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Seattle Startup AttoDx Wins NIH Funds for Ultrasensitive TB MDx


NEW YORK (GenomeWeb) – Start-up diagnostics firm AttoDx has been awarded National Institutes of Health funds to develop an ultrasensitive molecular diagnostic for difficult-to-detect cases of tuberculosis.

The one-year, Phase I Small Business Technology Transfer grant from the National Institute of Allergy and Infectious Disease totals $220,230. AttoDx will develop the assay in collaboration with Claremont BioSolutions and researchers at the University of Washington, initially for same-day diagnosis of smear-negative and extrapulmonary Mycobacterium tuberculosis from sputum and cerebrospinal fluid of HIV-coinfected patients.

The core technology of the diagnostic is molecular viability testing, or MVT, a technique licensed by AttoDx from UW. The patent-pending method amplifies ribosomal RNA precursors (pre-rRNA) which are rapidly upregulated when bacteria are nutritionally supplemented.

"If they're alive, they'll make pre-rRNA, and they'll make quite a large amount of it — that makes it easier to detect the bacteria," Jerry Cangelosi, consulting scientific director at AttoDx and a professor in the Department of Environmental and Occupational Health Sciences at UW told GenomeWeb in an interview.

The MVT method measures increased pre-rRNA copy number in response to nutritional stimulation. The ultimate product will consist of a single-use pre-PCR cartridge which will partition the sample and nutritionally stimulate half of it. From this, two cDNA samples result, and then these go into a qPCR system already present in a lab, Cangelosi explained.

MVT was originally designed to reduce false-positive results by estimating the proportion of living bacteria in a sample. However, a report in PLoS One last year showed that, for a diverse range of bacteria, stimulation-induced upregulation of pre-rRNA was so robust that the test was actually 5- to 10-fold more sensitive than standard PCR.

Moreover, the pre-rRNA are specific for different species of bacteria.

The pre-PCR cartridge will be spearheaded by Claremont BioSolutions, which "has a very nice, low-cost, battery-powered, sample processing system that can extract DNA and RNA very quickly and cheaply," Cangelosi said, noting that this element was described in a Journal of Clinical Microbiology study.

Claremont's miniaturized cell lysis system is also part of a number of projects at Keck Graduate Institute, and the firm has been funded in the past to develop it further for Clostridium difficile — a bacterium with a notoriously rugged external shell.

Cangelosi is also co-PI on a Keck grant to develop a point-of-care, lateral flow TB assay, also in collaboration with Claremont. That assay will be for adult, pulmonary TB, he said, while the newly funded 'ultrasensitive' test is for more challenging infections.

"There's plenty of cases, [such as] pediatric and extrapulmonary TB, where a clinician is pretty sure that a patient has tuberculosis but they're just not able to culture it, or see it by PCR," Cangelosi said. "Our product is designed for that situation — you would order this test when you really want the highest possible sensitivity."

When tuberculosis infects children or people with HIV, levels of bacteria are sometimes too low to detect by standard methods, and infections can also occur outside of the lungs.

Extrapulmonary TB is more common in people with HIV coinfection, and is not well detected by the commonly used Cepheid GeneXpert MTB/RIF test, as previously reported. That test has recently been shown to be less sensitive on samples from children.

There is also a particularly high rate of HIV and TB co-infection in Swaziland, and Cepheid's test was recently demonstrated to be unable to detect around 30 percent of infections in that country.

However, because of the estimated cost and technical complexity of the AttoDx MVT-based test, Cangelosi said the firm intends to initially market it in higher-income countries that also have high TB rates, such as China and India.

Those two countries account for almost 40 percent of the estimated global burden of tuberculosis according to the World Health Organization." These are countries that do have some clinical laboratory infrastructure … [our assay] is not going to be the cheapest possible TB test; it’s something that you order when you need the maximum sensitivity," Cangelosi said.

However, "AIDS-related, extrapulmonary, and pediatric TB — that's when the disease is most life-threatening," he said. "We don't envision stopping at tuberculosis, we envision going well beyond that, but TB is a good place to start because the need is so acute."