Abstract 2739: Panobinostat potentiates the cytotoxic effects of ara-C and daunorubicin on acute myeloid leukemia cells through suppressing the DNA damage response
Authors: Xie Chengzhi , Edwards Holly , Drenberg Christina , Baker Sharyn D. , Inaba Hiroto , Buck Steven A. , Taub Jeffrey W. , Ge Yubin 
Published: August 2012

Abstract
Acute myeloid leukemia (AML) remains a challenging disease to treat. Resistance to anthracycline [e.g., daunorubicin (DNR)] and ara-C-based chemotherapy is a major cause of treatment failure in this disease. Therefore, new therapies are urgently needed to improve treatment outcome of AML. Histone deacetylase (HDAC) inhibitors (HDACIs) are a promising new class of anti-cancer drugs, which induce differentiation, cell cycle arrest, and apoptosis in human leukemic cells, but not in normal cells. We previously demonstrated strikingly synergistic antileukemic interactions between valproic acid and ara-C in pediatric AML, accompanied by cooperative induction of DNA double-strand breaks (DSBs) and apoptosis. However, the precise molecular mechanisms remain to be identified. Recent studies with solid tumor cell lines established that HDACIs down-regulate expression of genes encoding critical DNA DSB repair proteins, which play critical roles in the DNA damage response (DDR), an important mechanism limiting chemotherapeutic efficacy. These results led to our hypothesis that HDACIs suppress the DDR in AML cells to enhance the cytotoxic effects of the DNA damaging chemotherapeutic drugs, DNR and ara-C. In four AML cell lines and ten diagnostic AML blast samples, treatment with the novel pan-HDACI, panobinostat, resulted in suppressed expression of BRCA1, CHK1, and RAD51 in a dose-dependent fashion, accompanied by dose-dependent induction of apoptosis or proliferation arrest. Treatments of the AML cell lines with ara-C or DNR resulted in induction of RAD51, BRCA1, and CHK1, which was abolished by co-administration of panobinostat. Importantly, panobinostat substantially enhanced ara-C- or DNR-induced DNA DSBs, as reflected in the induction of γH2AX, an established biomarker for DNA DSBs. This was accompanied by a dramatic synergistic induction of apoptosis. Whereas panobinostat alone resulted in G1 arrest, ara-C and DNR alone caused S-G2/M and G2/M arrest, respectively. The latter effects were abrogated when these agents were simultaneously administered with panobinostat. Interestingly, shRNA knockdown of BRCA1, CHK1, or RAD51 caused delayed repair of DNA DSBs induced by ara-C or DNR. Further, shRNA knockdown of BRCA1 or CHK1 resulted in significantly increased basal and ara-C- or DNR-induced apoptosis, along with substantially increased DNA DSBs and abrogated S and G2 checkpoints. In contrast, shRNA knockdown of RAD51 significantly enhanced DNR-induced DSBs and apoptosis, but had no impact on S and G2 checkpoints. These results support our hypothesis that suppression of the DDR by HDACIs represents a novel mechanism underlying the antileukemic activities of this class of drugs, and the synergy between HDACIs and DNA damaging agents in AML. Studies are underway evaluating in vivo efficacies of these drug combinations using xenograft NSG Mice.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2739. doi:1538-7445.AM2012-2739