Targeting anti-apoptotic drugs to failing cardiomyocytes
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115A Commerce Drive, Brookfield, CT, 06804
AbstractDESCRIPTION (provided by applicant): Heart failure is the major cause of morbidity and mortality worldwide. It is recognized that cardiomyocyte apoptosis plays an important role in heart failure. Intracellular proteases, known as caspases, play crucial rol e in initiation and execution of apoptosis. Experimental evidence indicates that timely administration of caspase inhibitors diminishes cardiomyocyte apoptosis and heart failure. However, clinical development of caspases inhibitors is difficult because as polar compounds they have difficulties in crossing cell membrane and eventually accumulate in the liver. We propose to test a hypothesis that early apoptotic cardiomyocytes can be rescued by targeted intracellular delivery of caspase inhibitors. As a targeting protein we propose to use annexin V, a protein that binds to phosphatidylserine, which is displayed on the surface of apoptotic cells, including apoptotic cardiomyocytes. Recent evidence indicates that annexin V is internalized by apoptotic ce lls, and therefore may be suitable for intracellular drug delivery. To increase the number of drug molecules that are delivered by annexin V, we propose to encapsulate caspase inhibitors into liposomes. For targeting to apoptotic cardiomyocytes, an nexin V conjugated to phospholipid will be inserted into the lipid membrane of drug-loaded liposomes. To avoid damaging annexin V by random conjugation, we will use a mutant annexin V expressed with N-terminal cysteine-containing tag for site-specific modi fication. Our preliminary data with this protein indicate that various payloads can be conjugated to this tag without affecting protein activity. In this exploratory project we will develop annexin V decorated liposomes and test their ability to de liver caspase inhibitors in amounts sufficient for rescuing cardiomyocytes from apoptotic injury. If our construct can rescue apoptotic cardiomyocytes in vitro, it provide experimental foundation and rational for testing this rescue strategy in vivo.
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