Project Grant R43AR079941

DEVELOPMENT OF DNA-LINKED MATRIX METALLO PROTEASE ACTIVITY PROBES IN OSTEOARTHRITIS RESEARCH - PROJECT SUMMARY CURRENTLY, MORE THAN 32 MILLION AMERICAN SUFFER FROM OSTEOARTHRITIS (OA), WHICH INCURRED HEALTHCARE COSTS OF ~$5 BILLION IN FY2019. THESE COSTS ARE HIGH LARGELY BECAUSE PHYSICIANS LACK EARLY DIAGNOSIS CAPABILITIES THAT WOULD PERMIT THE PRESCRIPTION OF PREVENTATIVE THERAPIES TO AT-RISK INDIVIDUALS. IN ADDITION, NO CONSENSUS BIOMARKER EXISTS FOR OA DIAGNOSIS DUE TO THE LACK OF VALIDATION AND POOR QUANTIFICATION, WHICH IMPEDES OA PHARMACEUTICAL DEVELOPMENT. THEREFORE, THERE IS AN UNMET NEED FOR NEW CLINICAL AND RESEARCH TOOLS FOR OA. ACTIVE MATRIX-METALLOPROTEASES (MMPS) PLAY SIGNIFICANT ROLES IN THE PATHOGENESIS OF OA BY DEGRADING EXTRACELLULAR MATRIX COMPONENTS. HIGH LEVELS OF CERTAIN MMPS WERE OBSERVED IN THE SYNOVIAL FLUID OF OA PATIENTS. THE EARLY DETECTION OF ELEVATED MMPS MAY BE OF CLINICAL BENEFIT AS IT WOULD PERMIT THE EARLIER IMPLEMENTATION OF THERAPEUTIC REGIMENS CAPABLE OF DELAYING OR PREVENTING DISEASE PROGRESSION. THESE MMPS BELONG TO A FAMILY OF ZINC- DEPENDENT PROTEASES IMPLICATED IN NUMEROUS DISEASE TYPES, SUCH AS CANCERS, CARDIOVASCULAR DISEASE, ARTHRITIS, AND AUTOIMMUNE DISEASES. CURRENT METHODS USED BY RESEARCHERS AND CLINICIANS TO MONITOR MMP ACTIVITIES LACK MANY DESIRABLE FEATURES SUCH AS SPECIFICITY, SENSITIVITY, AND ACCURACY. THE GOAL OF THIS PHASE I STUDY IS TO ADDRESS UNMET CLINICAL NEEDS BY DEVELOPING A HIGHLY SENSITIVE, SELECTIVE, AND MULTIPLEXED MMP ACTIVITY ASSAY USING DENCODA'S PROPRIETARY AND INNOVATIVE DNA-BASED ACTIVITY DETECTION SYSTEM USING OA AS A MODEL SYSTEM. THE USE OF NUCLEIC ACID-BASED TECHNOLOGIES IN LABORATORY AND CLINICAL ASSAYS IS WELL-KNOWN AND WELL-PROVEN IN MOST THE RECENT COVID-19 OUTBREAK. OUR ACTIVITY ASSAY USES MMP ACTIVE SITE PROBES TO ENCODE THE ACTIVITIES OF TARGET MMPS INTO DNA. AS SUCH, OUR PLATFORM REALIZED MULTIPLE ADVANTAGES OVER CURRENT ASSAYS, SUCH AS REPORTING ON THE SPECIFIC MMP ACTIVITIES AS OPPOSED TO LEVELS ALONE, COST EFFECTIVENESS, HIGH THROUGHPUT, MULTIPLEXING FORMAT, AND HIGH SENSITIVITY. IN AIM 1, WE WILL DEVELOP AND VALIDATE MULTIPLEXING DNA-LINKED MMP ACTIVITY PROBES SPECIFIC TO MMP-1 AND -9. SYNOVIAL FLUID FROM OA PATIENTS WILL BE USED FOR THIS VALIDATION. IN AIM 2, WE WILL FURTHER REDUCE THE COST OF OUR PROPRIETARY ASSAY SYSTEM. CURRENTLY, OUR PLATFORM REQUIRES ANTIBODIES SPECIFIC TO EACH INDIVIDUAL MMP. WE WILL AIM TO REDUCE COSTS VIA THE SELECTION OF MMP-1 AND -9-SPECIFIC BINDING MOLECULES USING IN VITRO SELECTION METHODS, NAMELY, MRNA DISPLAY AND DNA APTAMER SELECTION. OVERALL, THE KEY METRICS FOR SUCCESS FOR THIS PHASE I STUDY WILL BE THE DEMONSTRATION AND VALIDATION OF THE ASSAY SPECIFICITY WITH A DYNAMIC RANGE OF 5-LOG10 SCALE COMPARED TO NEGATIVE CONTROLS. THE SUCCESSFUL OUTCOME OF THESE STUDIES WILL VALIDATE THE FEASIBILITY OF OUR INNOVATIVE ASSAY FORMAT FOR SPECIFIC MMP ACTIVITY DETECTION. ULTIMATELY, OUR NOVEL DNA-BASED MMP ACTIVITY ASSAY PLATFORM WILL ADDRESS THE LIMITATIONS FACING OA DIAGNOSIS AND PHARMACEUTICAL DEVELOPMENT BY LOWERING HEALTHCARE COSTS AND REVEALING NEW AVENUES FOR NEXT GENERATION OA THERAPEUTICS.