Hylin-a1: A Pan-Inhibitor against Emerging and Re-Emerging Respiratory Viruses

Abstract

Pandemic and epidemic outbreaks of respiratory viruses are a challenge for public health and social care system worldwide, leading to high mortality and morbidity among the human populations. In light of the limited efficacy of current vaccines and antiviral drugs against respiratory viral infections and the emergence and re-emergence of new viruses, novel broad-spectrum antiviral drugs are needed for the prevention and treatment of these infections. Antimicrobial peptides with an antiviral effect, also known as AVPs, have already been reported as potent inhibitors of viral infections by affecting different stages of the virus lifecycle. In the present study, we analyzed the activity of the AVP Hylin-a1, secreted by the frog Hypsiboas albopunctatus, against a wide range of respiratory viruses, including the coronaviruses HCoV-229E and SARS-CoV-2, measles virus, human parainfluenza virus type 3, and influenza virus H1N1. We report a significant inhibitory effect on infectivity in all the enveloped viruses, whereas there was a lack of activity against the naked coxsackievirus B3. Considering the enormous therapeutic potential of Hylin-a1, further experiments are required to elucidate its mechanism of action and to increase its stability by modifying the native sequence.

Keywords: SARS-CoV-2; antimicrobial peptide; coronavirus; emerging infection; influenza; inhibitory peptide; respiratory viruses.

Figure 1

Hylin-a1 cytotoxic activity. Cells were treated with peptide at several concentrations ranging from 0.39 to 100 μM. After 2 and 24 h of treatment, cell viability was obtained by the spectrophotometer analysis of the MTT assay. Cells without peptide were used as a negative control (CTRL −), whereas DMSO was used as a positive control (CTRL +). **** p < 0.0001; ns: non-significant.

Figure 2

Antiviral activity against HCoV. Different assays (co-treatment, cell pre-treatment, post-treatment, and virus pre-treatment) were conducted in order to test the anti-HCoV activity. (A) Co-treatment; (B) cell pre-treatment; (C) post-treatment; (D) virus pre-treatment. Infected cells without peptide were used as controls (CTRL −). **** p < 0.0001; *** p < 0.001; ns: non-significant.

Figure 3

Antiviral activity against paramyxoviruses. Different assays were used to investigate the anti-MeV and anti-HPIV-3 activities. (A) Co-treatment; (B) cell pre-treatment; (C) post-treatment; (D) virus pre-treatment. Infected cells without peptide were used as controls (CTRL −). **** p < 0.0001; ns: non-significant.

Figure 4

Antiviral activity against RSV. (A) Plaque assay. Co-treatment was performed and viral plaques observed after 48 h. Infected cells without peptide were used as controls (CTRL –). (B) RSV F protein ELISA. The binding of cells to the F protein was analyzed both in the absence and presence of Hylin-a1. **** p < 0.0001; *** p < 0.001; ns: non-significant.

Figure 5

Antiviral activity against influenza virus. Different assays were carried out to verify the anti-influenza virus activity. (A) Co-treatment; (B) cell pre-treatment; (C) post-treatment; (D) virus pre-treatment. Infected cells without peptide were used as controls (CTRL –). **** p < 0.0001; ns: non-significant.