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Study IDs Saliva MicroRNA Differences in Those with Oral Cancer

NEW YORK (GenomeWeb Daily News) – In a paper appearing in the September issue of Clinical Cancer Research, an international research team reported that they have identified microRNAs that are differentially expressed in saliva samples from those with oral cancer.

The team, comprised of researchers from California and Serbia, used reverse transcriptase pre-amplification quantitative PCR to identify nearly 50 miRNAs in saliva samples from a dozen healthy individuals. When they assessed a subset of these in individuals with a type of oral cancer called oral squamous cell carcinoma, the team found four miRNAs that were differentially expressed, including two that were at significantly lower levels in oral cancer patients.

"[I]t is very appealing to think that we could detect a cancer-specific marker in a patient's saliva," Clinical Cancer Research Senior Editor Jennifer Grandis, an otolaryngology and pharmacology researcher at the University of Pittsburgh, who was not involved in the study, said in a statement. Grandis also emphasized the importance of finding ways to detect cancer without biopsies.

Oral squamous cell carcinoma, a cancer that originates in squamous cells in the lips and mouth, represents some 90 percent of all oral cancer cases. Only about half of individuals diagnosed with the disease survive five years or longer.

Because of its poor survival rate and lack of new treatments, researchers have been working towards finding early oral cancer detection methods, looking for transcriptomic, proteomic, and other patterns in saliva that could help distinguish individuals with oral squamous cell carcinoma from healthy individuals.

With the latest study, researchers rounded out the trio of oral cancer proteomic, transcriptomic, and miRNA profiles in saliva. Now, they are continuing to explore which of these could be most useful in a diagnostic setting.

"The key here is that we're not solely dependant on one analyte," senior author David Wong, director of the University of California at Los Angeles' Dental Research Institute, told GenomeWeb Daily News. "The real advance is that now we have three alphabets" for assessing oral cancer, he said.

To find miRNAs that are altered in oral cancer, Wong and his team used reverse transcriptase pre-amplification quantitative PCR, or RT-preamp-qPCR, to assess the levels of 314 miRNAs profiles from the miRBase database in saliva samples from six healthy controls.

In so doing, they identified 71 miRNAs that were present in at least two of the individuals tested. After evaluating these miRNAs in another six control individuals, the researchers found 47 commonly occurring miRNAs in the complete saliva sample and 52 in the saliva supernatant samples.

Of these, 13 miRNAs and 28 miRNAs were present in 11 out of 12 whole saliva and saliva supernatant samples, respectively.

"Together the data indicate that both the whole and supernatant saliva contain detectable amounts of miRNA, and there seemed to be a common set of miRNAs present in the saliva of healthy participants," Wong and his co-authors wrote.

The team then compared the miRNA profiles in the same 12 healthy individuals with those in saliva samples from 12 oral squamous cell carcinoma patients who were matched for gender, age, and smoking history. This comparison turned up four miRNAs that were differentially expressed in the oral cancer patients.

When the team tested these miRNAs in another 38 oral cancer patients and 38 controls, they pinpointed two miRNAs — miR-125a and miR-200a — that were found at statistically significant lower levels in the individuals with oral cancer than in the healthy controls.

At the moment, the researchers aren't focusing on the functions of these miRNAs, Wong explained, but are continuing to look for additional miRNAs that are differentially expressed in the saliva of those with oral cancer. "We're not ready to look into the biological function of these miRNAs at this time," Wong said.

Their results also indicated that supernatant saliva — the cell-free component of spit — is more diagnostically promising than whole saliva, which contains a heterogeneous mixture of saliva and cells, Wong explained. "The diagnostic component should be the cell-free component," he said, noting that cells in saliva could dilute or overpower the subtle differences that could be useful for diagnosing disease.

Wong said the team is eventually hoping to translate their findings into point-of-care technology that can be used to diagnose oral cancer in the clinic based on saliva samples.

For his part, Wong believes researchers are about two years away from developing a microchip for detecting oral cancer markers in saliva, though he conceded that many in the diagnostic field are waiting on definitive clinical data based on salivary diagnostics. Wong said he and his colleagues are currently doing pre-clinical salivary diagnostic trials in an academic setting.

"These findings suggest that the detection of miRNAs in saliva can be used as a non-invasive and rapid diagnostic tool for the diagnosis of oral cancer," the authors concluded.

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