Development and characterization of a potent and selective BLIMP1 protac as a novel, first-in-class therapy for multiple myeloma

Abstract

Abstract Introduction: B lymphocyte-induced maturation protein-1 (BLIMP1) encoded by the PRDM1 gene, is a transcriptional corepressor essential for plasma cell differentiation and survival. While BLIMP1's tumor suppressor function in lymphoma is well established, recent publications and genome-wide CRISPR dropout screens, including those from DepMap, have identified BLIMP1 as one of the top genetic dependencies in multiple myeloma. However, the role of BLIMP1 as a therapeutic target has remained elusive, as BLIMP1 has long been considered an undruggable target. Results: Through innovative screening and medicinal chemistry efforts, we identified potent and selective BLIMP1 ligands that were subsequently developed into first-in-class PROTACs. Western blot and proteomic analysis demonstrated these PROTACs are highly potent (induce over 90% maximal BLIMP1 degradation with a DC50 ≤ 1 nM) and remarkably selective over other PRDM family members. To understand downstream consequences of BLIMP1 loss, we performed RNA-seq analysis of myeloma lines treated with BLIMP1 PROTACs. This analysis revealed significant transcriptional changes in cell cycle genes (including downregulation of MYC and upregulation of CDKN2b), as well as stark modulation of the unfolded protein response pathway (downregulation of XBP1, IRE1 and ATF4). We next investigated the functional impact of BLIMP1 loss on multiple myeloma survival. Using inducible CRIPSR knockout systems, we found that BLIMP1 inhibits a subset of myeloma lines, distinguishing between BLIMP1-dependent and -independent backgrounds and corroborating DepMap data. Consistently, BLIMP1 PROTACs were only active in BLIMP1 genetically dependent myeloma lines without affecting BLIMP1-independent myeloma lines or other hematological cell types. Specifically, BLIMP1 PROTACs showed ≥50% growth inhibition in 11 out of 28 myeloma lines, with the most sensitive lines exhibiting G1 arrest and induction of apoptosis. We observed a strong concordance between the extent of BLIMP1 degradation and the depth of these cellular effects within these lines. Extending these findings in vivo, studies using three multiple myeloma xenograft models demonstrated a strong PK/PD correlation, with BLIMP1 PROTAC treatment resulting in 50–80% inhibition of tumor growth. Taken together, these results demonstrate that our potent and selective BLIMP1 PROTACs have robust anti-myeloma activity. Lastly, to determine whether BLIMP1 PROTAC responses could be further enhanced, we investigated combinations with myeloma standard-of-care agents. Co-treatment with dexamethasone resulted in a marked increase in cell death across various myeloma cell lines. While IMiDs effectively combined with VHL-based BLIMP1 PROTACs, combining IMiDs with CRBN-based BLIMP1 PROTACs was antagonistic, due to competition for CRBN binding. To overcome this effect, we developed BLIMP-IMiDs—one molecule designed to degrade both BLIMP1 and IMiD neosubstrates IKZF1 and IKZF3. These BLIMP-IMIDs showed striking anti-proliferative effects across a broad panel of myeloma lines and showed near-complete tumor growth inhibition in vivo. Conclusion: Our work is the first to identify BLIMP1 as a highly actionable target in myeloma. First-in-class BLIMP1 PROTACs demonstrate robust and selective BLIMP1 degradation and anti-myeloma activity as monotherapy and in combination with anti-myeloma agents. BLIMP-IMiDs provide a dual-targeting strategy that overcomes combination limitations, showing deep, broad anti-myeloma responses and highlighting BLIMP1-directed degradation as a promising therapy.