Modification of Alternative Splicing of Bcl-x Pre-mRNA in Prostate and Breast Cancer Cells

Abstract

There is ample evidence that deregulation of apoptosis results in the development, progression, and/or maintenance of cancer. Since many apoptotic regulatory genes (e.g. bcl-x) code for alternatively spliced protein variants with opposing functions, the manipulation of alternative splicing presents a unique way of regulating the apoptotic response. Here we have targeted oligonucleotides antisense to the 5′-splice site ofbcl-xL, an anti-apoptotic gene that is overexpressed in various cancers, and shifted the splicing pattern of Bcl-x pre-mRNA from Bcl-xL to Bcl-xS, a pro-apoptotic splice variant. This approach induced significant apoptosis in PC-3 prostate cancer cells. In contrast, the same oligonucleotide treatment elicited a much weaker apoptotic response in MCF-7 breast cancer cells. Moreover, although the shift in Bcl-x pre-mRNA splicing inhibited colony formation in both cell lines, this effect was much less pronounced in MCF-7 cells. These differences in responses to oligonucleotide treatment were analyzed in the context of expression of Bcl-xL, Bcl-xS, and Bcl-2 proteins. The results indicate that despite the presence of Bcl-x pre-mRNA in a number of cell types, the effects of modification of its splicing by antisense oligonucleotides vary depending on the expression profile of the treated cells. There is ample evidence that deregulation of apoptosis results in the development, progression, and/or maintenance of cancer. Since many apoptotic regulatory genes (e.g. bcl-x) code for alternatively spliced protein variants with opposing functions, the manipulation of alternative splicing presents a unique way of regulating the apoptotic response. Here we have targeted oligonucleotides antisense to the 5′-splice site ofbcl-xL, an anti-apoptotic gene that is overexpressed in various cancers, and shifted the splicing pattern of Bcl-x pre-mRNA from Bcl-xL to Bcl-xS, a pro-apoptotic splice variant. This approach induced significant apoptosis in PC-3 prostate cancer cells. In contrast, the same oligonucleotide treatment elicited a much weaker apoptotic response in MCF-7 breast cancer cells. Moreover, although the shift in Bcl-x pre-mRNA splicing inhibited colony formation in both cell lines, this effect was much less pronounced in MCF-7 cells. These differences in responses to oligonucleotide treatment were analyzed in the context of expression of Bcl-xL, Bcl-xS, and Bcl-2 proteins. The results indicate that despite the presence of Bcl-x pre-mRNA in a number of cell types, the effects of modification of its splicing by antisense oligonucleotides vary depending on the expression profile of the treated cells. reverse transcription-polymerase chain reaction base pair(s) poly(ADP-ribose) polymerase fluorescence-activated cell sorting Apoptosis, or programmed cell death, is a highly regulated process controlled by numerous genes that determine a proper response to death signals (1Adams J.M. Cory S. Science. 1998; 281: 1322-1326Crossref PubMed Scopus (4780) Google Scholar, 2Antonsson B. Martinou J.C. Exp. Cell Res. 2000; 256: 50-57Crossref PubMed Scopus (626) Google Scholar, 3Lowe S.W. Lin A.W. Carcinogenesis. 2000; 21: 485-495Crossref PubMed Scopus (1562) Google Scholar, 4Reed J.C. J. Clin. Oncol. 