Development of a virus-free sensor system to repurpose approved drugs for blocking Coronavirus replication
Lead Research Organisation:
University of Cambridge
Department Name: Pathology
Abstract
The COVID19 pandemic has highlighted the importance of timely discovery of antiviral therapeutics, especially when there is little information as to whether vaccines developed (and pending approval) enable a significant period of immunity. Therefore the development of cost effective, high-throughput sensor systems to rapidly repurpose approved drugs as antiviral agents is extremely desirable. We have developed a virus-free sensor system - pShiftSensor that can specifically screen for drugs to block SARS-CoV2 replication by inhibiting the ribosomal frameshift that is essential for production of the viral replicase. The sensor system is fluorescent-based, therefore permitting cost-effective high-throughput quantitate readout.
The system is based on the cis-element required for -1 programmed ribosomal frameshifting (-1 PRF), a non-canonical translation mechanism that is ubiquitous throughout all coronaviruses and used to synthesis viral proteins including the RNA-dependent RNA polymerase (RdRP), essential for viral replication. We have developed pShiftSensor-v1, which is a dual-reporter system that contains the SARS-CoV-2 -1PRF cassette (i.e. the slippery sequence UUUAAAC and the 3' pseudoknot). The system is bi-cistronic thereby avoiding expression artifacts due to differential delivery between the control and test reporter expression cassette. Through this proposal, we will upgrade our system to pShiftSensor-V2, which targets primary cell types such as small airway lung cells, the key entry cell type for SARS-CoV2 infection, and primary leukocytes, especially monocytes/macrophages and T-cells, essential for systemic infection. pShiftSensor-V2 will permit cost-effective, high-throughput cell-type specific screening of viral replication inhibitory compounds in a physiological environment.
The system is based on the cis-element required for -1 programmed ribosomal frameshifting (-1 PRF), a non-canonical translation mechanism that is ubiquitous throughout all coronaviruses and used to synthesis viral proteins including the RNA-dependent RNA polymerase (RdRP), essential for viral replication. We have developed pShiftSensor-v1, which is a dual-reporter system that contains the SARS-CoV-2 -1PRF cassette (i.e. the slippery sequence UUUAAAC and the 3' pseudoknot). The system is bi-cistronic thereby avoiding expression artifacts due to differential delivery between the control and test reporter expression cassette. Through this proposal, we will upgrade our system to pShiftSensor-V2, which targets primary cell types such as small airway lung cells, the key entry cell type for SARS-CoV2 infection, and primary leukocytes, especially monocytes/macrophages and T-cells, essential for systemic infection. pShiftSensor-V2 will permit cost-effective, high-throughput cell-type specific screening of viral replication inhibitory compounds in a physiological environment.
Organisations
People |
ORCID iD |
| Betty Chung (Principal Investigator) |
Publications
Albarnaz JD
(2022)
Molecular mimicry of NF-?B by vaccinia virus protein enables selective inhibition of antiviral responses.
in Nature microbiology
Thomas SE
(2022)
RNA structure mediated thermoregulation: What can we learn from plants?
in Frontiers in plant science
Zhang H
(2023)
Editorial: Plant RNA structure
in Frontiers in Plant Science
| Description | This study aims to develop a sensor system that enables large scale screening of compounds that can inhibit coronavirus replication in the context of primary cell, particular immune cells which are key cell types for viral dissemination. We have had significant setback due to unavailability of key reagents essential for the development of the sensor system, mostly due to the pandemic, and brexit. Notably, several key components are also utilised for mRNA-based vaccines and is now unavailable to purchase. Neverthless, we have re-established the system with alternative reagents and was able to reveal that the importance of targeting primary macrophages when identifying anti-viral compounds as have revealed that the behaviour of the sensor system we developed is different between cell lines (typically used for screening antiviral compounds), and primary cells but also between different primary cell types. |
| Exploitation Route | The final product could be utilised by a pharmaceutical company to screen for biologically relevant antiviral compounds (ie. uptake by primary cells of interest, as not all drugs are active in all cell types), not only for COVID but also HIV, alphavirus and japanese encephalitis virus. |
| Sectors | Chemicals,Healthcare,Pharmaceuticals and Medical Biotechnology |