[Monkey Pox] Rapid Research Response
Lead Research Organisation:
The Pirbright Institute
Department Name: Viral Immunology
Abstract
The project proposes a rapid response to the current monkeypox virus (MPXV) epidemic. It is led by the Pirbright Institute and the Centre for Virus Research - Glasgow - the two UKRI-funded institutes that lead on virus infections of animals and humans - and brings together relevant expertise from several other UK universities and institutions including the Universities of Cambridge, Oxford, Birmingham, Edinburgh and Surrey, Dstl, UKHSA, Guys and St Thomas NHS.
Between April and 18th July 2022, there have been 2137 confirmed cases of human monkeypox (MPX) in UK and the WHO has reported infections in all 5 WHO regions and 50 member states. The current epidemic is the largest ever known for MPXV. An urgent response to this growing epidemic is needed.
The consortium assembled proposes 6 inter-related work packages as follows.
1. Genomic characterisation of MPXV.
2. Examination of possible virus spillover from humans to UK animals
3. Study of the intrinsic and innate barriers to MPXV infection, and MPXV immune evasion strategies
4. Study of the immune response to MPXV infection and vaccination
5. Development of anti-viral drugs and monitoring for emergence of MPXV drug resistance
6. To develop point of care diagnostic tests for MPXV
WP 1. This will undertake sequencing of MPXV genomes isolated from humans in UK and monitor virus evolution and adaptation to humans. The sequencing will pay particular attention to the acquisition of genome mutations that might affect virus replication, transmission, virulence or drug resistance and links to WP5.
WP2. MPXV has a natural reservoir in rodents in parts of Africa and has a relatively broad host range that includes North American rodents, primates and humans. The widespread human infections provide a possible opportunity for human to animal transmission. This WP will evaluate this potential by examining the ability of MPXV to infect primary cells from a variety of UK animals.
WP3. This WP will evaluate the host response to infection by measuring the transcriptomic and proteomic responses to infection of human cells and testing the roles of specific host proteins in protecting against MPXV infection. Further, the ability of MPXV to counteract these defences will be tested building on what has been learnt from studies of related orthopoxviruses.
WP4. The immune response to MPXV infection of humans will be measured by determining the antibody and T cell responses. These will be compared with the responses to vaccination using the smallpox vaccine. A specific aim will be to identity signature T cell responses that are characteristic of MPXV infection. In addition to information vaccination programmes, the development of specific tests for immune monitoring will be undertaken.
WP5. This WP is concerned with the development of anti-MPXV drugs and builds on the development of CRUSH (COVID-19 Drug Screening and Resistance Hub) at CVR-Glasgow. Currently, 2 drugs are licensed for use against MPXV and these each target a specific virus protein, but mutation of these proteins can lead to drug resistance. The WP proposes to screen additional FDA-approved drugs that have activity against VACV for activity against MPXV. Cyclosporin A and non-immunosuppressive derivatives will be included since these target a proviral cellular protein, cyclophilin A, and therefore emergence of virus resistance is difficult.
WP 6. This will develop point of care (POC) diagnostic tests for MPXV. Currently, MPXV infection is confirmed by polymerase chain reaction (PCR), which is specific and sensitive but requires a specialist laboratory. A POC test (such as developed for SARS-CoV-2) would be of great benefit to speed diagnosis. Two approaches will be tried: a Lateral flow test (LAT) and a loop-mediated isothermal amplification (LAMP)-based assay.
Between April and 18th July 2022, there have been 2137 confirmed cases of human monkeypox (MPX) in UK and the WHO has reported infections in all 5 WHO regions and 50 member states. The current epidemic is the largest ever known for MPXV. An urgent response to this growing epidemic is needed.
The consortium assembled proposes 6 inter-related work packages as follows.
1. Genomic characterisation of MPXV.
2. Examination of possible virus spillover from humans to UK animals
3. Study of the intrinsic and innate barriers to MPXV infection, and MPXV immune evasion strategies
4. Study of the immune response to MPXV infection and vaccination
5. Development of anti-viral drugs and monitoring for emergence of MPXV drug resistance
6. To develop point of care diagnostic tests for MPXV
WP 1. This will undertake sequencing of MPXV genomes isolated from humans in UK and monitor virus evolution and adaptation to humans. The sequencing will pay particular attention to the acquisition of genome mutations that might affect virus replication, transmission, virulence or drug resistance and links to WP5.
