Abstract
Promotion of apoptosis may potentiate the sensitivity of tumor cells to chemotherapeutic agents, thus improving the efficacy of cancer treatment. The transfection of the proapoptotic bax gene, which results in the overexpression of bax protein, augments the growth inhibition of A253 cells by BNP1350. Increased drug response was associated with the induction of DNA fragmentation in the size of 30–200 Kb, generating a cleaved fragment of 18 kDa from full-length 21 kDa bax and the cleavage of PARP. A253/vec cells treated with 0.07 μM(IC50) of BNP1350 accumulated in G2 phase at 24 h after drug removal. In contrast, A253/Bax cells treated with an equimolar concentration of BNP1350 primarily displayed a G1 phase accumulation with a concurrent decrease in G2 phase. Certain cell cycle regulatory protein expression and activities were altered following drug exposure in both cell lines under similar conditions. Cdk2- and cdc2-associated H1 kinase activities were markedly increased in the A253/Bax cell line with marginal increased activity in the A253/vec cell line. A chk1 activity assay was performed with GST-cdc25C (200–256) or GST-cdc25CS216A (200–256) fusion proteins as the substrate. Increased chk1 activity was observed in the A253/vec cell line, with little change in the A253/Bax cell line, when exposed to equimolar concentrations of BNP1350 (0.07 μM). A Western blot of immunoprecipitated chk1 indicated that increased chk1 phosphorylation following DNA damage induced by BNP1350 was accompanied by the observed G2 accumulation in the A253/vec cell line, while only a slight increase in chk1 phosphorylation was seen in the A253/Bax cell line. A decreased expression of cdc25C was observed in the BNP1350-treated A253/Bax cells, but not in the A253/vec cell line. Following exposure to BNP1350, increased binding of 14-3-3 proteins to chk1 occurred in both cell lines, with more being observed in the A253/vec cell line. The data have shown that inhibition of the chk1 pathway accompanied by the abrogation of G2 arrest is involved in sensitizing A253 cells to BNP1350 by bax gene transfer. These findings suggest that bax gene transfer sensitizes A253 cells to BNP1350 through apoptosis promoting and G2/M DNA damage checkpoint regulatory pathways.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 50 print issues and online access
$259.00 per year
only $5.18 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
Abbreviations
- Topo I:
-
topoisomerase I
- SRB:
-
sulforhodamine B, cdk, cyclin-dependent kinase
- chk:
-
checkpoint kinase
- GST:
-
glutathione S-transferase
- PFGE:
-
pulsed-field gel electrophoresis
References
al-Khodairy F, Fotou E, Sheldrick KS, Griffiths DJ, Lehmann AR, Carr AM . 1994 Mol. Biol. Cell 5: 147–160
Arafat WO, Gomez-Navarro J, Xiang J, Barnes MN, Mahasreshti P, Alvarez RD, Siegal GP, Badib AO, Buchsbaum D, Curiel DT, Stackhouse MA . 2000 Mol. Ther. 1: 545–554
Brady HJ, Gil-Gomez G, Kirberg J, Berns AJ . 1996 EMBO J. 15: 6991–7001
Chan TA, Hwang PM, Hermeking H, Kinzler KW, Vogelstein B . 2000 Genes Dev. 14: 1584–1588
Chen L, Liu TH, Walworth NC . 1999 Genes Dev. 13: 675–685
Coll JL, Negoescu A, Louis N, Sachs L, Tenaud C, Girardot V, Demeinex B, Brambilla E, Brambilla C, Favrot M . 1998 Hum. Gene. Ther. 9: 2063–2074
Collins JA, Schandi CA, Young KK, Vesely J, Willingham MC . 1997 J. Histochem. Cytochem. 45: 923–934
Dubrez L, Goldwasser F, Genne P, Pommier Y, Solary E . 1995 Leukemia 9: 1013–1024
Elledge SJ . 1996 Science (Washington DC) 274: 1664–1672
Gil-Gomez G, Berns A, Brady HJ . 1998 EMBO J. 17: 7209–7218
Giaccia AJ, Shieh J, Cholon A, Brown JM . 1991 Mutat. Res. 263: 69–75
Goldwasser F, Shimizu T, Jackman J, Hoki Y, O'Connor PM, Kohn KW, Pommier Y . 1996 Cancer Res. 56: 4430–4437
Graves PR, Yu L, Schwarz JK, Gales J, Sausville EA, O'Connor PM, Piwnica-Worms H . 2000 J. Biol. Chem. 275: 5600–5605
Guo B, Yin MB, Toth K, Cao S, Azrak RG, Rustum YM . 1999 Oncol. Res. 11: 91–99
Guo B, Cao S, Toth K, Azrak RG, Rustum YM . 2000 Clin. Cancer Res. 6: 718–724
Hakem A, Sasaki T, Kozieradzki I, Penninger JM . 1999 J. Exp. Med. 189: 957–968
Hsu SL, Yin SC, Liu MC, Reichert U, Ho WL . 1999 Exp. Cell Res. 252: 332–341
Huang DC, O'Reilly LA, Strasser A, Cory S . 1997 EMBO J. 16: 4628–4638