1999; 17: 2941-2953Crossref PubMed Google Scholar); the relative levels of expression of pro- and anti-apoptotic genes appear to be particularly important (5Shinoura N. Yoshida Y. Asai A. Kirino T. Hamada H. Oncogene. 1999; 18: 5703-5713Crossref PubMed Scopus (64) Google Scholar, 6Kawakami K. Tsukuda M. Mizuno H. Nishimura G. Ishii A. Hamajima K. Anticancer Research. 1999; 19: 3927-3932PubMed Google Scholar, 7Taylor S.T. Hickman J.A. Dive C. J. Natl. Cancer Inst. 2000; 92: 18-23Crossref PubMed Scopus (89) Google Scholar, 8Liu Q.Y. Stein C.A. Clin. Cancer Res. 1997; 3: 2039-2046PubMed Google Scholar, 9Han J.S. Núñez G. Wicha M.S. Clarke M.F. Springer Semin. Immunopathol. 1998; 19: 279-288Crossref PubMed Scopus (16) Google Scholar). Deregulation of apoptosis, which contributes to the development, progression, and/or maintenance of cancer (3Lowe S.W. Lin A.W. Carcinogenesis. 2000; 21: 485-495Crossref PubMed Scopus (1562) Google Scholar, 4Reed J.C. J. Clin. Oncol. 1999; 17: 2941-2953Crossref PubMed Google Scholar, 10Wang D.G. Clin. Endocrinol. 1999; 51: 1-9Crossref PubMed Scopus (17) Google Scholar), is frequently caused by mechanisms that alter splicing of regulatory genes (11Mercatante D. Kole R. Pharmacol. Ther. 2000; 85: 237-243Crossref PubMed Scopus (69) Google Scholar, 12Jiang Z.H. Wu J.Y. Proc. Soc. Exp. Biol. Med. 1999; 220: 64-72Crossref PubMed Google Scholar). Therefore, this work focused on the modification of alternative splicing of bcl-x, a member of the bcl-2 family of apoptotic genes that play crucial roles in both inhibiting and activating the apoptotic response to cellular insults (1Adams J.M. Cory S. Science. 1998; 281: 1322-1326Crossref PubMed Scopus (4780) Google Scholar, 2Antonsson B. Martinou J.C. Exp. Cell Res. 2000; 256: 50-57Crossref PubMed Scopus (626) Google Scholar). Bcl-x is alternatively spliced to produce two distinct mRNAs and two variant proteins, Bcl-xL and Bcl-xS, that have antagonistic functions; the longer, 241-amino acid protein (Bcl-xL) inhibits apoptosis, whereas the shorter, 178-amino acid protein (Bcl-xS) activates it (13Boise L.H. González-Garcı́a M. Postema C.E. Ding L. Lindsten T. Turka L.A. Mao X. Núñez G. Thompson C.B. Cell. 1993; 74: 597-608Abstract Full Text PDF PubMed Scopus (2913) Google Scholar).Bcl-xL is highly expressed in many types of cancers, including multiple myeloma (14Tu Y. Renner S. Xu F. Fleishman A. Taylor J. Weisz J. Vescio R. Rettig M. Berenson J. Krajewski S. Reed J.C. Lichtenstein A. Cancer Res. 1998; 58: 256-262PubMed Google Scholar), small cell lung carcinoma (15Reeve J.G. Xiong J. Morgan J. Bleehen N.M. Br. J Cancer. 1996; 73: 1193-1200Crossref PubMed Scopus (91) Google Scholar), and breast cancer (16Olopade O.I. Adeyanju M.O. Safa A.R. Hagos F. Mick R. Thompson C.B. Recant W.M. Cancer J. Sci. Am. 1997; 3: 230-237PubMed Google Scholar). High Bcl-xL expression levels have been associated with an increased risk of metastasis in breast cancer (16Olopade O.I. Adeyanju M.O. Safa A.R. Hagos F. Mick R. Thompson C.B. Recant W.M. Cancer J. Sci. Am. 1997; 3: 230-237PubMed Google Scholar) as well as with an increased resistance to apoptosis induced by methotrexate and 5-fluorouracil (17Liu R. Page C. Beidler D.R. Wicha M.S. Núñez G. Am. J. Pathol. 1999; 155: 1861-1867Abstract Full Text Full Text PDF PubMed Scopus (73) Google Scholar) and etoposide and cisplatin (18Schmitt E. Cimoli G. Steyaert A. Bertrand R. Exp. Cell Res. 1998; 240: 107-121Crossref PubMed Scopus (44) Google Scholar). Additionally, an immunohistochemical study (19Krajewska M. Krajewski S. Epstein J.I. Shabaik A. Sauvageot J. Song K. Kitada S. Reed J.C. Am. J. Pathol. 