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A point-of-care assay to determine soldier dengue exposure and enable rapid, mass, cost-efficient dengue vaccination programs of military personnel

Description:

TECHNOLOGY AREA(S): Bio Medical 

OBJECTIVE: Develop a specific cost-effective point-of-care assay to rapidly identify dengue exposure history in the warfighter enabling force protection readiness and high-throughput mass dengue vaccination programs 

DESCRIPTION: Dengue virus (DENV) is a growing threat to tropical regions and the warfighter. It is a leading cause of fever in military deployed to tropical regions [Gibbons, EID, 2012], and the global incidence of dengue has dramatically increased in the last three decades [Messina, Trends Microbiol, 2014]. DENV infections may lead to loss of operational readiness and maybe life threatening and difficult to manage in austere settings [WHO, https://www.ncbi.nlm.nih.gov/books/NBK143157/, 2009]. The four dengue virus serotypes complicate risk management and readiness. First dengue infections are often asymptomatic and dramatically increase the chance of a more severe infection when exposed to a second dengue serotype [Soo, PLOS One, 2011]. Clinical trials have shown that dengue vaccines are less effective and potentially dangerous when administered to those who have not been previously exposed to dengue, as is the case with the majority of the US Military [Sridhar, NEJM, 2018; Gibbons, EID, 2012]. The World Health Organization now recommends that determining dengue exposure history (serostatus) is critical to inform what vaccine strategy should be used in order to maximize dengue protection and avoid vaccine related harm [Vannice, Vaccine, 2018]. Immunological assays which can detect prior exposure to dengue do exist, but they are not point-of-care [Welch, J Virol Methods, 2014]. Moreover, standard immunoassays such as ELISA lack specificity to DENV as they cross-react to other flaviviruses, particularly Zika virus (ZIKV), and may cross react with Yellow-Fever and Japanese Encephalitis vaccine-derived antibodies [Priyamvada, PNAS, 2016]. The current, approved, gold-standard to accurately determine dengue exposure history is to use plaque-reduction neutralizing assays (PRNT). However, PRNTs are cumbersome, require a specimen send-out to a central reference laboratory, have low reliability between laboratories, and are costly [Rainwater-Lovett, BMC ID, 2012]. The turn-around time for a PRNT result is too long when large-scale dengue vaccine programs are underway, particularly for time-sensitive mass deployments. Waiting for PRNT results would be unfeasible even for routine recruit basic training vaccination programs. A more specific ELISA assay has been recently developed, but this is non-FDA approved and there is limited, if any, clinical experience with its use [Balmaseda, PNAS, 2017]. This assay was designed primarily as a clinical ZIKV diagnostic assay. Moreover, this assay is not point-of-care and requires laboratory expertise for use and interpretation [Balmaseda, PNAS, 2017]. Other point-of-care diagnostic assays for dengue are designed to detect acute infections, not prior dengue exposure [Zainah, J Virol Methods, 2009] A rapid, point-of-care assay which measures dengue exposures of recruits would allow immediate determination of whether a warfighter can be vaccinated for dengue, and/or whether the warfighter requires a different dengue vaccine product or schedule. This would be vital for time-sensitive dengue vaccination programs of large volumes of troops before deployment when it would not be possible to obtain their serostatus in a feasible time frame. It would also remove the need to perform thousands to millions of expensive, time-demanding PRNT assays in existing Department of Defense laboratories which currently do not have the capacity to perform such a large volume of reference dengue diagnostic tests. A point of care dengue specific exposure device will reduce risk of any current or future dengue vaccine and pre-deployment dengue exposure testing will identify soldiers and general population that are at increased risk of severe disease when traveling or deploying to high risk dengue regions of the world. Of note this device would not be an acute disease diagnostic to identify pathogens causing fever and dengue disease. While current acute point-of-care febrile disease diagnostic platforms can estimate primary versus secondary dengue infection, they are calibrated and validated to do so only during an acute febrile illness and they cannot accurately detect prior dengue exposure in the asymptomatic host which would typically have far lower circulating levels of anti-flavivirus antibodies. 

PHASE I: By the end of Phase I the successful applicant will have: (i) Conceptualized the assay to include potential targets of dengue verse other related flavivirus specificity and demonstrate with design and data package supporting claims of specificity. (ii) Conceptualized and defined the target assay characteristics including a) biospecimen type (e.g. blood, sera), biospecimen volume required and proposed route of access (e.g. fingerstick, 1mL of sera derived from venipuncture); b) analyte of interest (e.g. antibody class) and method of detection; c) envisaged assay read out (e.g. colorimetric, digital); d) anticipated field operating characteristics including assay thermostability, cold chain requirements, necessary reagents, and operator skill required to perform and interpret the assay; and e) crude cost estimate for each assay unit (iii) Developed technological milestones for the full development of this assay (iv) Outlined a set of performance goals for the validation of this assay, including in-vitro validations and subsequent clinical validations (iv) Explicitly described the target protein(s), antigen(s) or antibody(s) or other analytical target, including their reproducible functional and structural characteristics While the awardee is expected to select an appropriate target analyte, they are strongly encouraged to co-ordinate the choice of antigen/antibody with the COR Specifically, the awardee will have: • Performed the assay in a research laboratory setting and demonstrated that it can be performed without any laboratory infrastructure (i.e., demonstrated the feasibility of point-of-care use). • Performed the assay in a research laboratory setting and demonstrated the feasibility of using only a relatively small amount of biospecimen which would be readily available in a point-of-care setting (using a small amount of biospecimen) to detect the analyte. • Evaluated the prototype product on a pilot panel of flavi-virus exposed and unexposed biological specimens available through the COR 

