Ofirnoflast, a novel inflammasome inhibitor, rewires transcriptional programs and shows clinical and preclinical efficacy in myeloid neoplasms Free

Myelodysplastic syndromes (MDS) and chronic myelomonocytic leukemia (CMML) are inflammatory myeloid neoplasms marked by ineffective erythropoiesis, chronic inflammation, and progression to acute myeloid leukemia (AML). Therapies directly targeting inflammation in these diseases remain limited. A key contributor to this pathology is aberrant activation of the NLRP3 inflammasome, which promotes proinflammatory cytokine release and disrupts hematopoietic differentiation. NIMA-related kinase 7 (NEK7) is an essential adaptor of NLRP3 that mediates inflammasome assembly. NEK7 also plays critical roles in mitosis and centrosome organization. Consequently, direct inhibition of NEK7 poses significant challenges due to off-target toxicity. Thus, there remains an urgent need for a therapeutic that selectively inhibits NEK7-NLRP3 interaction without impairing the physiological functions of NEK7. Here, we demonstrate that aberrant activation of the NEK7-NLRP3 inflammasome axis in malignant cells drives key pathogenic features of MDS and CMML, and identify ofirnoflast, a novel first-in-class allosteric inhibitor that selectively disrupts NEK7-NLRP3 binding, prevents inflammasome assembly, and halts disease progression while sparing NEK7's other essential functions.

Transcriptomic analysis of bone marrow CD34⁺ cells from MDS patients revealed elevated expression of inflammasome components, such as IL-18, CASPASE-1, and PYCARD, which correlated with shorter overall survival (low: n = 82; high: n = 58; P = 0.02) and increased AML transformation risk (P=0.009). In CMML, scRNA-seq showed overexpression of IL-18, CASPASE-1, and NLRP3, particularly within granulocyte-monocyte progenitors (P<0.0001) implicated in leukemic initiation.

Functionally, ofirnoflast inhibited the secretion of IL-1β and other inflammatory cytokines (e.g., TNF, IL-6, IL-18) by 5–10 fold across a spectrum of in vitro models, such as LPS-activated inflammatory monocytes (P<0.001), Tet2-mutant monocytes (P<0.001), CMML patient-derived cells (P<0.001), and myeloid leukemia cell lines (P=0.003), showing broad anti-inflammatory efficacy. Mechanistically, ofirnoflast suppressed Caspase-1 and Caspase-8 activity (critical effectors of the NLRP3 inflammasome) by approximately 2-fold (P = 0.006), thereby rescuing cleavage of GATA1, a key erythroid transcription factor, in primary MDS and leukemia cell lines. Restoration of GATA1 function led to a 6-fold reversal of the erythroid differentiation block in CMML and MDS patient-derived cells (P < 0.0001), directly addressing ineffective hematopoiesis.

To define the specificity of ofirnoflast, CRISPR-mediated NEK7 deletion abolished its anti-inflammatory effects in myeloid leukemia cells (P<0.0001), confirming NEK7 dependency. Furthermore, genetic ablation of IL1R1 in Tet2-mutant cells or pharmacologic inhibition of IL1R1 in primary MDS/CMML patient-derived cells with Anakinra abrogated the anti-leukemic effects of ofirnoflast, indicating that its therapeutic activity is primarily mediated through suppression of chronic inflammation driven by the IL-1β/IL1R1 axis.

In vivo, ofirnoflast treatment reduced leukemic burden by ~3-fold in both MDS/AML (n = 5, P = 0.005) and CMML (n = 7–9, P = 0.04) patient-derived xenograft models. scRNA-seq of treated xenografts revealed a dual mechanism of action: (1) suppression of inflammatory and proliferative signaling pathways (NF-κB, TNF, and cell cycle genes), and (2) induction of apoptosis and erythroid differentiation programs, corroborating both molecular and functional in vivo and in vitro findings.

Translating these insights to the clinic, a Phase 1 trial (n = 64; NCT05447546) in healthy volunteers showed that ofirnoflast significantly reduced serum levels of IL-1β, TNF-⍺, IL-6, and IL-18 to baseline values, compared to placebo (P < 0.001), providing early evidence of pharmacodynamic efficacy and tolerability in humans.

In summary, our integrated clinical, mechanistic, and translational studies establish ofirnoflast as a highly selective and potentially safer NEK7-NLRP3 disruptor, capable of restoring erythroid differentiation block and halting leukemic progression in MDS and CMML by targeting IL-1β–driven chronic inflammation without impairing NEK7's essential cellular functions. Mechanistically, by linking NLRP3 activation to ineffective hematopoiesis and leukemic transformation, our findings reveal a novel therapeutic strategy to improve outcomes in these high-risk myeloid neoplasms.