Abstract
Purpose :
Zika virus (ZIKV) is an emerging RNA virus that not only causes neurological but also ocular abnormalities in infants. In infected cells, ZIKV exploits autophagic processes to enhance its replication and spread. Thus, a range of autophagy inhibitors have emerged as promising strategies to combat RNA virus infections, with hydroxychloroquine (HCQ) being one of the most promising. In this study, we synthesized and assessed the antiviral activity of HCQ analogs and determined the underlying mechanisms.
Methods :
Human retinal pigment epithelial cells (ARPE-19), primary human trabecular meshwork cells (Pr. HTMC), and Vero cells were used in the study. The cytotoxic effects of the synthesized drugs were screened using MTT and LDH assays. The induction of autophagy was confirmed using immunofluorescence to detect LC3B puncta and western blot for the autophagy markers – LC3B-II and p62/SQSTM1. The viral replication was measured using a western blot to detect ZIKV protein NS3, immunofluorescence to detect envelope protein, and plaque assay to quantify progeny virions. The antiviral innate immune response and inflammatory genes were assessed by real-time PCR.
Results :
Among the ten synthesized compounds, two novel drug candidates, labeled GL-287 and GL-382, displayed potent antiviral effects against ZIKV infection in human ocular cells, primarily by inhibiting autophagy. The two analogs surpassed the antiviral efficacy of HCQ and other promising autophagy inhibitor drugs like ROC-325, GNS561, and DC661. Moreover, unlike HCQ, these drugs did not exhibit cytotoxicity in the ocular cells. Treatment with compounds 287 and 382 in ZIKV-infected cells increased the abundance of LC3 puncta, indicating an accumulation of autophagic vacuoles, which suggests a disruption of the autophagy process. Furthermore, compounds 287 and 382 effectively inhibited the ZIKV-induced antiviral inflammatory response (IL1b, IL-6, and TNFa) in ocular cells.
Conclusions :
Collectively, our study demonstrates the antiviral potential of novel compounds GL-287 and GL-382 against ZIKV via inhibiting autophagy.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.