Background MHC-E (HLA-E in humans, Qa-1 in mice) is a conserved, non-classical MHC class I molecule that is monomorphic, associates with β2-microglobulin (β2m), and presents canonical 9-mer peptides (VL9) from classical MHC-Ia signal peptides. MHC-E/VL9 complexes (NKG2A ligand) bind to the NKG2A/CD94 heterodimeric receptor on NK cells and cytotoxic T lymphocytes (CTL) to inhibit cytolytic activity. Antibodies to the NKG2A/CD94 receptor are currently being evaluated in pre-clinical and clinical studies. However, to date no studies have targeted the MHC-E/VL9 complex on tumor cells. We hypothesize that using a T-cell receptor mimicking (TCRm) antibody that binds to the MHC-E/VL9 complex can disrupt the NKG2A axis and may be a promising target for cancer immunotherapy.

Methods Here, we developed ABX002, a TCRm antibody that selectively binds to HLA-E presenting VL9 peptides. ABX002 has high affinity with a K_D ranging from 0.597 to 27.8 nM for various HLA-E/VL9 complexes. Enzyme-linked immunosorbent assays (ELISA) and flow cytometric analysis confirmed ABX002’s specificity, showing selective binding to HLA-E with VL9 peptides only. To assess the therapeutic efficacy of ABX002, a series of in vitro and in vivo experiments were conducted.

Results In vitro experiments showed ABX002 enhanced NKG2A+ NK cell-mediated lysis of target cells expressing HLA-E that were pulsed with VL9 peptides but not with control peptides, indicating the ability of ABX002 to disrupt the NKG2A-axis. Additionally, ABX002 blockade enhanced lytic activity of NKG2A+ NK and CTL to kill endogenously expressing HLA-E/VL9+ tumor cells.

A surrogate antibody (EXX-1) was used to target mouse Qa-1b/Qdm in pre-clinical settings. Tumor-bearing mice treated with EXX-1 showed significant tumor regression and improved survival rates compared to controls. Importantly, EXX-1 induced durable anti-tumor immunity and prevented tumor growth upon rechallenge. Notably, mice treated with anti-NKG2A antibody showed modest responses, suggesting differences in targeting the NKG2A ligand versus its receptor.

Additionally, EXX-1 antibody was used in combination therapy with anti-PD-(L)1 antibodies to treat tumor-bearing mice. These studies showed significantly enhanced tumor regression and improved survival rates compared to monotherapy, indicating that EXX-1 targeting the NKG2A ligand could improve the efficacy of PD-(L)1 blockade in mouse tumor models.

Conclusions Overall, our findings suggest targeting NKG2A ligand as a novel immunotherapy strategy. The precise specificity of TCRm antibodies, their ability to enhance effector-mediated cytotoxicity and induce durable immunity raise the prospect of using ABX002 as an immunotherapeutic agent. Further studies are warranted to explore the clinical applicability of ABX002 in various cancer types.

Ethics Approval All mouse experiments were controlled by the animal welfare committee (IACUC) of the University of Texas at Arlington under the guide for the care and use of laboratory animal, in accordance with the national research council.