A Novel Small Molecule Drug-Based Approach for Treating Beta-Globinopathies

Liu, Jun

Beta-globinopathies, such as β-thalassemia and sickle cell disease, present significant global health challenges. Existing treatments, including γ-globin inducers, blood transfusions, gene therapies, and bone marrow transplantation, face hurdles of cost, efficacy, and tolerability. In addition to the known function of BCL11A, recent research underscores the crucial role of ZBTB7A/LRF, another transcriptional factor, in suppressing γ-globin during the erythroid globin switch. While transcription factors have historically been deemed "undruggable”, recent advancements in therapeutic modalities, like molecular glue degraders, have spurred interest in targeting them. Leveraging nearly two decades of expertise in transcription factor biology, access to compound libraries, and collaborations, we identified and characterized a novel small molecule drug that is capable of downregulating both ZBTB7A and BCL11A, two known γ-globin repressors.

Through in silico and phenotypic screens, we discovered SH6, which was originally characterised as a novel small molecule ZBTB7A degrader but later found to also downregulate BCL11A levels. The globin induction effect of SH6 was evaluated and validated using various erythroid cell models, including HUDEP2 cells, normal CD34 cell-derived erythroid cells and sickle cell patient iPSC models. γ-globin induction was assessed by qPCR, flow cytometry, and clinical grade HPLC analyzer. Mechanistic studies explored SH6's dependence on the proteasome machinery and the Cereblon E3 ligase system to explore the drug's mechanism of action as a molecular glue degrader. Synergistic effects with other disease-modifying agents, such as hydroxyurea, decitabine, and pomalidomide, were investigated.

Treatment with SH6 (0.25uM) led to a 6-fold increase in γ-globin expression in HUDEP2 cells compared to DMSO, surpassing other γ-globin-inducing treatments and without causing significant toxicity or erythroid differentiation. In healthy donor CD34 stem cell-derived erythroid cells, SH6 induced significant functional hemoglobin F without affecting differentiation markers at 0.25uM (11% vs 34% HbF, compared to DMSO by HPLC). Preliminary evidence suggests SH6's potential efficacy in hydroxyurea-unresponsive sickle cell patients, with increased trend of γ-globin expression observed with flow cytometry. SH6 effectively downregulated/degraded ZBTB7A/BCL11A in a proteasome-dependent manner within 24 hours of treatment, and its effect is abrogated in a CRBN-E3 ligase knockout HUDEP2 cell line. Synergistic effects with Decitabine were notable, showing a 4-fold increase in mRNA expression compared to SH6 alone and a 9-fold increase compared to Decitabine alone. Additionally, SH6 demonstrated the unique ability to reactivate more "primitive" forms of globins, epsilon and zeta globins, suggesting potential relevance for treating both alpha and beta thalassemia.

Disclosures: No relevant conflicts of interest to declare.

See more of: 113. Sickle Cell Disease, Sickle Cell Trait, and Other Hemoglobinopathies, Excluding Thalassemias: Basic and Translational: Identification of New Molecular Targets to Modulate Sickle Cell Disease
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172 A Novel Small Molecule Drug-Based Approach for Treating Beta-Globinopathies