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SIRT1 deacetylase promotes acquisition of genetic mutations for drug resistance in CML cells.

Wang Z, Yuan H, Roth M, Stark JM, Bhatia R, Chen WY - Oncogene (2012)

Bottom Line: The tyrosine kinase inhibitor imatinib effectively treats CML, but acquired resistance can develop because of BCR-ABL mutations.SIRT1 knockdown also suppresses de novo genetic mutations of hypoxanthine phosphoribosyl transferase gene in CML and non-CML cells upon treatment with DNA damaging agent camptothecin.These results reveal a previously unrecognized role of SIRT1 for promoting mutation acquisition in cancer, and have implication for targeting SIRT1 to overcome CML drug resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Biology, Beckman Research Institute, City of Hope, Duarte, CA, USA.

ABSTRACT
BCR-ABL transforms bone marrow progenitor cells and promotes genome instability, leading to development of chronic myelogenous leukemia (CML). The tyrosine kinase inhibitor imatinib effectively treats CML, but acquired resistance can develop because of BCR-ABL mutations. Mechanisms for acquisition of BCR-ABL mutations are not fully understood. Using a novel culture model of CML acquired resistance, we show that inhibition of SIRT1 deacetylase by small molecule inhibitors or gene knockdown blocks acquisition of BCR-ABL mutations and relapse of CML cells on tyrosine kinase inhibitors. SIRT1 knockdown also suppresses de novo genetic mutations of hypoxanthine phosphoribosyl transferase gene in CML and non-CML cells upon treatment with DNA damaging agent camptothecin. Although SIRT1 can enhance cellular DNA damage response, it alters functions of DNA repair machineries in CML cells and stimulates activity of error-prone DNA damage repair, in association with acquisition of genetic mutations. These results reveal a previously unrecognized role of SIRT1 for promoting mutation acquisition in cancer, and have implication for targeting SIRT1 to overcome CML drug resistance.

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Influence of HR repair factors for CML acquired resistance(a) NBS1 acetylation upon SIRT1 knockdown. NBS1 was immunoprecipitated and analyzed by Western blot with anti-acetylated lysine antibody, followed by NBS1 antibody. FLAG antibody was used for immunoprecipitation control. (b) NBS1 and RAD51 knockdown in KCL-22 cells. (c) Effects of RAD51 and NBS1 knockdown on KCL-22 cell growth. (d) NBS1 and RAD51 knockdown KCL-22 cells were enriched with puromycin selection and subjected to relapse assay on 2.5 μM imatinib. (e) RAD51 and NBS1 knockdown reduced BCR-ABL mutation by soft agar clonogenic assay. Left panel, resistant colonies with imatinib treatment. Right panel, plating control without imatinib.
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Figure 5: Influence of HR repair factors for CML acquired resistance(a) NBS1 acetylation upon SIRT1 knockdown. NBS1 was immunoprecipitated and analyzed by Western blot with anti-acetylated lysine antibody, followed by NBS1 antibody. FLAG antibody was used for immunoprecipitation control. (b) NBS1 and RAD51 knockdown in KCL-22 cells. (c) Effects of RAD51 and NBS1 knockdown on KCL-22 cell growth. (d) NBS1 and RAD51 knockdown KCL-22 cells were enriched with puromycin selection and subjected to relapse assay on 2.5 μM imatinib. (e) RAD51 and NBS1 knockdown reduced BCR-ABL mutation by soft agar clonogenic assay. Left panel, resistant colonies with imatinib treatment. Right panel, plating control without imatinib.

Mentions: SIRT1 also deacetylates HR repair factor NBS1 (Nijmegen Breakage Syndrome), a component of MRN (MRE11-RAD50-NBS1) complex,19 and regulates recruitment of NBS1 and RAD51 to DNA damage foci for repair.18 We found that SIRT1 knockdown increased NBS1 acetylation in KCL-22 cells (Figure 5a). Knockdown of NBS1 or RAD51 moderately affected cell growth (Figure 5b,c), but suppressed BCR-ABL mutations and CML cell relapse on imatinib (Figure 5d,e). These results suggest that SIRT1 also regulates HR repair machineries for BCR-ABL mutation acquisition.


