Dynamic, results-driven scientific research advisor with an outstanding track record in leading and conducting health research. Over twelve years’ experience managing a premier pharmacology laboratory, partnering with the Branch Chief to set the strategic and policy framework of the National Cancer Institute (NCI) laboratory and achieve stated outcomes. Provides scientific leadership and strategic direction of the research portfolio and priorities of NCI. Supports and oversees the scientific aspects of the work performed by the research teams, and recommends strategies to build their research capacity. Develops and implements strategies to engage external researchers, including collaborative research proposals. Extensive experience working with cross-functional scientific and research teams. Excellent influencing and interpersonal skills. Unique ability to embrace different points of view and values the contributions of others. A global understanding of organizations and culture that goes beyond traditional boundaries of geography. Supervisors: Dr. Yves Pommier Address: Bethesda, Maryland, United States
Poly(ADP-Ribose) polymerase (PARP) inhibitors represent a promising class of novel anticancer age... more Poly(ADP-Ribose) polymerase (PARP) inhibitors represent a promising class of novel anticancer agents. In the present study, we explored the strategy of combining the classical topoisomerase I (Top1) inhibitor camptothecin (CPT) and the novel PARP inhibitor ABT-888 in human cancer cells. ABT-888 potentiated cell killing by CPT. By colony forming assay, the survival ratio of CPT-treated colon cancer HT-29 cells was 52%, while the survival fraction were decreased to 35% when HT-29 cells were co-treated with CPT and ABT-888. By measuring γH2AX, a DNA double-strand break marker, we found that ABT-888 enhanced the γH2AX level induced by CPT. The enhanced γH2AX response in the presence of ABT-888 affected both replication-related γH2AX but also transcription-related γH2AX. In osteosarcoma U2OS cells, ABT-888 also enhanced γH2AX in both EdU (5-ethynyl-2′- deoxyuridine) positive and negative cells. Even in non-replicating human primary lymphocytes, ABT-888 enhanced CPT-induced γH2AX foci (1.3 ± 1.2 vs. 5.1 ± 3.1 γH2AX foci/per cell in the absence and presence of co-treatment with ABT-888, respectively). Proteasome inhibition with MG-132 blocked the γH2AX induction by CPT, and PARP inhibition could not overcome the block, which demonstrates that PARP acts downstream from the proteasome. Neutral Comet assays showed that the enhanced γH2AX response was related to DNA breaks. Alkaline elution assays revealed that ABT-888 increased CPT-induced DNA single-strand breaks (SSBs) but without a corresponding increase in DNA protein crosslinks indicating that ABT-888 was inducing DNA breaks in addition to the Top1-DNA cleavage complexes trapped by CPT. These additional DNA breaks were dependent on XPF/ERCC1 endonuclease. Furthermore, after removing CPT, reversal of γH2AX in the absence or presence of ABT-888 showed the same reversal rate, although there was more γH2AX in the presence of ABT-888, suggesting that PARP inhibition did not affect the repair of double-strand breaks. Moreover, PARP inhibition enhanced DNA-PK and ATM activation with increased formation of Rad51 foci. Therefore, we propose that PARP inhibition forces the cells to activate alternative endonucleolytic and recombination pathways in response to Top1-mediated DNA damage. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2973.
