The Expression and Purification of a Bioactive Inflammatory Cytokine for Structural and Drug Discovery Applications
Dr. Lisa Warner
It is projected that 271,270 people in the U.S. will be diagnosed with breast cancer in 2019. Of those people with metastatic breast cancer, only 27% will live to see the 5-year mark. The pleiotropic IL-6 inflammatory cytokine Oncostatin M (OSM) has been shown to influence the metastasis of breast cancer. OSM signals through the LIF/JAK/STAT3 pathway and is secreted by immune cells associated with inflammatory response. When OSM collects at a stationary tumor site, causing chronic inflammation, the mobility of cancer cells is stimulated allowing for metastasis to occur. This overall aim of this research is to purify large quantities of OSM so that structural and drug discovery analyses can be conducted to further understand how OSM interacts with small molecule inhibitors (SMI’s). Successful optimization of an OSM expression scheme in E. coli that yields high expression by using a cleavable maltose binding protein (MBP) affinity tag for initial capture has been achieved. After removal of the MBP with tobacco etch virus protease, OSM is further purified by anion exchange and size exclusion chromatography. Future work will include testing a new affinity tagged (6His) version of the MBP construct, X-ray crystallography and co-crystallization with small molecule inhibitors that will work to shut down OSM and inhibit cancer cells from moving throughout the body. Thousands of lives could be saved by producing an SMI based oral medication that will hinder the metastasis of breast cancer cells, allowing time for tumor removal and returning the quality of life to patients.
Rushing, Brittany R.; Grantham, Briana R.; Woodbury, Luke; Mass, Olga; and Warner, Lisa, "The Expression and Purification of a Bioactive Inflammatory Cytokine for Structural and Drug Discovery Applications" (2020). 2020 Undergraduate Research Showcase. 162.