PHASE II: The Phase II deliverables will include: (i) Construction and demonstration of the operation of the assay prototype (ii) A detailed plan for clinical validation (iii) Performance of a clinical validation of the assay on archived or prospectively collected bio-specimens from humans and higher order animals with known exposure to DENV, ZIKV, other flaviviruses and flavivirus vaccines already determined by gold-standard methods. This validation will include metrics of assay validity and reliability, with estimates of uncertainty around these metrics. This validation must address the broad genetic and antigenic diversity of DENV by global location. The expected performance parameters would be a sensitivity greater than 90% and a specificity greater than 90%, although the target performance characteristics may depend on the setting of use (see Phase III) and the pre-test probability of disease exposure and can be coordinated and refined with the COR. 

PHASE III: The expected Phase III end-state is an FDA approved, low-cost, point-of-care, closed-system, easy-to-use and easy-to-interpret assay which can be used on a relatively low volume of easily accessible biospecimen. The transfer from research to operational capacity would occur via the biotechnological industry pathway, such that appropriate scale up and feasible unit costs can be accommodated. This end-product would likely be used in vaccination clinics during basic recruit training and/or in vaccination clinics as part of large-scale pre-deployment soldier readiness programs. It is envisaged that this point-of-care test that could be operated by a nurse or other healthcare professional in the office without the need for laboratory expertise. The specific indication would to immediately determine a soldier’s dengue serostatus, permitting an on-the-spot decision about which vaccine product/schedule they need to receive (including a decision whether it is safe for the soldier to receive any dengue vaccine at all). A similar consumer group may be civilians presenting to a travel clinic for pre-travel dengue risk advice and vaccination. This end-product would also be critical for civilian population dengue vaccination programs and we would envisage it would be used on a population scale in dengue endemic regions to facilitate widespread dengue vaccination programs which are projected to greatly reduce the overall burden of dengue in many tropical regions, but which are currently restricted by host serostatus safety concerns. Current guidance from the WHO for the current single licensed dengue vaccine (which is now licensed in over 20 countries) is that the host dengue sero-status should be determined before vaccination [Vannice, Vaccine, 2018]. However, this may be logistically and financially prohibitive in many lower-resource regions, even if recently developed ELISA platforms are extensively validated and approved for diagnostic use [Turner, Trans R Soc Trop Med Hyg. 2018]. This envisaged product would therefore ‘unlock’ the full potential of currently licensed, and perhaps future dengue vaccines, to substantially reduce the burden of this disease. 

REFERENCES: 

1: Balmaseda A, Stettler K, Medialdea-Carrera R, Collado D et al. Antibody-based assay discriminates Zika virus infection from other flaviviruses. Proc Natl Acad Sci U S A. 2017 Aug 1

2: 114(31):8384-8389. doi: 10.1073/pnas.1704984114. Epub 2017 Jul 17.

3:  Gibbons RV, Streitz M, Babina T, Fried JR. Dengue and US military operations from the Spanish-American War through today. Emerging infectious diseases. 2012

4: 18(4):623-30

5:  Priyamvada L, et al. Human antibody responses after dengue virus infection are highly cross-reactive to Zika virus. Proc Natl Acad Sci USA. 2016

6: 113:7852–7857

7:  Sridhar S, Luedtke A, Langevin E, Zhu M, Bonaparte M, Machabert T, Savarino S, Zambrano B, Moureau A, Khromava A, Moodie Z, Westling T, Mascareñas C, Frago C, Cortés M, Chansinghakul D, Noriega F, Bouckenooghe A, Chen J, Ng SP, Gilbert PB, Gurunathan S, DiazGranados CA. Effect of Dengue Serostatus on Dengue Vaccine Safety and Efficacy. N Engl J Med. 2018 Jul 26

8: 379(4):327-340

9:  Vannice KS, Wilder-Smith A, Barrett ADT, Carrijo K, Cavaleri M, de Silva A, Durbin AP, Endy T, Harris E, Innis BL, Katzelnick LC, Smith PG, Sun W, Thomas SJ, Hombach J. Clinical development and regulatory points for consideration for second-generation live attenuated dengue vaccines. Vaccine. 2018 Jun 7

10: 36(24):3411-3417

KEYWORDS: Dengue Virus, Flaviviruses, Zika Virus, Dengue Vaccines, Diagnostic Assay, Immunoassay, Serostatus, Soldier Lethality 

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