MYC/GSPT1 protein degradation in refractory/relapsed T-cell prolymphocytic leukemias

Abstract

Abstract Background: T-cell prolymphocytic leukemia (T-PLLs) is a rare, mature T-cell-derived, aggressive hematologic malignancy. T-PLLs are one of the most challenging hematologic malignancies based on refractoriness to conventional chemotherapy and targeted therapies, including alemtuzumab. Chromosome rearrangement and overexpression of TCL-1 are hallmarks of T-PLL, accounting for approximately 95% of cases. Nearly 50% of cases carry chromosome 8 gains, leading to upregulation of the oncogene c-Myc. Recently, we discovered a feedforward loop of c-Myc and GSPT1 (eRF3a), a key translation termination factor that promotes translation in leukemias, and established highly effective anti-leukemia/lymphoma activities of GT19630, the first-in-class MYC/GSPT1 proteolysis targeting chimera in MYC-driven hematologic malignancies (Nishida et al. biorxiv 2025 650490, under revision). We hypothesized that c-Myc/GSPT1 translation is activated in T-PLL cells and targeting c-Myc/GSPT1 by protein degradation is effective in relapsed/refractory T-PLLs. Method: We evaluated single-cell levels of baseline protein translation using the O-propargyl-puromycin (OPP) nascent protein synthesis assay and multicolor flow cytometry in T-PLL and normal peripheral blood mononuclear cells (PBMCs). We investigated the activity of GT19715 in T-PLL patient-derived xenograft (PDX) models established from relapsed/refractory T-PLL patients in vitro and in vivo. Single-cell mass cytometry (CyTOF) was applied to determine distinct surface and intracellular molecular features of primary and PDX T-PLL cells. Results: c-Myc and GSPT1 protein levels were increased in T-PLL cells compared to PBMCs. Compared to CD3+ normal T-cells from healthy donors (N = 3), CD7+ T-PLL cells in primary T-PLL samples (N = 5) exhibited significantly increased baseline protein translation levels determined by OPP protein synthesis flow cytometry (Mean fluorescence intensity: 532.3 ± 88.1 vs 7,090 ± 1,193 for CD3+ T-cells vs CD7+ T-PLL cells, respectively, P = 0.0062), suggesting increased c-Myc/GSPT1 protein translation in T-PLL cells compared to normal PBMCs. GT19715 degraded c-Myc and GSPT1 proteins at 1 nM and induced cell death in PDX T-PLL cells (IC50 < 1 nM). GT19715 dramatically reduced circulating T-PLL cells and significantly prolonged survival in mice carrying PDX T-PLL cells (median OS 1.0 vs 9.3 weeks for vehicle- and GT19715-treated groups, respectively, P < 0.0001). After euthanizing mice, we found spleen and liver packed with T-PLL cells in vehicle control mice while only minimal remaining tumor cells in liver peri-centrolobular regions, suggesting sustained tumor suppression in vivo. CyTOF analysis revealed high levels of CD2, CD4, CD7, TCL1 and ZAP70 in primary and PDX T-PLL cells compared to normal bone marrow T cells. Primary and PDX T-PLL samples share a distinct cell population with elevated levels of CD26, CD28, CD69, and CD45RA. Interestingly, these cells exhibit elevated TCL-1, ZAP70, BCL-2, MCL-1, p-AKT, p-NFkB and p-STAT5 levels with partially high Ki-67, features that suggest identification of a clone driving propagation and therapy resistance. Conclusion: We identified increased protein translation of c-Myc and GSPT1 in T-PLL cells compared to normal PBMCs. Targeted protein degradation of c-Myc and GSPT1 is highly effective in relapsed/refractory T-PLL in vitro and in vivo, with over 900% extension of survival in a clinically relevant, PDX T-PLL model, providing rationale for clinical investigations.