-tumor response without long-term genomic risks.

Optimizing CAR design for NK cells requires a different approach than for T cells. Traditional CAR constructs with CD3ζ signaling domains may not fully harness NK cell cytotoxicity, prompting the development of NK-specific CARs incorporating signaling domains like DAP12, DNAM-1, or 2B4. Co-expression of cytokines like IL-15 supports NK cell expansion and survival, addressing one of the major limitations of NK-based therapies. These modifications are particularly relevant for off-the-shelf NK cell products from iPSCs or umbilical cord blood, which require additional engineering for sustained efficacy.

Key Activation Pathways

CAR-T and CAR-NK cells rely on distinct signaling pathways for activation, persistence, and cytotoxic function. CAR-T cells activate through the CD3ζ chain, triggering downstream signaling via Lck kinase and ZAP-70 phosphorylation. This cascade activates NF-κB, MAPK, and PI3K-Akt pathways, enhancing proliferation, cytokine secretion, and cytotoxicity. Costimulatory domains like CD28 and 4-1BB refine these signals, with CD28 promoting rapid expansion and effector function, while 4-1BB enhances metabolic fitness and long-term survival.

NK cells do not rely on a single receptor but integrate signals from activating and inhibitory receptors. CAR-NK cells engage pathways involving DAP12, FcRγ, and DNAM-1, which activate key intermediates like SYK and PLCγ, leading to calcium mobilization and cytotoxic granule release. Unlike CAR-T cells, which depend on antigen presentation via MHC, NK cells use receptors like NKG2D to detect stress-induced ligands, allowing them to target tumor cells that evade T cell recognition.

Cellular Receptor Profiles

The receptor landscape of T and NK cells defines their ability to target cancer. CAR-T cells rely on an engineered receptor designed to bind a specific tumor-associated antigen, such as CD19 in B-cell malignancies. This specificity enhances tumor targeting but limits CAR-T cells to antigen-expressing cancer cells, creating vulnerabilities to antigen-negative escape variants. Endogenous T cell receptors (TCRs) remain functional, meaning CAR-T cells can still be influenced by MHC interactions and tumor microenvironment factors that suppress function.

NK cells, in contrast, possess a diverse array of activating and inhibitory receptors that dictate their response to malignant cells. Activating receptors like NKG2D, DNAM-1, and natural cytotoxicity receptors (NCRs) recognize stress-induced ligands on tumor cells. Inhibitory receptors, particularly killer-cell immunoglobulin-like receptors (KIRs) and NKG2A, prevent NK cells from attacking healthy tissues. This receptor diversity allows NK cells to recognize tumors that downregulate MHC molecules to evade T cell detection. CAR-NK therapies often incorporate NK-specific signaling domains like DAP12 or 2B4 to enhance cytotoxic responses while maintaining safety.