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Report

Teaser, summary, work performed and final results

Periodic Reporting for period 1 - TOPO-TB (TOPO-TB)

Teaser

Summary

WHO, with its 2015 UN Millennium Development Goals (MDG), defined targets for TB incidence and prevalence towards 2025 and 2035. As part of the post-2015 global TB strategyâ€™s first pillar on integrated, patient-centred care and prevention, the early diagnosis of TB is emphasized and inclusion of drug susceptibility testing (DST) is now targeted as a universal standard for patient care, including for both new and previously treated patients. By specifically addressing the TB diagnosis market and social needs set by 2015 MDG, the TOPO-TB project will secure a broader TB detection, reduce current screening delays and accelerate TB treatment, therefore contributing to improved clinical decisions. Furthermore, due to its point-of-care format and low-cost, the TOPO-TB project will also contribute to increase diagnosis at the primary health care level, where the majority of patients seek medical attention. All these project outcomes
will directly contribute to the achievement of 2015 MDG TB goals and therefore address current EU and global challenges for infectious diseases control.

Tuberculosis (TB) remains a major global health problem, ranking as the second leading cause of death caused by an infectious disease worldwide, after the human immunodeficiency virus (HIV). The latest estimates from WHO state 9 million new TB cases and 1.5 million TB deaths in 2013. As a result of global disease control programs implemented over the last decade, the number of TB cases is slowly declining each year. However, given that most deaths are preventable TB mortality is unacceptably high and efforts to combat it must be accelerated. Wide health care access to diagnosis, and correct treatment administration, are still unmet needs which dramatically delay TB elimination strategies. In 2013, about 64% of the total amount of people who developed TB was reported as newly diagnosed cases, yet the remaining 3 million cases were either not diagnosed or diagnosed but not reported to national TB programs.

Why is the TOPO-technology interesting - highlights:

ïƒ˜ - Powerful and very simple extraction method
ïƒ˜ - Only live pathogens are detected (no risk of false positive due to previous infections)
ïƒ˜ - Isothermal reaction
ïƒ˜ - Internal control possible
- Highly sensitive and specific detection due to the high copy number of the specific enzyme which generates numerous products that are further amplified before they are detected.
ïƒ˜ - Mutation independent
ïƒ˜ - TOBO-TB allows detection of TB in non-sputum samples

TOPO-TB design, test and concept validation. Based on the scientific results a viable and optimal business model for key markets through verification of our product and commercialization cost assumptions. The outcomes of this study will be 1) feasibility of concept fully proven with regard to TOPO-TB sensitivity and specificity range in comparison to state of the art technologies; 2) end-user requirements and market segment definition; 3) business model; 4) delineation of commercial and production supply chain of partners; and 5) financial risk analysis for each key market. The ultimate goal is to achieve, by 2018, a pilot launch in markets where TB incidence is the highest.

The TOPO-TB business model is created using the Business Model Canvas frame (a depiction of the model is attached below)

Work performed

The technology platform â€“ TOPO detection - presents a new concept that relies on an essential DNA modifying pathogen expressed enzyme (TopI) that is used as biomarker for infections. The use of essential enzymes as biomarkers ensures that only live pathogens are detected. Also, the fact that the assay detects pathogens directly rather than the immune response against pathogens in patients, adds further to optimize clinical decisions, since general health, time window after infection, vaccinations, age, gender or pregnancy will not affect the outcome of the test. Indeed early detection is very important in terms of treatment efficiency.
High sensitivity is anticipated due to the massive signal amplification inherent to the enzyme activity, as demonstrated previously . This is achieved by each target enzyme converting many (in theory infinite numbers) of substrate molecules to specific products, which are directly detected. The chosen enzyme target, the type I topoisomerases , is highly abundant in the plasmodium and mycobacteria cells with estimated copy numbers in the range of 100.000 in the examined cells , adding further to the expected sensitivity. Moreover, the target enzyme play essential roles for the pathogens rendering the risk for mutant stains that cannot be detected by TOPO detection extremely low or even zero.
The generic TOPO diagnostic platform is based on enzyme release using microfluidics, target identification and signal amplification followed by visual detection or read-out on paper. The concept can be integrated in one device suitable for point of care testing. Device requirement specifications will be validated in collaboration with FIND/WHO.

The TB diagnostics technology landscape has dramatically improved over the last years with several fully integrated tests aiming to reach the market. Nevertheless, despite the latest technological advancements and increasing diagnostic tools adoption, available methods are generally very costly and require extensive training skills, preventing their widespread adoption. In particular, the lack of both 1) a rapid, sputum-based, molecular test for microscopy centers and 2) a biomarker-based, low-cost, non-sputum-based test remain key priorities for an effective TB diagnostics.

Headlines of the results obtained - for details see attached slides:

The technology is slightly optimized in order to increse sensitivity (the basic principle is the same)

The TOPO-TB technology allows quantative detection

Different mycobacterial strains were all detected successfully using the TOPO technology

The TOPO-TB assay is specific to mycobacteria

Mycobacteria can be detected in samples mimicking saliva from TB patients

Conclusion: The TOPO-TB assay is fast, sensitive, specific and can be finally adapted to low respurce settings. The WHO requirements are fulfilled including the opportunity to detect TB from non-sputum samples.

Final results

There are several novel technologies out on the market, so why a need for a new one? Todayâ€™s technologies are first and foremost developed for the research laboratories and larger well established clinical laboratories. This translates into more expensive and technically demanding platforms. Existing established point-of-care technologies are fast and easy to perform but generally lack in sensitivity and specificity. A cost-efficient technology with high performance characteristics is still missing especially for applications e.g. malaria, tuberculosis and other common diseases in low-resource areas. We foresee Zymonostics technology TOPO to be a strong candidate to fill that gap.

Market trends:
The opportunity to reduce mortality caused by TB and even eliminate the disease is possible. Several global organizations (UNITAID, FIND) supervised by WHO are actively working towards gaining better management of the diseases by providing education and working with the industry to provide better tools. According the many studies conducted the highest priority for TB diagnostics would be to gain access to 1) rapid sputum-based molecular diagnostic test to replace the microscopic smear tests done today and 2) a rapid, very easy (even instrument free) biomarker based non-sputum sample test for decentralized testing, this test should also be suitable for children and extrapulmonary TB (EPTB).

Summary:
TOPO-TB holds the potential to fulfill these two high priority requirements. Our technology is not only isothermal, but requires no instrumentation for signal amplification. The technology as such works on various samples; non-invasive and invasive. TOPO-TB is also expected to be superior with regard to detection limit compared to the current gold standard tests and in contrast to LAMP it is simpler and relying on enzyme â€œpowerâ€ (due to the low detection limit of the TOPO-technology we expect to be able to diagnose TB without any cultivation steps). Finally, the TOPO-TB biomarker/target is a DNA modifying enzyme which is strictly required for the survival of M. tuberculosis. Hence, TOPO-TB is naturally less prone to false negatives than current NAATs, although maintaining specificity, sensitivity and simplicity.