SIRT1 deacetylase promotes acquisition of genetic mutations for drug resistance in CML cells.

Wang Z, Yuan H, Roth M, Stark JM, Bhatia R, Chen WY - Oncogene (2012)

Influence of HR repair factors for CML acquired resistance(a) NBS1 acetylation upon SIRT1 knockdown. NBS1 was immunoprecipitated and analyzed by Western blot with anti-acetylated lysine antibody, followed by NBS1 antibody. FLAG antibody was used for immunoprecipitation control. (b) NBS1 and RAD51 knockdown in KCL-22 cells. (c) Effects of RAD51 and NBS1 knockdown on KCL-22 cell growth. (d) NBS1 and RAD51 knockdown KCL-22 cells were enriched with puromycin selection and subjected to relapse assay on 2.5 μM imatinib. (e) RAD51 and NBS1 knockdown reduced BCR-ABL mutation by soft agar clonogenic assay. Left panel, resistant colonies with imatinib treatment. Right panel, plating control without imatinib.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3376246&req=5

Figure 5: Influence of HR repair factors for CML acquired resistance(a) NBS1 acetylation upon SIRT1 knockdown. NBS1 was immunoprecipitated and analyzed by Western blot with anti-acetylated lysine antibody, followed by NBS1 antibody. FLAG antibody was used for immunoprecipitation control. (b) NBS1 and RAD51 knockdown in KCL-22 cells. (c) Effects of RAD51 and NBS1 knockdown on KCL-22 cell growth. (d) NBS1 and RAD51 knockdown KCL-22 cells were enriched with puromycin selection and subjected to relapse assay on 2.5 μM imatinib. (e) RAD51 and NBS1 knockdown reduced BCR-ABL mutation by soft agar clonogenic assay. Left panel, resistant colonies with imatinib treatment. Right panel, plating control without imatinib.
Mentions: SIRT1 also deacetylates HR repair factor NBS1 (Nijmegen Breakage Syndrome), a component of MRN (MRE11-RAD50-NBS1) complex,19 and regulates recruitment of NBS1 and RAD51 to DNA damage foci for repair.18 We found that SIRT1 knockdown increased NBS1 acetylation in KCL-22 cells (Figure 5a). Knockdown of NBS1 or RAD51 moderately affected cell growth (Figure 5b,c), but suppressed BCR-ABL mutations and CML cell relapse on imatinib (Figure 5d,e). These results suggest that SIRT1 also regulates HR repair machineries for BCR-ABL mutation acquisition.

Bottom Line: The tyrosine kinase inhibitor imatinib effectively treats CML, but acquired resistance can develop because of BCR-ABL mutations.SIRT1 knockdown also suppresses de novo genetic mutations of hypoxanthine phosphoribosyl transferase gene in CML and non-CML cells upon treatment with DNA damaging agent camptothecin.These results reveal a previously unrecognized role of SIRT1 for promoting mutation acquisition in cancer, and have implication for targeting SIRT1 to overcome CML drug resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Biology, Beckman Research Institute, City of Hope, Duarte, CA, USA.

ABSTRACT
BCR-ABL transforms bone marrow progenitor cells and promotes genome instability, leading to development of chronic myelogenous leukemia (CML). The tyrosine kinase inhibitor imatinib effectively treats CML, but acquired resistance can develop because of BCR-ABL mutations. Mechanisms for acquisition of BCR-ABL mutations are not fully understood. Using a novel culture model of CML acquired resistance, we show that inhibition of SIRT1 deacetylase by small molecule inhibitors or gene knockdown blocks acquisition of BCR-ABL mutations and relapse of CML cells on tyrosine kinase inhibitors. SIRT1 knockdown also suppresses de novo genetic mutations of hypoxanthine phosphoribosyl transferase gene in CML and non-CML cells upon treatment with DNA damaging agent camptothecin. Although SIRT1 can enhance cellular DNA damage response, it alters functions of DNA repair machineries in CML cells and stimulates activity of error-prone DNA damage repair, in association with acquisition of genetic mutations. These results reveal a previously unrecognized role of SIRT1 for promoting mutation acquisition in cancer, and have implication for targeting SIRT1 to overcome CML drug resistance.

Show MeSH
Related in: MedlinePlus