Tools for Assessing the Protective Efficacy of TB Vaccines in Humans: in vitro Mycobacterial Growth Inhibition Predicts Outcome of in vivo Mycobacterial Infection

Tanner, Rachel; Satti, Iman; Harris, Stephanie A.; O’Shea, Matthew K.; Cizmeci, Deniz; O’Connor, Daniel; Chomka, Agnieszka; Matsumiya, Magali; Wittenberg, Rachel; Minassian, Angela M.; Meyer, Joel N.; Fletcher, Helen A.; McShane, Helen

doi:10.3389/fimmu.2019.02983

Cited by 28 publications

(47 citation statements)

References 68 publications

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“…Additional work should aim to further improve assay sensitivity, evaluate performance in trials of novel TB vaccine candidates and interrogate immune mechanisms of control. In the human direct MGIA, mycobacterial growth inhibition has been variously associated with ratio of monocytes to lymphocytes 27 , 28 , 53 , polyfunctional CD4 + T-cells 54 , B-cell and IgG1 responses 28 , cytokines associated with trained immunity 27 and distinct transcriptomic profiles 55 ; similar mechanisms may contribute in the macaque 56 .…”

Section: Discussionmentioning

confidence: 99%

A non-human primate in vitro functional assay for the early evaluation of TB vaccine candidates

et al. 2021

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We present a non-human primate mycobacterial growth inhibition assay (MGIA) using in vitro blood or cell co-culture with the aim of refining and expediting early tuberculosis vaccine testing. We have taken steps to optimise the assay using cryopreserved peripheral blood mononuclear cells, transfer it to end-user institutes, and assess technical and biological validity. Increasing cell concentration or mycobacterial input and co-culturing in static 48-well plates compared with rotating tubes improved intra-assay repeatability and sensitivity. Standardisation and harmonisation efforts resulted in high consistency agreements, with repeatability and intermediate precision <10% coefficient of variation (CV) and inter-site reproducibility <20% CV; although some systematic differences were observed. As proof-of-concept, we demonstrated ability to detect a BCG vaccine-induced improvement in growth inhibition in macaque samples, and a correlation between MGIA outcome and measures of protection from in vivo disease development following challenge with either intradermal BCG or aerosol/endobronchial Mycobacterium tuberculosis (M.tb) at a group and individual animal level.

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Section: Discussionmentioning

confidence: 99%

A non-human primate in vitro functional assay for the early evaluation of TB vaccine candidates

et al. 2021

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“…In order to explore a potential functional role for the antibodies detected, we applied an unbiased sum-of-the-parts mycobacterial growth inhibition assay (MGIA) that we have previously optimised, standardised and harmonised as a potential surrogate of TB vaccine-induced protection [45, 70]. Using this assay, we (and others) have demonstrated ability to detect a BCG vaccine-induced response and an association between the outcome of the direct MGIA and protection from in vivo mycobacterial infection [44, 71–74]. Various immune mechanisms have been proposed to contribute to mycobacterial control in the direct MGIA including polyfunctional CD4+ T cells and trained innate immunity [44, 72, 73].…”

Section: Discussionmentioning

confidence: 99%

“…ELISAs were performed as previously described [44]. The antigens used were purified protein derivative (PPD) from M.tb (Statens Serum Institut (SSI), Denmark) at a concentration of 5μg/ml; whole BCG SSI at a concentration of 5×10 5 CFU/ml, M.tb H37Rv cell membrane fraction, culture filtrate or whole cell lysate all at a concentration of 2μg/ml; or lipoarabinomannan (LAM) at a concentration of 1μg/ml (BEI resources repository).…”

Section: Methodsmentioning

confidence: 99%

“…MGIAs were performed as previously described [44], co-culturing 3×10 6 PBMC and ~500 CFU BCG Pasteur in a volume of 480μl RPMI (containing 2mM l-glutamine and 25mM HEPES), plus 120μl autologous serum per well of a 48-well-plate for 96 hours at 37°C and 5% CO 2 . At the end of the culture period, co-cultures were added to 2ml screw-cap tubes and centrifuged at 12,000rpm for 10 minutes.…”

Section: Methodsmentioning

confidence: 99%

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Induction of specific antibodies, IgG-secreting plasmablasts and memory B cells following BCG vaccination

