"Stealth" CAR-NK Cells Can Evade Immune Detection And Reduce Side Effects

In recent years, CAR-T cell therapy has achieved remarkable success in treating blood cancers. However, it suffers from a critical weakness: the weeks-long and expensive preparation process, leaving many patients waiting for treatment and missing the optimal treatment window. Even more frustrating is that even when using a patient's own immune cells, these "modified warriors" can sometimes be accidentally attacked by other immune cells in the body, leading to the failure of the treatment.

Now, in a study published in the international journal Nature Communications, titled "Selective HLA knockdown and PD-L1 expression prevent allogeneic CAR-NK cell rejection and enhance safety and anti-tumor responses in xenograft mice," scientists from institutions including MIT and Harvard Medical School have successfully engineered "stealth" CAR-NK cells, effectively evading the host immune system and providing a more sustained anti-cancer effect.

NK cells (natural killer cells) are the human immune system's first line of defense, specialized for recognizing and eliminating cancerous and virally infected cells. Compared to CAR-T cells, CAR-NK cells offer a unique advantage. They do not cause severe graft-versus-host disease, meaning that donor cells can be safely used across different individuals. In this article, researchers set out to address the challenge of donor cell rejection. By deeply studying the interactions between immune cells, they discovered a key mechanism: CD8+ T cells are the primary cell type mediating allogeneic rejection, and this process relies on HLA class I proteins on the surface of NK cells.

Researcher Professor Rizwan Romee explained, "This is like giving the immune system a target to attack. If we can hide this target, the donor cells can become 'invisible' in the patient's body."

The research team therefore devised a solution: a "three-in-one" genetic modification strategy for NK cells. First, they identified a specific shRNA (small interfering RNA) that precisely reduces the expression of HLA-ABC on the NK cell surface while leaving another key protein, HLA-E, unaffected. HLA-E can bind to the inhibitory receptor NKG2A on host NK cells, preventing them from being accidentally killed by autoimmune cells. This "precise knockout" acts like a disguise for the NK cell's "recognition tag," concealing the target from CD8⁺ T cell attack while retaining the "protective signal" against the body's own NK cells.

Even more ingeniously, the research team integrated this "stealth component" with a chimeric antigen receptor (CAR) and an immunomodulatory protein (PD-L1 or single-chain HLA-E) into the same lentiviral vector, enabling the transformation to be completed with a single transfection. For example, CD19-CAR-NK cells targeting lymphoma can precisely recognize the CD19 protein on the cancer cell surface through the CAR, inhibit CD8⁺ T cell activity through PD-L1, or further enhance their "stealth" effect through single-chain HLA-E. This one-step production process significantly simplifies the process and lays the foundation for large-scale production of off-the-shelf cells.

Figure 1. HLA-ABC-reduced allogeneic NK cells evade killing by host CD8+ T cells. (Liu F, et al., 2025)

The effectiveness of this "stealth system" was validated in a humanized mouse model. Researchers injected mice with human lymphoma cells, followed by infusions of both standard CAR-NK cells and engineered "stealth" CAR-NK cells. Results showed that the standard CAR-NK cells were completely eliminated by the mice's CD8⁺T cells within two weeks, leading to rapid tumor recurrence. However, the "stealth" CAR-NK cells were still detectable in significant numbers three weeks later, maintaining their potent cancer-killing ability. Tumors in the mice shrank significantly, with some even disappearing completely. Even more surprising, the engineered cells also upregulated the expression of cytotoxic genes like granzyme B and perforin, while reducing the activation of exhaustion-related genes, suggesting a more sustained cancer-fighting effect.

Equally crucial is the breakthrough in safety. Cytokine release syndrome (CRS), a common and fatal side effect of CAR-T therapy, was significantly mitigated in this system. Research has found that "stealth" CAR-NK cells can reduce the release of inflammatory factors such as IL-6, IFNγ, and TNF, which are the primary culprits in CRS. In mouse serum, the engineered cells reduced levels of key inflammatory factors like IL-6 by over 60%, with some factors completely undetectable, addressing a major safety concern of immunotherapy.

The research team noted that "stealth" CAR-NK cells targeting CD19 (lymphoma) and mesothelin (ovarian cancer) have demonstrated efficacy in preclinical models and will move forward with clinical trials. Because NK cells have enhanced penetration into solid tumors, they may also have potential applications in the treatment of solid cancers such as lung and breast cancer. Even more exciting, they are exploring the use of similar technology to engineer NK cells for the treatment of autoimmune diseases such as lupus. Through their "stealth" and immune-modulating properties, NK cells can modulate abnormal immune responses while avoiding host elimination.

Reference

Liu F, et al. Selective HLA knockdown and PD-L1 expression prevent allogeneic CAR-NK cell rejection and enhance safety and anti-tumor responses in xenograft mice. Nature Communications, 2025, 16(1): 8809.