The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells
The only currently available approach to early efficacy testing of tuberculosis (TB) vaccine candidates is in vivo preclinical challenge models. These typically include mice, guinea pigs and non-human primates (NHPs), which must be exposed to virulent M.tb in a ‘challenge’ experiment following vacci...
Main Authors: | , , , , , , , , , , , , , , , |
---|---|
Format: | Journal article |
Language: | English |
Published: |
F1000Research
2021
|
_version_ | 1797087819747819520 |
---|---|
author | Tanner, R Hoogkamer, E Bitencourt, J White, A Boot, C Sombroek, CC Harris, SA O'Shea, MK Wright, D Wittenberg, R Sarfas, C Satti, I Verreck, FAW Sharpe, SA Fletcher, HA McShane, H |
author_facet | Tanner, R Hoogkamer, E Bitencourt, J White, A Boot, C Sombroek, CC Harris, SA O'Shea, MK Wright, D Wittenberg, R Sarfas, C Satti, I Verreck, FAW Sharpe, SA Fletcher, HA McShane, H |
author_sort | Tanner, R |
collection | OXFORD |
description | The only currently available approach to early efficacy testing of tuberculosis (TB) vaccine candidates is in vivo preclinical challenge models. These typically include mice, guinea pigs and non-human primates (NHPs), which must be exposed to virulent M.tb in a ‘challenge’ experiment following vaccination in order to evaluate protective efficacy. This procedure results in disease development and is classified as ‘Moderate’ in severity under EU legislation and UK ASPA licensure. Furthermore, experiments are relatively long and animals must be maintained in high containment level facilities, making them relatively costly. We describe an in vitro protocol for the direct mycobacterial growth inhibition assay (MGIA) for use in the macaque model of TB vaccine development with the aim of overcoming some of these limitations. Importantly, using an in vitro assay in place of in vivo M.tb challenge represents a significant refinement to the existing procedure for early vaccine efficacy testing. Peripheral blood mononuclear cell and autologous serum samples collected from vaccinated and unvaccinated control animals are co-cultured with mycobacteria in a 48-well plate format for 96 hours. Adherent monocytes are then lysed to release intracellular mycobacteria which is quantified using the BACTEC MGIT system and colony-forming units determined relative to an inoculum control and stock standard curve. We discuss related optimisation and characterisation experiments, and review evidence that the direct NHP MGIA provides a biologically relevant model of vaccine-induced protection. The potential end-users of the NHP MGIA are academic and industry organisations that conduct the assessment of TB vaccine candidates and associated protective immunity using the NHP model. This approach aims to provide a method for high-throughput down-selection of vaccine candidates going forward to in vivo efficacy testing, thus expediting the development of a more efficacious TB vaccine and offering potential refinement and reduction to the use of NHPs for this purpose. |
first_indexed | 2024-03-07T02:41:06Z |
format | Journal article |
id | oxford-uuid:aa7b0714-d38c-41ff-8d03-4151a50f3d20 |
institution | University of Oxford |
language | English |
last_indexed | 2024-03-07T02:41:06Z |
publishDate | 2021 |
publisher | F1000Research |
record_format | dspace |
spelling | oxford-uuid:aa7b0714-d38c-41ff-8d03-4151a50f3d202022-03-27T03:15:30ZThe in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cellsJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:aa7b0714-d38c-41ff-8d03-4151a50f3d20EnglishSymplectic ElementsF1000Research2021Tanner, RHoogkamer, EBitencourt, JWhite, ABoot, CSombroek, CCHarris, SAO'Shea, MKWright, DWittenberg, RSarfas, CSatti, IVerreck, FAWSharpe, SAFletcher, HAMcShane, HThe only currently available approach to early efficacy testing of tuberculosis (TB) vaccine candidates is in vivo preclinical challenge models. These typically include mice, guinea pigs and non-human primates (NHPs), which must be exposed to virulent M.tb in a ‘challenge’ experiment following vaccination in order to evaluate protective efficacy. This procedure results in disease development and is classified as ‘Moderate’ in severity under EU legislation and UK ASPA licensure. Furthermore, experiments are relatively long and animals must be maintained in high containment level facilities, making them relatively costly. We describe an in vitro protocol for the direct mycobacterial growth inhibition assay (MGIA) for use in the macaque model of TB vaccine development with the aim of overcoming some of these limitations. Importantly, using an in vitro assay in place of in vivo M.tb challenge represents a significant refinement to the existing procedure for early vaccine efficacy testing. Peripheral blood mononuclear cell and autologous serum samples collected from vaccinated and unvaccinated control animals are co-cultured with mycobacteria in a 48-well plate format for 96 hours. Adherent monocytes are then lysed to release intracellular mycobacteria which is quantified using the BACTEC MGIT system and colony-forming units determined relative to an inoculum control and stock standard curve. We discuss related optimisation and characterisation experiments, and review evidence that the direct NHP MGIA provides a biologically relevant model of vaccine-induced protection. The potential end-users of the NHP MGIA are academic and industry organisations that conduct the assessment of TB vaccine candidates and associated protective immunity using the NHP model. This approach aims to provide a method for high-throughput down-selection of vaccine candidates going forward to in vivo efficacy testing, thus expediting the development of a more efficacious TB vaccine and offering potential refinement and reduction to the use of NHPs for this purpose. |
spellingShingle | Tanner, R Hoogkamer, E Bitencourt, J White, A Boot, C Sombroek, CC Harris, SA O'Shea, MK Wright, D Wittenberg, R Sarfas, C Satti, I Verreck, FAW Sharpe, SA Fletcher, HA McShane, H The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells |
title | The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells |
title_full | The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells |
title_fullStr | The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells |
title_full_unstemmed | The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells |
title_short | The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells |
title_sort | in vitro direct mycobacterial growth inhibition assay mgia for the early evaluation of tb vaccine candidates and assessment of protective immunity a protocol for non human primate cells |
work_keys_str_mv | AT tannerr theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT hoogkamere theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT bitencourtj theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT whitea theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT bootc theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sombroekcc theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT harrissa theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT osheamk theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT wrightd theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT wittenbergr theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sarfasc theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sattii theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT verreckfaw theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sharpesa theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT fletcherha theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT mcshaneh theinvitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT tannerr invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT hoogkamere invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT bitencourtj invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT whitea invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT bootc invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sombroekcc invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT harrissa invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT osheamk invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT wrightd invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT wittenbergr invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sarfasc invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sattii invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT verreckfaw invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT sharpesa invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT fletcherha invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells AT mcshaneh invitrodirectmycobacterialgrowthinhibitionassaymgiafortheearlyevaluationoftbvaccinecandidatesandassessmentofprotectiveimmunityaprotocolfornonhumanprimatecells |