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Karius Eyeing Sepsis, Transplant Patients as Key Markets for Infectious Disease Sequencing Test

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NEW YORK (GenomeWeb) – Karius, a three-year-old startup with a commercial test to diagnose infectious diseases by sequencing cell-free pathogen DNA in blood, is beginning to narrow its focus on the most promising immediate applications for its assay.

At the Infectious Diseases Society of America's annual IDWeek conference earlier this month, Karius Cofounder and CEO Mickey Kertesz said that the company believes that the test, called the Karius Digital Culture, is already proving its worth in two key areas: identifying the cause of sepsis and diagnosing infections in immunocompromised patients, especially bone marrow and stem cell transplant recipients.

"Within immunocompromised patients, especially in pediatrics, you don't have the blood volume or time to mess around with various hypotheses and tests," Kertesz said. Meanwhile, in the case of sepsis, early data is "even more encouraging than we thought it would be," he added, with some of the firm's academic collaborators showing the utility of Karius Digital Culture as a first-line test.

Karius, based in Redwood City, California, spun out of Stanford University in 2014 and began offering its assay to select hospitals earlier this year. Physicians who order the assay send a 5-mL blood sample from a patient to Karius' CLIA-certified and CAP-accredited laboratory, where plasma cell-free DNA is extracted and sequenced using Illumina systems — currently "the fastest, cheapest way to obtain genetic data," Kertesz said.

The company then uses proprietary methods to remove human sequences from the data — "on the sequencer [the sample] is strongly enriched for bacterial, viral, and fungal DNA," Kertesz said — and aligns the remaining reads against a database of more than 1,250 viruses, bacteria, and eukaryotic pathogens. The test currently has a turnaround time of approximately one business day from receipt of sample and costs around $2,000.

At IDWeek, several of Karius' academic collaborators presented early data and anecdotal evidence validating the Karius Digital Culture for several applications, chief among them identifying the culprits of septic and post-transplant infections. In fact, the company's assay was one of the most widely presented technologies at the meeting, as it was at the annual meeting of the American Society for Microbiology in June. But, Kertesz noted at IDWeek, "Where we are now is a lot farther than even two or three months ago."

In one late-breaking abstract at IDWeek, Simone Thair, a postdoc in the Emergency Medicine department at Stanford who has no financial ties to Karius, presented early data from the SEP-SEQ Trial, a prospective study designed to clinically validate the company's test for pathogen detection in sepsis.

In the study, Thair and her Stanford colleagues, alongside several Karius scientists, enrolled 350 patients admitted to the emergency department with signs and symptoms of sepsis, and collected plasma samples from the patients at the time of initial blood culture.

The researchers then compared results of the test to initial blood cultures and all subsequent microbiological tests performed during the patients' first seven days of hospitalization. Karius identified an organism in 210 of the 350 patient samples, or 60 percent, compared to blood culture, which yielded a positive result in just 63 patients, or 18 percent. Meanwhile, 131 out of 350 patients, or about 38 percent, had a potential infectious etiology identified using a composite reference standard of all microbiology testing performed within seven days of hospitalization.

The researchers also set up an independent panel of three infectious disease specialists not affiliated with Stanford or Karius to adjudicate all of the cases. After the subjects were evaluated by this expert panel, 182 of the 210, or about 87 percent, of the positive Karius test results were judged to be consistent with a potential cause of sepsis.

"That means you have 182 of the 350 patients with a plausible cause of sepsis being identified," Thair said in an interview following the meeting. "[Compared] to 63 blood cultures – you've got almost a three-fold improvement. With sepsis this is very encouraging, largely because we know that 30 to 40 percent of patients who die from sepsis never had a positive blood culture. For those patients, having some way to identify what is going on with them is absolutely imperative and could be a total game changer."

The researchers are currently preparing a paper on the trial to submit for peer-reviewed publication.

Thair noted that despite the test's promise, it is not yet able to be a standalone assay for diagnosing septic infections. "This isn't meant to replace blood culture, but it could definitely provide valuable insight for those patients where everything is coming back negative, and we don't know what's wrong, and then they unfortunately die," Thair said.

Another benefit of the test, she added, is that "in our particular cohort, there were quite a few organisms that Karius was able to call out that are possible causes of sepsis and are considered fastidious, hence, they are very difficult to culture. For those patients where we know we're probably not going to get a blood culture back, or it's going to take forever to get one, this test is capable of identifying those cases."

Meanwhile, in a separate Karius-sponsored presentation at IDWeek, Lauge Farnaes, a research fellow at the Rady Children's Institute for Genomic Medicine in San Diego, shared a few success stories using the test at Rady Children's Hospital to diagnose infections in immunocompromised pediatric patients.

One 5-year-old female patient with pre-B-cell acute lymphoblastic leukemia was admitted with a fever and neutropenia and found via blood culture testing to have Klebsiella bacteremia. She was treated with antibiotics and showed a blood count recovery, but during treatment her fever recurred even though she was otherwise symptom-free.

As part of a standard hematology workup, her physicians decided to send a blood sample to Karius. Even though all other infectious organism tests were negative, the Karius test revealed the presence of the fungus Rhizopus oryzae, which "really surprised everyone involved," Farnaes said during the presentation.

