A multinational research team have developed a three-drug combination of orally available, safe-in-human antiviral agents that may effectively inhibit enterovirus replication in human cell and organoid cultures.

In the study, published in Cellular and Molecular Life Sciences, the researchers demonstrated that the combination of pleconaril, AG7404, and mindeudesivir worked synergistically to target different stages of the viral life cycle.

Enteroviruses, a subgroup of picornaviruses, can infect various human organs and cause diseases ranging from the common cold to more severe conditions such as meningitis, myocarditis, pancreatitis, hand-foot-and-mouth disease (HFMD), sepsis, poliomyelitis, and type 1 diabetes. Recent outbreaks of echovirus 11 (EV11), Enterovirus A71 (EVA71), and CVA13 have caused serious illnesses and deaths in Europe, Asia, and Africa, respectively.

"Currently, there are no approved treatments for enterovirus infections," noted Erlend Ravlo, of the of the Norwegian University of Science and Technology, and colleagues.

They tested the combination against a broad range of enteroviruses, including CVA13, CVB5, CVB6, EV1, EV6, EV7, and EV11. The testing was conducted in various human cell types including lung (A549 and HE), eye (RPE), cervical (HeLa), skeletal (RD), and pancreatic (MiaPaca-2 and β-cells) cells.

The combination demonstrated effectiveness without detectable cytotoxicity. "The cocktail protected human HeLa and RD cells against CVA13, CVB6, and EV7 for 72 h," noted researchers. This protection was particularly noteworthy as "the triple drug combination delayed the emergence of antiviral drug resistance, distinguishing it from monotherapy or two-drug cocktails."

Of particular significance was the triple combination's efficacy in human organoid models, which provided a more accurate simulation of viral diseases compared with traditional cell cultures. The drug cocktail protected human pancreatic, eye, lung, heart, and brain organoids from enterovirus-mediated cell death.

In pancreatic beta-cell experiments, the researchers observed that "the drug treatment did not affect cell viability, glucose, or insulin levels," suggesting the potential application for treating enterovirus-triggered type 1 diabetes. Additionally, the combination "preserved the contraction rhythm of infected heart organoids," indicating minimal impact on cardiac function.

Pleconaril targets the viral VP1 protein and prevents virus entry into host cells; AG7404 inhibits the viral 3C protease responsible for processing the viral polyprotein; and mindeudesivir attenuates replication and transcription of viral RNA, noted the researchers.

This multitarget approach contributed to the combination's synergistic effect, enabling lower doses of each component while maintaining efficacy and reducing the risk of drug resistance emerging.

The time-of-addition experiments demonstrated that the combination could be effective even when administered postinfection. "The combination of pleconaril, AG7404, and mindeudesivir can be added between 2 h before and 12 h after infection and still effectively inhibit viral replication," they noted.

All three components of this combination have previously completed clinical trials for other applications and established safety profiles in humans, potentially accelerating the pathway to clinical use against enterovirus infections.

The "combination consistently exhibits additive or synergistic effects across a range of enteroviruses in vitro, underscoring its potential as a broad-spectrum antiviral cocktail," and highlights the need for further animal studies and clinical trials, concluded researchers.

They reported no relevant financial or non-financial interests to disclose.