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Molecular TB Assay From PBD Biotech Shows Promise in Detecting Latent-to-Active Disease Risk

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NEW YORK – A blood-based assay that combines phage-induced lysis of infectious bacteria and molecular detection of bacterial DNA has shown promise in predicting which individuals with latent tuberculosis might progress to active disease. 

At the European Congress of Clinical Microbiology and Infectious Diseases (ECCMID) on Tuesday, Jee Whang Kim of the Leicester Tuberculosis Research Group in the UK presented unpublished data from a small study of the Actiphage TB test from Nottingham, UK-based PBD Biotech. 

In the study, the researchers showed for the first time that live Mycobacterium tuberculosis bacteria could be detected in the blood of individuals determined to have had latent TB using radiological imaging. The study results further suggested that Actiphage was significantly more specific in detecting risk for full-blown tuberculosis than the gold standard, interferon-gamma release assays (IGRAs), with similar sensitivity. 

Actiphage uses a bacteriophage called mycobacteriophage D29 to detect and lyse M. tuberculosis in a blood sample, thereby releasing bacterial DNA that can be subsequently detected using real-time PCR. According to PBD Biotech CEO Jane Theaker, the assay workflow comprises isolating white blood cells that contain viable mycobacteria from the blood sample, then incubating the white blood cells in a media that simultaneously and osmotically lyses the cells and acts as a suitable media for the bacteriophage to infect and lyse the viable mycobacteria. 

This phage infection in turn releases the bacterial DNA, which is cleaned up and undergoes PCR testing for gene IS6110. 

The test was previously shown to be able to accurately detect active TB infection in human patients, with some evidence for the detection of latent TB when bacteria levels in the blood are extremely low. 

Detecting latent TB is paramount for controlling disease outbreaks, and IGRAs such as those from Qiagen and PerkinElmer’s Oxford Immunotec have proven to be the gold standard for this. However, it would also be valuable for both outbreak mitigation and early treatment to have a diagnostic test to accurately predict which latent TB cases will progress to active TB, something that IGRA’s aren’t as good at doing. 

About 5 percent to 10 percent of latent TB cases progress to active TB, and studies have shown that those with recently acquired infections are at the greatest risk because the immune system has not yet controlled the infection. Since IGRA tests measure a host immune response, they may not pick up extremely low levels of bacteria, which is usually the case in latent patients at risk of progressing. 

Building off the previous pilot study, the Leicester researchers hypothesized that Actiphage should be able to identify a subset of patients with latent tuberculosis who show features of bacterial infection associated with increased risk of progression. 

In their study, they used 18FDG PET-CT scanning to assess active inflammation associated with latent tuberculosis and correlated that with early Actiphage results, repeating the scan after three months to characterize disease progression and to reveal target sites for additional sampling. They also sought to compare the performance of Actiphage with that of IGRA. 

The investigators used Actiphage, IGRA (Qiagen’s QuantiFeron Gold Plus), and PET-CT to test and analyze 21 healthy adults who had been in the same household as someone with pulmonary TB. The PET-CT findings were classified as positive, indeterminate, or negative based on pre-specified criteria. Subjects with positive PET-CT findings were investigated for active TB, and those with negative scans had a repeat scan after three months to characterize infection trajectory. All participants were followed for 12 months and received no TB prevention therapy, the researchers said. 

The team was able to isolate M. tuberculosis in four subjects with latent TB infection. Actiphage also detected bacteria in 6 out of 7 subjects with what was classified as incipient TB — that is, subjects who are harboring viable bacteria and are likely to progress to full-blown disease barring treatment, but who do not yet show signs or symptoms. 

When compared with IGRA, Actiphage showed a sensitivity and specificity of 68.8 percent and 80 percent, respectively, for predicting FDG uptake in PET-CT — a sign of disease progression — compared with 81 percent and 20 percent using IGRA. In addition, 10 participants had persistent or progressive infection revealed by follow-up PET-CT, with six receiving TB treatment. When including prospective outcome, Actiphage testing at baseline had sensitivity and specificity of 80 percent and 64 percent in discriminating between subjects with resolving and progressive infection. 

After the ECCMID meeting, PBD Biotech's Theaker noted that progression to active TB was not the endpoint of interest, as only one person developed this. “We were looking at the risk of developing TB,” Theaker said. “IGRA picked up all positive PET-CT scans but was also positive in people with negative PET-CT scans. Actiphage was positive in all people that had positive PET-CT scans with incipient infection, but negative in those with positive PET-CT scans that did not have incipient TB. In that respect, Actiphage was both very specific and very sensitive for identifying people at risk of developing TB.” 

One caveat to the study is that the utility of FDG uptake in PET-CT is itself still not fully established for determining disease progression, though it has been shown in several studies to be associated with positive or negative treatment outcome. 

Theaker said that PBD Biotech is now planning further trials of Actiphage in South Africa, Zambia, and the UK to test the assay and develop it further. She also noted that the results of the ECCMID study have been submitted for publication in an undisclosed peer-reviewed journal.