Chemical strategies to using small molecules to stimulate hypoxia inducible factors (HIFs) activity and trigger a hypoxic response under normoxic conditions, such as iron chelators and inhibitors of prolyl hydroxylase domain (PHD) enzymes, have broad-spectrum activities and off-target effects. Here we disclose VH298, a potent VHL inhibitor that stabilizes HIF-α and elicits a hypoxic response via a different mechanism, that is the blockade of the VHL:HIF-α protein–protein interaction downstream of HIF-α hydroxylation by PHD enzymes. We show that VH298 engages with high affinity and specificity with VHL as its only major cellular target, leading to selective on-target accumulation of hydroxylated HIF-α in a concentration- and time-dependent fashion in different cell lines, with subsequent upregulation of HIF-target genes at both mRNA and protein levels. VH298 represents a high-quality chemical probe of the HIF signalling cascade and an attractive starting point to the development of potential new therapeutics targeting hypoxia signalling.
SummaryWidespread resistance to first-line TB drugs is a major problem that will likely only be resolved through the development of new drugs with novel mechanisms of action. We have used structure-guided methods to develop a lead molecule that targets the thioesterase activity of polyketide synthase Pks13, an essential enzyme that forms mycolic acids, required for the cell wall of Mycobacterium tuberculosis. Our lead, TAM16, is a benzofuran class inhibitor of Pks13 with highly potent in vitro bactericidal activity against drug-susceptible and drug-resistant clinical isolates of M. tuberculosis. In multiple mouse models of TB infection, TAM16 showed in vivo efficacy equal to the first-line TB drug isoniazid, both as a monotherapy and in combination therapy with rifampicin. TAM16 has excellent pharmacological and safety profiles, and the frequency of resistance for TAM16 is ∼100-fold lower than INH, suggesting that it can be developed as a new antitubercular aimed at the acute infection.PaperClip
The
von Hippel–Lindau tumor suppressor protein is the substrate
binding subunit of the VHL E3 ubiquitin ligase, which targets hydroxylated
α subunit of hypoxia inducible factors (HIFs) for ubiquitination
and subsequent proteasomal degradation. VHL is a potential target
for treating anemia and ischemic diseases, motivating the development
of inhibitors of the VHL:HIF-α protein–protein interaction.
Additionally, bifunctional proteolysis targeting chimeras (PROTACs)
containing a VHL ligand can hijack the E3 ligase activity to induce
degradation of target proteins. We report the structure-guided design
and group-based optimization of a series of VHL inhibitors with low
nanomolar potencies and improved cellular permeability. Structure–activity
relationships led to the discovery of potent inhibitors 10 and chemical probe VH298, with dissociation constants <100 nM,
which induced marked HIF-1α intracellular stabilization. Our
study provides new chemical tools to probe the VHL-HIF pathways and
new VHL ligands for next-generation PROTACs.
The authors propose that the capture of rocuronium by Org 25969 causes the rapid reversal of neuromuscular block. The reversal can be explained by the rapid transfer of free rocuronium from the effect compartment (neuromuscular junction) to the central compartment, in which it is bound to Org 25969. This explains the increase in total plasma concentration of rocuronium (free and bound to Org 25969).
Introduction. Glycine is one of the major inhibitory neurotransmitters in the spinal cord and brain stem of vertebrates. 1 The inhibitory actions of glycine are mediated by the strychnine-sensitive glycine receptor (ssGlyR), a ligand-gated chloride channel distributed throughout the spinal cord and brain stem. 2 Glycine is also known to potentate the action of glutamate acting as an essential co-agonist on postsynaptic N-methyl-D-aspartate (NMDA) receptors. 3 Synaptic levels of glycine are believed to be controlled by high-affinity glycine transporters. These transporters are members of a large family of sodium/chloride-dependent transporters, which are composed of single oligomeric proteins containing 12 hydrophobic membrane-spanning domains. 4 Molec-
A potent, non-cytotoxic indazole sulfonamide was identified by
high-throughput screening of >100,000 synthetic compounds for activity
against Mycobacterium tuberculosis (Mtb). This
non-cytotoxic compound did not directly inhibit cell wall biogenesis but
triggered a slow lysis of Mtb cells as measured by release of
intracellular green fluorescent protein (GFP). Isolation of resistant mutants
followed by whole-genome sequencing showed an unusual gene amplification of a 40
gene region spanning Rv3371 to Rv3411c and in
one case a potential promoter mutation upstream of guaB2
(Rv3411c) encoding inosine monophosphate dehydrogenase
(IMPDH). Subsequent biochemical validation confirmed direct inhibition of IMPDH
by an uncompetitive mode of inhibition and growth inhibition could be rescued by
supplementation with guanine, a bypass mechanism for the IMPDH pathway. Beads
containing immobilized indazole sulfonamides specifically interacted with IMPDH
in cell lysates. X-ray crystallography of the IMPDH-IMP-inhibitor complex
revealed that the primary interactions of these compounds with IMPDH were direct
pi-pi interactions with the IMP substrate. Advanced lead compounds in this
series with acceptable pharmacokinetic properties failed to show efficacy in
acute or chronic murine models of tuberculosis (TB). Time-kill experiments
in vitro suggest that sustained exposure to drug
concentrations above MIC for 24 hours were required for a cidal effect, levels
that have been difficult to achieve in vivo. Direct measurement
of guanine levels in resected lung tissue from tuberculosis infected animals and
patients revealed 0.5–2 mM concentrations in caseum and normal lung
tissue. The high lesional levels of guanine and the slow lytic, growth-rate
dependent, effect of IMPDH inhibition pose challenges to developing drugs
against this target for use in treating TB.
With
the emergence of multidrug-resistant strains of Mycobacterium
tuberculosis there is a pressing need for new oral drugs
with novel mechanisms of action. Herein, we describe the identification
of a novel morpholino–thiophenes (MOT) series following phenotypic
screening of the Eli Lilly corporate library against M. tuberculosis strain H37Rv. The design, synthesis, and structure–activity
relationships of a range of analogues around the confirmed actives
are described. Optimized leads with potent whole cell activity against
H37Rv, no cytotoxicity flags, and in vivo efficacy in an acute murine
model of infection are described. Mode-of-action studies suggest that
the novel scaffold targets QcrB, a subunit of the menaquinol cytochrome c oxidoreductase, part of the bc1-aa3-type cytochrome c oxidase complex that is responsible for driving oxygen-dependent
respiration.
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