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HDAC inhibitors potentiate the activity of the BCR/ABL kinase inhibitor KW-2449 in imatinib-sensitive or -resistant BCR/ABL+ leukemia cells in vitro and in vivo.

ABSTRACT: The purpose of this study was to determine whether histone deacetylase (HDAC) inhibitors (HDACI) such as vorinostat or entinostat (SNDX-275) could increase the lethality of the dual Bcr/Abl-Aurora kinase inhibitor KW-2449 in various Bcr/Abl(+) human leukemia cells, including those resistant to imatinib mesylate (IM).Bcr/Abl(+) chronic myelogenous leukemia (CML) and acute lymphoblastic leukemia (ALL) cells, including those resistant to IM (T315I, E255K), were exposed to KW-2449 in the presence or absence of vorinostat or SNDX-275, after which apoptosis and effects on signaling pathways were examined. In vivo studies combining HDACIs and KW2449 were carried out by using a systemic IM-resistant ALL xenograft model.Coadministration of HDACIs synergistically increased KW-2449 lethality in vitro in multiple CML and Ph(+) ALL cell types including human IM resistant cells (e.g., BV-173/E255K and Adult/T315I). Combined treatment resulted in inactivation of Bcr/Abl and downstream targets (e.g., STAT5 and CRKL), as well as increased reactive oxygen species (ROS) generation and DNA damage (?H2A.X). The latter events and cell death were significantly attenuated by free radical scavengers (TBAP). Increased lethality was also observed in primary CD34(+) cells from patients with CML, but not in normal CD34(+) cells. Finally, minimally active vorinostat or SNDX275 doses markedly increased KW2449 antitumor effects and significantly prolonged the survival of murine xenografts bearing IM-resistant ALL cells (BV173/E255K).HDACIs increase KW-2449 lethality in Bcr/Abl(+) cells in association with inhibition of Bcr/Abl, generation of ROS, and induction of DNA damage. This strategy preferentially targets primary Bcr/Abl(+) hematopoietic cells and exhibits enhanced in vivo activity. Combining KW-2449 with HDACIs warrants attention in IM-resistant Bcr/Abl(+) leukemias.

SUBMITTER: Nguyen T

PROVIDER: S-EPMC3096723 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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HDAC inhibitors potentiate the activity of the BCR/ABL kinase inhibitor KW-2449 in imatinib-sensitive or -resistant BCR/ABL+ leukemia cells in vitro and in vivo.

Nguyen Tri T Dai Yun Y Attkisson Elisa E Kramer Lora L Jordan Nicholas N Nguyen Nguyen N Kolluri Nikhil N Muschen Markus M Grant Steven S

Clinical cancer research : an official journal of the American Association for Cancer Research 20110407 10

<h4>Purpose</h4>The purpose of this study was to determine whether histone deacetylase (HDAC) inhibitors (HDACI) such as vorinostat or entinostat (SNDX-275) could increase the lethality of the dual Bcr/Abl-Aurora kinase inhibitor KW-2449 in various Bcr/Abl(+) human leukemia cells, including those resistant to imatinib mesylate (IM).<h4>Experimental design</h4>Bcr/Abl(+) chronic myelogenous leukemia (CML) and acute lymphoblastic leukemia (ALL) cells, including those resistant to IM (T315I, E255K) ...[more]

PMID: 21474579

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Project description:BackgroundWith the widespread clinical application of tyrosine kinase inhibitors (TKIs), an increasing number of chronic myeloid leukaemia (CML) patients have developed resistance or intolerance to TKIs. BCR/ABL is the oncoprotein of CML. HSP90 is an essential chaperone of BCR/ABL and plays an important role in protein folding and the function of BCR/ABL. Therefore, inhibiting the chaperone function of HSP90 may be an effective strategy for CML treatment and to overcome TKI resistance.MethodsThe effect of KW-2478 on CML cell viability, apoptosis and cell cycle progression was detected by CCK-8 assay or flow cytometry. The levels of BCR/ABL, HSP90 and other signalling proteins were detected by western blots. The mitochondrial membrane potential was detected by flow cytometry combined with JC-1 staining. The interaction between BCR/ABL and HSP90α was detected by coimmunoprecipitation. The effect of KW-2478 on BCR/ABL carcinogenesis in vivo was investigated in CML-like mouse models.ResultsKW-2478 inhibited growth and induced apoptosis of CML cells. KW-2478 inhibited the chaperone function of HSP90α and then weakened the BCR/ABL and MAPK signalling pathways. This treatment also caused an increase in p27 and p21 expression and a decrease in cyclin B1 expression, which led to G2/M phase arrest. The mitochondrial pathway was primarily responsible for KW-2478-induced apoptosis. KW-2478 had a synergistic effect with imatinib in growth inhibition. Notably, KW-2478 had a stronger effect on growth inhibition, apoptosis induction and cell cycle arrest of K562/G01 cells than K562 cells. KW-2478 could effectively prolong the mouse lifespan and alleviate disease symptoms in CML-like mouse models.ConclusionsThis finding demonstrated that KW-2478 had anticancer properties in imatinib-sensitive and imatinib-resistant CML cells and illustrated the possible mechanisms. This study provides an alternative choice for CML treatment, especially for TKI-resistant patients with BCR/ABL amplification and TKI-intolerant patients.