1996; 148: 1567-1576PubMed Google Scholar) showed that Bcl-x, presumably Bcl-xL, was expressed in 100% of prostate tumors and that its immunointensity was stronger in higher grade metastases.Bcl-xS antagonizes the pro-survival properties of Bcl-xL (13Boise L.H. González-Garcı́a M. Postema C.E. Ding L. Lindsten T. Turka L.A. Mao X. Núñez G. Thompson C.B. Cell. 1993; 74: 597-608Abstract Full Text PDF PubMed Scopus (2913) Google Scholar, 20Minn A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google Scholar) and appears to induce apoptosis. For example, high levels of Bcl-xS induced apoptosis in cancer cells from patients with colon and stomach cancers (21Clarke M.F. Apel I.J. Benedict M.A. Eipers P.G. Sumantran V. González-Garcı́a M. Doedens M. Fukunaga N. Davidson B. Dick J.E. Minn A.J. Boise L.H. Thompson C.B. Wicha M. Núñez G. Proc. Natl. Acad. Sci. U. S. A. 1995; 92: 11024-11028Crossref PubMed Scopus (127) Google Scholar) as well as in human mammary tumors in nude mice (22Ealovega M.W. McGinnis P.K. Sumantran V.N. Clarke M.F. Wicha M.S. Cancer Res. 1996; Google Scholar). of Bcl-xS apoptosis in an breast cancer cell the cells apoptotic treatment with the and etoposide V.N. M.W. Núñez G. Clarke M.F. Wicha M.S. Cancer Res. 1995; Google from this showed that antisense oligonucleotides be to the splicing of various genes in cell H. S. Kole R. Proc. Natl. Acad. Sci. U. S. A. 1996; PubMed Scopus Google Scholar, H. D. Kole R. 1999; PubMed Scopus Google Scholar, F. K. S. K. S. Kole R. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, Kole J. J.A. Kole R. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). that the alternative 5′-splice site in of Bcl-x with an antisense oligonucleotide shift splicing from Bcl-xL to Bcl-xS the of pro-apoptotic Bcl-xS protein and the of its anti-apoptotic that this shift in splicing be and that it induced apoptotic in the prostate cancer cell in the breast cancer cell splicing from Bcl-xL to Bcl-xS and of apoptosis were much less in the responses to oligonucleotide treatment were in the of cell of the two cell have of the that alternative splicing of Bcl-x pre-mRNA two with antagonistic and an oligonucleotide antisense to the 5′-splice site of Bcl-xL to shift splicing from the anti-apoptotic splice Bcl-xL, to the pro-apoptotic splice This approach be to antisense of Bcl-xL N.M. Oncogene. 1999; 18: PubMed Scopus Google Scholar, S. R. J. U. J. Cancer. 2000; PubMed Scopus Google Scholar) by a in Bcl-xL to a in the of the antagonistic Bcl-xS, the effects of the In an oligonucleotide targeted to the 5′-splice site of Bcl-x which to an in the Bcl-xS splice variant and a in the Bcl-xL splice was a of apoptosis in PC-3 cells an oligonucleotide targeted to the which the expression of the Bcl-xL of PC-3 and MCF-7 cells with the oligonucleotide to a and shift in splicing of Bcl-x pre-mRNA from the Bcl-xL to and to a in the of Bcl-xS of the Bcl-xL the Bcl-xL protein in the cells in the There be two for this the the Bcl-xL and the Bcl-xS splice variant. This from to the Bcl-xS the Bcl-xL protein be in the cells high its was The in PC-3 Bcl-xS apoptosis and/or cell death despite the of anti-apoptotic Since Bcl-xS to Bcl-xL and inhibits its anti-apoptotic A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google