Selection of Aptamers for CED-9/Bc1-2 Family Cell Death Regulators and Their Application in Study of Apoptosis Regulation and Drug Design for Breast Cancer

摘要

Programmed cell death (apoptosis) plays an essential role in maintaining the physiological balance of appropriate cell numbers by opposing uncontrolled cell proliferation. The pathway of programmed cell death appears to be highly conserved from C. elegans to humans, suggesting that studies of programmed cell death in C. elegans can provide important information for understanding how cell death is regulated and executed in humans. Moreover, novel means developed in C. elegans to modulate programmed cell death may also be applied to humans for better detection, prevention as well as treatment of human diseases caused by abnormal apoptosis (e.g. cancer, autoimmune disorders, and neurodegenerative diseases). In this study, we are employing the technique of SELEX (Systematic evolution of ligands by exponential amplification) to identify small RNA aptamers with high binding specificity and affinity for key cell death regulators, including CED-9 and CEDA-4 from C. elegans and Bcl-2 and Bcl-xL from humans. We hope to use these RNA aptamers to probe how Bcl-2 family proteins regulate programmed cell death in both C. elegans and mammalian cells. Importantly, if these RNA aptamers can be used to modulate apoptosis in C. elegans or mammalian cells, they may provide important insights into devising new diagnostic and therapeutic drugs to treat cancer and various apoptosis- related diseases. So far, we have successfully obtained RNA aptamers that bind CED-9 with Kds of approximately 10 nM. This high binding affinity will allow us to further study the effects of these aptamers in regulating apoptosis using both in vitro and in vivo assays: We have also conducted several rounds of SELEX experiments on CEDA and Bcl-xL and have obtained candidate molecules with increasing binding affinity in vitro to these two proteins.