Hypoxia-inducible factors (HIFs) are master transcriptional regulators, central to cellular survival in hypoxia and frequently activated within malignancy. Whilst malignant context directs the role of HIFs within oncogenesis, these mechanisms are not well characterised. Applying the JAK2V617F myeloproliferative neoplasms (MPNs) oncogene-driver model, in which HIF-1α is stabilised in normoxia (20% O2), we sought to determine whether the modality of HIF-1 activation directs its function. Through direct analysis of hypoxia-activated vs. JAK2V617F-activated HIF-1 at the chromatin, we define a JAK2V617F-HIF-1 regulon that diverges from canonical HIF/hypoxia targets. In a cohort of 172 JAK2V617F-MPN patients, we observe significant association of the JAK2V617F-HIF-1 regulon, but not canonical HIF-1 gene signatures, with disease severity, progression, and patient survival. We further define a subset gene signature (HIF1-MPN-BP) significantly associated with spontaneous transformation to blast phase MPNs. Finally, we identify that JAK2V617F-induced HIF-1α stabilisation is mediated via PIM1 kinase. Our findings demonstrate that HIF-1 activation by the JAK2V617F-PIM1 axis significantly alters HIF-1 transcription function, desensitising HIF-1 activity to cellular oxygen levels, and restricting the HIF-1 regulon to a set of disease-associated target genes within JAK2V617F-MPNs. These findings restore the potential for specific therapeutic targeting of HIF-1 by delineating malignant activation from the physiological hypoxic response.

Somatic mutations in PHF6 (PHD finger protein 6) are common in hematologic malignancies and confer worse overall survival in acute myeloid leukemia patients. These mutations are predominantly frameshift or nonsense variants, resulting in a truncated PHF6 protein. However, the specific role of truncated PHF6 in leukemogenesis remains unclear. Here, we generated a transgenic mouse model, Phf6R274XTg, expressing a patient-derived truncated Phf6 mutation specifically in hematopoietic lineages. Unlike Phf6 knock-out mice, which do not develop spontaneous diseases, Phf6R274XTg mice developed a spectrum of spontaneous hematologic malignancies that recapitulate key features of PHF6-mutated hematologic malignancies in patients. Expression of PHF6aa1-273 led to expansion of the hematopoietic stem cell/progenitor cell (HSC/HPC) pool and promoted myeloid-skewed differentiation. The Phf6R274X mutation also induced substantial transcriptional dysregulation in HSC/HPCs. Single-cell RNA-seq analysis revealed a unique HSC trajectory and enhanced myeloid-biased differentiation in Phf6R274XTg HSC/HPCs. Additionally, truncation of PHF6 altered genome-wide H3K27ac occupancy via enhanced activity of KAT6B acetyltransferase. Treatment with CTx-648, a KAT6A/KAT6B inhibitor, restored HSC function in Phf6R274XTg mice and prolonged the survival of leukemic Phf6R274XTg mice. These findings demonstrate a gain-of-function effect for truncated PHF6aa1-273 in driving leukemogenesis and highlight KAT6B as a promising therapeutic target in PHF6 truncation-associated hematologic malignancies.