Activity of a novel bcl-2/bcl-xL-bispecific antisense oligonucleotide against tumors of diverse histologic origins.
BACKGROUND
Increased expression of the anti-apoptotic proteins Bcl-2 and Bcl-xL is involved in the development and progression of many tumors. We recently reported that the bcl-2/bcl-xL-bispecific antisense oligonucleotide 4625 induces apoptosis in lung carcinoma cells. To further assess the therapeutic potential of oligonucleotide 4625, we investigated its effect on a series of human tumor cell lines of diverse histologic origins in vitro and in vivo.


METHODS
Oligonucleotide 4625-mediated inhibition of bcl-2 and bcl-xL expression in vitro was measured in breast carcinoma cells with the use of reverse transcription-polymerase chain reaction (PCR), real-time PCR, and western blotting. Cytotoxicity was assessed in several different cell lines by measurement of tumor cell growth, propidium iodide uptake, and nuclear apoptosis. The in vivo activity of oligonucleotide 4625 was determined by the inhibition of growth of established tumor xenografts in nude mice, immunohistochemical staining of Bcl-2 and Bcl-x proteins in the tumors, and western blotting of tumor lysates. Apoptosis in tumor xenografts was detected with the use of in situ TUNEL (i.e., terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-digoxigenin nick end labeling) staining. All statistical tests are two-sided.


RESULTS
In breast carcinoma cells, oligonucleotide 4625 treatment reduced bcl-2 and bcl-xL messenger RNA levels in a dose-dependent manner. At 600 nM:, oligonucleotide 4625 reduced Bcl-2 and Bcl-xL protein levels to 25% (95% confidence interval [CI] = 16% to 34%) and 20% (95% CI = 14% to 26%), respectively, of the levels in untreated cells and it decreased viability in all cell lines mainly by inducing apoptosis. In vivo, oligonucleotide 4625 statistically significantly inhibited the growth of breast and colorectal carcinoma xenografts by 51% (95% CI = 28% to 74%) and 59% (95% CI = 44% to 74%), respectively, relative to those treated with control oligonucleotide 4626; it also reduced Bcl-2 and Bcl-xL protein levels and induced tumor cell apoptosis.


CONCLUSIONS
The bcl-2/bcl-xL-bispecific antisense oligonucleotide 4625 merits further study as a novel compound for cancer therapy.
Division of Oncology, Department of Internal Medicine, University Hospital, Zurich, Switzerland.
O
O
Gautschi
S
S
Tschopp
R A
RA
Olie
S H
SH
Leech
A P
AP
Simões-Wüst
A
A
Ziegler
B
B
Baumann
B
B
Odermatt
J
J
Hall
R A
RA
Stahel
U
U
Zangemeister-Wittke
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000188:drug therapy; D002277:Carcinoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D015179:Colorectal Neoplasms / Q000188:drug therapy; D004305:Dose-Response Relationship, Drug; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D008175:Lung Neoplasms / Q000188:drug therapy; D008297:Male; D008545:Melanoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D009841:Oligonucleotides / Q000494:pharmacology* / Q000627:therapeutic use; D016376:Oligonucleotides, Antisense; D020320:Oligoribonucleotides, Antisense / Q000494:pharmacology* / Q000627:therapeutic use; D011471:Prostatic Neoplasms / Q000188:drug therapy; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors* / Q000187:drug effects / Q000235:genetics; D012333:RNA, Messenger / Q000187:drug effects; D012334:RNA, Neoplasm / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012878:Skin Neoplasms / Q000188:drug therapy; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured; D051020:bcl-X Protein
D000970:Antineoplastic Agents; C494811:BCL2L1 protein, human; C494812:Bcl2l1 protein, mouse; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D020320:Oligoribonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D051020:bcl-X Protein; C423963:oligonucleotide 4625