Project Grant F31HL184422

RNA APTAMERS FOR RNA DRUG DELIVERY TO CARDIAC CELLS - PROJECT SUMMARY RECENT ADVANCEMENTS IN SCIENTIFIC RESEARCH HAVE REVEALED THE IMMENSE POTENTIAL OF RNA-BASED DRUGS, OFFERING THERAPEUTIC OUTCOMES THAT OVERCOME THE LIMITATIONS OF CONVENTIONAL DRUGS. HOWEVER, THE DELIVERY OF RNA THERAPEUTICS, ESPECIALLY TO CARDIAC CELLS, REMAINS A CHALLENGE. IMPROVEMENTS IN THE DELIVERABILITY OF RNA INCLUDE LIPID NANOPARTICLES (LNPS), A KEY CLASS OF DRUG DELIVERY SYSTEM THAT INCLUDES NANOPARTICLES APPROVED BY THE FDA. BUT LNPS CARRYING PAYLOADS FOR THE CARDIOVASCULAR SYSTEM OFTEN DELIVER TO OFF-TARGET ORGANS LIKE THE LIVER, KIDNEY, AND SPLEEN, LIMITING THE DOSAGE DELIVERED TO CARDIAC CELLS AND THE THERAPEUTIC EFFECT. ANOTHER APPROACH IS THE CONJUGATION OF THE RNA DRUG TO A CHEMICAL GROUP OR POLYMER TO ENHANCE CELLULAR UPTAKE VIA ENDOCYTOSIS, SUCH AS THE SUGAR MOLECULE GALNAC (N-ACETYLGALATOSAMINE), WHICH TARGETS THE ASIALOGLYCOPROTEIN RECEPTOR (ASGPR) IN THE LIVER. WHILE THIS APPROACH OFFERS A STRAIGHT-FORWARD AND CHEAP SOLUTION FOR TARGETED DRUG DELIVERY, IT REQUIRES KNOWLEDGE OF RECEPTOR SUBSTRATE RELATIONSHIPS AND CELL-TYPE SPECIFIC SURFACE RECEPTORS AND SUBSTRATES; THERE IS NO ESTABLISHED RECEPTOR SUBSTRATE FOR DELIVERY TO CARDIAC CELLS. RNA APTAMERS ARE A CLASS OF NUCLEIC ACID-BASED MOLECULES THAT ARE EMERGING AS A PROMISING TOOL FOR TARGETED DRUG DELIVERY. HERE, WE HAVE UTILIZED CELL-INTERNALIZING SELEX TO IDENTIFY RNA SEQUENCES THAT ENTER PRIMARY MOUSE CARDIOMYOCYTES (CM) OR CARDIAC FIBROBLASTS (CF). THE GOAL OF THIS PROJECT IS TO IDENTIFY CM AND CF SPECIFIC RNA APTAMERS TO DELIVER RNA THERAPEUTICS TO TREAT CARDIAC HYPERTROPHY AND FIBROSIS. WE DEMONSTRATED THE INTERNALIZATION OF THE TOP-HIT CM AND CF RNA APTAMERS WITH RT-QPCR AND FLUORESCENCE CONFOCAL MICROSCOPY. WE FUNCTIONALLY TESTED THE CM APTAMER'S ABILITY TO DELIVER AN ANTISENSE OLIGONUCLEOTIDE (ASO) THAT SUCCESSFULLY DOWNREGULATED MALAT1 IN CMS. IN AIM 1, I WILL FURTHER CHARACTERIZE THE IDENTIFIED APTAMERS BY VALIDATING THE SPECIFICITY OF THE APTAMERS USING BIOCHEMICAL MEASUREMENT AND FLUORESCENT IMAGING. I WILL ALSO IDENTIFY THE APTAMERS' TARGET RECEPTOR TO REVEAL RECEPTOR-SUBSTRATE RELATIONSHIPS IN CARDIAC CELLS. IN AIM 2, I WILL ESTABLISH THE APTAMERS' ABILITY TO DELIVER ASOS THAT MANAGE FIBROSIS AND HYPERTROPHY IN VITRO WITH PMCF AND PMCM. THIS PROJECT WILL LAY THE FOUNDATIONAL GROUNDWORK FOR A NOVEL TARGETED DELIVERY PLATFORM TO CARDIAC CELL TYPES IMPLICATED IN HEART DISEASE.