Scholar), to that the is or that this is the for the pro-apoptotic properties of A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google appears that the apoptotic response of MCF-7 cells to oligonucleotide treatment is to the of Bcl-xS protein by the shift in the that the of Bcl-xS protein to apoptotic regulatory that its be the and of the genes that be for the apoptotic resistance of MCF-7 cells and a shift in alternative splicing of Bcl-x pre-mRNA was in breast cancer cell R. and R. this work was in in a lung cell treated with N.M. 1999; 17: PubMed Scopus (189) Google Scholar). cell lines, as well as the gene is in PC-3 L. 1993; PubMed Scopus Google Scholar, Cancer Res. 51: Google Scholar) and Y. J. Cancer. 1999; PubMed Scopus Google Scholar) in N.M. 1999; 17: PubMed Scopus (189) Google Scholar) or MCF-7 Y. J. Cancer. 1999; PubMed Scopus Google Scholar) and Bcl-2 is in PC-3 Q.Y. Stein C.A. Clin. Cancer Res. 1997; 3: 2039-2046PubMed Google Scholar), Y. J. Cancer. 1999; PubMed Scopus Google Scholar), and N.M. 1999; 17: PubMed Scopus (189) Google Scholar) cells and high in MCF-7 cells Y. J. Cancer. 1999; PubMed Scopus Google Scholar). treatment with to significant apoptosis in PC-3 whereas a in Bcl-2 in MCF-7 cells to levels in PC-3 cells apoptosis. appears that the and bcl-2 genes play a in the of the effects on cellular In this it is that Bcl-xL was in 100% of prostate Bcl-2 was in (19Krajewska M. Krajewski S. Epstein J.I. Shabaik A. Sauvageot J. Song K. Kitada S. Reed J.C. Am. J. Pathol. 1996; 148: 1567-1576PubMed Google Scholar). in breast as well as in breast cancer cell lines, the levels of Bcl-2 were higher of J.M. M. Krajewski S. S. E. Reed J.C. Cancer Res. 1998; PubMed Scopus Google responses of PC-3 and MCF-7 cells to the shift in Bcl-x pre-mRNA splicing to of to of a apoptotic and to cell death by colony In this cell death in both PC-3 and MCF-7 cell lines, the of the of MCF-7 cells and of PC-3 cells the treatment with oligonucleotide for MCF-7 cells and for PC-3 that in a of Bcl-xL to Bcl-xS of the in of the of the oligonucleotide of and for MCF-7 and PC-3 a This is by of the oligonucleotide in MCF-7 cells. In to PC-3 in which oligonucleotide in the in MCF-7 the of the in it was to a process Res. N. Y. 1996; PubMed Scopus Google that cells in response to a shift in the to the in of the although Bcl-xL is expressed in a number of cell types, including cell S. M. Shabaik A. S. L. Reed J.C. Cancer Res. Google Scholar), the apoptotic of the oligonucleotide be in cells with a gene expression profile to the prostate cancer PC-3 cell In a of the oligonucleotide with by cisplatin in cells N.M. 1999; 17: PubMed Scopus (189) Google Scholar) types of cells to apoptosis. of cellular by the oligonucleotide be a in the of and in the of cancer Apoptosis, or programmed cell death, is a highly regulated process controlled by numerous genes that determine a proper response to death signals (1Adams J.M. Cory S. Science. 1998; 281: 1322-1326Crossref PubMed Scopus (4780) Google Scholar, 2Antonsson B. Martinou J.C. Exp. Cell Res. 2000; 256: 50-57Crossref PubMed Scopus (626) Google Scholar, 3Lowe S.W. Lin A.W. Carcinogenesis. 2000; 21: 485-495Crossref PubMed Scopus (1562) Google Scholar, 4Reed J.C. J. Clin. Oncol. 1999; 17: 2941-2953Crossref PubMed Google Scholar); the relative levels of expression of pro- and anti-apoptotic genes appear to be particularly important (5Shinoura N. Yoshida Y. Asai A. Kirino T. Hamada H. Oncogene. 