UAB (University of Alabama at Birmingham) researchers have developed a new, inexpensive and efficient method for producing and studying a type of human papillomavirus (HPV) that causes cervical cancer.
The process could speed understanding of how the virus functions and causes diseases, and lead to new prevention or treatment options.
In findings reported on-line and in print in January in Genes & Development, the UAB team detailed a process for producing HPV-18 in the laboratory. Previously, the virus had proven resistant to propagation in a lab setting, making it extremely difficult for scientists to study the virus and its effects on the host cells that it infects.
"The old method for propagating papillomaviruses in the lab for study was compromised by several factors," said Louise Chow, Ph.D., professor of biochemistry and molecular genetics at UAB and a study co-author. "We could only look at the viral DNA gene by individual gene, which gave us little insight into how the entire virus coordinated its replication program or how it interacted with the host cells and tissues that had been infected."
The new method, which Chow and study co-author Thomas Broker, Ph.D., professor of biochemistry and molecular genetics, have been developing for over 20 years, for the first time allows researchers to reproduce the entire infection cycle of HPV-18 in primary human skin cells, called keratinocytes. The breakthrough is the result of several years of intensive and creative efforts by graduate students Hsu-Kun (Wayne) Wang and Aaron Duffy, coauthors of the publication. Scientists now can observe how the virus behaves in the same cells it would infect in a human body.
"This system provides marvelous opportunities to understand how HPV works on a very basic, molecular level," Broker said. "The ongoing research will reveal promising targets for drug design, better understanding of how the new prophylactic HPV vaccines block infection and, for the first time, offer real hope that we can find a way to combat a virus that potentially affects 80 to 85 percent of women in the world, through therapeutic treatment of established infections."
Chow and Broker say the old method for producing HPV cells suffered from several factors, including the need to use bacterially derived recombinant DNA to introduce the viral DNA into skin cells. The presence of bacterial DNA sequences altered the HPV DNA so that it no longer closely resembled the natural viral structures found in human infections. Their new method generates circular viral genomes in the cells with high efficiency, and these can go on to replicate autonomously, without complications associated with the usual gene transfer vectors.
Another stumbling block was the need to use immortalized host cells to grow HPV. Immortalized cells have been genetically altered to live indefinitely and they do not differentiate properly into full-thickness skin tissue. While the use of immortalized cells as a host allowed the viral genome to be studied gene by gene, immortalized cells do not resemble the actual primary cells found in the body, preventing meaningful observations of how HPV functions in normal human host cells.