BOSTON -- More and more mammographies are beingperformed in the U.S., as women's awareness of breast-cancerrisk increases. That spells good news for diagnosis and therapy,but, paradoxically, bad news of sorts for research.
"Because of the increased use of mammography," observedmolecular oncologist Suzanne Fuqua, "we are now findingsmaller and smaller lesions. And because of that," she toldBioWorld, "researchers such as myself are faced with theproblem of finding technologies amenable to very, very smalltissue samples."
Fuqua described her problem, and its latest solution, at a one-day seminar here last week on "The Molecular Basis for GeneExpression." Sponsored by Applied Biosystems, a division ofPerkin-Elmer Corp., this was the last of eight such high-techroad shows on the same topic. Between May 3 and 13, it playedCosta Mesa, Calif., Seattle, Chicago, Research Triangle Park, N.C.,Washington, D.C., Philadelphia, New York and Boston.
Fuqua, who is an assistant professor at the University of TexasHealth Science Center in San Antonio, told some 200biotechnology scientists and executives at the seminar that sheand her team are tackling two breast cancer puzzlers.
-- Why some patients benefit from anti-estrogen therapy, somedon't, and some switch from responding to resisting thehormone treatment;
-- How to track, and eventually intervene in, the earlyprogression of the disease, for which there are currently nomolecular-level clues.
"Many of the oncogenes and suppresser genes that have beenisolated to date may not be involved in the very earliest stagesof breast cancer," Fuqua explained. "I think that is where thefuture is in this field, to be able to tell someone with a familyhistory of breast cancer, or who comes up with a lump or abump and has it taken out: 'Will you or will you not progress tofully developed breast cancer.' "
Estrogen wreaks its oncogenic mayhem on a breast-cancertumor cell by connecting with a surface receptor, which in turnturns on a growth stimulating factors. The preferred hormonalchemotherapeutic is tamoxifen, an estrogen mimic that blocksthe receptor.
Tamoxifen's clinical use, Fuqua reported, has significantlyreduced recurrence and mortality. But there is no reliable wayof differentiating between women who have tumor genes thatrespond to the drug and those who don't.
San Antonio, she told BioWorld, has one of the world's largestand oldest tissue banks of deep-frozen breast-tumor biopsyspecimens, numbering tens of thousands of samples. Thisresource enables her to look for gene variation in hundreds ofvariant tumors.
Her search has been aided in recent years by ever-more-advanced gene machines. Some of these she helped develop asa field scientist collaborating with ABS, "working out thechemistry of isolating RNA on their nucleic acid extractor."
In early-stage breast cancer, Fuqua said, "our goal is to isolatechanges in tumor-suppresser genes, which mutate to becometumor-promoters." She described the difficulties of analyzingRNA expression by differential hybridization and subtractivehybridization. Fuqua turned from these technologies late lastyear to differential display (DD), invented by Arthur Pardee ofBoston's Dana Farber Cancer Center, and announced in August1992.
"I think the most promising technology available today isdifferential display," Fuqua said, "because you can use verysmall amounts of sample to go through a large number ofbreast cancers. A lot of the old technologies, although just assensitive, utilize much larger amounts of starting material,samples of which are not available any more."
DD's technology is "very simple." To compare, say, cells from ahormone-responding tumor with a resister involves isolatingtotal RNA (not, tediously, as in past methods, only messengerRNA), make a cDNA copy, and then, using "the beautiful ideathat Pardee invented, add a primer to prime that reversetranscriptase process, anchoring it to the polyadenylated tailpresent on mRNA." Finally, Fuqua goes on, amplify this cDNAcopy by PCR, and put in a tag, such as a radiolabeled nucleotide,usually 32P.
"For us researchers of child-bearing age," Fuqua told heraudience, "handling radiotoxic 32P every day is bad news." Sheran experiments with other isotopes, and settled on 35S, whichhas reduced risk. But her team's technical virtuosity didn't stopthere. They worked out "one of the most exciting adaptations ofthe method -- fluorescently labeling the primers in non-toxicdifferential colors, for display on the gel. This uses machinerysuch as ABS's automated DNA sequencer for all thevisualization and quantitation."
So far, the San Antonio group has isolated portions of genesdetermining benign vs. malignant cells, and those with full-blown or half-blown tamoxifen resistance. "Our job now," Fuquaadded, "is to isolate the entire gene, and move into largeprospective or retrospective clinical trials to determine theirultimate significance in breast cancer."
She said she hopes to have the genes cloned, and a preliminaryscreen of their clinical importance, by the end of this year.
-- David N. Leff Science Editor
(c) 1997 American Health Consultants. All rights reserved.