1999; 18: 5703-5713Crossref PubMed Scopus (64) Google Scholar, 6Kawakami K. Tsukuda M. Mizuno H. Nishimura G. Ishii A. Hamajima K. Anticancer Research. 1999; 19: 3927-3932PubMed Google Scholar, 7Taylor S.T. Hickman J.A. Dive C. J. Natl. Cancer Inst. 2000; 92: 18-23Crossref PubMed Scopus (89) Google Scholar, 8Liu Q.Y. Stein C.A. Clin. Cancer Res. 1997; 3: 2039-2046PubMed Google Scholar, 9Han J.S. Núñez G. Wicha M.S. Clarke M.F. Springer Semin. Immunopathol. 1998; 19: 279-288Crossref PubMed Scopus (16) Google Scholar). Deregulation of apoptosis, which contributes to the development, progression, and/or maintenance of cancer (3Lowe S.W. Lin A.W. Carcinogenesis. 2000; 21: 485-495Crossref PubMed Scopus (1562) Google Scholar, 4Reed J.C. J. Clin. Oncol. 1999; 17: 2941-2953Crossref PubMed Google Scholar, 10Wang D.G. Clin. Endocrinol. 1999; 51: 1-9Crossref PubMed Scopus (17) Google Scholar), is frequently caused by mechanisms that alter splicing of regulatory genes (11Mercatante D. Kole R. Pharmacol. Ther. 2000; 85: 237-243Crossref PubMed Scopus (69) Google Scholar, 12Jiang Z.H. Wu J.Y. Proc. Soc. Exp. Biol. Med. 1999; 220: 64-72Crossref PubMed Google Scholar). Therefore, this work focused on the modification of alternative splicing of bcl-x, a member of the bcl-2 family of apoptotic genes that play crucial roles in both inhibiting and activating the apoptotic response to cellular insults (1Adams J.M. Cory S. Science. 1998; 281: 1322-1326Crossref PubMed Scopus (4780) Google Scholar, 2Antonsson B. Martinou J.C. Exp. Cell Res. 2000; 256: 50-57Crossref PubMed Scopus (626) Google Scholar). Bcl-x is alternatively spliced to produce two distinct mRNAs and two variant proteins, Bcl-xL and Bcl-xS, that have antagonistic functions; the longer, 241-amino acid protein (Bcl-xL) inhibits apoptosis, whereas the shorter, 178-amino acid protein (Bcl-xS) activates it (13Boise L.H. González-Garcı́a M. Postema C.E. Ding L. Lindsten T. Turka L.A. Mao X. Núñez G. Thompson C.B. Cell. 1993; 74: 597-608Abstract Full Text PDF PubMed Scopus (2913) Google Scholar). Bcl-xL is highly expressed in many types of cancers, including multiple myeloma (14Tu Y. Renner S. Xu F. Fleishman A. Taylor J. Weisz J. Vescio R. Rettig M. Berenson J. Krajewski S. Reed J.C. Lichtenstein A. Cancer Res. 1998; 58: 256-262PubMed Google Scholar), small cell lung carcinoma (15Reeve J.G. Xiong J. Morgan J. Bleehen N.M. Br. J Cancer. 1996; 73: 1193-1200Crossref PubMed Scopus (91) Google Scholar), and breast cancer (16Olopade O.I. Adeyanju M.O. Safa A.R. Hagos F. Mick R. Thompson C.B. Recant W.M. Cancer J. Sci. Am. 1997; 3: 230-237PubMed Google Scholar). High Bcl-xL expression levels have been associated with an increased risk of metastasis in breast cancer (16Olopade O.I. Adeyanju M.O. Safa A.R. Hagos F. Mick R. Thompson C.B. Recant W.M. Cancer J. Sci. Am. 1997; 3: 230-237PubMed Google Scholar) as well as with an increased resistance to apoptosis induced by methotrexate and 5-fluorouracil (17Liu R. Page C. Beidler D.R. Wicha M.S. Núñez G. Am. J. Pathol. 1999; 155: 1861-1867Abstract Full Text Full Text PDF PubMed Scopus (73) Google Scholar) and etoposide and cisplatin (18Schmitt E. Cimoli G. Steyaert A. Bertrand R. Exp. Cell Res. 1998; 240: 107-121Crossref PubMed Scopus (44) Google Scholar). Additionally, an immunohistochemical study (19Krajewska M. Krajewski S. Epstein J.I. Shabaik A. Sauvageot J. Song K. Kitada S. Reed J.C. Am. J. Pathol. 