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Out-of-hospital cardiac arrest is a leading cause of death in the US, with a mortality rate over 90%. Preclinical studies demonstrate that cooling during cardiopulmonary resuscitation (CPR) is highly beneficial, but can be challenging to implement clinically. No medications exist for improving long-term cardiac arrest survival. We have developed a 20- amino acid peptide, TAT-PHLPP9c, that mimics cooling protection by enhancing AKT activation via PH domain leucine-rich repeat phosphatase 1 (PHLPP1) inhibition. Complementary studies were conducted in mouse and swine. C57BL/6 mice were randomized into blinded saline control and peptide-treatment groups. Following a 12-minute asystolic arrest, TAT- PHLPP9c was administered intravenously during CPR and significantly improved the return of spontaneous circulation, mean arterial blood pressure and cerebral blood flow, cardiac and neurological function, and survival (4 hour and 5 day). It inhibited PHLPP-NHERF1 binding, enhanced AKT but not PKC phosphorylation, decreased pyruvate dehydrogenase phosphorylation and sorbitol production, and increased ATP generation in heart and brain. TAT-PHLPP9c treatment also reduced plasma taurine and glutamate concentrations after resuscitation. The protective benefit of TAT-PHLPP9c was validated in a swine cardiac arrest model of ventricular fibrillation. In conclusion, TAT-PHLPP9c may improve neurologically intact cardiac arrest survival without the need for physical cooling., Introduction Out-of-hospital cardiac arrest (CA) affects 350,000 people annually, with an overall survival rate of less than 10% (1). Cardiopulmonary resuscitation (CPR), defibrillation, and targeted temperature management (TTM) using active [...]

Induction of fetal hemoglobin (HbF) has been shown to be a viable therapeutic approach to treating sickle cell disease and potentially other β-hemoglobinopathies. To identify targets and target-modulating small molecules that enhance HbF expression, we engineered a human umbilical-derived erythroid progenitor reporter cell line (HUDEP2_HBG1_HiBiT) by genetically tagging a HiBiT peptide to the carboxyl (C)-terminus of the endogenous HBG1 gene locus, which codes for γ-globin protein, a component of HbF. Employing this reporter cell line, we performed a chemogenomic screen of approximately 5000 compounds annotated with known targets or mechanisms that have achieved clinical stage or approval by the US Food and Drug Administration (FDA). Among them, 10 compounds were confirmed for their ability to induce HbF in the HUDEP2 cell line. These include several known HbF inducers, such as pomalidomide, lenalidomide, decitabine, idoxuridine, and azacytidine, which validate the translational nature of this screening platform. We identified avadomide, autophinib, triciribine, and R574 as novel HbF inducers from these screens. We orthogonally confirmed HbF induction activities of the top hits in both parental HUDEP2 cells as well as in human primary CD34+ hematopoietic stem and progenitor cells (HSPCs). Further, we demonstrated that pomalidomide and avadomide, but not idoxuridine, induced HbF expression through downregulation of several transcriptional repressors such as BCL11A, ZBTB7A, and IKZF1. These studies demonstrate a robust phenotypic screening workflow that can be applied to large-scale small molecule profiling campaigns for the discovery of targets and pathways, as well as novel therapeutics for sickle cell disease and other β-hemoglobinopathies. [ABSTRACT FROM AUTHOR]

The actin filament-binding and filament-severing activities of the aplyronine, kabiramide, and reidispongiolide families of marine macrolides are located within the hydrophobic tail region of the molecule. Two synthetic tail analogues of aplyronine C (SF-01 and GC-04) are shown to bind to G-actin with dissociation constants of (285±33) and (132±13) nM, respectively. The crystal structures of actin complexes with GC-04, SF-01, and kabiramide C reveal a conserved mode of tail binding within the cleft that forms between subdomains (SD) 1 and 3. Our studies support the view that filament severing is brought about by specific binding of the tail region to the SD1/SD3 cleft on the upper protomer, which displaces loop-D from the lower protomer on the same half-filament. With previous studies showing that the GC-04 analogue can sever actin filaments, it is argued that the shorter complex lifetime of tail analogues with F-actin would make them more effective at severing filaments compared with plasma gelsolin. Structure-based analyses are used to suggest more reactive or targetable forms of GC-04 and SF-01, which may serve to boost the capacity of the serum actin scavenging system, to generate antibody conjugates against tumor cell antigens, and to decrease sputum viscosity in children with cystic fibrosis.