Phosphatidylinositol 4-Kinase III Beta Is Essential for Replication of Human Rhinovirus and Its Inhibition Causes a Lethal Phenotype In Vivo

Abstract

Human rhinovirus (HRV) is the predominant cause of the common cold, but more importantly, infection may have serious repercussions in asthmatics and chronic obstructive pulmonary disorder (COPD) patients. A cell-based antiviral screen against HRV was performed with a subset of our proprietary compound collection, and an aminothiazole series with pan-HRV species and enteroviral activity was identified. The series was found to act at the level of replication in the HRV infectious cycle. In vitro selection and sequencing of aminothiazole series-resistant HRV variants revealed a single-nucleotide mutation leading to the amino acid change I42V in the essential HRV 3A protein. This same mutation has been previously implicated in resistance to enviroxime, a former clinical-stage antipicornavirus agent. Enviroxime-like compounds have recently been shown to target the lipid kinase phosphatidylinositol 4-kinase III beta (PI4KIII␤). A good correlation between PI4KIII␤ activity and HRV antiviral potency was found when analyzing the data over 80 compounds of the aminothiazole series, covering a 750-fold potency range. The mechanism of action through PI4KIII␤ inhibition was further demonstrated by small interfering RNA (siRNA) knockdown of PI4KB, which reduced HRV replication and also increased the potency of the PI4KIII␤ inhibitors. Inhibitors from two different structural classes with promising pharmacokinetic profiles and with very good selectivity for PI4KIII␤ were used to dissociate compound-related toxicity from target-related toxicity. Mortality was seen in all dosing groups of mice treated with either compound, therefore suggesting that short-term inhibition of PI4KIII␤ is deleterious.

Rhinovirus Uses a Phosphatidylinositol 4-Phosphate/Cholesterol Counter-Current for the Formation of Replication Compartments at the ER-Golgi Interface

“…The Lipid Kinase PI4K3b Is Required for HRV Replication HRV serotypes from species A and B require PI4Ks for infection, as suggested by studies with chemical inhibitors (Spickler et al, 2013;van der Schaar et al, 2013). To determine which PI4Ks were required for HRV infection and replication, we used a pool of small interfering RNAs (siRNA) directed against the known PI4Ks.…”

“…There are two classes of PI 4-kinases (class 2 and 3 PI4K). Class 3 PI4K has been implicated in replication of hepatitis C virus and picornaviruses (Hsu et al, 2010;Reiss et al, 2011;Spickler et al, 2013;van der Schaar et al, 2013). PI4K2a effectively functions on endosomes; PI4K2b is involved in the transmission of stress signals, and producing PI4,5P 2 at the plasma membrane and other membranes; and PI4K3a has been implicated in the formation of ER exit sites for cargo export from ER to Golgi (Balla, 2013).…”

GBF1- and ACBD3-Independent Recruitment of PI4KIIIβ to Replication Sites by Rhinovirus 3A Proteins

“…Recently, we and others found that replication of all enteroviruses, including HRV, is abolished in the presence of PI4KIII␤ inhibitors (2,16). Although this indicates that the kinase is also a crucial host factor for HRV, it is as yet unknown whether HRV also actively recruits PI4KIII␤ to replication sites and whether this is mediated via its 3A protein, since the absence of a 3A-GBF1 interaction for HRV could affect recruitment of downstream factor PI4KIII␤.…”

A Complex Comprising Phosphatidylinositol 4-Kinase IIIβ, ACBD3, and Aichi Virus Proteins Enhances Phosphatidylinositol 4-Phosphate Synthesis and Is Critical for Formation of the Viral Replication Complex

“…PI4P binds to viral RNA polymerase 3D to facilitate viral RNA synthesis. PI4KB activity is required for the replication of the genomes of other enteroviruses such as enterovirus 71 and human rhinovirus (20,21).…”