Program: Oral and Poster Abstracts
Session: 605. Molecular Pharmacology, Drug Resistance – Lymphoid and Other Diseases: Poster I
Methods: In vitro WST-1 cell viability assays were carried out on TOM1 cells (non-ABL1 mutant, imatinib sensitive Ph+ ALL) and NALM1 cells (non-ABL1 mutant, imatinib resistant CML lymphoid blast crisis) with imatinib and entinostat (a HDACi). Synergy was assessed using Calcusyn software. Western blots were performed assessing BCL6 expression and acetylation, and expression of downstream effectors of apoptosis. Two separate in vivo xenograft mouse experiments were performed transplanting TOM1 and NALM1 cells into Nod SCID Gamma (NSG) mice. Cohorts of TOM1 mice were treated with imatinib 50mg/kg BID, entinostat 15mg/kg QD, imatinib plus entinostat combination, or vehicle control. In the NALM1 mice we added a higher dose imatinib cohort (100 mg/kg BID) due to known imatinib resistance.
Results: In vitro, there was substantially more synergy of the imatinib/entinostat combination in imatinib-resistant NALM1 cells vs. the imatinib-sensitive TOM1 cells. Average Combination Index (CI) values in TOM1 cells across multiple entinostat and imatinib doses was 1.2 (CI: =1 suggest additive effect, <1 = synergy, >1 = antagonism), while the CI in NALM1 cells at the same dose combinations was 0.53. We noted BCL6 upregulation and decreased BCL6 acetylation – signs correlating with resistance – in Western blots of NALM1 and TOM1 cells treated with imatinib, while exposure to entinostat caused increased acetylation of BCL6 and increased expression of downstream tumor suppressors.
In the imatinib-sensitive TOM1 xenograft trial, the combination displayed a significant reduction in bone marrow leukemic blast involvement versus control following 6 weeks of dosing as measured by flow cytometry (36.9% mean decrease, p=0.001). There was a trend toward decreased bone marrow involvement between the combination treatment and other active treatment arms. There was no difference in peripheral blood blast percentage between arms. In the imatinib-resistant NALM1 xenograft trial, the combination showed a significant decrease in peripheral blood blast percentage in the combination arms versus all other arms after only two weeks of therapy (p=0.0008).
Conclusions: Upregulation of activated BCL6 is a known mechanism of resistance in Ph+ ALL that may be abrogated by acetylation of BCL6 with HDACi, as our in-vitro data suggests. Further, we have shown in xenograft models of Ph+ acute lymphoblastic leukemia that combination therapy with HDACi + imatinib, even in imatinib-resistant leukemia, has significant activity. Interestingly, the combination appears more active in resistant disease than in imatinib-sensitive disease. This combination could prove a viable strategy to attenuate imatinib- (and perhaps other TKI-) resistance in Ph+ ALL relapse, particularly in cases not driven by ABL1 kinase domain mutations.
Disclosures: No relevant conflicts of interest to declare.
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