1996; 148: 1567-1576PubMed Google Scholar) showed that Bcl-x, presumably Bcl-xL, was expressed in 100% of prostate tumors and that its immunointensity was stronger in higher grade Bcl-xS antagonizes the pro-survival properties of Bcl-xL (13Boise L.H. González-Garcı́a M. Postema C.E. Ding L. Lindsten T. Turka L.A. Mao X. Núñez G. Thompson C.B. Cell. 1993; 74: 597-608Abstract Full Text PDF PubMed Scopus (2913) Google Scholar, 20Minn A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google Scholar) and appears to induce apoptosis. For example, high levels of Bcl-xS induced apoptosis in cancer cells from patients with colon and stomach cancers (21Clarke M.F. Apel I.J. Benedict M.A. Eipers P.G. Sumantran V. González-Garcı́a M. Doedens M. Fukunaga N. Davidson B. Dick J.E. Minn A.J. Boise L.H. Thompson C.B. Wicha M. Núñez G. Proc. Natl. Acad. Sci. U. S. A. 1995; 92: 11024-11028Crossref PubMed Scopus (127) Google Scholar) as well as in human mammary tumors in nude mice (22Ealovega M.W. McGinnis P.K. Sumantran V.N. Clarke M.F. Wicha M.S. Cancer Res. 1996; Google Scholar). of Bcl-xS apoptosis in an breast cancer cell the cells apoptotic treatment with the and etoposide V.N. M.W. Núñez G. Clarke M.F. Wicha M.S. Cancer Res. 1995; Google Scholar). from this showed that antisense oligonucleotides be to the splicing of various genes in cell H. S. Kole R. Proc. Natl. Acad. Sci. U. S. A. 1996; PubMed Scopus Google Scholar, H. D. Kole R. 1999; PubMed Scopus Google Scholar, F. K. S. K. S. Kole R. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, Kole J. J.A. Kole R. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). that the alternative 5′-splice site in of Bcl-x with an antisense oligonucleotide shift splicing from Bcl-xL to Bcl-xS the of pro-apoptotic Bcl-xS protein and the of its anti-apoptotic that this shift in splicing be and that it induced apoptotic in the prostate cancer cell in the breast cancer cell splicing from Bcl-xL to Bcl-xS and of apoptosis were much less in the responses to oligonucleotide treatment were in the of cell of the two cell have of the that alternative splicing of Bcl-x pre-mRNA two with antagonistic and an oligonucleotide antisense to the 5′-splice site of Bcl-xL to shift splicing from the anti-apoptotic splice Bcl-xL, to the pro-apoptotic splice This approach be to antisense of Bcl-xL N.M. Oncogene. 1999; 18: PubMed Scopus Google Scholar, S. R. J. U. J. Cancer. 2000; PubMed Scopus Google Scholar) by a in Bcl-xL to a in the of the antagonistic Bcl-xS, the effects of the In an oligonucleotide targeted to the 5′-splice site of Bcl-x which to an in the Bcl-xS splice variant and a in the Bcl-xL splice was a of apoptosis in PC-3 cells an oligonucleotide targeted to the which the expression of the Bcl-xL of PC-3 and MCF-7 cells with the oligonucleotide to a and shift in splicing of Bcl-x pre-mRNA from the Bcl-xL to and to a in the of Bcl-xS of the Bcl-xL the Bcl-xL protein in the cells in the There be two for this the the Bcl-xL and the Bcl-xS splice variant. This from to the Bcl-xS the Bcl-xL protein be in the cells high its was The in PC-3 Bcl-xS apoptosis and/or cell death despite the of anti-apoptotic Since Bcl-xS to Bcl-xL and inhibits its anti-apoptotic A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google Scholar), to that the is or that this is the for the