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GlycoPath to Develop MALDI Mass Spec Test for Liver Fibrosis

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NEW YORK – With seed funding from mass spectrometry vendor Bruker, diagnostics outfit GlycoPath is working to develop MALDI-based assays analyzing protein glycosylation patterns that can indicate disease.

Specifically, the Charleston, South Carolina-based company is working on a test for diagnosing liver fibrosis in patients at risk for the disease due to conditions including hepatitis B and C and nonalcoholic steatohepatitis (NASH). The goal is to make the test commercially available through a CLIA facility as early as 2023, said Richard Drake, GlycoPath's CEO and a professor of proteomics at the Medical University of South Carolina.

The company's researchers use antibody capture arrays to pull down glycoproteins of interest, followed by enzymatic digestion to release the glycans attached to these proteins. They then profile the glycans using MALDI mass spectrometry, looking for changes in protein glycosylation patterns indicative of disease.

Protein glycosylation, in which sugars get attached to various amino acids, is a common post-translational modification and an important phenomenon in the biology of various diseases, including many cancers. Given this, researchers have long considered glycoproteins as protein biomarkers, and more than half of cancer protein markers currently approved by the US Food and Drug Administration, in fact, are glycoproteins.

Glycoproteomic analysis has been challenging, however, due in large part to the wide variety of possible glycosylations, called glycans, and the complexity of different glycan structures.

Drake said, though, that the understanding of glycan structures had advanced to the point where researchers are able to profile them to detect disease and disease progression.

"We pretty much know how they are made and processed," he said. "Even the most complex glycans, there is an underlying, essentially mathematical component to it. You can do the math of the possible different sugar residues that could be present. We still see some surprises here and there, but for the most part, the possible [structures] are pretty well defined right now."

This regularity allows researchers to determine the presence of different glycans on a protein via mass spectrometry.

"Essentially, we're looking at [a] barcode," Drake said.

Called GlycoTyper, GlycoPath's liver fibrosis test will look at changes in glycosylation on immunoglobulin G, a protein whose glycosylation patterns are particularly well studied given its role in antibody therapeutics.

"It turns out that with immunoglobulin glycans there is a 'right' pattern that indicates health," Drake said. "If the structures are too short, say the glycans are truncated, then the IgGs don't bind to their receptor very well. So if you are detecting that, that is an indication that something is going wrong. And if they get more decorated with extra fucosylation and sialylation, that becomes more of an inflammatory-type structure."

"We know what normal should look like, and so we are looking for the differences," he said.

Drake said that looking at the glycan patterns of the different IgG subtypes, as opposed to total IgG, provides an additional level of specificity, helping the test better distinguish liver fibrosis from other conditions and between early and late stages of the disease.

GlycoPath envisions the test being useful for detecting and monitoring liver fibrosis in patients with viral hepatitis and, particularly in the US, NASH, or in patients who are deemed at high risk of developing NASH due to other conditions or lifestyle factors.

The test "would be a quick way to monitor and also to inform clinical decisions as far as how much extra testing a patient should undergo," Drake said. He noted that imaging approaches like ultrasound and MRI are commonly used to evaluate patients with suspected liver fibrosis. Biopsy is also used, though he said the field is moving away from that given the expensive and invasive nature of the approach and the challenges of effectively biopsying obese patients.

GlycoPath said last month that it had received an undisclosed amount of seed funding from Bruker, a portion of which it is using to put together a validation cohort to further refine its assay.

So far, GlycoPath has run the assay on around 100 samples, including around 25 controls, 30 early-stage fibrosis patients, and 35 later-stage fibrosis patients.Drake said that in that work, the test was able to distinguish between healthy, early-stage, and late-stage patients with an accuracy in the 90 percent range. The company is now working to evaluate the assay in another roughly 120 samples spanning the same three groups.

It is also exploring the potential of the assay to monitor patients for the development of liver cancer. Drake said that the number of glycoproteins already used as biomarkers opened a potential opportunity for the company to add value by providing more fine-grained glycosylation information.

He gave the example of the prostate cancer biomarker prostate-specific antigen (PSA).

"We could glyco-type PSA out of urine in men at risk for or being monitored for prostate cancer," he said, noting that this could potentially improve that marker's specificity or sensitivity.

Drake has collaborated with Bruker in the past, particularly on imaging mass spectrometry workflows analyzing glycans in tissue. He said the company was using several different Bruker platforms for its work and was currently working to adapt its assay for the  SmartFlex instrument, which is at the core of Bruker's MALDI Biotyper system.

Drake said that GlycoPath could also commercialize assays based on his MALDI imaging work with Bruker in the future.

"That is certainly a component down the line, it's just one of those things that is going to take longer to get to market," he said, noting that this remains a focus on his academic work. He and his MUSC colleagues have used MALDI imaging to analyze prostate cancer, investigating how different glycan profiles are associated with tumor aggressiveness and the path of disease progression.

Drake said that following the planned validation work, GlycoPath would seek to raise Series A funding, though he did not specify a target amount.