Chimeric antigen receptor (CAR) T cell immunotherapy aims to redirect a patient’s T cells to recognize and destroy malignant cells. These cells are genetically altered to express CARs that combine an antigen recognition domain with intracellular T cell signaling components to deliver T cell specificity and functionality. By using CAR T cells, researchers can specifically target antigen-expressing cells and then initiate signaling pathways that trigger effector T cell function. In theory this seems great, right? However, CAR T cell therapy exhibits on-target off-tumor (OTOT) toxicity. In OTOT toxicity, therapeutics, drugs, or cells, do exactly what they’re meant to do, just in the wrong place or on the wrong cell. Therefore, controlling OTOT toxicity during CAR T cell therapy is one of the most important current challenges for optimal success of this new treatment strategy. Meeting this goal was exactly the aim of the new study by Dr. Stanley Riddell, a professor in the Translational Science and Therapeutics Division, and led by Dr. Tamer Basel Shabaneh, a postdoctoral fellow in Dr. Riddell’s lab.
An attractive option for CAR T cell therapy is targeting the human epidermal growth factor receptor 2 (HER2), which, while expressed at low levels in normal epithelial cells, is overexpressed in several epithelial cancers. However, HER2-targeted CAR T cells pose a risk for OTOT toxicity because it is expressed in normal cells. In this new study, Basel examined different HER2 CARs to identify which one does not elicit toxicity in normal cells but does kill the target cells. To test this, Basel “developed an immunocompetent tumor model of CAR T-cell therapy targeting murine HER2 (mHER2) and examined the effect of CAR affinity, T-cell dose, and lymphodepletion on safety and efficacy.” This was done because “murine models have employed strategies to express human HER2 (hHER2) but fail to recapitulate physiologic expression levels and tissue locations of HER2 in humans.” The research team engrafted the mice with murine lung cancer cells that were transduced with the ectodomain of mHER2, for high expression of HER2. Using this model, Basel found that “the development and severity of OTOT toxicity is influenced by HER2 CAR affinity, cell dose, and intensity of lymphodepletion.”
Structurally, the antigen-targeting portion of the CAR is an extracellular single-chain variable immunoglobulin fragment (scFv) that is then fused to intracellular signaling domains, which allows for T cell activation upon ligand binding. To identify a good candidate for designing the CAR extracellular domain, Basel screened for anti-mHER2 antibodies based on affinity and specificity. Once the antibodies were identified, they were used to construct the mHER2 CAR T cells. Selecting a low and high affinity mHER2, the team generated the CAR T cells and evaluate their ability to promote T-cell proliferation, cytokine production and cytotoxicity when these CAR T cells were co-cultured with mHER2+ tumor cell in vitro, in addition to their safety and efficacy in vivo models.
Surprisingly, the team found that the low affinity mHER2 CAR T cells targeted tumors cells that expressed high levels of HER2, without toxicity to low-HER2 expressing normal cells. In contrast, high affinity mHER2 CAR T cells elicited toxicity and did not have anti-tumor activity. “These data indicate that despite superior recognition of HER2 in vitro, high-affinityHER2-targeted CAR T elicited overt, moderate-to-severe toxicity, acquired an exhaustion signature, and failed toimprove antitumor activity against HER2+B16 F10 tumors in vivo,” the authors stated. Moreover, the team found that high-affinity HER2-targeted CAR T-induced toxicity correlated with CAR T dose, lymphodepletion intensity, and systemic proliferation of CAR T and it was independent of the presence of HER2 on the tumors.
Basel’s study demonstrated that “targeting tumors that express high levels of HER2 can be achieved using low-affinity CAR T cells,” without “eliciting discernible toxicity, identifying a potential safe approach for treating HER2-high tumors.” The authors hope that this study shows the “general principles to guide the development of safe CAR T may be elucidated from preclinical models.”
Fred Hutch/University of Washington/Seattle Children’s Cancer Consortium member Dr. Stanley Riddell contributed to this work.
This work was supported by grants from the National Institute of Health, the Department on Defense, and the Lyell immunopharma.
Shabaneh TB, Stevens AR, Stull SM, Shimp KR, Seaton BW, Gad EA, Jaeger-Ruckstuhl CA, Simon S, Koehne AL, Price JP, Olson JM, Hoffstrom BG, Jellyman D, Riddell SR. Systemically administered low-affinity HER2 CAR T cells mediate antitumor efficacy without toxicity. J Immunother Cancer. 2024 Feb 7;12(2):e008566