Preclinical Screening Platform Identifies Azatadine‐Dimaleate as a Potent Repurposed Therapeutic Against SARS‐CoV‐2 Infection

The emergence of SARS-CoV-2 posed a major global public health threat, necessitating urgent development of therapeutics. Despite vaccine availability, continuous emergence of viral variants with enhanced transmissibility and immune escape capabilities, and consequential impacts on health services, requires effective antiviral therapeutics. Drug repurposing offers an expeditious strategy to identify therapeutics with established safety profiles. We implemented a comprehensive three-tiered validation approach, screening 2,570 compounds against SARS-CoV-2 in vitro, followed by ex vivo validation in well-differentiated primary human bronchial epithelial cell (WD-PBEC) cultures, and rigorous in vivo assessment. This methodical progression identified Azatadine-Dimaleate, a H1-receptor antagonist, as an exceptional candidate with consistent efficacy across all systems. Azatadine-Dimaleate demonstrated potent antiviral activity- EC50: 4.0 µM (95% CI: 3.2-4.8 µM), reducing viral replication by ~5,000-fold at 25 µM in epithelial cultures and lowering peak viral titers in WD-PBECs by 1.4 log₁₀, and 2.33 log₁₀ at 48 and 96 hpi, respectively, compared to controls. There was also a concomitant reduction in expression of interferons and pro-inflammatory genes, including IL-6. Combination with Remdesivir synergistically enhanced antiviral activity, reducing the EC50 of both drugs by > 60%. In the K18-hACE2 transgenic mouse model, Azatadine-Dimaleate significantly reduced weight loss (4% vs. 12%, p ≤ 0.05), decreased viral loads, and halved viral antigen expression in lung tissues. Unlike many candidates that faltered in complex models, Azatadine-Dimaleate maintained efficacy across all platforms. These findings support its clinical evaluation, alone or in combination with Remdesivir, as a versatile therapeutic with strong potential to address current and emerging SARS-CoV-2 variants.