Increased PKR Expression Promotes MDS Evolution to Acute Leukemia in the NHD13 Mouse Model

Protein kinase R (PKR) is an interferon-inducible serine-threonine kinase that is activated in response to inflammatory cytokines and cellular stressors. When over-expressed in murine hematopoietic tissues, PKR results in peripheral blood (PB) and bone marrow (BM) features consistent with myelodysplastic syndrome (MDS). We hypothesized that PKR can cooperate with MDS driver mutations to promote MDS and acute leukemia. To test this, we crossed the Nup98-HoxD13 (NHD13) transgenic model of MDS with our TgPKR and PKR-KO mice to generate NHD13-TgPKR and NHD13-KO mice. These mice were aged alongside NHD13 and wild type (WT) controls. Using RT-PCR, we evaluated PKR expression in NHD13 mice. 9/13 NHD13 mice developed acute leukemia with leukocytosis and/or increased BM blasts whereas 4/13 had MDS only without PB or BM evidence of acute leukemia. Non-leukemic NHD13 mice had PKR expression levels below WT controls (RQ: 0.412 to 0.744). Alternatively, PKR expression in the leukemic group was significantly higher than WT controls and non-leukemic mice (RQ: 3.3 vs. 1.0 and 0.5, respectively) and was associated with shortened survival (mean: 304 vs. 345 days; p < 0.05). We then compared NHD13-TgPKR vs. NHD13-KO mice and discovered significantly lower white blood cells (WBC) counts in the NHD13-TgPKR group (2.7 vs. 5.6 x 10⁶ cells/ml, p <0.05; n=8/group) with significant differences in lymphocytes and neutrophils. With aging, NHD13-KO mice lived significantly longer than NHD13 mice (352.1 vs. 314.3 days, p <0.05) and NHD13-TgPKR mice (273.9 days, p <0.05). There were minor differences in the frequency of acute leukemia between the groups (NHD13-TgPKR: 58.3% vs. NHD13-KO: 41.6%). Next, we evaluated the frequency of somatic mutations in PB reticulocytes using the PIG-A assay that we have previously validated for measuring genomic instability of BM derived cells. We discovered an increased frequency of somatic mutations, indicative of greater genomic instability of the MDS HSPC, in 9-month-old NHD13-TgPKR (3.64%) vs. WT (<0.01%), NHD13 (0.26%), and NHD13-KO (0.18%) mice (p <0.05). Finally, the pharmacologic inhibition of PKR in NHD13 mice improved the WBC count (1.6 vs. 3.6 x 10⁶ cells/ml, p <0.05) and significantly reduced genomic instability (0.11 vs. 0.39%, p <0.05). Collectively, these data indicate that increased PKR expression is associated with the development of acute leukemia and shortened survival in the NHD13 mouse model. Further, NHD13-TgPKR mice, when compared to NHD13-KO mice, have a more aggressive MDS phenotype with deeper PB cytopenias, increased BM blasts, and shortened survival resulting from increased genomic instability. These data indicate that PKR can cooperate with MDS driver mutations and suggests that PKR may be a promising target for future MDS therapies, particularly in patients with high-level PKR expression.