Optimization of 4-Aminopiperidines as Inhibitors of Influenza A Viral Entry That Are Synergistic with Oseltamivir

Abstract

Vaccination is the most prevalent prophylactic means for controlling seasonal influenza infections. However, an effective vaccine usually takes at least 6 months to develop for the circulating strains. Therefore, new therapeutic options are needed for the acute treatment of influenza infections to control this virus and prevent epidemics/pandemics from developing. We have discovered fast-acting, orally bioavailable acylated 4-aminopiperidines with an effective mechanism of action targeting viral hemagglutinin (HA). Our data show that these compounds are potent entry inhibitors of influenza A viruses. We present docking studies that suggest an HA binding site for these inhibitors on H5N1. Compound 16 displayed a significant decrease of viral titer when evaluated in the infectious assays with influenza virus H1N1 (A/Puerto Rico/8/1934) or H5N1 (A/Vietnam/1203/2004) strains and the oseltamivir-resistant strain with the most common H274Y mutation. In addition, compound 16 showed significant synergistic activity with oseltamivir in vitro.

Structure of avian influenza hemagglutinin in complex with a small molecule entry inhibitor

“…Specific types of intermolecular interactions include hydrophobic (residues HA1-H38, HA1-Q40, HA2-W21, and HA2-I45) and polar interactions between the chlorine atoms of CBS1117 and side chains of residues HA1-T325 and HA2-T49. The importance of the polar interaction between the benzyl 2-chlorine and the HA2-T49 side chain hydroxyl is in agreement with the SAR analysis of this series of compounds, which suggested that halogen substitutions to the benzyl ring were essential for compound activity (Gaisina et al, 2020;Hussein et al, 2020). Finally, we note that characterization of the H5 HA-CBS1117 interaction by WaterLOGSY NMR confirmed compound binding to H5 HA ( Fig S2A).…”

“…CBS1117 has greater potency against group 1 HA strains, including circulating H1 HA and avian H5 HA (Gaisina et al, 2020;Hussein et al, 2020). Our laboratories and others have noted that HA fusion inhibitors generally exhibit group specificity.…”

“…Our laboratories and others have noted that HA fusion inhibitors generally exhibit group specificity. Examples include group 1 fusion inhibitors MBX2329 (EC 50~0 .5 μM against H1 HA versus EC 50 > 100 μM against H3 HA [Basu et al, 2014]), MBX2546 (EC 50~0 .3 μM against H1 HA versus EC 50 > 100 μM against H3 HA [Basu et al, 2014]), and CBS1117 (EC 50~3 μM against H5 HA versus EC 50 > 50 μM against H3 HA [Gaisina et al, 2020]). In contrast, Arbidol is an interesting exception in that it targets a wide range of enveloped viruses including both group 1 and 2 HA of influenza (i.e., it is a promiscuous binder), albeit at relatively low potency against influenza (EC 50 > 8 μM [Brancato et al, 2013;Kadam & Wilson, 2017]).…”

“…For example, the piperidine/piperazine and chlorobenzyl/benzyl rings of the two compounds ( Fig 6A) show significant overlap in the structure alignment of the complexes (Fig 6B), with similar protein interactions and modes of actions. However, in the case of CBS1117, the linker between the piperidine and benzyl rings and the The activity of the compounds has been previously described (Gaisina et al, 2020). To generate this figure, the compounds 16 and 26 were manually aligned with CBS1117.…”

“…Recently, our laboratories have described the discovery of a HA fusion inhibitor compound with a 4-aminopiperidine scaffold from an HTS screen of~20,000 compounds (Hussein et al, 2020). The best hit, termed CBS1117, exhibited EC 50 = 3.0 μM and low toxicity (CC 50 > 100 μM) in the pseudotype virus assay in A549 cells infected with influenza HA from H5N1 (Gaisina et al, 2020;Hussein et al, 2020). In this work, we characterize the binding of CBS1117 to avian H5 HA by x-ray crystallography, NMR, and mutagenesis and discuss new insights into the compound's mechanism of action and group specific activity.…”