Objective:The aim was to determine whether failing human hearts have increased sensitivity to the inotropic and toxic effects of ouabain, and to examine alterations in NdK-ATPase that might explain the observed higher ouabain sensitivity. Methods: For contractility studies, a total of 57 trabeculae were isolated from two nonfailing (death from head injury) and 10 terminally failing, explanted human hearts. After the experiment, each trabecula was inspected under the light microscope for morphological alterations consistent with heart failure. Samples for biochemical and molecular studies were obtained from five non-failing and 13 failing hearts. Total NdK-ATPase was measured in desoxycholate treated homogenates and expressed per unit of tissue wet or dry weight, DNA, protein, or myosin. Interference from residual bound digoxin due to previous therapy was excluded. The expression of the three ci isoforms was studied at both the mRNA level using northern blots and the protein level by analysis of dissociation kinetics of the [3H]ouabain-enzyme complex. Results: Trabeculae showing morphological alterations and decreased contractility were sensitive to lower concentrations of ouabain (3-100 nM) than control trabeculae (100-1000 nM); the inotropic ECSo and the minimum toxic concentration were both reduced.[3H]Ouabain binding was significantly lower (p<

Complement factor D (FD), a highly specific S1 serine protease, plays a central role in the amplification of the alternative complement pathway (AP) of the innate immune system. Dysregulation of AP activity predisposes individuals to diverse disorders such as age-related macular degeneration, atypical hemolytic uremic syndrome, membranoproliferative glomerulonephritis type II, and paroxysmal nocturnal hemoglobinuria. Previously, we have reported the screening efforts and identification of reversible benzylamine-based FD inhibitors (1 and 2) binding to the open active conformation of FD. In continuation of our drug discovery program, we designed compounds applying structure-based approaches to improve interactions with FD and gain selectivity against S1 serine proteases. We report herein the design, synthesis, and medicinal chemistry optimization of the benzylamine series culminating in the discovery of 12, an orally bioavailable and selective FD inhibitor. 12 demonstrated systemic suppression of AP activation in a lipopolysaccharide-induced AP activation model as well as local ocular suppression in intravitreal injection-induced AP activation model in mice expressing human FD.

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