In Cancer Cells, Silenced Genes Reveal Vulnerabilities
Last week, for instance, a team from NCI reported a novel way to find genes that a cancer cell needs to survive. The genes might not contain mutations or other alterations associated with cancer, but inhibiting them could potentially help control the disease.
The method is a genetic screen that uses RNA interference (RNAi) - a technology for silencing genes - to identify genes that, when silenced, cause cancer cells to die or stop dividing.
In a demonstration of the strategy, reported online in Nature, the researchers identified three genes that could be potential therapeutic targets for a type of lymphoma. None of the genes had previously been linked to cancer.
"This genetic screen could lead to a new realm of therapeutic targets beyond the small set of genes we have already identified," says lead researcher Dr. Louis Staudt of NCI's Center for Cancer Research (CCR).
The traditional way of searching for cancer drug targets is to identify genes that are consistently mutated or deregulated in cancer cells. This has produced, for instance, the leukemia drug imatinib (Gleevec) and the breast cancer drug trastuzumab (Herceptin).
But only so many genes are consistently altered in cancer cells. The genetic screen creates a new class of potential targets: genes involved in a cellular process, or pathway, that is necessary for a cancer cell's survival.
"We call it an Achilles heel genetic screen because it identifies the pathways in the cancer cell that are most vulnerable to attack," Dr. Staudt says, noting that it is technically called a loss-of-function RNAi genetic screen.
Previous genetic screens involving RNAi technology have identified genes that, when silenced, spur the growth of cancer cells. This method, in contrast, identifies genes that cancer cells cannot live without.
"Large-scale RNAi screens are not particularly new, but what Dr. Staudt has now shown is that you can use them for what we call negative selection," says Dr. René Bernards of the Netherlands Cancer Institute, who has developed RNAi genetic screens for cancer.
Negative selection refers to identifying potential targets in cells that have gone missing from a population of cultured cells.
"The study is an important proof of concept, and we're going to hear more about this in the future," adds Dr. Bernards.
The technological advance made by Dr. Staudt's team was an on/off switch that controls the silencing of genes in living cells. Until now, delivering certain types of short hairpin interfering RNA molecules, or shRNAs, into cells could kill the cells immediately.
In their demonstration experiment, the researchers silenced 2,500 genes in 2 types of diffuse large B cell lymphoma cells - activated B cell-like (ABC) and germinal center B cell-like (GCB).
"Our hypothesis was that we should find different genes that are required for the proliferation or survival of the lymphomas because these are very different diseases clinically," says Dr. Vu N. Ngo, also of CCR, who led the experiment.
The first step was to grow cultures of lymphoma cells. Each cell then received a modified virus containing genetic code for producing a single shRNA. Three weeks later, the researchers added drugs to the cell cultures to trigger the production of shRNAs, thereby silencing one gene per cell.
Next, the researchers determined which cells and genes had been eliminated from the populations using microarray technology and molecular tags attached to the shRNAs.
Further research showed that three of the missing genes - CARD11, MALT1 and BCL10 - are required to turn on a pathway that is continuously activated in patients with the ABC type of lymphoma but not the GCB type.
"The genetic screen revealed a new mechanism in this lymphoma that we didn't know about before," says Dr. Staudt, noting that genetic screens often yield surprises.
His laboratory intends to screen cell cultures representing all types of lymphomas and eventually all types of human cancers.
The method, he suggests, could be used to create a new classification of cancers based not on the type of cancer, but on which pathways inside a cancer cell are critically required for its proliferation or survival.
The results illustrate the concept that some genetic mutations are toxic only when they occur with other mutations or in the absence of gene activity. Knowing which interactions are toxic in specific cancers could greatly help in developing therapies.
An article about how discovering context-specific genetic interactions could lead to new cancer drugs was published in Science in 1997 by Drs. Leland Hartwell and Stephen Friend of the Fred Hutchinson Cancer Research Center.
"This was long before any tools had been developed to detect these genetic interactions," notes Dr. Bernards.
Now that the tools are available, they will be used, he says, because there are not enough good drug targets out there to treat the major forms of cancer in the next decade.
"I am absolutely convinced that the next Gleevec will come out of a genetic screen like this one," Dr. Bernards adds.