Rhinoviruses are the pathogens most often responsible for the common
cold, and are a frequent cause of exacerbations in asthma, chronic obstructive
pulmonary disease and cystic fibrosis. Here we report discovery of IMP-1088, a
picomolar dual inhibitor of the human N-myristoyltransferases
NMT1 and NMT2, and use it to demonstrate that pharmacological inhibition of host
cell N-myristoylation rapidly and completely prevents
rhinoviral replication without inducing cytotoxicity. Identification of
cooperative binding between weak-binding fragments led to rapid inhibitor
optimization through fragment reconstruction, structure-guided fragment linking,
and conformational control over linker geometry. We show that inhibition of
co-translational myristoylation of a specific virus-encoded protein (VP0) by
IMP-1088 potently blocks a key step in viral capsid assembly, delivering low
nanomolar antiviral activity against multiple rhinovirus strains, poliovirus and
foot-and-mouth disease virus, and protection of cells against virus-induced
killing, highlighting the potential of host myristoylation as a drug target in
picornaviral infections.
The promising drug target N-myristoyltransferase (NMT) catalyses an essential protein modification thought to occur exclusively at N-terminal glycines (Gly). Here, we present highresolution human NMT1 structures co-crystallised with reactive cognate lipid and peptide substrates, revealing high-resolution snapshots of the entire catalytic mechanism from the initial to final reaction states. Structural comparisons, together with biochemical analysis, provide unforeseen details about how NMT1 reaches a catalytically competent conformation in which the reactive groups are brought into close proximity to enable catalysis. We demonstrate that this mechanism further supports efficient and unprecedented myristoylation of an N-terminal lysine side chain, providing evidence that NMT acts both as N-terminallysine and glycine myristoyltransferase.
Cancer gene therapy requires the design of non-viral vectors that carry genetic material and selectively deliver it with minimal toxicity. Non-viral vectors based on cationic natural polymers can form electrostatic complexes with negatively-charged polynucleotides such as microRNAs (miRNAs). Here we investigated the physicochemical/biophysical properties of chitosan–hsa-miRNA-145 (CS–miRNA) nanocomplexes and the biological responses of MCF-7 breast cancer cells cultured in vitro. Self-assembled CS–miRNA nanocomplexes were produced with a range of (+/−) charge ratios (from 0.6 to 8) using chitosans with various degrees of acetylation and molecular weight. The Z-average particle diameter of the complexes was <200 nm. The surface charge increased with increasing amount of chitosan. We observed that chitosan induces the base-stacking of miRNA in a concentration dependent manner. Surface plasmon resonance spectroscopy shows that complexes formed by low degree of acetylation chitosans are highly stable, regardless of the molecular weight. We found no evidence that these complexes were cytotoxic towards MCF-7 cells. Furthermore, CS–miRNA nanocomplexes with degree of acetylation 12% and 29% were biologically active, showing successful downregulation of target mRNA expression in MCF-7 cells. Our data, therefore, shows that CS–miRNA complexes offer a promising non-viral platform for breast cancer gene therapy.
The Sonic Hedgehog
(Shh) signaling pathway plays a critical role
during embryonic development and cancer progression. N-terminal palmitoylation
of Shh by Hedgehog acyltransferase (Hhat) is essential for efficient
signaling, raising interest in Hhat as a novel drug target. A recently
identified series of dihydrothienopyridines has been proposed to function via this mode of action; however, the lead compound in this
series (RUSKI-43) was subsequently shown to possess cytotoxic activity
unrelated to canonical Shh signaling. To identify a selective chemical
probe for cellular studies, we profiled three RUSKI compounds in orthogonal
cell-based assays. We found that RUSKI-43 exhibits off-target cytotoxicity,
masking its effect on Hhat-dependent signaling, hence results obtained
with this compound in cells should be treated with caution. In contrast,
RUSKI-201 showed no off-target cytotoxicity, and quantitative whole-proteome
palmitoylation profiling with a bioorthogonal alkyne-palmitate reporter
demonstrated specific inhibition of Hhat in cells. RUSKI-201 is the
first selective Hhat chemical probe in cells and should be used in
future studies of Hhat catalytic function.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.