pro-apoptotic properties of A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google appears that the apoptotic response of MCF-7 cells to oligonucleotide treatment is to the of Bcl-xS protein by the shift in the that the of Bcl-xS protein to apoptotic regulatory that its be the and of the genes that be for the apoptotic resistance of MCF-7 cells and a shift in alternative splicing of Bcl-x pre-mRNA was in breast cancer cell R. and R. this work was in in a lung cell treated with N.M. 1999; 17: PubMed Scopus (189) Google Scholar). cell lines, as well as the gene is in PC-3 L. 1993; PubMed Scopus Google Scholar, Cancer Res. 51: Google Scholar) and Y. J. Cancer. 1999; PubMed Scopus Google Scholar) in N.M. 1999; 17: PubMed Scopus (189) Google Scholar) or MCF-7 Y. J. Cancer. 1999; PubMed Scopus Google Scholar) and Bcl-2 is in PC-3 Q.Y. Stein C.A. Clin. Cancer Res. 1997; 3: 2039-2046PubMed Google Scholar), Y. J. Cancer. 1999; PubMed Scopus Google Scholar), and N.M. 1999; 17: PubMed Scopus (189) Google Scholar) cells and high in MCF-7 cells Y. J. Cancer. 1999; PubMed Scopus Google Scholar). treatment with to significant apoptosis in PC-3 whereas a in Bcl-2 in MCF-7 cells to levels in PC-3 cells apoptosis. appears that the and bcl-2 genes play a in the of the effects on cellular In this it is that Bcl-xL was in 100% of prostate Bcl-2 was in (19Krajewska M. Krajewski S. Epstein J.I. Shabaik A. Sauvageot J. Song K. Kitada S. Reed J.C. Am. J. Pathol. 1996; 148: 1567-1576PubMed Google Scholar). in breast as well as in breast cancer cell lines, the levels of Bcl-2 were higher of J.M. M. Krajewski S. S. E. Reed J.C. Cancer Res. 1998; PubMed Scopus Google responses of PC-3 and MCF-7 cells to the shift in Bcl-x pre-mRNA splicing to of to of a apoptotic and to cell death by colony In this cell death in both PC-3 and MCF-7 cell lines, the of the of MCF-7 cells and of PC-3 cells the treatment with oligonucleotide for MCF-7 cells and for PC-3 that in a of Bcl-xL to Bcl-xS of the in of the of the oligonucleotide of and for MCF-7 and PC-3 a This is by of the oligonucleotide in MCF-7 cells. In to PC-3 in which oligonucleotide in the in MCF-7 the of the in it was to a process Res. N. Y. 1996; PubMed Scopus Google that cells in response to a shift in the to the in of the although Bcl-xL is expressed in a number of cell types, including cell S. M. Shabaik A. S. L. Reed J.C. Cancer Res. Google Scholar), the apoptotic of the oligonucleotide be in cells with a gene expression profile to the prostate cancer PC-3 cell In a of the oligonucleotide with by cisplatin in cells N.M. 1999; 17: PubMed Scopus (189) Google Scholar) types of cells to apoptosis. of cellular by the oligonucleotide be a in the of and in the of cancer have of the that alternative splicing of Bcl-x pre-mRNA two with antagonistic and an oligonucleotide antisense to the 5′-splice site of Bcl-xL to shift splicing from the anti-apoptotic splice Bcl-xL, to the pro-apoptotic splice This approach be to antisense of Bcl-xL N.M. Oncogene. 1999; 18: PubMed Scopus Google Scholar, S. R. J. U. J. Cancer. 