Shed glycans as possible markers for ovarian cancer
more closely resembled those of the cell supernatant samples. Again, most of Early detection is critical for the survivthe oligosaccharides were found in the al of ovarian cancer patients. If ovar10% and 40% acetonitrile fractions. ian cancer is not detected until a late After the researchers identified some stage, the prognosis is bleak. Because of the oligosaccharides by MS, they resymptoms often are not recognizable alized they had obtained an u nusual until the disease has progressed to a result: Oligomers of hexuronic acid late stage, clinicians must rely on a test (HexA) were present in both cancer that measures the level of a mucin glyand control sera. Lebrilla says that this coprotein called CA125 in the blood. study may be one of the first to report Currently, the CA125 test is the only one the existence of HexA oligomers in huapproved by the U.S. Food and Drug man serum. “When we do our analysis, Administration for the diagnosis of I think we’re seeing a lot more than othovarian cancer. Although it has been er [researchers] because their methods used for years, it is not very are a lot more specific,” he sensitive or specific, and says. For instance, many rethe high number of false searchers studying glycans negatives puts women’s do derivatization reactions, lives in jeopardy. Therebut some oligosaccharides fore, many researchers now are not amenable to that are on the hunt for new biotype of reaction, so they markers that could be used are not analyzed. Although in a diagnostic test for ovarthe new method is less sensitive than derivatizaian cancer. tion methods, it “allows us In this issue of JPR (pp 1626–1635), Carlito Lebrilla to cast a wider net,” says Lebrilla. and colleagues at the University of California, DaWhen the glycan comvis, report a new method positions of the cancer for isolating possible bioand control samples were Serum glycoproteins Glycan markers markers for ovarian cancompared, a total of 15 pocer. With the method, the tential biomarkers were researchers found several found. Lebrilla says that the High-carb biomarkers. Oligosaccharides harvested from O-linked glycoresearchers are still charglycans that were uniquely proteins could serve as biomarkers for ovarian cancer. expressed in ovarian canacterizing the glycans that d istinguish the cancer sera cer patients. Most investigators are seeking markeach fraction from one cell line to anfrom the control sera. Although b-elimers for ovarian cancer by breaking open other, with some variations. ination releases mostly O-linked olicells and studying the proteins that are But would the procedure work with gosaccharides, it can also release some inside, says Lebrilla, but he and his colhuman serum? When the researchers N-linked ones. “We were looking for laborators took a different tack. Glyapplied the method to five serum sammucins, which are mainly O-linked, but cosylation patterns change when cells ples from ovarian cancer patients and the structures that we’re getting could become cancerous, and some glycoprofive serum samples from control subbe either N- or O-linked. We may be getteins, such as CA125, are shed from tujects, the results differed from the preting mixtures of both,” he says. mors and detected in the blood. “We vious analysis. Oligosaccharides were According to Lebrilla, the researchthought it just seemed like a natural concentrated in the 20% fraction, ers already have completed an expandthing—[to] look at things that come whereas peptides were abundant in the ed version of this study with 75 patients off the cell rather than the cells them10% and 40% fractions. Lebrilla says and control subjects. In addition, they selves,” he says. In particular, the rethat glycoproteins were abundant in have used the method to identify posearchers homed in on the mucin famthe media of the cultured cells, but hutential markers for prostate and breast ily of glycoproteins. The mucin CA125 man serum is much more complex and cancer and have compared these canmay not be the best marker for ovarian contains many additional proteins and didates with the potential ovarian cancancer, but other mucins may be more peptides. “Peptides ionize so much betcer markers. “Glycosylation is really sensitive and specific, they reasoned. ter than carbohydrates that any signifripe at this moment for looking at difIn the method, the supernatant of icant amount of peptides will swamp ferent types of markers,” says Lebrilcultured cells or human serum is diaout your carbohydrate signals,” he exla. “I think in the next year or so, you’re lyzed and lyophilized. A b-elimination plains. When the researchers added a going to start seeing a lot more glycan reaction is performed on the dried samprotease step to the protocol (after SPE markers for diseases.” ple to release O-linked oligosacchaelution), the oligosaccharide profiles —Katie Cottingham CARLITO LEBRILLA
rides, which then are preconcentrated by solid-phase extraction (SPE). The oligosaccharide-enriched sample is dialyzed again and analyzed by MALDI FTMS. The researchers tested the protocol on the supernatants of four ovarian cancer cell lines. They eluted oligosaccharides from the SPE matrix with 10%, 20%, and 40% aqueous acetonitrile, producing three fractions. The majority of the oligosaccharides were found in the 10% and 40% acetonitrile fractions. Significant overlap existed among the oligosaccharides that were found in
Journal of Proteome Research • Vol. 5, No. 7, 2006 1525
