SBIR-STTR Award

Dengue fever and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) have emerged as the most important arthropod-borne viral diseases of humans. There are four distinct dengue virus types (DEN-1, DEN-2, DEN-3, and DEN-4), each capable of causing disease in humans. The conserved 3°_-noncoding sequences of four dengue virus serotypes have been successfully utilized to develop as a TaqMan-based RT-PCR (funded by MIDRP STO A/L) to quantitatively identify dengue viruses from different regions of the world. The 0bjective of this phase I proposal is to transit the mature real-time reverse transcriptase polymerase chain reaction (RT-PCR) technology of the dengue 3°_-noncoding region based assay system (develop by Walter Reed Army Institute of Research) into a field deployable and user-friendly diagnostics device, so that the dengue diagnostics technology can be used in the dengue-endemic areas where the US military may be deployed in the future. Access Bio will optimize and scale up production of the fluorogenic RT-PCR reagent kits for serotype-specific dengue virus detection. We will develop lyophilized kit components, then evaluate the kit sensitivity, specificity, and stability using various concentrations of cultured dengue virus or dengue virus cDNA. Access Bio has developed a proprietary fluorescence technology that we will prove for this application. According to the Center for Disease Control, as of 1997 dengue became the most important mosquito-borne viral disease affecting humans. Its global distribution is comparable to that of malaria and an estimated 2.5 billion people live in areas at risk for epidemic transmission. Each year, tens of millions of cases of dengue fever occur and, depending on the year, up to hundreds of thousands of cases of dengue hemorrhagic fever (DHF). In addition, increased travel by airplane provides the ideal mechanism for transporting dengue viruses between population centers of the tropics, resulting in a constant exchange of dengue viruses and other pathogens. After successful research in the phase I study and followed by extensive research in phase II, this system could be immediately commercialized through the US military, the Center for Disease Control, US public health agencies, the Pan American Health Organization, and health organizations in countries where dengue is endemic. If this technology is feasible, we will license out the products developed to partner larger diagnostics companies for marketing. Given the increased the spread of dengue virus epidemic transmission concerns, the products developed to detect serotype specific dengue virus will likely see considerable market opportunity.

Keywords:
Dengue Virus, Flavivirus, Dengue Serotype, Fluorogenic Rt-Pcr, Ready To Use, Lyophilized Reagent Pads, Time-Resolved Fluorescence, Lateral Flow Immuno

The first objective of Phase II is to validate the ready-to-use DNA-based dengue diagnostics kit under laboratory conditions using culture-derived dengue viruses as well as clinically confirmed dengue infected samples from different geographic origins. These prototype products will be developed for military and civilian settings which have ideal laboratory conditions. The second objective of Phase II is to develop a point-of-care rapid detection system for detecting dengue viruses with a ready-to-use RT-PCR kit. Such systems provide stable and convenient PCR result analysis without gel electrophoresis. This system will be useful at the non-ideal laboratory conditions and in field conditions.. This method is unique, specific, and easy to operate in a field-deployable system that can provide rapid results in a few minutes with one or two simple steps. It is considered as a very convenient point-of-care test since it does not require a sophisticated facility, equipment, and highly trained personnel to perform the assay. This system will achieve comparable results to the fluorogenic RT-PCR. When an expensive real time PCR machine is not available, this kit will provide comparable test results with a low priced conventional PCR machine.

Keywords:
Dengue Virus, Flavivirus, Dhf/Dss, Ready-To-Use Kit, Fluorogenic Rt-Pcr, Serotype Specific, Dengue S