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

J Thorac Cardiovasc Surg. 2002 Jun;123(6):1191-8. doi: 10.1067/mtc.2002.121684.

Abstract

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-xl and the proapoptotic proteins Bax and Bak. We have previously shown pharmacologic inhibition of bcl-xl expression in malignant pleural mesothelioma can lead to apoptosis, so we sought to determine whether antisense oligonucleotides directed at bcl-xl 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-xl antissense oligonecleotides directed near the messenger RNA initiation sequence with and without a liposomal delivery system. Untreated cells and bcl-xl sense oligonucleotides were controls. Cell viability was measured by colorimetric assay, and apoptosis was evaluated with Hoechst staining and sub-G(1) fluorescence-activated cell sorter analysis.

Results: Bcl-xl 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(1) 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-xl gene product engender apoptosis in mesothelioma cell lines. The therapeutic potential of inhibiting expression of this protein in mesothelioma should be evaluated.

MeSH terms

  • Apoptosis*
  • Colorimetry
  • Down-Regulation
  • Flow Cytometry
  • Gene Expression Regulation, Neoplastic*
  • Humans
  • Mesothelioma / genetics
  • Mesothelioma / metabolism
  • Mesothelioma / therapy*
  • Oligonucleotides, Antisense / therapeutic use*
  • Pleural Neoplasms / genetics
  • Pleural Neoplasms / metabolism
  • Pleural Neoplasms / therapy*
  • Proto-Oncogene Proteins c-bcl-2 / metabolism*
  • RNA, Messenger / metabolism
  • bcl-X Protein

Substances

  • BCL2L1 protein, human
  • Oligonucleotides, Antisense
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Messenger
  • bcl-X Protein