Cetylpyridinium chloride (CPC) reduces zebrafish mortality from influenza infection: Super-resolution microscopy reveals CPC interference with multiple protein interactions with phosphatidylinositol 4,5-bisphosphate in immune function

Abstract

The COVID-19 pandemic raises significance for a potential influenza therapeutic compound, cetylpyridinium chloride (CPC), which has been extensively used in personal care products as a positively-charged quaternary ammonium antibacterial agent. CPC is currently in clinical trials to assess its effects on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) morbidity. Two published studies have provided mouse and human data indicating that CPC may alleviate influenza infection, and here we show that CPC (0.1 μM, 1 h) reduces zebrafish mortality and viral load following influenza infection. However, CPC mechanisms of action upon viral-host cell interaction are currently unknown. We have utilized super-resolution fluorescence photoactivation localization microscopy to probe the mode of CPC action. Reduction in density of influenza viral protein hemagglutinin (HA) clusters is known to reduce influenza infectivity: here, we show that CPC (at non-cytotoxic doses, 5–10 μM) reduces HA density and number of HA molecules per cluster within the plasma membrane of NIH-3 T3 mouse fibroblasts. HA is known to colocalize with the negatively-charged mammalian lipid phosphatidylinositol 4,5-bisphosphate (PIP 2 ); here, we show that nanoscale co-localization of HA with the PIP 2 -binding Pleckstrin homology (PH) reporter in the plasma membrane is diminished by CPC. CPC also dramatically displaces the PIP 2 -binding protein myristoylated alanine-rich C-kinase substrate (MARCKS) from the plasma membrane of rat RBL-2H3 mast cells; this disruption of PIP 2 is correlated with inhibition of mast cell degranulation. Together, these findings offer a PIP 2 -focused mechanism underlying CPC disruption of influenza and suggest potential pharmacological use of this drug as an influenza therapeutic to reduce global deaths from viral disease.

Cholesterol Regulation of Membrane Proteins Revealed by Two-Color Super-Resolution Imaging

“…Using proteins allows for genetically encoding the fluorescent label needed for super-resolution [ 94 ]. Genetically encoded PIP 2 sensors are available for PALM imaging [ 43 , 95 ]. PIP 2 is typically measured on the inner leaflet of the plasma membrane, which is not accessible in live cells.…”

The Development and Application of Opto-Chemical Tools in the Zebrafish

“…Many photoactivatable florescent proteins and synthetic dyes have been widely used for super-resolution imaging including Dendra2, mEos2, Dronpa, and rhodamine-derived dyes [ 246 , 247 ]. Super-resolution imaging using these photoactivable fluorophores has also been applied in various zebrafish studies [ 248 , 249 , 250 , 251 ].…”

Virucidal efficacy of cetylpyridinium chloride and benzydamine hydrochloride fixed combination lozenges against influenza A virus: A review

“…CPC также обладает противовирусным действием [8,33,40,41]. Доказана эффективность CPC против респираторных вирусных патогенов.…”

“…CPC снижал смертность рыбок данио от гриппозной инфекции [40]. При этом снижение смертности рыбок от гриппа коррелировало со снижением вирусной нагрузки.…”