After completing her antibiotics, they ran the test again to confirm the result and got the same organism back. As such, her doctors ordered a chest CT, which revealed a very localized lung lesion and enabled the physicians to make a definitive diagnosis. The patient was treated with different antibiotics for the fungal infection, and she was allowed to continue with her previously prescribed chemotherapy for pre-B-ALL.

"This infection would have most certainly progressed to a point where it was unresectable, which likely would have had a really negative outcome for her," Farnaes said. "Immunocompromised patients with pulmonary nodules have been some of our highest [diagnostic] yield and [Karius] has been very useful in that sense, but also for more traditional infections."

In another case, a 5-year-old with pneumonia started antibiotics for suspected methicillin-resistant Staphylococcus aureus, but her condition deteriorated and she required hospitalization. A CT scan also revealed that she had some sort of colonization in her lungs. However, despite several epidemiological clues — her mother had latent tuberculosis, the girl had consumed unpasteurized dairy products, the family owned three parrots, and the patient had not traveled outside the San Diego area — her doctors were unable to determine the organism in her lungs.

Enter the Karius test, which revealed that the patient simply had a very bad case of Streptococcus pneumoniae. "We were able to get rid of the MRSA drugs, and treat the patient while hospitalized with a more targeted antibiotic," Farnaes recalled. "The Karius testing really allowed us to do our antibiotic stewardship as we're supposed to and not just do the broadest antibiotics possible."

Finally, Farnaes recalled the case of a 9-year-old male who had lived in American Samoa and presented with a fever, tachycardia, and "massive vegetation" on his prosthetic mitral valve. After multiple rounds of antibiotics and multiple negative blood culture tests, his physicians sent a blood sample to Karius. At the same time, they removed a piece of his heart valve, which by this point had suffered severe damage and was rendered insufficient.

The Karius Digital Culture came back with a diagnosis of Kingella kingae, a Gram-negative aerobic coccobacilli, which the doctors knew they could treat with a specific antibiotic.

"But, again, we were new to [the Karius] test, so the question is, do you trust this test enough that you are going to totally narrow his antibiotics for a specific organism based on the results?" Farnaes said. "We were a little uncomfortable with that at first."

To bolster their confidence, they performed Kingella-specific PCR on a piece of the removed heart valve, which confirmed a positive diagnosis and allowed them to treat the patient for that specific infection.

"In all these cases, we were able to make pretty significant changes to the management of patients, really increase our antimicrobial stewardship, and, in some cases, save lives and avoid procedures," Farnaes said.

In the first two months of using the test, Rady Children's sent 33 distinct samples to Karius. Farnaes said that an informal survey of everyone in his division that had ordered the test revealed that it informed management in 39 percent of patients.

"Part of the reason that was only 39 percent is we still don't really know what the negative predictive value of this test is," Farnaes said. "Sometimes when we screen for an infection, we send it and it's negative. As we get more experience with the test, we're going to have a better idea of [negative predictive value]. So, I suspect that clinical usefulness number is going to increase with time."

Farnaes did note a few challenges using the test. In cases of bowel infection, for instance, "we get a lot of translocation; and you can get a lot of bacteria coming out of the bowel, so we have found that it can be difficult to interpret the results in that case." Furthermore, his clinic works with a lot of neonates, "so the sample volume can be challenging," he said.

Nevertheless, overall, "we've found this very useful," he said. "We're still trying to figure out exactly who the right patients to use it for are. But a … very significant part of the time it is helping with the management of patients."

In another IDWeek presentation, clinical researchers from the University of California, San Francisco and Karius presented early data from the Diagnosis of Infection in Stem Cell Transplant Patients Over Time (DISCOVER) trial, which prospectively enrolled 20 patients.

Cytomegalovirus, one of the most common and important complications after stem cell transplantation, was identified in 12 of 20 patients. Patient samples were taken within a day of each other for Karius plasma testing and CMV qPCR over multiple time points, and positive agreement between Karius and qPCR was about 82 percent. However, when values below the lower limit of quantitation for qPCR were excluded, positive agreement between the tests was 100 percent.

In addition, the authors noted, Karius identified four viruses, and detected pathogens such as S. aureus and Chlamydia trachomatis approximately eight days and 30 days prior to targeted testing for those organisms, respectively.

The researchers concluded that although further work is needed to determine the lower limits of detection for the Karius test, using it to monitor stem-cell transplant patients for infection "could permit earlier detection of pathogens, enabling earlier targeted therapy for this vulnerable population."

In August Karius raised $50 million in a Series A financing round, money that it said it would use to support new and ongoing clinical trials, scale up its laboratory capacity, and continue commercialization of its test.

At IDWeek, CEO Kertesz said that the company has also recently hired several new employees, including Todd Whitson, who has been named chief commercial officer after serving in that same role at CareDx, and prior to that, as VP of commercial operations for Ariosa Diagnostics, now a part of Roche.

Kertesz also said that the company will be "doubling down" on R&D and clinical studies to drive adoption and acceptance of the Digital Culture assay. Of note, the ability to detect RNA viruses in addition to DNA viruses "is on the roadmap," Kertesz said.

Karius will continue to focus on the US market in the near term, offering its test as an LDT through its clinical laboratory. A regulatory filing with the US Food and Drug Administration is not immediately in the cards, but it is possible, Kertesz said.

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