Why do some people who have never smoked a cigarette in their lives develop lung cancer, while others who have smoked heavily since their teen-age years never show signs of the disease?
Scientists suspect that the interaction between an individual's genetic makeup and exposures to environmental toxicants determines the probability of getting a cancer.
Dr. Helmut Zarbl, an investigator in the Hutch's Program in Cancer Biology, aims to determine if particular genes are sensitive to the actions of chemical toxicants and how these genes are involved in the development of cancer.
Long-term exposure to toxicants - such as asbestos or tobacco smoke - can induce mutations (changes in the DNA sequence) in exposed cells, resulting in altered gene functions. In addition, environmental toxicants, also called mutagens, lead to altered patterns of gene expression in these cells that may allow for the rapid growth of mutated or pre-malignant cells.
To identify the genes that are sensitive to the effect of these chemicals, Zarbl has taken advantage of DNA microarray technology, which allows scientists to monitor the expression of thousands of genes at a time.
DNA arrays are glass slides onto which thousands of individual genes or portions of genes have been affixed. Researchers extract RNA - a molecule synthesized from active genes - from cells in a tissue sample, label it with a fluorescent dye and drop it onto the glass slide. The RNAs that adhere to their respective genes on the slide indicate which genes, in a given cell type at a given time, are expressed, revealing a molecular fingerprint.
"By comparing the molecular fingerprints from cells exposed to different doses of chemical toxicants, the sensitivity of a specific gene to that toxicant can be monitored," said Zarbl, an investigator in both the Human Biology and Public Health Sciences divisions.
Expression level after exposure
The genes that are sensitive to a particular toxicant can be identified by an increase or decrease in their expression level after exposure. The patterns of expression induced by different toxicants can be used to classify the environmental agents on the basis of the mechanisms of action. These substances can be ranked or assigned a place on a scale of relative risk for causing cancer.
In addition to the molecular fingerprinting of toxicants involved in cancer development, Zarbl collaborates with a number of investigators at the Hutch and the University of New Mexico to obtain molecular profiles of human tumors using DNA microarray technology.
With these profiles, the pattern of gene expression in tumors can be correlated with the tumor phenotype, clinical outcomes and response to specific therapeutic interventions. The hope is that an understanding of different patterns of gene expression will some day make it possible to tailor specific therapies to individual patients on the basis of the molecular changes in their tumor.
Zarbl also is developing a DNA microarray-based technology that will make it possible to simultaneously measure both mutations and expression levels of many specific genes implicated in cancer risk.
"This high throughput analysis may lead to the identification of combinations of mutations or genetic polymorphisms in individuals that may be predisposed to cancer," Zarbl said.
Dr. Barbara Trask, director of the Human Biology Division, said the work is an important new tool for understanding the interplay between environment and cancer.
"Helmut's unusual capacity to straddle the worlds of technology and biology is yielding powerful new methods for studying how exposure to chemical toxicants can cause cancer," she said.
Another method Zarbl uses to detect mutations within a population relies on the properties of the DNA.
The structure of DNA resembles a ladder with two strands of bases held together by bonds. Under high temperature, the bonds - analogous to rungs of a ladder - will break, separating the two strands. If a DNA sequence contains a mutation, the mutation will affect the way the DNA melts and the strands may not separate completely. By mapping out where the strands failed to separate, the location of the gene mutation can be identified.
This technique, called Constant Denaturant Capillary Electrophoresis (CDCE), makes it possible to simultaneously screen a large number of individuals for a specific mutation. Smaller pools of DNA can be screened to identify individuals carrying the specific mutation.
"Application of the CDCE method will be useful in screening populations for specific mutations that contribute to particular types of diseases, including cancer," Zarbl said.
His collaborators at the Massachusetts Institute of Technology and Northeastern University already have validated CDCE by screening large populations for mutations in the highly-studied HPRT gene. HPRT mutations in T lymphocytes used widely as biomarkers of environmental exposure and effect.
Zarbl's lab is developing CDCE assays in collaboration with several Hutch investigators to detect mutant genes involved in colon cancer and neurological diseases and to look for variants in HIV, the virus that causes AIDS.
[Dr. Karen Spratt is a postdoctoral fellow in the Kemp lab in the Human Biology Division.]