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But this week in the journal Nature*, a team of scientists report that in the future, there may be a new way around this old problem. Using DNA microarray technology--a powerful new research tool that can record the expression patterns of thousands of genes at once--the group was able to show that, as currently defined, diffuse large B-cell lymphoma, the most common form of non-Hodgkin's lymphoma (NHL), is actually two distinct diseases.
Louis Staudt, M.D., Ph.D., a scientist at the National Cancer Institute (NCI) and a senior author on the paper, said this finding helps to explain why about 40 percent of patients with this type of NHL can be cured with standard chemotherapy regimens, while other patients who seemingly have the same disease often relapse. "It's a case of mistaken identity," said Staudt. "The tumor cells might look very similar, but this study offers strong evidence that their molecular engines work very differently."
This week's paper, one of the first of its kind in medical research, also adds to the growing interest among scientists to create molecular profiles of common cancers. "This finding offers one of the first glimpses into how cancer will be diagnosed in the future," said NCI Director Richard Klausner, M.D. "It will be based on well-characterized biological differences among tumor cells that tell us more precisely how aggressive a tumor will be and how best to treat it."
Added Patrick Brown, M.D., Ph.D., a senior author on the paper and a scientist with the Howard Hughes Medical Institute at Stanford School of Medicine, "A DNA microarray is like a microscope for watching a living genome in action. They allow us for the first time to see a detailed picture of the gene expression program in each tumor. This program specifies the design of the cell and the script for its behavior."
In approaching this study, Staudt and his colleagues began by mining data stored in the NCI's Tumor Gene Index database, a publicly available catalogue of genes expressed in various normal and cancerous cells. According to R. Eric Davis, Ph.D., an NCI scientist and a leading author on the paper, he and his collaborators began by teasing out of the database thousands of genes that are uniquely expressed in immune B cells, an indication that they are important to the biology of these cells.
"We arrayed on a specialized glass-slide chip more than 15,000 of these unique genes with about 3,000 other genes that are involved in various cancers or in the immune system," said Ash Alizadeh, a scientist at Stanford University School of Medicine and one of the lead authors on the study. "This customized microarray, which is slightly larger than a penny and contains over 18,000 genes in all, is known as a 'Lymphochip.'" The principal members of the Stanford laboratory team are Brown, Alizadeh, Michael Eisen, Ph.D. (a lead author), and David Botstein, Ph.D.
Alizadeh said that by collecting and then tagging gene transcripts with a flourescent dye, they can later see on the Lymphochip which of the known arrayed genes are active in normal or cancerous B cells. Depending on the distinct color and intensity of the flourescent glow of each arrayed gene, they also can determine at which level each gene is expressed. "The idea was to profile the genetic activity of B cells at various stages of their development," said Alizadeh. "This gave us a point of comparison when we performed the same profiles in cancerous B cells."
Alizadeh said that he and his collaborators performed their first analysis on three common types of B-cell cancers: follicular lymphoma, chronic lymphocytic leukemia, and diffuse large B-cell lymphoma, or DLBCL.
But as the experiments proceeded, the scientists discovered something interesting. The DLBCL samples showed distinct differences in their expression of hundreds of genes, suggesting that DLBCL might be more than one cancer. To test this hypothesis, the scientists focused their attention on the expression of genes that are known to be important when B cells reside in the
so-called germinal center of the lymph node, a key point in their maturation process.
The analysis showed two distinct patterns of gene expression, signifying that they were looking at two subtypes of the cancer. In one of the subtypes, called "GC B-like," the expression of germinal center genes was largely congruent with that of normal B cells. But in the second subtype, known as "activated B-like," these genes were expressed at low or undetectable levels. This finding-that the subtypes express germinal center genes differently--pointed to major biological differences among them.
Staudt said he and his colleagues next wondered whether these differences in the two subtypes influenced a patient's prognosis, further supporting the idea that these were biologically distinct forms of DLBCL.
In a pilot study of 42 previously untreated DLBCL patients who were being treated with anthracycline-based chemotherapeutic regimens, the scientists discovered significant differences in five-year survival rates among the subtypes. For patients with "GC B-like," three-fourths were alive at the five-year mark; for "activated B-like," fewer than one fourth were still living.
"Genetic profiling allows scientists to define distinct molecular subtypes of common tumors with unique clinical and biologic behaviours," said Wing Chan, M.D., an author on the paper and a scientist at the University of Nebraska Medical Center. "In the future, this hopefully will allow us to develop better, more targeted treatment for patients."
Ronald Levy, M.D., an author on the paper and a scientist at the Stanford University Medical Center, agrees. "As this finding shows, genetic profiling of tumors will offer extra power in predicting which patients will do well and which will do poorly," he said. "This more precise diagnosis of a developing cancer should help in the future in more accurately guiding a patient's treatment decisions."
According to Staudt, a follow-up study is already under way to verify and extend the group's finding, a standard practice in science. Diffuse large B-cell lymphoma is diagnosed in over 25,000 Americans each year. At a time when many cancers are decreasing in incidence in the United States, this lymphoma is increasing, making it a growing public health concern.
Staudt also stressed that this work involved an extremely productive collaboration involving scientists at institutions throughout the United States. These include: Stanford University School of Medicine in Palo Alto, Calif.; Research Genetics in Huntsville, Ala; University of Nebraska Medical Center in Omaha, Neb.; Johns Hopkins School of Medicine in Baltimore; and Walter Reed Army Medical Center in Washington, D.C.
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