Project description:BACKGROUND:Chronic myeloid leukemia (CML) pathogenesis is mainly driven by the oncogenic breakpoint cluster region-Abelson murine leukemia viral oncogene homolog 1 (BCR-ABL) fusion protein. Since BCR-ABL displays abnormal constitutive tyrosine kinase activity, therapies using tyrosine kinase inhibitors (TKis) such as imatinib represent a major breakthrough for the outcome of CML patients. Nevertheless, the development of TKi resistance and the persistence of leukemia stem cells (LSCs) remain barriers to cure the disease, justifying the development of novel therapeutic approaches. Since the activity of histone deacetylase (HDAC) is deregulated in numerous cancers including CML, pan-HDAC inhibitors may represent promising therapeutic regimens for the treatment of CML cells in combination with TKi. RESULTS:We assessed the anti-leukemic activity of a novel hydroxamate-based pan-HDAC inhibitor MAKV-8, which complied with the Lipinski's "rule of five," in various CML cells alone or in combination with imatinib. We validated the in vitro HDAC-inhibitory potential of MAKV-8 and demonstrated efficient binding to the ligand-binding pocket of HDAC isoenzymes. In cellulo, MAKV-8 significantly induced target protein acetylation, displayed cytostatic and cytotoxic properties, and triggered concomitant ER stress/protective autophagy leading to canonical caspase-dependent apoptosis. Considering the specific upregulation of selected HDACs in LSCs from CML patients, we investigated the differential toxicity of a co-treatment with MAKV-8 and imatinib in CML versus healthy cells. We also showed that beclin-1 knockdown prevented MAKV-8-imatinib combination-induced apoptosis. Moreover, MAKV-8 and imatinib co-treatment synergistically reduced BCR-ABL-related signaling pathways involved in CML cell growth and survival. Since our results showed that LSCs from CML patients overexpressed c-MYC, importantly MAKV-8-imatinib co-treatment reduced c-MYC levels and the LSC population. In vivo, tumor growth of xenografted K-562 cells in zebrafish was completely abrogated upon combined treatment with MAKV-8 and imatinib. CONCLUSIONS:Collectively, the present findings show that combinations HDAC inhibitor-imatinib are likely to overcome drug resistance in CML pathology.

Project description:Previously, it has been stated that the BCR-ABL fusion-protein is sufficient to induce Chronic Myeloid Leukemia (CML), but additional genomic-changes are required for disease progression. Hence, we profiled control and tyrosine kinase inhibitors (TKI) alone or in combination with other drug-treated CML-samples in different phases, categorized as drug-sensitive and drug-resistant on the basis of BCR-ABL transcripts, the marker of major molecular-response. Molecular-profiling was done using the molecular-inversion probe-based-array, Human Transcriptomics-Array2.0, and Axiom-Biobank genotyping-arrays. At the transcript-level, clusters of control, TKI-resistant and TKI-sensitive cases were correlated with BCR-ABL transcript-levels. Both at the gene- and exon-levels, up-regulation of MPO, TPX2, and TYMS and down-regulation of STAT6, FOS, TGFBR2, and ITK lead up-regulation of the cell-cycle, DNA-replication, DNA-repair pathways and down-regulation of the immune-system, chemokine- and interleukin-signaling, TCR, TGF beta and MAPK signaling pathways. A comparison between TKI-sensitive and TKI-resistant cases revealed up-regulation of LAPTM4B, HLTF, PIEZO2, CFH, CD109, ANGPT1 in CML-resistant cases, leading to up-regulation of autophagy-, protein-ubiquitination-, stem-cell-, complement-, TGFβ- and homeostasis-pathways with specific involvement of the Tie2 and Basigin signaling-pathway. Dysregulated pathways were accompanied with low CNVs in CP-new and CP-UT-TKI-sensitive-cases with undetectable BCR-ABL-copies. High CNVs (previously reported gain of 9q34) were observed in BCR-ABL-independent and -dependent TKI, non-sensitive-CP-UT/AP-UT/B-UT and B-new samples. Further, genotyping CML-CP-UT cases with BCR-ABL 0-to-77.02%-copies, the identified, rsID239798 and rsID9475077, were associated with FAM83B, a candidate for therapeutic resistance. The presence of BCR-ABL, additional genetic-events, dysregulated-signaling-pathways and rsIDs associated with FAM83B in TKI-resistant-cases can be used to develop a signature-profile that may help in monitoring therapy.

Dataset Information

HDAC inhibitors potentiate the activity of the BCR/ABL kinase inhibitor KW-2449 in imatinib-sensitive or -resistant BCR/ABL+ leukemia cells in vitro and in vivo.

Publications

HDAC inhibitors potentiate the activity of the BCR/ABL kinase inhibitor KW-2449 in imatinib-sensitive or -resistant BCR/ABL+ leukemia cells in vitro and in vivo.

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