Batracylin (8-aminoisoindolo [1,2-b]quinazolin-10(12H)-one; NSC320846) is an investigational clin... more Batracylin (8-aminoisoindolo [1,2-b]quinazolin-10(12H)-one; NSC320846) is an investigational clinical anticancer agent. Previous animal studies showed activity against solid tumors and Adriamycin-resistant leukemia. We initially sought to test the proposed Top2-mediated DNA cleavage activity of batracylin and identify potential biomarkers for activity. COMPARE analysis in the NCI-60 cell lines showed batracylin activity to be most closely related to the class of Top2 inhibitors. The 50% growth inhibition (GI50) value for batracylin in HT29 colon carcinoma cells was 10 μmol/L. DNA-protein cross-links, consistent with Top2 targeting, were measured by alkaline elution. DNA single-strand breaks were also detected and found to be protein associated. However, only a weak induction of DNA double-strand breaks was observed. Because batracylin induced almost exclusively DNA single-strand breaks, we tested batracylin as a Top1 inhibitor. Batracylin exhibited both Top1- and Top2α/β-mediated DNA cleavage in vitro and in cells. The phosphorylation of histone (γ-H2AX) was tested to measure the extent of DNA damage. Kinetics of γ-H2AX “foci” showed early activation with low μmol/L concentrations, thus presenting a useful early biomarker of DNA damage. The half-life of γ-H2AX signal reversal after drug removal was consistent with reversal of DNA-protein cross-links. The persistence of the DNA-protein complexes induced by batracylin was markedly longer than by etoposide or camptothecin. The phosphorylated DNA damage–responsive kinase, ataxia telangiectasia mutated, was also found activated at sites of γ-H2AX. The cell cycle checkpoint kinase, Chk2, was only weakly phosphorylated. Thus, batracylin is a dual Top1 and Top2 inhibitor and γ-H2AX could be considered a biomarker in the ongoing clinical trials. [Cancer Res 2007;67(20):9971–9]
Di-2-pyridylketone-4,4,-dimethyl-3-thiosemicarbazone (Dp44mT) is being developed as an iron chela... more Di-2-pyridylketone-4,4,-dimethyl-3-thiosemicarbazone (Dp44mT) is being developed as an iron chelator with selective anticancer activity. We investigated the mechanism whereby Dp44mT kills breast cancer cells, both as a single agent and in combination with doxorubicin. Dp44mT alone induced selective cell killing in the breast cancer cell line MDA-MB-231 when compared with healthy mammary epithelial cells (MCF-12A). It induces G1 cell cycle arrest and reduces cancer cell clonogenic growth at nanomolar concentrations. Dp44mT, but not the iron chelator desferal, induces DNA double-strand breaks quantified as S139 phosphorylated histone foci (γ-H2AX) and Comet tails induced in MDA-MB-231 cells. Doxorubicin-induced cytotoxicity and DNA damage were both enhanced significantly in the presence of low concentrations of Dp44mT. The chelator caused selective poisoning of DNA topoisomerase IIα (top2α) as measured by an in vitro DNA cleavage assay and cellular topoisomerase-DNA complex formation. Heterozygous Nalm-6 top2α knockout cells (top2α+/−) were partially resistant to Dp44mT-induced cytotoxicity compared with isogenic top2α+/+ or top2β−/− cells. Specificity for top2α was confirmed using top2α and top2β small interfering RNA knockdown in HeLa cells. The results show that Dp44mT is cytotoxic to breast cancer cells, at least in part, due to selective inhibition of top2α. Thus, Dp44mT may serve as a mechanistically unique treatment for cancer due to its dual ability to chelate iron and inhibit top2α activity. [Cancer Res 2009;69(3):948–57]
Poly(ADP-Ribose) (PAR) polymerase (PARP) inhibitors represent an important class of novel antican... more Poly(ADP-Ribose) (PAR) polymerase (PARP) inhibitors represent an important class of novel anticancer agents. The present study explores the molecular rationale for combining veliparib (ABT-888) with camptothecin (CPT) and its clinical derivatives, topotecan and irinotecan. ABT-888 inhibited PAR induction by CPT and increased CPT-induced cell killing and DNA damage measured by histone gamma-H2AX and comet assays. Notably, increased DNA breaks by ABT-888 was not associated with a corresponding increase of topoisomerase I cleavage complexes (Top1cc). Also, ABT-888 failed to further enhance the gamma-H2AX response in tyrosyl-DNA-phosphodiesterase (TDP1) knockout cells, which indicates that PARP inhibition no longer enhances Top1-induced DNA damage once TDP1 is inactivated. To explore alternative endonuclease pathway for the repair of Top1cc upon PARP inhibition, we tested the role of the 3′-flap endonuclease XPF-ERCC1. XPF-ERCC1 inactivation reduced the ABT-888-induced gamma-H2AX response both in non-replicating and replicating cells, and further enhanced the antiproliferative effect of ABT-888 in CPT-treated cells. Together, our findings demonstrate that XPF-ERCC1 acts as an alternative endonuclease pathway for the repair of Top1-induced DNA damage. Its function becomes critical when the PARP-TDP1 excision repair pathway is inactivated. Our study suggests the rationale for combining Top1 and PARP inhibitors in tumors with XPF-ERCC1 deficiency. 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 4691. doi:1538-7445.AM2012-4691
Supplemental information contains three figures. Figure s1 displays plots of in vitro inhibition ... more Supplemental information contains three figures. Figure s1 displays plots of in vitro inhibition plotted versus IC50 values for inhibitors as measured for three individual cell lines. Figure S2 displays the low rate probability and religation inhibition measures of inhibition plotted versus the average normalized IC50 values for each of the four inhibitors. Figure S3 displays the fraction of relaxed DNA measured in the ensembled supercoil relaxation assay as a function of the inhibitor concentration for the four inhibitors.
