In the United States and most other developed countries, lung cancer is the leading cause of cancer-related death. Notoriously hard to detect in its early stages, the overall five-year survival rate for the disease is only 14 percent.
Maik Hüttemann, Ph.D., associate professor of molecular medicine and genetics in Wayne State University’s School of Medicine, along with research associate Jeffrey Doan, Ph.D. and Ph.D. student Christopher Sinkler, are working to improve those odds by developing a screening technique for a gene that may be indicative of lung cancer. This patented platform technology utilizes probe ligation and rolling circle amplification to screen for any target gene in many different types of diseases.
To begin, though, the research has centered on COX4-1 and COX4-2, genes encoding variants of a subunit of the important enzyme cytochrome c oxidase (or COX), the “pacemaker of aerobic metabolism.”
In healthy cells, the primary type of metabolism is aerobic oxidative phosphorylation in the mitochondria. This type of metabolism most efficiently produces adenosine triphosphate (ATP), biological energy that fuels most of the body’s processes. In cancer cells, however, the predominant type of metabolism is glycolysis, an anaerobic process. Products of glycolysis and associated pathways are required for the rapidly proliferating cells.
“The switch from aerobic to glycolytic metabolism reflects the cancer cell’s need for building blocks for DNA, lipid, and protein biosynthesis,” Hüttemann says.
In an ongoing study, Hüttemann and colleagues found that the expression of lung-specific COX4-2 is dramatically downregulated in lung cancer. This was measured against the somatic form of the gene, COX4-1, which is expressed at similar levels in cancer and non-cancer tissue. The finding holds for various types of lung cancer – approximately 80 samples were tested – including a cell culture model simulating carcinogenesis in vivo.
“In that model, COX4-2 expression is downregulated from the earliest stages,” he says. “And, this makes sense because we have shown that COX4-2 expression makes cells more aerobic, which is the opposite of what the cancer wants. Therefore, our results suggest that COX4-2 expression is a mechanism-based marker for lung cancer.”
Hüttemann’s current work aims to expand on these findings by investigating expression levels of COX4-2 with greater accuracy. He is developing probe ligation and rolling circle amplification techniques to detect levels of both COX4-1 and COX4-2 expression. (Decreased COX4-2 expression may indicate the presence of cancer.)
The techniques entail Hüttemann’s lab synthesizing DNA segments, or “probes,” that recognize and attach to the target gene transcript. Using DNA polymerase and a circular DNA template, DNA generation amplifies the circular DNA 1000-fold. Fluorescent molecules are then attached to the amplified DNA.
“Using the fluorescence, the signal of each probe is amplified, enabling our lab to locate and quantify the levels of COX4-1 and COX4-2 expression with much greater accuracy than we ever could before,” Hüttemann says.
Hüttemann’s group is currently working to further develop the techniques into a robust working assay in the lab. Once this is achieved, the next step will be to test it using clinical specimens from lung cancer patients.
Improved ability to measure the expression levels of the two COX-subunit genes may result in a test for the early screening of lung cancer.
“Only 15 percent of lung cancer tumors are detected in time to do something about it,” Hüttemann says. “Our hope with this assay is to develop a robust screening method. In particular, individuals at high risk for developing lung cancer, including smokers, would benefit from such a test.”
Although lung cancer is Hüttemann’s primary interest, he also aims to develop the assay as a platform technology to screen for expression levels of any gene.
“Although this step is further into the future, if we can successfully develop the technology to detect expression of a gene of interest and amplify its signal, our ability to screen for specific gene expression will increase dramatically,” Hüttemann says. “This, in turn, may help in the early detection of not only different kinds of cancers, but perhaps other diseases such as neurodegenerative disorders, diabetes, and innumerable other health problems with a genetic footprint.”
To learn more, visit http://genetics.wayne.edu/faculty/huttemann/index.php.