Monoclonal Sandwich ELISA for Serum Erythroferrone for Clinical Assessment of Ineffective Erythropoiesis

Project Summary Abstract ErythroferroneERFEthe erythroid regulator of hepcidin has recently been discovered and characterized as a hormone expressed in erythroblasts in the marrowERFE responds to erythropoietinEPOsignaling from the EPO receptors in the marrow that stimulate ERFE synthesis and release into the plasmaERFE then acts as a suppressor of hepcidin expression in the liverHepcidin is the master regulator of iron absorption from the diet and iron recycling of red blood cellsRBCsby macrophagesThusERFE as a suppressor of hepcidinacts to increase iron absorption from the diet and plasma iron concentrations by freeing iron from stores in hepatocytesERFE was discovered in phlebotomized mice as a hormone involved with stress erythropoiesis following hemorrhage and was shown to be highly elevated from near undetectable levels at baseline in mouse models ofthalassemia and ineffective erythropoiesisIntrinsic LifeSciences licensed patent pending ERFE technology from UCLA and has subsequently developed a small suite of monoclonal antibodiesmAbsdirected at ERFEUsing a pair of these mAbs we produced a mAb sandwich research use onlyRUOELISA and began technical validation and clinical assessment of the kitThis proposal presents Preliminary Data supporting the technical quality of the ERFE test as well as its potential clinical utilityWe recapitulated the mouse phlebotomy experiments by sampling serum ERFE before and after plasma apheresis in blood donors and established the preliminary normal range for ERFE in humans in serum samples from first time blood donorsWe also established a preliminary lower limit of detectionLLODand lower limit of quantitationLLOQofandng mlThe median value of the normal range was observed to beng mlThis suggests that the ERFE sandwich ELSA is sensitive and suitable for commercial developmentWe confirmed clinical significance of the ERFE sandwich ELISA with de identified samples fromthalassemia and X linked sideroblastic anemiaXLSApatientsThe PI is requesting Direct to Phase II SBIR funding under PARto continue our exciting research and development efforts on the ERFE monoclonal sandwich ELISA over the next two yearsOur experience to date with the ERFE assay indicates that our research strategy is feasible and we expect the outcome of our Phase II RandD studies will be a fully automated clinical ERFE laboratory developed testLDTvalidated under Clinical Laboratory Improvement AmendmentsCLIAand College of American PathologistsCAPguidelinesThe CLIA CAP validated LDT would be available to healthcare providers in the US and would guide development and validation of a RUO kit for basic biology studies of ERFE in various disease statesAn additional deliverable will be a set of Design Controls necessary to manufacture a monoclonal sandwich IVD for ERFE that is suitable for FDA clearanceWe believe that our ERFE IVD will gain widespread clinical use for detection of ineffective erythropoiesis and as an important clinical diagnostic test forthalassemia syndromes