TGen LymeSeq Assay Moving Forward With Financial Backing From Advocacy Group

NEW YORK (GenomeWeb) – Sometimes it pays to chat with your seatmate on an airplane. When a concerned parent of a child with Lyme disease happened to be seated next to the Translational Genomics Research Institute's director of translational research, Daniel Van Hoff, on a recent flight, a discovered shared interest in improving diagnosis and treatment for the illness ultimately led to a projected half a million dollars in support for a new TGen sequencing-based test for Borrelia species and co-transmitted pathogens.

The concerned parent was Tammy Crawford, now the executive director of an advocacy group called Focus On Lyme. Her organization has raised funding through donations, events, and auctions — currently totaling $301,730, and with a goal of $500,000 — which is now slated to support testing of TGen's new assay on patient samples.

Called LymeSeq, the platform-agnostic assay uses amplicon, or focused, sequencing, similar in some ways to exon sequencing, Paul Keim, director of the pathogen genomics division at TGen's Pathogen and Microbiome Institute in Flagstaff, Arizona, and developer of the test, said in an interview.

There are approximately 30,000 new cases of Lyme disease each year, according to the US Centers of Disease Control and Prevention, concentrated in the Northeast and upper Midwest.

However, monitoring of the clinical situation has been hampered overall by the quality of currently used diagnostics, Keim said.

"What is Lyme and what isn't Lyme is really based on tests right now that aren't really all that good," he said. 

Clinicians currently tend to use a two-tiered serological approach to Lyme testing, which combines ELISA and Western blotting, Keim said. However, this assay has been shown to be about as sensitive as a coin flip, in part because there are many strains and species of Borrelia, phases of infection, and consequences of having other tick-borne co-infections, not to mention underlying patient genetics affecting how the illness is manifest.

"It is a complex disease that needs a lot more study and better diagnostics," Keim said.

Crawford said her family's relief at finally obtaining a diagnosis for her then 19-year-old daughter Jessica's severe illness using standard Lyme testing led to disappointment when an infectious disease expert they visited said he did not have faith in the Lyme disease test results they'd received. The expert said Jessica did not have Lyme and several other doctors went further, telling Crawford that the illness did not exist, she said.

"I became very frustrated, and because [Lyme disease] is not clearly understood as a disease, none of the treatment is covered by insurance," Crawford said in an email. "I was fortunate to be able to care for my daughter, but millions cannot. I decided to look to science to do better."

After making the in-flight TGen connection, Crawford said she was encouraged to contact Keim, whose previous work included using a similar sequencing-based approach to develop a tuberculosis diagnostic capable of detecting hetero-resistance down to as little as 0.1 percent drug resistant bacteria in a sputum sample.

Keim agreed to begin developing the Lyme assay and Crawford funded the initial round of development herself with a $75,000 donation, she said. Now, her advocacy group is working to raise additional funds needed for further development by hosting scientific conferences, galas, and charity golf tournaments.

The diagnostic value of the resulting LymeSeq test is particularly apparent compared to real-time PCR, Keim said. The sequencing test, for which the lab is currently using the Illumina MiSeq, is quantitative and also has a similar sensitivity as qPCR, in that it can detect single nucleic acid changes.

But, "In addition to that, it can be highly multiplexed ... in the order of 300 to 400 different targeted regions in one assay," Keim said.

In the case of Lyme, this allows the test to detect common pathogens and parasites  co-transmitted by ticks along with Borrelia, such as babesia parasites, anaplasma bacteria, and infections like tularemia, ricketsia, and erlichia. One recent study estimated that there is co-transmission of additional pathogens along with Borrelia about 45 percent of the time.

These co-infections complicate the clinical manifestation of Lyme, and "being able to diagnose these other diseases at the same time is critically important," Keim said.

The resulting sequence of targeted regions also validates and verifies that the pathogen detected is truly Borrelia, and not the result of a promiscuous PCR primer.

The test further enables species identification, which is important because different species appear to cause differing clinical presentation, as seems to be the case for a species discovered last year at the Mayo clinic, for example, which causes severe illness. 

Keim, who is also a microbiologist at Northern Arizona University, noted that the entire field of Lyme research is funded extensively through philanthropic efforts.

"It's an underfunded area, relative to other infectious diseases," he said, adding, "It's become an area of a lot of advocacy, very reminiscent of what happened with HIV in the early days."

But Keim said he hopes advocate funding for Lyme disease will soon lead to more interest from federal funders. 

The TGen assay will next be further validated and optimized in the lab, followed by testing in patient samples, and Keim suggested it may be commercialized in the future, either through licensing, a lab-developed test, or even through a 510(k) approval, but that decision is a ways off. However, he also noted that the assay could be also be useful for tick surveillance. 

Lyme disease assays from other firms, meanwhile, are also on the path to commercialization. Ceres Nanosciences announced last month it had raised $3 million recently for a proteomics-based test to detect Lyme antigens in urine and that it was working to move the test, which it has been offering through a limited access program for more than a year, to the point-of-care market, including doctors' offices and pharmacy clinics.

Oxford Immunotec, meanwhile, acquired Imugen last year for $22.2 million in part based on Immugen's strategy to focus on immune-regulated conditions, including tick-borne diseases like Lyme, and its antibody and PCR tests.

The Pathogen and Microbiome Institute at TGen is also currently working on a project to track Clostridium difficile "from bed to bed and ambulance to ambulance," Keim said, enabled in part by declines in cost of multiplexed sequencing. The institute is also planning to release news related to commercialization of an assay for Valley Fever in the near future, he said.