WP2. MPXV has a natural reservoir in rodents in parts of Africa and has a relatively broad host range that includes North American rodents, primates and humans. The widespread human infections provide a possible opportunity for human to animal transmission. This WP will evaluate this potential by examining the ability of MPXV to infect primary cells from a variety of UK animals.
WP3. This WP will evaluate the host response to infection by measuring the transcriptomic and proteomic responses to infection of human cells and testing the roles of specific host proteins in protecting against MPXV infection. Further, the ability of MPXV to counteract these defences will be tested building on what has been learnt from studies of related orthopoxviruses.
WP4. The immune response to MPXV infection of humans will be measured by determining the antibody and T cell responses. These will be compared with the responses to vaccination using the smallpox vaccine. A specific aim will be to identity signature T cell responses that are characteristic of MPXV infection. In addition to information vaccination programmes, the development of specific tests for immune monitoring will be undertaken.
WP5. This WP is concerned with the development of anti-MPXV drugs and builds on the development of CRUSH (COVID-19 Drug Screening and Resistance Hub) at CVR-Glasgow. Currently, 2 drugs are licensed for use against MPXV and these each target a specific virus protein, but mutation of these proteins can lead to drug resistance. The WP proposes to screen additional FDA-approved drugs that have activity against VACV for activity against MPXV. Cyclosporin A and non-immunosuppressive derivatives will be included since these target a proviral cellular protein, cyclophilin A, and therefore emergence of virus resistance is difficult.
WP 6. This will develop point of care (POC) diagnostic tests for MPXV. Currently, MPXV infection is confirmed by polymerase chain reaction (PCR), which is specific and sensitive but requires a specialist laboratory. A POC test (such as developed for SARS-CoV-2) would be of great benefit to speed diagnosis. Two approaches will be tried: a Lateral flow test (LAT) and a loop-mediated isothermal amplification (LAMP)-based assay.
Technical Summary
The 200 kb MPXV genome is a challenging sequencing target due to repeated sequences near each genomic terminus and low GC (33%) content. A fast, reliable and low-cost method will be developed that can accurately reconstruct the genome for evolutionary analysis, genomic epidemiology and functional character-isation.
A panel of cell lines from a range of rodent and livestock species available at Pirbright will be used to examine the host range of MPXV. Virus growth in cell culture will be measured using classical methods and live-cell analysis systems.
We will determine MPXV evolution of tropism and hIFN-I antagonism, to predict virus human-to-human transmission.
We will evaluate the long-term immunity induced by MPXV infection in humans and define differences between vaccine-induced responses: immunodominance pattern, functional immunophenotype and if T cell responses to MPXV viral immediate early proteins exhibit better viral control than other specificities. Also, associations with antibody responses and clinical outcome will be evaluated.
We will establish and optimise antiviral screening pipelines to assess the antiviral efficacy of tecovirimat and brincidofovir, drugs with known activity against orthopoxviruses. We assess 26 FDA-approved drugs and a novel cyclosporine A derivative that inhibit VACV in vitro. Also, current MPXV isolates will be tested in cell culture for the rapid testing of circulating strains of MPXV for tecovirimat-resistance.
We will develop two rapid point of care diagnostic tests. Lateral flow Assay (LFA). An orthopoxvirus (OPV) LFA has been developed at Dstl, comprising 4 Mabs all reactive with old world OPVs. Making use of novel bead types we will provide data packs to enable rapid progress through regulatory pathways for licensure.
Loop-mediated isothermal amplification (LAMP)-based assay. LAMP is a rapidly maturing technology providing PCR levels of sensitivity and specificity without the need for thermal cycling.
A panel of cell lines from a range of rodent and livestock species available at Pirbright will be used to examine the host range of MPXV. Virus growth in cell culture will be measured using classical methods and live-cell analysis systems.
We will determine MPXV evolution of tropism and hIFN-I antagonism, to predict virus human-to-human transmission.
We will evaluate the long-term immunity induced by MPXV infection in humans and define differences between vaccine-induced responses: immunodominance pattern, functional immunophenotype and if T cell responses to MPXV viral immediate early proteins exhibit better viral control than other specificities. Also, associations with antibody responses and clinical outcome will be evaluated.
