Abstract 3511: A T cell engager (TCE), HB134, targeting LILRB4 and CD3, exhibits potent anti-tumor activity with a favorable safety profile for monocytic AML

Abstract Background: Acute myeloid leukemia (AML), is a heterogeneous and highly aggressive malignancy, characterized by poor prognosis and limited treatment options. Monocytic AML, including M4 and M5 subtypes, presents a significant clinical challenge due to its dismal prognosis and high relapse rates post-transplant. The leukocyte immunoglobulin-like receptor B4 (LILRB4) is overexpressed in monocytic AML but is absent in normal hematopoietic stem cells, highlighting its potential as a therapeutic target. Here, we report the development of a novel TCE targeting LILRB4 and CD3, HB134, for the treatment of monocytic AML. Methods: The HB134 TCE was generated by fusing an anti-LILRB4 nanobody and an anti-CD3 Fab to a human IgG1 with a silent Fc region. Surface plasmon resonance, enzyme-linked immunosorbent assay, and flow cytometry were used to measure HB134’s binding activities to LILRB4 and CD3. Luciferase-based cellular reporter assays and a primary T cell-dependent cellular cytotoxicity (TDCC) assay using peripheral blood mononuclear cells (PBMCs) were employed to assess HB134’s T cell activation activity. Additionally, in vivo anti-tumor efficacy and pharmacokinetic (PK) profile were evaluated in mouse models. Results: HB134 exhibited sub nanomolar high binding affinity for human LILRB4 and low binding affinity to human CD3. In the cellular luciferase reporter assays, in the presence of LILRB4-positive monocytic AML cells (THP-1 or MOLM-13 cell line), HB134 induced robust T cell activation; in contrast, no T cell activation was observed in the presence of LILRB4-negative cells, indicting HB134 can specifically induce T cell activation in a LILRB4-engagement-dependent manner. In the TDCC assays using PBMCs, HB134 induced potent cytotoxicity against LILRB4-positive THP-1 and MOLM-13 cells with limited induction of cytokine release syndrome-related proinflammatory cytokines, including IL-6 and TNF-α. In an immunodeficient mouse model engrafted with human PBMCs and LILRB4-positive THP-1-luciferase cells, HB134 showed potent anti-tumor efficacy. Additionally, HB134 demonstrated a favorable PK profile in BALB/c mice. Conclusions: HB134, a TCE designed to engage T cells via CD3 and specifically target LILRB4, exhibits potent anti-tumor activity and favorable PK and safety profiles, positioning HB134 as a promising therapeutic candidate for treating LILRB4-positive monocytic AML. Citation Format: Juan Liu, Fangfang Ren, Chunmei Liu, Jing Shao, Fang Yang, Jianhe Chen, Bin Ye, Jianhua Sui. A T cell engager (TCE), HB134, targeting LILRB4 and CD3, exhibits potent anti-tumor activity with a favorable safety profile for monocytic AML [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3511.