Antisense RNA down-regulation of bcl-xL Expression in prostate cancer cells leads to diminished rates of cellular proliferation and resistance to cytotoxic chemotherapeutic agents.
bcl-xL is a M(r) 26,000 bcl-2 homologue that is highly expressed in prostate cancer cells. In previous studies, the down-regulation of its expression by antisense oligonucleotides led to resistance. In this work, the 445-bp 5' terminus of the bcl-xL cDNA was cloned in the antisense orientation and stably transfected into DU145 and LNCaP prostate cancer cells. In the DU145 (and to a lesser extent the LNCaP) transfectants, phenotypic changes (versus mock-transfected cells) included an increase in doubling time (from 36 to 175 h) in the clone in which bcl-xL protein expression was 25% of control. The transfectants did not demonstrate characteristic apoptotic changes, as demonstrated by 4',6-diamidino-2-phenylindole staining, lack of either DNA laddering, caspase-3 activation, or poly(ADP)ribose and lamin cleavage, and the absence of a significant sub-G(0) population. Cell cycle analysis demonstrated an increase in a tetraploid population (from 28% to 66%), as well as the appearance of a hypertetraploid population. Levels of cIAP-1 protein were almost undetectable in the mock cells but increased at least 25-fold in the DU145 transfectants. The down-regulation of bcl-xL in both DU145 (and to a much lesser extent in LNCaP) cells led to their resistance to cytotoxic agents, including docetaxel, mitoxantrone, etoposide, vinblastine, and carboplatin. Reversion of bcl-xL expression in stable DU145 transfectants to nearly the levels found in the mock-transfected cells was accomplished by retroviral infection of the cells with a bcl-xL sense cDNA under control of a prolific promoter. This led to a dramatic increase in the growth rate and in BrdUrd incorporation, as well as a sharp decrease in the expression of cIAP-1 protein. Overall, these findings highlight the adaptability of prostate cancer cells to loss of bcl-xL and suggest that in addition to its prosurvival role, bcl-xL protein may also be involved in the regulation of the rate of cellular proliferation.
Vilenchik
Maria
M
Department of Medicine, Columbia University, College of Physicians and Surgeons, New York, New York 10032, USA.
Raffo
Anthony J
AJ
Benimetskaya
Luba
L
Shames
David
D
Stein
C A
CA
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000235:genetics; D002455:Cell Division / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D006801:Humans; D008297:Male; D011003:Ploidies; D011471:Prostatic Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis* / Q000235:genetics; D016372:RNA, Antisense / Q000235:genetics*; D014162:Transfection; D051636:X-Linked Inhibitor of Apoptosis Protein; D051020:bcl-X Protein
D000970:Antineoplastic Agents; C494811:BCL2L1 protein, human; D004273:DNA, Neoplasm; D011506:Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016372:RNA, Antisense; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; D051020:bcl-X Protein