Bitencourt

Wilkie

Álvarez

et al. 2021

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Many tuberculosis (TB) vaccine candidates are designed as a boost to BCG; an understanding of the BCG-induced immune response is therefore critical, and the opportunity to relate this to circumstances where BCG does protect may direct the design of more efficacious vaccines. While the T cell response to BCG vaccination has been well-characterised, little is known about the B cell and antibody response. We demonstrate BCG vaccine-mediated induction of specific antibodies in different human populations and macaque species which represent important preclinical models for TB vaccine development. We observe a strong correlation between antibody titres in serum versus plasma with modestly higher titres in serum. We also report for the first time the rapid and transient induction of antibody-secreting plasmablasts following BCG vaccination, together with a robust and durable memory B cell response in humans. Finally, we demonstrate a potential contribution of the antibody response to BCG vaccine-mediated control of mycobacterial growth in vitro. Taken together, our findings indicate that the humoral immune response in the context of BCG vaccination merits further attention to determine whether TB vaccine candidates could benefit from the induction of humoral as well as cellular immunity.

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“…We have previously optimised, standardised and harmonised the direct MGIA for use with PBMC from humans and non-human primates (NHPs), and we and others have applied it to demonstrate a BCG vaccine-induced improvement in mycobacterial control, and confirm a biologically relevant response that correlates with protection from experimental in vivo mycobacterial infection (Fletcher et al, 2013;Smith et al, 2016;Joosten et al, 2018;Prabowo et al, 2019;Tanner et al, 2019;Tanner et al, 2020;Tanner et al, 2021). In parallel, the murine direct MGIA has been applied to detect a response to BCG and other TB vaccine candidates (Marsay et al, 2013;Yang et al, 2016;Zelmer et al, 2016;Jensen et al, 2017) and a preliminary protocol made available on BioRxiv (Zelmer et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

Assessment of the reproducibility and inter-site transferability of the murine direct splenocyte mycobacterial growth inhibition assay (MGIA)

Tanner

Zelmer

Painter

et al. 2021

Preprint

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Tuberculosis (TB) vaccine candidates must be tested for safety and efficacy using preclinical challenge models prior to advancement to human trials, because of the lack of a validated immune correlate or biomarker of protection. New, unbiased tools are urgently needed to expedite the selection of vaccine candidates at an early stage of development and reduce the number of animals experimentally infected with virulent Mycobacterium tuberculosis (M.tb). In recent years, there has been a concerted effort to develop standardised functional ex vivo mycobacterial growth inhibition assays (MGIAs) as a potential surrogate read-out of vaccine efficacy. We have previously described a direct MGIA for use with mouse splenocytes. In the current study, we set out to systematically compare co-culture conditions for the murine direct splenocyte MGIA with respect to both intra-assay repeatability and inter-site reproducibility. Common sample sets were shared between laboratory sites and reproducibility and sensitivity to detect a BCG-vaccine induced response were assessed. Co-culturing 5x10^6 splenocytes in 48-well plates resulted in improved reproducibility and superior sensitivity to detect a vaccine response compared with standing or rotating sealed 2ml screw-cap tubes. As the difference between naive and BCG vaccinated mice was not consistently detected across both sample sets at both sites, we sought to further improve assay sensitivity by altering the multiplicity of infection (MOI). Cell viability at the end of the co-culture period was improved when splenocyte input number was reduced, with the highest viability for the condition of 3x10^6 splenocytes in 48-well plates. This cell input was also associated with the greatest sensitivity to detect a BCG vaccine-mediated MGIA response using an M.tb inoculum. Based on our findings, we recommend optimal co-culture conditions in a move towards aligning direct MGIA protocols and generating a cross-species consensus for early evaluation of TB vaccine candidates and biomarker studies.

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Tools for Assessing the Protective Efficacy of TB Vaccines in Humans: in vitro Mycobacterial Growth Inhibition Predicts Outcome of in vivo Mycobacterial Infection

Cited by 28 publications

References 68 publications

A non-human primate in vitro functional assay for the early evaluation of TB vaccine candidates

A non-human primate in vitro functional assay for the early evaluation of TB vaccine candidates

Induction of specific antibodies, IgG-secreting plasmablasts and memory B cells following BCG vaccination

Assessment of the reproducibility and inter-site transferability of the murine direct splenocyte mycobacterial growth inhibition assay (MGIA)

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