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Northwestern Researchers Develop RT-qPCR Assay for Depression Biomarkers, Seek Industry Partners

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NEW YORK (GenomeWeb) — Scientists at Northwestern University have developed a blood-based biomarker assay of clinical depression. The assay uses reverse-transcriptase quantitative PCR to detect levels of RNA transcripts that correlate with a depressive episode as well as with remission after cognitive behavioral therapy.

The researchers claim that the assay may also be able to predict which patients with depression will not respond to CBT, which could ultimately aid physicians in treatment selection. They have filed patents on the assay and are seeking partners to develop it further.

A small study validating the assay in 64 adults was published online in the journal Translational Psychiatry this week, as covered in GenomeWeb Daily News.

In an interview with PCR Insider, Eva Redei, a professor of psychiatry at Northwestern University's Feinberg School of Medicine and a lead investigator on the study, explained that nine of the 20 transcripts measured were diagnostic for depression, and "three of those nine seem to be diagnostic for vulnerability," as they continued to show expression differences in patients whose depression went into remission after CBT.

The biomarker signature requires further validation, Redei said, but expression levels of some of the transcripts in the panel appear to be correlated with major depression, as diagnosed by a Mini-international neuropsychiatric interview and PHQ-9 questionnaire. "I don't know whether [the transcripts] are causative or not, of course, but they are definitely reflective of what happens" in the disease, she said.

The work grew out of previous microarray studies on animal models. The group did genome-wide expression analyses of blood samples from rats bred for anxiety and depression traits, as well as rats subjected to depression phenotype-inducing stresses. They also showed these transcripts expressed in the hippocampus and amygdala of the animal models.

Combining data from the models of both endogenous and environmental stress-related depression yielded a set of candidate transcripts. "We took those that had the same expression differences in the brain and in the blood," Redei explained.

Then, in a pilot study of early-onset major depression, the group examined blood samples from 28 unmedicated young adults, half of whom were suffering from major depressive disorder. They discovered 11 transcripts that were differentially expressed in youths with MDD versus those without the disorder.

The current Translational Psychiatry study examined the biomarker panels using blood from 32 depressed adults with varying medication status, before and after 18 weeks of cognitive therapy, compared with 32 controls.

The transcripts examined in the current study are also known to be expressed in human brain, Redei said, but added that they have not done expression analyses there, in part because she doubts the RNA biomarker signature would persist postmortem.

Results in the study were reported as change in RT-qPCR cycle threshold in a total of 20 transcripts of interest versus a control gene, 18s rRNA. Regarding this choice, Redei said the group had tried five or six other housekeeping genes, but found this one to be the most stably expressed in blood.

Whole blood was collected into PAXgene Blood RNA tubes from Qiagen, which have the advantage of making RNA stable at room temperature. RNA was extracted using the PAXgene Blood RNA kit.

For the RT-qPCR, the researchers used TaqMan RT reagents from Thermo Fisher Scientific's Applied Biosystems, and qPCR was carried out using SYBR Green on the ABI 7900 platform. Each transcript was measured in triplicate for every sample, in separate RT-qPCR reactions.

In the future, the group hopes to make the assay "as simple as possible," Redei said, including possibly multiplexing the reaction and simplifying the extraction process. She said she is aware of other RNA collection and sample prep methods for whole blood that have come on the market since the experiments began, but has not tried any of these yet.

The biological function of many of the transcripts and their relationship to major depression is not known. "Some of them we know nothing about, they haven't been described for a specific function," Redei said. "Others clearly have cellular function ... not specific to neurons or to the brain, but they are present in the brain as well ... In general, you can say they have very basic cellular function," she said.

Now, the group is looking to develop this test into a commercial product, and seeking investment and partners, Redei said.

Two patents have been filed on the assay, with Northwestern as the assignee. "The goal is to partner to move this as far as possible into the clinic," Redei said. "There are [other assays] coming behind it, so I would like to focus on [those] ... but then this one can move on. For that, I absolutely need partners [and] money, that's the bottom line," she said.

Redei envisions developing this assay into a US Food and Drug Administration-approved diagnostic, rather than a laboratory-developed test. "If it's FDA approved, then any laboratory can do it," she said.

The release of the paper this week spurred a flurry of press engagements for Redei, including four television interviews on Tuesday. She told PCR Insider that this is not her favorite part of being a researcher, but she hopes it will spread the word about the biomarker panel and move it closer to commercialization.

"I hope it is going to result in licensing, investing, or any other way that moves it forward," she said. "If it only exists as a paper in my drawer, what good does it do?"

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