Hope from HOXB13, New Variant Implicated in Prostate Cancer Risk
Advances in targeted sequencing technology help scientists track down a rare variant linked to significantly increased risk of prostate cancer.
Scientists at the Translational Genomics Research Institute (TGen), the University of Michigan, Johns Hopkins, University of North Carolina, and Wake Forest recently published a novel variant linked to increased risk of familial prostate cancer that they hope will one day be used as a diagnostic tool in the clinic. The study, published in January 2012 in the New England Journal of Medicine, analyzed families that had first-degree relatives diagnosed with prostate cancer, focusing on a region of chromosome 17 that has in the past been implicated in this kind of prostate cancer. The researchers found a variant (G84E) in the HOXB13 gene that appears to increase a carrier’s risk for prostate cancer by an order of magnitude.
John Carpten, Ph.D., Director of the Integrated Cancer Genomics Division at TGen, was one of the scientists involved in the work — and after years spent studying familial prostate cancer, he says that a new technology acquired by TGen enabled the team to find a variant where earlier methods of studying this genetic region had been unsuccessful.
Carpten got started in prostate cancer research back in the mid ’90s. “The crux of my postdoctoral fellowship was to identify inherited risk factors for prostate cancer, particularly familial prostate cancer where there was clustering of three or more affected men with prostate cancer within a nuclear family,” he says. At the time, that meant using “a series of genetic markers spaced about 10 million bases apart across the genome to identify regions of linkage.” As Carpten’s career progressed — he went on to become an investigator at the National Human Genome Research Institute at NIH before joining TGen in 2003 — he branched out to other forms of cancer, but prostate cancer continued to be an area of interest.
That theme has been helped along by a longtime collaboration with Kathleen Cooney, M.D., at the University of Michigan Health System. Over time, Cooney has recruited hundreds of families that have at least two or three first-degree relatives who have been diagnosed with prostate cancer; studies of these families implicated the Q arm of chromosome 17 in the disease. “Kathy and I have collaborated through the years to refine that region of linkage to try to minimize it down to the smallest region possible where a gene might reside that would be associated with elevated risk in those families,” Carpten says. The next step was to track down variants in the genetic region that might be playing a role in prostate cancer susceptibility, but the tools available at the time — PCR and Sanger sequencing — “limited our abilities to efficiently screen more broadly across larger sets of exons in larger cohorts,” Carpten says.
The turning point for getting this particular study up and running came when TGen purchased the RainDance RDT 1000, a platform used for targeted sequencing. “After my introduction to the RainDance technology,” Carpten says, “one of my lab members, Christiane Robbins, became an expert in the use of RainDance exon enrichment and sequencing using next-generation sequencing technologies.” He contacted Cooney at Michigan “and told her we had a technology up and running at TGen which would allow us to actually PCR amplify all of the exons in her region of interest in a single assay— about 200 genes, about 2,500 exons. We could amplify and sequence them all using RainDance and next-generation sequencing technologies.” They teamed up, submitted a supplement to a prostate cancer research grant Cooney had been awarded from the National Cancer Institute, and got the green light for the new project.
The difference between the RainDance platform and earlier approaches, Carpten says, really came down to flexibility and ease of customization. “The primary thing was the flexibility — that we weren’t locked into any particular panel, that we could actually customize a panel of exons specifically for our purpose,” he notes. “The flexibility of RainDance allowed us to design primers specifically around those genes that were of interest to us, and that included all of the genes that mapped to our region on chromosome 17 as well as some other interesting candidate prostate cancer genes throughout the genome.” He also credits the RainDance team for “their help in designing the assays and quality control. It was a great experience.”
With the technology in hand, the success of the project came down to building the best collaboration to perform the study. TGen would handle the targeted sequencing. Michigan’s Cooney would contribute her expertise on chromosome 17 as well as her family collection, and more families came from William Isaacs at Johns Hopkins University. “Bill is very, very well known in this space and has done as much as anyone in the world in hereditary prostate cancer research,” says Carpten, adding that Isaacs also provided key insight into study design. Other critical components of the study came from partners at the University of North Carolina, led by Ethan Lange, a statistical genetics expert who “spoke a lot into the design … and which were the most powered families, which was the best individual within the family to sequence,” Carpten says. Finally, Jianfeng Xu headed up the group at Wake Forest University, which handled validation work for variants detected earlier in the process, genotyping them in “several thousand population-based controls and several thousand sporadic prostate cancer cases,” according to Carpten.
The G84E variant discovered by the team is not unlike BRCA1 or other cancer-linked mutations already being used in the clinic. “The variant is exceedingly rare, but the effect is exceptionally high,” Carpten says. “Not that many people have it, but if you have it your risk of prostate cancer goes way up.” In the paper, the authors note that the mutation was found in just 0.6 percent of people diagnosed with late-onset, non-familial prostate cancer, compared with 3.1 percent in men with early-onset familial prostate cancer.
The clinical implications of this led the collaborators to publish in a medical, rather than a purely scientific, journal. “From our perspective,” Carpten says, “the translational aspect of the work far outweighs the impact of another cool sequencing study.” The variant is more than just another genetic mutation: it’s “a possible novel early diagnostic test for prostate cancer,” he says. “If we can identify men within families that are at extremely high risk of developing prostate cancer, perhaps we can think about early surveillance, early monitoring, and early detection — and possibly save those men’s lives.”
Aside from his high hopes for the clinical utility of this risk variant, Carpten also believes that the study serves as validation of the target enrichment and next-gen sequencing approach used by his team. While there are more genetic regions of interest for which Carpten says “I would love to be able to apply the same approach,” the bigger picture is paving the path for other researchers. “We hope that others will follow our lead in applying this approach to identify other genes associated with hereditary prostate cancer,” he adds.
“The important take-home message is that there are probably rare variants out there that play a significant role in cancer predisposition, and the availability of technologies such as RainDance and next-generation sequencing now allow us to survey large numbers of genes and exons to potentially identify those rare variants that we have not historically been able to capture using array-based technologies,” Carpten says.
— Meredith Salisbury
Germline Mutations in HOXB13 and Prostate-Cancer Risk