2000; PubMed Scopus Google Scholar) by a in Bcl-xL to a in the of the antagonistic Bcl-xS, the effects of the In an oligonucleotide targeted to the 5′-splice site of Bcl-x which to an in the Bcl-xS splice variant and a in the Bcl-xL splice was a of apoptosis in PC-3 cells an oligonucleotide targeted to the which the expression of the Bcl-xL of PC-3 and MCF-7 cells with the oligonucleotide to a and shift in splicing of Bcl-x pre-mRNA from the Bcl-xL to and to a in the of Bcl-xS of the Bcl-xL the Bcl-xL protein in the cells in the There be two for this the the Bcl-xL and the Bcl-xS splice variant. This from to the Bcl-xS the Bcl-xL protein be in the cells high its was The in PC-3 Bcl-xS apoptosis and/or cell death despite the of anti-apoptotic Since Bcl-xS to Bcl-xL and inhibits its anti-apoptotic A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google Scholar), to that the is or that this is the for the pro-apoptotic properties of A.J. Boise L.H. Thompson C.B. J. Biol. Chem. 1996; 271: 6306-6312Abstract Full Text Full Text PDF PubMed Scopus (189) Google Scholar). appears that the apoptotic response of MCF-7 cells to oligonucleotide treatment is to the of Bcl-xS protein by the shift in the that the of Bcl-xS protein to apoptotic regulatory that its be the and of the genes that be for the apoptotic resistance of MCF-7 cells and For a shift in alternative splicing of Bcl-x pre-mRNA was in breast cancer cell R. and R. this work was in in a lung cell treated with N.M. 1999; 17: PubMed Scopus (189) Google Scholar). cell lines, as well as the gene is in PC-3 L. 1993; PubMed Scopus Google Scholar, Cancer Res. 51: Google Scholar) and Y. J. Cancer. 1999; PubMed Scopus Google Scholar) in N.M. 1999; 17: PubMed Scopus (189) Google Scholar) or MCF-7 Y. J. Cancer. 1999; PubMed Scopus Google Scholar) and Bcl-2 is in PC-3 Q.Y. Stein C.A. Clin. Cancer Res. 1997; 3: 2039-2046PubMed Google Scholar), Y. J. Cancer. 1999; PubMed Scopus Google Scholar), and N.M. 1999; 17: PubMed Scopus (189) Google Scholar) cells and high in MCF-7 cells Y. J. Cancer. 1999; PubMed Scopus Google Scholar). treatment with to significant apoptosis in PC-3 whereas a in Bcl-2 in MCF-7 cells to levels in PC-3 cells apoptosis. appears that the and bcl-2 genes play a in the of the effects on cellular In this it is that Bcl-xL was in 100% of prostate Bcl-2 was in (19Krajewska M. Krajewski S. Epstein J.I. Shabaik A. Sauvageot J. Song K. Kitada S. Reed J.C. Am. J. Pathol. 1996; 148: 1567-1576PubMed Google Scholar). in breast as well as in breast cancer cell lines, the levels of Bcl-2 were higher of J.M. M. Krajewski S. S. E. Reed J.C. Cancer Res. 1998; PubMed Scopus Google Scholar). The responses of PC-3 and MCF-7 cells to the shift in Bcl-x pre-mRNA splicing to of to of a apoptotic and to cell death by colony In this cell death in both PC-3 and MCF-7 cell lines, the of the of MCF-7 cells and of PC-3 cells the treatment with oligonucleotide for MCF-7 cells and for PC-3 that in a of Bcl-xL to Bcl-xS of the in of the of the oligonucleotide of and for MCF-7 and PC-3 a This is by of the oligonucleotide in MCF-7 cells. In to PC-3 in which oligonucleotide in the in MCF-7 the of the in it was to a process Res. N. Y. 1996; PubMed Scopus Google Scholar). The that cells in response to a shift in the to the in of the although Bcl-xL is expressed in a number of cell types, including cell S. M. Shabaik A. S. L. Reed J.C. Cancer Res. Google Scholar), the apoptotic of the oligonucleotide be in cells with a gene expression profile to the prostate cancer PC-3 cell In a of the oligonucleotide with by cisplatin in cells N.M. 1999; 17: PubMed Scopus (189) Google Scholar) types of cells to apoptosis. of cellular by the oligonucleotide be a in the of and in the of cancer for