Session: 604. Molecular Pharmacology and Drug Resistance: Myeloid Neoplasms: Poster III
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Acute Myeloid Malignancies, AML, Combination therapy, Diseases, Treatment Considerations, Myeloid Malignancies, Biological Processes, Molecular biology, Multi-systemic interactions
In this study, MOLM-13 and THP-1 cells were employed to establish ABT-199-resistant AML cell lines, named MOLM-13 R and THP-1 R, with distinct genomic backgrounds (FLT3-ITD mutant in MOLM-13 vs. TP53 mutant in THP-1), which are more likely to develop adaptive resistance to venetoclax-containing combinations. Our results indicate that usnic acid combined with ABT-199 effectively restored ABT-199 sensitivity to drug-resistant cells. The drug resistance reversal fold for 1.5 μM usnic acid in MOLM-13 R cells was 9.7, versus 9.5 for 4 μM usnic acid in THP-1 R cells. The combination therapy also demonstrated prominent anti-leukemic effects in the xenograft model. The combined administration reduced the average proportion of human CD45+ leukemic cells from 43.17% to 11.50%. RNA-seq showed that response of EIF2AK1 (HRI) to heme deficiency was markedly enriched in the combination group by reactome enrichment analysis. HRI, an eIF2α kinase, can induce eIF2-mediated integrated stress response. To examine the effects on downstream ISR signaling, real-time PCR assay and western blot were conducted in MOLM-13 R and THP-1 R cells administered the study drugs for 24 h. The combination of usnic acid and ABT-199 boosted the expression of the integrated stress response (ISR)-associated genes (ATF4, CHOP, and NOXA), CHOP protein levels and eIF2α phosphorylation at serine 51. ISRIB, a chemical inhibitor of the ISR, could rescue combination therapy-induced apoptosis, upregulation of ISR-associated genes. These findings demonstrate that ABT-199 and usnic acid jointly induce the ISR, which subsequently triggers cell death.
Furthermore, our findings showed that treatment with ABT-199 combined with usnic acid markedly increased NOXA protein amounts, and reduced anti-apoptotic factor MCL-1 protein levels. The downregulation of MCL-1 was linked to an increase in MCL-1 phosphorylation at serine 159 and subsequent destruction by the proteasome. MOLM-13 R cells overexpressing MCL-1 exhibited dramatically weakened apoptosis induced by usnic acid and ABT-199 co-administration, highlighting the significance of MCL-1 in the impact of ABT-199-usnic acid. In addition, ISRIB was able to reverse the inhibition of MCL-1 protein, and impair the degradation process in the combination group.
Overall, addition of usnic acid to ABT-199 results in an enhancement of its inhibitory effect on resistant AML in cell culture and animal experiments, because of enhanced MCL-1 downregulation resulting from ISR hyperactivation. These findings provide compelling evidence that combined treatment with usnic acid and ABT-199 constitutes a novel approach for AML cases that are more likely to develop resistance to ABT-199 and should be further investigated in clinical settings.
Disclosures: No relevant conflicts of interest to declare.
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