Robert A. Olie

Induction of apoptosis in lung-cancer cells following bcl-xL anti-sense treatment.

Over‐expression of the anti‐apoptotic protein bcl‐xL is frequently found in lung cancer where it potentially contributes to tumor development, progression and drug resistance. To override the apoptotic block in lung‐adenocarcinoma and small‐cell‐lung‐cancer (SCLC) cells caused by over‐expression of bcl‐xL, an anti‐sense oligodeoxynucleotide was designed targeting a sequence unique to the bcl‐xL coding region and not shared by the pro‐apoptotic splice variant bcl‐xS. Moreover, to improve the biophysical properties of the anti‐sense compound, 2`‐methoxy‐ethoxy modifications were made to selected deoxy‐ribose residues. The resulting gapmer oligonucleotide 4259 was tested on lung‐adenocarcinoma and SCLC cell lines in vitro. Treatment of the adenocarcinoma cell lines A549 and NCI‐H125 and the SCLC cell lines SW2 and NCI‐H69 with 600 nM 4259 reduced bcl‐xL levels by 70 to 90%. In the lung‐adenocarcinoma cell lines, apoptosis was induced, as indicated by caspase‐3‐like protease activation and nuclear condensation and fragmentation. In contrast, in the SCLC cell lines, no induction of apoptosis could be demonstrated. These findings imply that bcl‐xL is a more critical survival factor for lung adenocarcinomas than for SCLC, and suggest the use of oligonucleotide 4259 for therapy of this major sub‐type of lung cancer. Int. J. Cancer 86:570–576, 2000.

Bcl-xl antisense treatment induces apoptosis in breast carcinoma cells.

Upregulated expression of bcl‐xL is involved in the initiation and progression of breast cancer by inhibiting tumor cell apoptosis. Here we describe the use of the 2`‐O‐methoxy‐ethoxy antisense oligonucleotide 4259 targeting nucleotides 687–706 of the bcl‐xL mRNA, a sequence that does not occur in the pro‐apoptotic bcl‐xS transcript, to restore apoptosis in estrogen‐dependent and independent breast carcinoma cells. The antisense effect of oligonucleotide 4259 was examined on the mRNA and protein level using real‐time PCR and Western blot analysis, respectively, and the induction of cell death was investigated in viability and apoptosis assays. Treatment of MCF7 cells with oligonucleotide 4259 at a concentration of 600 nM for 20 hr decreased bcl‐xL mRNA and protein levels by more than 80% and 50%, respectively. This resulted in the induction of apoptosis characterized by mitochondrial cytochrome c release, decrease of mitochondrial transmembrane potential, and the appearance of condensed nuclei in approximately 40% of cells. Moreover, oligonucleotide 4259 efficiently downregulated bcl‐xL expression and decreased cell growth in the breast carcinoma cell lines T‐47D, ZR‐75‐1, and MDA‐MB‐231. Our data emphasize the importance of bcl‐xL as a survival factor for breast carcinoma cells and suggest that oligonucleotide 4259 deserves further investigations for use in breast cancer therapy. Int. J. Cancer 87:582–590, 2000.