A Novel SAHA-Bendamustine Hybrid Induces Apoptosis of Leukemia Cells

Introduction Hybrid anticancer drugs are of great therapeutic interests as they can potentially overcome the flaws of conventional chemotherapy drugs and improve their efficacy. Histone deacetylase inhibitors (HDACi) and DNA damaging agents have showed synergistic effects in recent studies. In this study, we reported a novel hybrid NL-101 that combines chemo-active groups from suberoylanilide hydroxamic acid (SAHA) and bendamustine, the typical HDACi and alkylating agent respectively.The anticancer effect of NL-101 and its possible mechanisms were investigated in human leukemia cell lines and primary leukemia cells.

Methods MTT assay was performed to determine the proliferation of Kasumi-1 and NB4 cells treated with NL-101. Cell cycle distribution and apoptosis rate were detected by flow cytometry. Western-blot analysis was used to analyze the level of acetylated H3 as well as apoptotic-related proteins including γ-H2AX, PARP, caspase-3, Bax, Bcl-2 and Bcl-xL. Bone marrow mononuclear cells of AML patients were isolated by density gradient centrifugation. Wright staining and Western blot were performed to determine the inducing apoptosis effect.

Results NL-101 inhibited the proliferation of leukemia cell lines Kasumi-1 and NB4 cells with similar IC₅₀ to that of SAHA. Cell cycle analysis indicated that NL-101 induced S phase arrest. As expected, apoptotic cell death was observed in response to NL-101 treatment. After treatment with 2 µmol/L NL-101 for 48 hours, the apoptosis rate of Kasumi-1 and NB4 cells were (60.19±12.01)% and (49.43±11.61)%, respectively. Western blot analysis showed that NL-101 exposure could induce the accumulation of acetylated Histone H3 and γ-H2AX as the biomarker of DNA double-strand breaks. Anti-apoptotic protein Bcl-xL involved in mitochondrial death pathway was also decreased. Moreover, NL-101 induced apoptosis with a low micromolar IC₅₀ in various leukemia cell lines but not in nonmalignant cell line HEK293. The efficacy of NL-101 was also tested in human primary leukemia cells and all the treated samples exhibited apoptosis confirmed by the morphological examination and expression of apoptotic markers.

Conclusions The novel SAHA-bendamustine hybrid NL-101 inhibited the proliferation and induced apoptotic cell death of leukemia cell lines and primary leukemia cells. It presented the properties of both HDAC inhibition and DNA damaging. Down-regulation of Bcl-xL was also involved in the apoptosis induction. These results indicated that NL-101 might be a potential compound for the treatment of leukemia.