Antisense therapy for malignant mesothelioma with oligonucleotides targeting the bcl-xl gene product

Objective: Malignant pleural mesothelioma is resistant to conventional therapies and to apoptosis. The bcl-2 family genes are major determinants of apoptotic homeostasis. Malignant pleural mesothelioma lines and tumors rarely express the antiapoptotic Bcl-2 protein but routinely express the antiapoptotic protein Bcl-x1 and the proapoptotic proteins Bax and Bak. We have previously shown pharmacologic inhibition of bcl-x1 expression in malignant pleural mesothelioma can lead to apoptosis, so we sought to determine whether antisense oligonucleotides directed at bcl-x1 messenger RNA would engender apoptosis. possibly through a forced imbalance of bcl-2 family proteins. Methods: Malignant pleural mesothelioma lines REN (epithelial) and I-45 (sarcomatous) were exposed to modified bcl-x1 antissense oligonecleotides directed near the messenger RNA initiation sequence with and without a liposomal delivery system. Untreated cells and bcl-x1 sense oligonucleotides were controls. Cell viability was measured by colorimetric assay, and apoptosis was evaluated with Hoechst staining and sub-G, fluorescence-activated cell sorter analysis, Results: Bcl-x1 protein expression after antisense oligonucleotides was downwardly regulated in both cell lines relative to sense oligonucleotides (>65%). Significant cellular killing in both the I-45 and REN cell lines was achieved with antisense oligonucleotides (compared with sense oligonucleotides) without (P=.003 and .006, respectively) and with (P=.006 and .0005, respectively) liposomal delivery. Hoechst staining and sub-G, fluorescence-activated cell sorter analysis demonstrated apoptosis to be the mechanism of cellular death. Use of a liposomal delivery system increased therapeutic effect and allowed lower doses of antisense oligonucleotides. Conclusion: Antisense oligonucleotides directed at the bcl-x1 gene product engender apoptosis in mesothelioma cell lines. The therapeutic potential of inhibiting expression of this protein in mesothelioma should be evaluated.