title: Nanomolar CFTR inhibition by pore-occluding divalent polyethylene glycol-malonic acid hydrazides.
abstract: Inhibitors of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel have potential application as antisecretory therapy in cholera. We synthesized mono- and divalent CFTR inhibitors consisting of a malonic acid hydrazide (MalH) coupled via a disulfonic stilbene linker to polyethylene glycols (PEGs; 0.2-100 kDa). IC50 values for CFTR inhibition were 10-15 microM for the monovalent MalH-PEGs, but substantially lower for divalent MalH-PEG-MalH compounds, decreasing from 1.5 to 0.3 microM with increasing PEG size and showing positive cooperativity. Whole-cell patch-clamp showed voltage-dependent CFTR block with inward rectification. Outside-out patch-clamp showed shortened single-channel openings, indicating CFTR pore block from the extracellular side. Luminally added MalH-PEG-MalH blocked by >90% cholera toxin-induced fluid secretion in mouse intestinal loops (IC50 approximately 10 pmol/loop), and greatly reduced mortality in a suckling mouse cholera model. These conjugates may provide safe, inexpensive antisecretory therapy.
authors.affiliation: Department of Medicine and Physiology, 1246 Health Sciences East Tower, University of California, San Francisco, CA 94143-0521, USA.
authors.forename: N D
authors.initials: ND
authors.lastname: Sonawane
authors.forename: Dan
authors.initials: D
authors.lastname: Zhao
authors.forename: Olga
authors.initials: O
authors.lastname: Zegarra-Moran
authors.forename: Luis J V
authors.initials: LJ
authors.lastname: Galietta
authors.forename: A S
authors.initials: AS
authors.lastname: Verkman
mesh_terms: D000818:Animals; D002772:Cholera Toxin / Q000737:chemistry; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D006801:Humans; D006834:Hydrazines / Q000737:chemistry*; D020128:Inhibitory Concentration 50; D009682:Magnetic Resonance Spectroscopy; D008314:Malonates / Q000737:chemistry*; D051379:Mice; D008807:Mice, Inbred BALB C; D008956:Models, Chemical; D018408:Patch-Clamp Techniques; D011092:Polyethylene Glycols / Q000737:chemistry*; D051381:Rats; D013267:Stilbenes / Q000737:chemistry
chemical_list: D006834:Hydrazines; D008314:Malonates; D013267:Stilbenes; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D011092:Polyethylene Glycols; D002772:Cholera Toxin; C030290:malonic acid