Proof of concept and mechanistic evaluation of lbl-034, a potent GPRC5D-targeting T cell engager with affinity-optimized, sterically engineered anti-CD3 arm, in a multiple myeloma xenograft mouse model

Abstract Introduction T-cell mediated therapies have demonstrated promising efficacy in multiple myeloma (MM), exemplified by the recent US approval of talquetamab for relapsed/refractory multiple myeloma (RRMM). LBL-034 is a novel, differentiated bispecific antibody (fully humanized IgG1) that targets GPRC5D and CD3. Designed as a T-cell engager (TCE), LBL-034 incorporates two bivalent GPRC5D-binding domains with high avidity and a low-affinity, sterically masked anti-CD3 single-chain variable fragment (scFv). In vitro studies show that LBL-034 induces conditional T cell activation, characterized by the weak CD3 binding and minimal cytokine release in the absence of GPRC5D-expressing target cells. These properties position LBL-034 as a promising TCE for the treatment of MM. Currently, LBL-034 is under investigation in a Phase Ⅰ/Ⅱ clinical trial in patients with RRMM (NCT06049290). Methods A murine model of MM was established by implanting MM1.s cells, which exhibit high GPRC5D expression, along with human peripheral blood mononuclear cells (PBMCs) in humanized NSG mice. Fifteen days post-tumor inoculation, mice were randomized into different treatment groups. Each group received intravenous administration of either PBS (control), LBL-034, or talquetamab at doses of 0.05 mg/kg, 0.3 mg/kg or 1.5 mg/kg. Tumor growth inhibition was calculated as the percentage difference in mean tumor volumes between treated and control group. LBL-034 exposure in serum and tumor tissues was quantified using an ELISA-based ligand binding assay. Pharmacodynamics (PD) assessments of T cell subtypes within the tumor were performed using immunohistochemistry (IHC) and flow cytometry. Multicolor flow cytometry panels (hCD45, hCD3, hCD4, hCD8, hCD25, hCD19, hCD138, hGPRC5D) were used to detect MM1.s cells in serum or tumor samples. IHC staining was employed to evaluate tumor-infiltrating T cells (hCD3, hCD4, hCD8) and GPRC5D-positive MM1.s cells. Additionally, serum levels of IFN-γ and soluble BCMA (sBCMA) were measured using commercial assay kits. Results LBL-034 exposure at 1.5 mg/kg in tumor tissues was approximately 10% of that measured in serum, aligning with exposure levels observed at clinically efficacious doses above 400 µg/kg in humans. In serum, LBL-034 demonstrated a longer half-life than talquetamab (23.2 h vs 17.4 h). Efficacy outcomes favored LBL-034, with complete tumor regressions observed in 3 of 4 treatment groups, whereas talquetamab achieved no complete regression at 0.3 mg/kg. Both antibodies produced 100% complete tumor regression at 1.5 mg/kg. Notably, LBL-034 induced tumor responses earlier, as soon as Day 5 compared to Day 7 with talquetamab. A dose-dependent reduction in sBCMA levels was also observed, correlating well with the efficacy outcomes. In terms of cytokine release, LBL-034 induced significantly lower levels of serum IFN-γ—approximately 3.0-fold and 4.6-fold lower at 0.3 mg/kg and 1.5mg/kg compared to talquetamab, indicating a lower risk of cytokine release syndrome (CRS). This reduction reflects LBL-034's unique molecular design, which enables conditional T cell activation. Following the initial 1.5 mg/kg dose of LBL-034, there was a marked activation of CD4+ (12.0-fold) and CD8+ (18.5-fold) T cells, along with increased CD3+ (3.2-fold), CD4+ (4.0-fold) and CD8+ (3.1-fold) T cells infiltration into tumors, as measured by both flow cytometry and IHC. In addition, GPRC5D+MM1.s cells were reduced by 98.1% after the second LBL-034 dose at 1.5 mg/kg, as confirmed by flow cytometry and further supported by IHC analysis. Conclusions LBL-034 demonstrated complete tumor regression at 1.5 mg/kg and effectively validated its mechanism of action in the MM xenograft mouse model. Compared to talquetamab, LBL-034's unique molecular design facilitated a faster anti-tumor response, reduced risk of CRS, and an extended half-life. These preclinical findings have been further supported by results from the ongoing Phase Ⅰ/Ⅱ clinical trial.