Discovery of a novel, highly potent and orally bioavailable pyrrolidinone indole series of irreversible Myeloperoxidase (MPO) inhibitors.

Myeloperoxidase (MPO) is a heme-containing peroxidase from phagocytic cells, which plays an important role in the innate immune response. The primary anti-microbial function of MPO is achieved by catalyzing the oxidation of halides by hydrogen peroxide (H2O2). Upon activation of phagocytes, MPO activity is detectable in both phagosomes and extracellularly, where it can remain or transcytose into interstitial compartments. Activated MPO leads to oxidative stress and tissue damage in many inflammatory states, including cardiovascular disease. Starting from a low molecular weight (LMW) high throughput screening (HTS) hit, here we report the discovery of a novel pyrrolidinone indole (IN-4) as a highly potent MPO inhibitor. This compound displays similar in vitro potency across peroxidation, plasma and NETosis assays. In a dilution/dialysis study, <5% of the original MPO activity was detected post-incubation of MPO with IN-4, suggesting irreversible enzyme inhibition. A fast MPO inactivation rate (kinact/Ki) and low partition ratio (k3/k4) make IN-4 kinetic properties attractive for an MPO inhibitor. This compound also displays significant selectivity over the closely related thyroid peroxidase (TPO), and is selective for extracellular MPO over intracellular (neutrophil) MPO. Moreover, IN-4 shows good exposure, low clearance and high oral bioavailability in mice, rats and dogs. The high in vitro MPO activity and high oral exposure observed with IN-4 result in a dose-dependent inhibition of MPO activity in three mouse models of inflammation. In conclusion, IN-4 is a novel, potent, mechanism-based and selective MPO inhibitor, which may be used as superior therapeutic agent to treat multiple inflammatory conditions, including cardiovascular disease.

Insights into the red algae and eukaryotic evolution from the genome of Porphyra umbilicalis (Bangiophyceae, Rhodophyta).

Porphyra umbilicalis (laver) belongs to an ancient group of red algae (Bangiophyceae), is harvested for human food, and thrives in the harsh conditions of the upper intertidal zone. Here we present the 87.7-Mbp haploid Porphyra genome (65.8% G + C content, 13,125 gene loci) and elucidate traits that inform our understanding of the biology of red algae as one of the few multicellular eukaryotic lineages. Novel features of the Porphyra genome shared by other red algae relate to the cytoskeleton, calcium signaling, the cell cycle, and stress-tolerance mechanisms including photoprotection. Cytoskeletal motor proteins in Porphyra are restricted to a small set of kinesins that appear to be the only universal cytoskeletal motors within the red algae. Dynein motors are absent, and most red algae, including Porphyra, lack myosin. This surprisingly minimal cytoskeleton offers a potential explanation for why red algal cells and multicellular structures are more limited in size than in most multicellular lineages. Additional discoveries further relating to the stress tolerance of bangiophytes include ancestral enzymes for sulfation of the hydrophilic galactan-rich cell wall, evidence for mannan synthesis that originated before the divergence of green and red algae, and a high capacity for nutrient uptake. Our analyses provide a comprehensive understanding of the red algae, which are both commercially important and have played a major role in the evolution of other algal groups through secondary endosymbioses.

Allosteric LFA-1 inhibitors modulate natural killer cell function.

Natural killer (NK) cells are believed to play an important role in a variety of disease pathologies, including transplant rejection and autoimmunity. None of the therapeutic modalities currently available are known to potently interfere with NK cell activity. Here we demonstrate for the first time that low molecular weight inhibitors of the integrin lymphocyte function-associated antigen-1 (LFA-1) readily block NK cell adhesion, activation, and NK cell-mediated cytolysis in vitro, in contrast to other immunosuppressive agents. These effects were independent of the type of allosteric mechanism by which LFA-1 inhibition was achieved. In addition, we describe a simple, nonradioactive whole-blood assay that should be suitable to monitor NK cell activation in clinical practice. Taken together, our study underlines the importance of LFA-1 in NK cell effector functions and indicates that allosteric LFA-1 inhibitors may become important tools to further elucidate the therapeutic potential of NK cell modulation in immunological diseases.