We will establish and optimise antiviral screening pipelines to assess the antiviral efficacy of tecovirimat and brincidofovir, drugs with known activity against orthopoxviruses. We assess 26 FDA-approved drugs and a novel cyclosporine A derivative that inhibit VACV in vitro. Also, current MPXV isolates will be tested in cell culture for the rapid testing of circulating strains of MPXV for tecovirimat-resistance.
We will develop two rapid point of care diagnostic tests. Lateral flow Assay (LFA). An orthopoxvirus (OPV) LFA has been developed at Dstl, comprising 4 Mabs all reactive with old world OPVs. Making use of novel bead types we will provide data packs to enable rapid progress through regulatory pathways for licensure.
Loop-mediated isothermal amplification (LAMP)-based assay. LAMP is a rapidly maturing technology providing PCR levels of sensitivity and specificity without the need for thermal cycling.
Organisations
Publications
Chen Y
(2023)
Perspective on the application of genome sequencing for monkeypox virus surveillance.
in Virologica Sinica
Albarnaz J
(2023)
Quantitative proteomics defines mechanisms of antiviral defence and cell death during modified vaccinia Ankara infection
in Nature Communications
Wellington D
(2022)
T cells are ready for the fight against monkeypox.
in Cell host & microbe
Zhao Y
(2023)
TRIM5a restricts poxviruses and is antagonized by CypA and the viral protein C6.
in Nature
| Description | Although the number of mpox infection cases in non-endemic countries is decreasing, it is still crucial to establish robust and accurate genomic workflows to support future outbreak responses. Such workflows are essential to ensure a prompt and effective response. Moreover, the analysis of well-characterized cohorts of patients we are conducting can provide valuable insights into various aspects of the disease, such as the estimation of viral co-infections, determining if disease severity is affected by viral genomic markers, and understanding intra-host evolution, and transmission. Such insights can significantly accelerate our ability to respond rapidly to future outbreaks and take necessary measures to prevent their spread. The work has generated exciting data. Specifically, we have shown that protein C6 from vaccinia virus (VACV) and closely related orthologues from monkeypox virus (MPXV) and variola virus (VARV) bind to and induce degradation of cellular proteins TRIM5a, HDAC4 (2) and HDAC5. The consequence of this targetted degradation is that the restriction imposed by these host factors on these viruses is negated. As a second defense against TRIM5a, these viruses also recruit cyclophilin A to the virus capsid and the proviral activity of CypA is reversed by cyclosporine A and non-immunosuppressive derivatives. These drugs were shown to be anti-viral for orthopoxviruses including MPXV and given that they target a cellular protein, the emergence of drug resistant virus, as has already happened with tecovirimat, is unlikely: clinical testing against MPXV is warranted. |
| Exploitation Route | crucial to establish robust and accurate genomic workflows to support future outbreak responses. Such workflows are essential to ensure a prompt and effective response. Moreover, the analysis of well-characterized cohorts of patients we are conducting can provide valuable insights into various aspects of the disease, such as the estimation of viral co-infections, determining if disease severity is affected by viral genomic markers, and understanding intra-host evolution, and transmission. Such insights can significantly accelerate our ability to respond rapidly to future outbreaks and take necessary measures to prevent their spread. Collectively, these studies will lead to a better understanding of host restriction factors that inhibit MPXV replication and whether these factors and virus antagonism of them influences the host range of MPXV. As exemplified by the detailed study of TRIM5a, the increased understanding of host restriction and virus countermeasures can lead to the identification of anti-viral drugs for MPXV and other poxviruses. |
| Sectors | Healthcare |
| Description | We are facilitating technology transfer in various ways. Firstly, we provided a comprehensive one-week training to the coordinator of the clinical virology laboratory at the College of Medicine, University of Ibadan, Nigeria, aimed at supporting local mpox sequencing. Additionally, we are planning a similar training event with a virology clinical fellow from Guy's and St Thomas Trust (UK). Lastly, we will soon share our wet lab and bioinformatic pipeline optimizations with colleagues from Madrid and Barcelona, which will enable them to improve their processes. While our achievements have not been published yet, we have shared them during collaborative meetings to further our collective goals. |
| First Year Of Impact | 2022 |
| Sector | Healthcare |
| Impact Types | Policy & public services |