Project Grant R01DK140369

PROTEIN INTERACTIONS THAT CONTROL CALCIUM OXALATE STONE FORMATION - KIDNEY STONES REMAIN AN ENIGMATIC DISEASE DESPITE DECADES OF WORK BY MANY INVESTIGATORS WITH NO REAL ADVANCEMENT IN CALCIUM OXALATE (CAOX) STONE TREATMENT FOR MORE THAN 20 YEARS, BEYOND THE FOCUS ON LOWERING URINARY SUPERSATURATION WITH RESPECT TO CAOX CRYSTALS. WHILE THE RECURRENCE RATE CAN BE REDUCED, THE DISEASE CANNOT BE CURED WITH CURRENT THERAPIES. WITH PREVALENCE OF APPROXIMATELY 10% IN MOST DEVELOPED COUNTRIES AND INCREASING IN RECENT YEARS, RESEARCH THAT COULD DEFINE THE MECHANISMS UNDERLYING STONE FORMATION IS INCREASINGLY RELEVANT. ORGANIC MATERIALS (MAINLY PROTEINS) APPEAR TO COAT ALL CRYSTAL SURFACES IN CAOX STONES, AND THE PRESENCE OF PROTEINS APPEARS TO BE CRITICAL TO STONE FORMATION. THESE PROTEIN CRYSTAL AGGREGATES FORM SPONTANEOUSLY (SELF- ASSEMBLE) TO FORM DENSE SOLID OBJECTS WITH SUBSTANTIAL MECHANICAL STRENGTH. NO SINGLE PROTEIN IS CRITICAL TO THIS PROCESS. WE HAVE FOCUSED FOR MANY YEARS ON MIXTURES OF STRONG POLYANIONS (PA) WITH STRONG POLYCATIONS (PC), WHICH SELF AGGREGATE AT LOW CONCENTRATIONS AND INDUCE CAOX CRYSTAL AGGREGATION WHEN MIXED IN ROUGHLY EQUAL PROPORTIONS. DIRECT MEASUREMENTS OF PROTEIN COMPOSITIONS IN STONE MATRIX HAVE DEMONSTRATED ENRICHMENT OF PA AND PC IN STONES, BUT ALSO DEMONSTRATED A MAJOR FRACTION OF MANY WEAKLY IONIC PROTEINS THAT EXHIBIT PREFERENTIAL ADSORPTION TO STONE MATRIX COMPARED TO URINE. UNFORTUNATELY, THE SHEER NUMBER OF PROTEINS COMPRISING STONE MATRIX OBSCURES THE NATURE OF THEIR ROLE IN STONE FORMATION, YET THESE BROADLY CONSTITUTED MIXTURES CLEARLY YIELD HARD, DENSE AGGREGATES WHEN MIXED WITH CAOX CRYSTALS. THE PROTEIN MIXTURES FOUND IN OTHER KIDNEY STONE TYPES ARE SIMILAR IN MOST PROTEINS, AND OTHER MINERALIZED TISSUES, LIKE HUMAN BONE AND MOLLUSK SHELLS, CONTAIN SIMILARLY BROAD PROTEIN MIXTURES WITH MANY COMMON FEATURES. FURTHERMORE, STUDIES OF PA/PC MIXTURES IN POLYMER SCIENCE SHOW THAT THESE SYSTEMS PHASE SEPARATE (AGGREGATE), BUT DON'T ACHIEVE THE DENSE SOLID CONCENTRATIONS OF POLYMERS OBSERVED IN STONES. WE HYPOTHESIZE THAT CAOX STONE FORMATION IS DEPENDENT ON A COMPLEX MIXTURE OF PROTEINS, INCLUDING STRONGLY ANIONIC, STRONGLY CATIONIC, AND WEAKLY CHARGED PROTEINS THAT INTERACT WITH CAOX CRYSTALS TO PRODUCE THE DENSE PROTEIN-CRYSTAL AGGREGATES (COMPOSITES) WITH HIGH MECHANICAL STRENGTH THAT ARE KIDNEY STONES. WE WILL USE THE ESTABLISHED PROTEIN COMPOSITIONS OF CAOX STONE MATRIX AS A STARTING POINT FOR MAKING CONTROLLED MIXTURES OF WELL CHARACTERIZED POLYMERS AND PROTEINS AND STUDYING THEIR INTERACTIONS WITH CAOX CRYSTALS TO UNDERSTAND THE SELF-ASSEMBLY OF STONE LIKE COMPOSITES, AS WELL AS THEIR MECHANICAL PROPERTIES. OUR STUDY TEAM IS IDEALLY SUITED TO THIS TASK, INCLUDING A CLINICIAN WITH POLYMER PHYSICAL CHEMISTRY TRAINING AND KIDNEY STONE EXPERIENCE AND AN EXPERT IN CHEMICAL ENGINEERING AND POLYELECTROLYTE PHASE BEHAVIOR. THE PROPOSED WORK INCLUDES 3 SPECIFIC AIMS. 1: TEST A RANGE OF PROTEIN/POLYMER COMPOSITIONS FOR THEIR ABILITY TO ALTER CAOX CRYSTALLIZATION PROCESSES (NUCLEATION, GROWTH, AND AGGREGATION). 2: MEASURE THE MECHANICAL STRENGTH OF CAOX CRYSTAL AGGREGATES SPONTANEOUSLY FORMED WITH THESE MIXTURES. 3: PREPARE MACROSCOPIC COMPOSITES (SIMULATED STONES) FOR MECHANICAL TESTING TO BRIDGE UNDERSTANDING TO CLINICAL DISEASE.