Compromised antigen binding and signaling interfere with bispecific CD19 and CD79a chimeric antigen receptor function.

Therapy with CD19-directed chimeric antigen receptor (CAR) T cells has transformed the treatment of advanced B-cell malignancies. However, loss of or low antigen expression can enable tumor escape and limit the duration of responses achieved with CAR T-cell therapy. Engineering bispecific CAR T cells that target 2 tumor antigens could overcome antigen-negative escape. We found that CD79a and b, which are heterodimeric components of the B-cell receptor, were expressed on 84.3% of lymphoma cases using immunohistochemistry, and 87.3% of CD79ab-positive tumors also coexpressed CD19. We generated 3 bispecific permutations: tandem, bicistronic, and pooled products of CD79a-CD19 or CD79b-CD19 CAR T cells and showed that bispecific CAR T cells prevented the outgrowth of antigen-negative cells in a CD19-loss lymphoma xenograft model. However, tandem and bicistronic CAR T cells were less effective than monospecific CD19 or CD79a CAR T cells for the treatment of tumors that only expressed CD19 or CD79, respectively. When compared with monospecific CAR T cells, T cells expressing a tandem CAR exhibited reduced binding of each target antigen, and T cells expressing a bicistronic CAR vector exhibited reduced phosphorylation of downstream CAR signaling molecules. Our study showed that despite added specificity, tandem and bicistronic CAR T cells exhibit different defects that impair recognition of tumor cells expressing a single antigen. Our data provide support for targeting multiple B-cell antigens to improve efficacy and identify areas for improvement in bispecific receptor designs.

Inhibition of LSD1 with Bomedemstat Sensitizes Small Cell Lung Cancer to Immune Checkpoint Blockade and T-Cell Killing.

PURPOSE: The addition of immune checkpoint blockade (ICB) to platinum/etoposide chemotherapy changed the standard of care for small cell lung cancer (SCLC) treatment. However, ICB addition only modestly improved clinical outcomes, likely reflecting the high prevalence of an immunologically "cold" tumor microenvironment in SCLC, despite high mutational burden. Nevertheless, some patients clearly benefit from ICB and recent reports have associated clinical responses to ICB in SCLC with (i) decreased neuroendocrine characteristics and (ii) activation of NOTCH signaling. We previously showed that inhibition of the lysine-specific demethylase 1a (LSD1) demethylase activates NOTCH and suppresses neuroendocrine features of SCLC, leading us to investigate whether LSD1 inhibition would enhance the response to PD-1 inhibition in SCLC. EXPERIMENTAL DESIGN: We employed a syngeneic immunocompetent model of SCLC, derived from a genetically engineered mouse model harboring Rb1/Trp53 inactivation, to investigate combining the LSD1 inhibitor bomedemstat with anti-PD-1 therapy. In vivo experiments were complemented by cell-based studies in murine and human models. RESULTS: Bomedemstat potentiated responses to PD-1 inhibition in a syngeneic model of SCLC, resulting in increased CD8+ T-cell infiltration and strong tumor growth inhibition. Bomedemstat increased MHC class I expression in mouse SCLC tumor cells in vivo and augmented MHC-I induction by IFNγ and increased killing by tumor-specific T cells in cell culture. CONCLUSIONS: LSD1 inhibition increased MHC-I expression and enhanced responses to PD-1 inhibition in vivo, supporting a new clinical trial to combine bomedemstat with standard-of-care PD-1 axis inhibition in SCLC.

Immunogenic Chemotherapy Enhances Recruitment of CAR-T Cells to Lung Tumors and Improves Antitumor Efficacy when Combined with Checkpoint Blockade.

Adoptive therapy using chimeric antigen receptor-modified T cells (CAR-T cells) is effective in hematologic but not epithelial malignancies, which cause the greatest mortality. In breast and lung cancer patients, CAR-T cells targeting the tumor-associated antigen receptor tyrosine kinase-like orphan receptor 1 (ROR1) infiltrate tumors poorly and become dysfunctional. To test strategies for enhancing efficacy, we adapted the KrasLSL-G12D/+;p53f/f autochthonous model of lung adenocarcinoma to express the CAR target ROR1. Murine ROR1 CAR-T cells transferred after lymphodepletion with cyclophosphamide (Cy) transiently control tumor growth but infiltrate tumors poorly and lose function, similar to what is seen in patients. Adding oxaliplatin (Ox) to the lymphodepletion regimen activates tumor macrophages to express T-cell-recruiting chemokines, resulting in improved CAR-T cell infiltration, remodeling of the tumor microenvironment, and increased tumor sensitivity to anti-PD-L1. Combination therapy with Ox/Cy and anti-PD-L1 synergistically improves CAR-T cell-mediated tumor control and survival, providing a strategy to improve CAR-T cell efficacy in the clinic.