Modifying Lung Cancer Apoptosis Through Alternative RNA Splicing

Lung cancer currently causes the most cancer-related deaths nationally and is a particularly heavy burden for Kentucky. More effective treatment strategies are essential to better control this disease. Defects in the pathways controlling cell death, also known as apoptosis, contribute both to the formation of tumors and the frequent failure of anti-cancer treatments. While most tumor cells retain the ability to undergo apoptosis, they have an altered balance between positive and negative apoptosis factors and thus survive signals that should induce death. Many apoptosis factor genes encode multiple gene products with opposite functions through the process of alternative RNA splicing. Thus, changes in alternative RNA splicing regulation, an important mechanism of gene regulation known to be altered in cancer cells, is likely to be a contributing factor in altering cellular apoptotic responses. Previous work with the Bcl-x gene has shown that an oligonucleotide that enhances pro-apoptotic Bcl-xS and decreases anti-apototic Bcl-xL mRNA levels makes the cells more sensitive to chemotherapy agents and radiation treatment. This provides a proof-of-principle that targeting alternative splicing of apoptosis genes alters the sensitivity of cells to death induced by anti-cancer treatments. I propose that modifying the splice patterns of multiple apoptosis factors to enhance expression of their pro-apoptotic forms in lung cancer cells will sensitize the cells to anti-cancer treatments; a combination of effective oligonucleotides, used in conjunction with standard treatments for lung cancer, will enhance treatment of drug-resistant tumors and lower the effective treatment dose to be less toxic to the patient. To test this hypothesis, this grant will characterize the RNA splicing patterns and expression levels of a set of apoptosis genes in a panel of lung tumor cells and normal lung, design and test oligonucleotides to redirect splice patterns of target apoptosis genes, and test oligonucleotide-transfected cells for increased sensitivity to drugs and radiation. At the end of two years I plan to have identified a set of genes whose processing and/or expression can be altered with oligonucleotides to sensitize lung cancer cells to anti-cancer treatments. In the future, this work will be expanded to include preclinical animal models and will move toward translation applications.