Poster Presentation 12th Australasian Virology Society Meeting 2024

Targeted protein degraders of SARS-CoV-2 main protease demonstrate superior antiviral activity over enzymatic inhibition alone (#261)

Baolong Pan 1 , Philip E Thompson 1 , Simon J Mountford 1 , Danielle E Anderson 2 , Kate E Jarman 3 , Danielle R Tilmanis 3 , Belinda Maher 4 , Jake Shortt 4 , Sam A Greenall 4 , Nadia Warner 2
  1. Medicinal Chemistry, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia
  2. VIDRL at The Doherty Institute, Melbourne, VIC, Australia
  3. Walter and Eliza Hall Institute, Parkville, VIC, Australia
  4. Department of Medicine, Monash University, Clayton, VIC, Australia

Originally developed as oncology therapeutics, targeted protein degraders (TPDs) (also known as proteolysis targeting chimeras or PROTACs), are bifunctional small-molecule drugs composed of two active ligands which simultaneously engage both a target protein and a host cell ubiquitin ligase recruiter, triggering target protein ubiquitination and degradation via the proteasome.

In contrast to conventional inhibition-based drugs, TPDs do not require high affinity binding for their effect, and have been demonstrated to maintain efficacy against variant protein, including chemotherapy-resistant forms.

We hypothesise that this unique feature could provide a new avenue to more effective, broader acting antiviral drugs with negligible selection of drug resistance.

The aim of this work was to establish a high-efficiency synthesis approach to generate TPDs from established viral protein inhibitors, and to determine whether TPDs possess superior antiviral potency using SARS-CoV-2 as a viral model.

Using a novel bead-based chemical synthesis approach, we adapted the clinically approved SARS-CoV-2 Mpro inhibitor nirmatrelvir into a panel of TPDs and found three effective compounds utilising VHL or IAP ubiquitin ligase ligands. Precisely matched non-degrading controls were also synthesised.

A Dox-inducible HiBit-tagged Mpro nanoluciferase reporter cell line was generated and used to confirm these TPDs could degrade wt and nirmatrelvir-resistant Mpro protein in a concentration, time and host proteasomal dependent fashion whereas non-degrading controls and nirmatrelvir could not.

Using SARS-CoV-2 infection of Calu3 and Vero/TMPRSS/2 cells we confirmed that the TPDs had higher antiviral potency than their non degrading controls.

This work demonstrates the feasibility of generating TPDs from established viral protein inhibitors, and confirms TPDs possess superior antiviral potency and maintained efficacy against drug-resistant virus, overcoming a major limitation of conventional direct antiviral approaches.

The synthesis, screening and antiviral validation approach developed here can be expanded to rapidly develop antiviral TPDs to any virus once a binding ligand has been identified.