Jackson JR, Gilmartin A, Imburgia C, Winkler JD, Marshall LA, Roshak A . 2000 Cancer Res. 60: 566–572
Jacotot E, Ferri KF, Kroemer G . 2000 Pathol. Biol. 48: 271–279
Krajewski S, Blomqvist C, Franssila K, Krajewska M, Wasenius VM, Niskanen E, Nordling S, Reed JC . 1995 Cancer Res. 55: 4471–4478
Laemmli UK . 1970 Nature (Lond.) 227: 680–685
Lind EF, Wayne J, Wang QZ, Staeva T, Stolzer A, Petrie HT . 1999 J. Immunol. 162: 5374–5379
Linette GP, Li Y, Roth K, Korsmeyer SJ . 1996 Proc. Natl. Acad. Sci. USA 93: 9545–9552
Lopez-Girona A, Furnari B, Mondesert O, Russell P . 1999 Nature (Lond.) 397: 172–175
McPake CR, Tillman DM, Poquette CA, George EO, Houghton JA, Harris LC . 1998 Oncol. Res. 10: 235–244
Oberhammer F, Wilson JM, Dive C, Morris ID, Hickman JA, Wakeling AE, Walker PR, Sikorska M . 1993 EMBO J. 12: 3679–3684
O'Connell MJ, Raleigh JM, Verkade HM, Nurse P . 1997 EMBO J. 16: 545–554
Ohi R, Gould KL . 1999 Curr. Opin. Cell Biol. 11: 267–273
Panadero A, Yin MB, Voigt W, Rustum YM . 1995 Oncol. Res. 7: 73–81
Peng CY, Graves PR, Thoma RS, Wu Z, Shaw AS, Piwnica-Worms H . 1997 Science (Washington DC) 277: 1501–1505
Peng CY, Graves PR, Ogg S, Thoma RS, Byrnes MJ, Wu Z, Stephenson MT, Piwnica-Worms H . 1998 Cell Growth Differ. 9: 197–208
Raleigh JM, O'Connell MJ . 2000 J. Cell Sci. 113: 1727–1736
Roshak A, Gilmartin A, Imburgia C, Marshall LA, Jackson JR . 1999 Proc. Am. Assoc. Cancer Res. 40: 413–414
Sard L, Accornero P, Tornielli S, Delia D, Bunone G, Campiglio M, Colombo MP, Gramegna M, Croce CM, Pierotti MA, Sozzi G . 1999 Proc. Natl. Acad. Sci. USA 96: 8489–8492
Schwartz DC, Cantor CR . 1984 Cell 37: 67–75
Shinoura N, Saito K, Yoshida Y, Hashimoto M, Asai A, Kirino T, Hamada H . 2000 Cancer Gene Ther. 7: 739–748
Skehan P, Storeng R, Scudiero D, Monks A, McMahon J, Vistica D, Warren JT, Bokesch H, Kenney S, Boyd MR . 1990 J. Natl. Cancer Inst. 82: 1107–1112
Strobel T, Swanson L, Korsmeyer S, Cannistra SA . 1996 Proc. Natl. Acad. Sci. USA. 93: 14094–14099
Strobel T, Kraeft SK, Chen LB, Cannistra SA . 1998 Cancer Res. 58: 4776–4781
Sugimoto C, Fujieda S, Seki M, Sunaga H, Fan GK, Tsuzuki H, Borner C, Saito H, Matsukawa S . 1999 Int. J. Cancer 82: 860–867
Tai YT, Lee S, Niloff E, Weisman C, Strobel T, Cannistra SA . 1998 J. Clin. Oncol. 16: 2583–2590
Tsai LH, Takahashi T, Caviness Jr VS, Harlow E . 1993 Development 119: 1029–1040
Vogelbaum MA, Tong JX, Higashikubo R, Gutmann DH, Rich KM . 1998 J. Neurosurg. 88: 99–105
Walworth N, Davey S, Beach D . 1993 Nature (Lond.) 363: 368–371
Walworth NC, Bernards R . 1996 Science (Washington DC) 271: 353–356
Wan S, Capasso H, Walworth NC . 1999 Yeast 15: 821–828
Xiang J, Chao DT, Korsmeyer SJ . 1996 Proc. Natl. Acad. Sci. USA 93: 14559–14563
Xiang J, Gomez-Navarro J, Arafat W, Liu B, Barker SD, Alvarez RD, Siegal GP, Curiel DT . 2000 J. Gene Med. 2: 97–106
Yin MB, Toth K, Cao S, Guo B, Frank C, Slocum HK, Rustum YM . 1999 Int. J. Cancer 83: 341–348
Yin MB, Guo B, Panadero A, Frank C, Wrzosek C, Slocum HK, Rustum YM . 1999 Exp. Cell Res. 247: 189–199
Yin MB, Guo B, Vanhoefer U, Azrak RG, Minderman H, Frank C, Wrzosek C, Slocum HK, Rustum YM . 2000 Mol. Pharmacol. 57: 453–459
Acknowledgements
We are grateful to Yolanda Sanchez for providing us with GST-cdc25C (200–256) and GST-cdc25CS216A (200–256). We also thank Geri Wagner for her secretarial assistance and BioNumerik Pharmaceuticals Inc. for providing us with BNP1350. This work was supported in part by project grant CA 65761 and a Comprehensive Cancer Center Support Grant CA 16056 from the National Cancer Institute, Bethesda, MD. G Hapke, was supported by grant HA3116/1-1 from the Deutsche Forschungsgemeinschaft, Bonn, Germany.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Yin, Mb., Hapke, G., Guo, B. et al. The Chk1-Cdc25C regulation is involved in sensitizing A253 cells to a novel topoisomerase I inhibitor BNP1350 by bax gene transfer. Oncogene 20, 5249–5257 (2001). https://doi.org/10.1038/sj.onc.1204686
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/sj.onc.1204686
Keywords
This article is cited by
-
Gossypin Induces G2/M Arrest in Human Malignant Glioma U251 Cells by the Activation of Chk1/Cdc25C Pathway
Cellular and Molecular Neurobiology (2012)