Poly(ADP-Ribose) polymerase (PARP) inhibitors represent a promising class of novel anticancer age... more Poly(ADP-Ribose) polymerase (PARP) inhibitors represent a promising class of novel anticancer agents. In the present study, we explored the strategy of combining the classical topoisomerase I (Top1) inhibitor camptothecin (CPT) and the novel PARP inhibitor ABT-888 in human cancer cells. ABT-888 potentiated cell killing by CPT. By colony forming assay, the survival ratio of CPT-treated colon cancer HT-29 cells was 52%, while the survival fraction were decreased to 35% when HT-29 cells were co-treated with CPT and ABT-888. By measuring γH2AX, a DNA double-strand break marker, we found that ABT-888 enhanced the γH2AX level induced by CPT. The enhanced γH2AX response in the presence of ABT-888 affected both replication-related γH2AX but also transcription-related γH2AX. In osteosarcoma U2OS cells, ABT-888 also enhanced γH2AX in both EdU (5-ethynyl-2′- deoxyuridine) positive and negative cells. Even in non-replicating human primary lymphocytes, ABT-888 enhanced CPT-induced γH2AX foci (1.3 ± 1.2 vs. 5.1 ± 3.1 γH2AX foci/per cell in the absence and presence of co-treatment with ABT-888, respectively). Proteasome inhibition with MG-132 blocked the γH2AX induction by CPT, and PARP inhibition could not overcome the block, which demonstrates that PARP acts downstream from the proteasome. Neutral Comet assays showed that the enhanced γH2AX response was related to DNA breaks. Alkaline elution assays revealed that ABT-888 increased CPT-induced DNA single-strand breaks (SSBs) but without a corresponding increase in DNA protein crosslinks indicating that ABT-888 was inducing DNA breaks in addition to the Top1-DNA cleavage complexes trapped by CPT. These additional DNA breaks were dependent on XPF/ERCC1 endonuclease. Furthermore, after removing CPT, reversal of γH2AX in the absence or presence of ABT-888 showed the same reversal rate, although there was more γH2AX in the presence of ABT-888, suggesting that PARP inhibition did not affect the repair of double-strand breaks. Moreover, PARP inhibition enhanced DNA-PK and ATM activation with increased formation of Rad51 foci. Therefore, we propose that PARP inhibition forces the cells to activate alternative endonucleolytic and recombination pathways in response to Top1-mediated DNA damage. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2973.
Batracylin (8-aminoisoindolo [1,2-b]quinazolin-10(12H)-one; NSC320846) is an investigational clin... more Batracylin (8-aminoisoindolo [1,2-b]quinazolin-10(12H)-one; NSC320846) is an investigational clinical anticancer agent. Previous animal studies showed activity against solid tumors and Adriamycin-resistant leukemia. We initially sought to test the proposed Top2-mediated DNA cleavage activity of batracylin and identify potential biomarkers for activity. COMPARE analysis in the NCI-60 cell lines showed batracylin activity to be most closely related to the class of Top2 inhibitors. The 50% growth inhibition (GI50) value for batracylin in HT29 colon carcinoma cells was 10 μmol/L. DNA-protein cross-links, consistent with Top2 targeting, were measured by alkaline elution. DNA single-strand breaks were also detected and found to be protein associated. However, only a weak induction of DNA double-strand breaks was observed. Because batracylin induced almost exclusively DNA single-strand breaks, we tested batracylin as a Top1 inhibitor. Batracylin exhibited both Top1- and Top2α/β-mediated DNA cleavage in vitro and in cells. The phosphorylation of histone (γ-H2AX) was tested to measure the extent of DNA damage. Kinetics of γ-H2AX “foci” showed early activation with low μmol/L concentrations, thus presenting a useful early biomarker of DNA damage. The half-life of γ-H2AX signal reversal after drug removal was consistent with reversal of DNA-protein cross-links. The persistence of the DNA-protein complexes induced by batracylin was markedly longer than by etoposide or camptothecin. The phosphorylated DNA damage–responsive kinase, ataxia telangiectasia mutated, was also found activated at sites of γ-H2AX. The cell cycle checkpoint kinase, Chk2, was only weakly phosphorylated. Thus, batracylin is a dual Top1 and Top2 inhibitor and γ-H2AX could be considered a biomarker in the ongoing clinical trials. [Cancer Res 2007;67(20):9971–9]
