Synthesis and pharmacodynamics evaluation of cyclobutenamide derivatives against influenza a virus in vitro and in vivo.

Influenza A virus (IAV) constitutes a continuous threat to human health, and existing antiviral drugs face limitations such as drug resistance and secondary inflammatory damage. In order to identify safe, highly effective, and multi-mechanistic candidates against IAV, a series of cyclobutenamide derivatives were synthesized via the Tf2NH-catalyzed Ficini [2 + 2] cycloaddition of ynamides with enones. Biological evaluation identified compound 3 h as the most promising biological hit, which exhibited high inhibitory activity against IAV (EC50 = 2.01 ± 0.47 μM) and an excellent selectivity index (SI = 169.84). Mechanistic studies revealed that compound 3 h possesses a dual efficacy of antivirus and host protection: it not only effectively suppresses viral replication, but also significantly inhibits virus-induced apoptosis and the release of pro-inflammatory cytokines by modulating host immune signaling pathways, thereby mitigating cytokine storm and oxidative stress damage. In the IAV-infected mouse model, weight loss and pulmonary pathological damage were significantly alleviated by compound 3 h treatment, and viral load and inflammatory cytokine levels in lung tissues were effectively reduced. Furthermore, compound 3 h demonstrated good biological safety both in vitro and in vivo. These results suggest that the cyclobutenamide compound 3 h is a highly promising biological hit for the treatment of IAV infection.