Di-2-pyridylketone-4,4,-dimethyl-3-thiosemicarbazone (Dp44mT) is being developed as an iron chela... more Di-2-pyridylketone-4,4,-dimethyl-3-thiosemicarbazone (Dp44mT) is being developed as an iron chelator with selective anticancer activity. We investigated the mechanism whereby Dp44mT kills breast cancer cells, both as a single agent and in combination with doxorubicin. Dp44mT alone induced selective cell killing in the breast cancer cell line MDA-MB-231 when compared with healthy mammary epithelial cells (MCF-12A). It induces G1 cell cycle arrest and reduces cancer cell clonogenic growth at nanomolar concentrations. Dp44mT, but not the iron chelator desferal, induces DNA double-strand breaks quantified as S139 phosphorylated histone foci (γ-H2AX) and Comet tails induced in MDA-MB-231 cells. Doxorubicin-induced cytotoxicity and DNA damage were both enhanced significantly in the presence of low concentrations of Dp44mT. The chelator caused selective poisoning of DNA topoisomerase IIα (top2α) as measured by an in vitro DNA cleavage assay and cellular topoisomerase-DNA complex formation. Heterozygous Nalm-6 top2α knockout cells (top2α+/−) were partially resistant to Dp44mT-induced cytotoxicity compared with isogenic top2α+/+ or top2β−/− cells. Specificity for top2α was confirmed using top2α and top2β small interfering RNA knockdown in HeLa cells. The results show that Dp44mT is cytotoxic to breast cancer cells, at least in part, due to selective inhibition of top2α. Thus, Dp44mT may serve as a mechanistically unique treatment for cancer due to its dual ability to chelate iron and inhibit top2α activity. [Cancer Res 2009;69(3):948–57]
Poly(ADP-Ribose) (PAR) polymerase (PARP) inhibitors represent an important class of novel antican... more Poly(ADP-Ribose) (PAR) polymerase (PARP) inhibitors represent an important class of novel anticancer agents. The present study explores the molecular rationale for combining veliparib (ABT-888) with camptothecin (CPT) and its clinical derivatives, topotecan and irinotecan. ABT-888 inhibited PAR induction by CPT and increased CPT-induced cell killing and DNA damage measured by histone gamma-H2AX and comet assays. Notably, increased DNA breaks by ABT-888 was not associated with a corresponding increase of topoisomerase I cleavage complexes (Top1cc). Also, ABT-888 failed to further enhance the gamma-H2AX response in tyrosyl-DNA-phosphodiesterase (TDP1) knockout cells, which indicates that PARP inhibition no longer enhances Top1-induced DNA damage once TDP1 is inactivated. To explore alternative endonuclease pathway for the repair of Top1cc upon PARP inhibition, we tested the role of the 3′-flap endonuclease XPF-ERCC1. XPF-ERCC1 inactivation reduced the ABT-888-induced gamma-H2AX response both in non-replicating and replicating cells, and further enhanced the antiproliferative effect of ABT-888 in CPT-treated cells. Together, our findings demonstrate that XPF-ERCC1 acts as an alternative endonuclease pathway for the repair of Top1-induced DNA damage. Its function becomes critical when the PARP-TDP1 excision repair pathway is inactivated. Our study suggests the rationale for combining Top1 and PARP inhibitors in tumors with XPF-ERCC1 deficiency. 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 4691. doi:1538-7445.AM2012-4691
Supplemental information contains three figures. Figure s1 displays plots of in vitro inhibition ... more Supplemental information contains three figures. Figure s1 displays plots of in vitro inhibition plotted versus IC50 values for inhibitors as measured for three individual cell lines. Figure S2 displays the low rate probability and religation inhibition measures of inhibition plotted versus the average normalized IC50 values for each of the four inhibitors. Figure S3 displays the fraction of relaxed DNA measured in the ensembled supercoil relaxation assay as a function of the inhibitor concentration for the four inhibitors.
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