Episode 338: Recent developments in TB vaccines.
Last week, both Science (Ep 338-1) and Nature (Ep 338-2) announced that -finally- a large phase III trial ( with 26,000 adults) will be launched in Africa and South-East Asia to test a candidate subunit vaccine, originally developed by Areas (a non-profit) and now owned by GSK (Ep 338-3).
It is a recombinant protein subunit vaccine (M72), combined with one of the potent GSK adjuvants (AS01E ) that is also used in their RTS,S malaria and recombinant zoster vaccine.
This vaccine showed about 50 % efficacy in a phase 2 trial to prevent reactivation from latent TB.
Should we be excited about that result? We rejected the CureVac RNA vaccine against SARS-CoV-2, because it was “only” 50 % efficacious and now WHO is in favor of further developing any TB vaccine with 50 % efficacy….
But…TB is not COVID….
In this Episode, I will first give some background on TB and the classical BCG vaccine, which will provide the context to understand the development of novel vaccine concepts and then elaborate on some examples of vaccines aiming at prevention of infection (POI) versus prevention of disease (POD).
Par 1 General background on TB: clinical and epidemiological aspects
Ep 338-4: Master course by G Vanham and O Goovaerts 2017 for those of you who are not familiar with TB
Ep 338-5: Kaufmann in Essential Tuberculosis 2021 provides a complex, but realistic picture of the clinical evolution
TB represents as complex continuum from Mtb infection to active TB disease,
which can progress forward, reverse backward, or stand still at any point of the infection process.
Up to 10% of cured individuals suffer from TB recurrence within 12 months in high-endemic areas, because
- reinfection = infection with a new strain of Mtb
- or relapse = reappearance of the original strain, which had evaded elimination by drug treatment
There is evidence of insufficiency of the immune response induced by natural Mtb infection during both latent TB infection (LTBI) and active TB to fully protect against disease.
Note: Ep 338-15: Hilary Whitworth Methods 2013: IGRAs – The gateway to T cell based TB diagnosis
TST = Tuberculin Skin Test: involves intradermal injection of Mtb antigens and subsequent observation (at
48–72 h) for the induction of cutaneous induration, resulting from a delayed-type hypersensitivity response to the antigens ;
IGRA = Interferon-gamma release assay: detect a T cell IFN-γ response in vitro following overnight stimulation of PBMCs or whole blood with TB antigens
In both cases, the antigens are specific for MTB ( and different from the vaccine strain BCG), therefore these tests show that TB infection has occurred, but indirectly (through the T cell-mediated sensitization to TB antigens
Ep 338-6: Swati Gupta Critic Rev Therapeut Drug Carrier Systems 2012
A schematic view on clinical TB
HIV is a strong stimulus to reactivate T from LTBI, because of CD4+ T cell dysfunction and depletion.
Innate and adaptive immune responses
First line treatment of TB
Note: RIF = Rifampycin; INH = Isoniazid; PZA = pyrazinamide; EMB = Ethambutol; SM = Streptomycin
Ep 338-7: WHO Key facts on TB April 2023:
1.7 billion people or a quarter of the population on this globe lives with latent TB infection
Ep 338-8: WHO Detailed Report 2022
Most affected: Africa, especially Central and South; Asia: South and South-East; Peru and Bolivia
Undernourishment, HIV, alcohol, smoking and diabetes are very important risk factors !
Multi-drug resistance (MDR) mainly India, Pakistan, China, Russia, Philippines, Indonesia, South-Africa and Nigeria
Success rate in multidrug resistant cases slowly improving
Mortality by TB
TB mortality is slowly declining, especially in HIV (+) people
Par 2 Immunology of TB is very very complex and true correlate of protection still enigmatic
Ep 338-9: Maria Romana Cells MDPI 2023: Structural view at vaccine development against MTB
Figure 1. A simplified sketch of Mtb-induced immune reaction.
(A) Upon MTB infection, macrophages produce cytokines which recruit other immune cells to form the Mtb granuloma. In these structures, macrophages can differentiate in other cell types, like epithelioid cells, giant cells, and foamy macrophages.
(B) Antigen presenting cells (APCs) expose antigens on their MHC molecules, thus bringing them to T cells CD4 and CD8. Cytokine production by APCs induce T cell differentiation in various directions (Th1, Th2, Th17 CD4+ T cells as well as cytolytic CD8+ T cells
But which of these responses contributes to protection?
Ep 338-10: Janez Ferluga Immunobiology 2020 Natural Immunity to MTB
Fig. 1. Mtb invasion and granuloma formation:
(A) Mtb entering through airway mucosa are phagocytosed by alveolar Mφs;
(B) These alveolar Mφs then release various cytokines recruiting NK cells, DCs, γδT cells, and innate invariant T cells (orange in colour), providing first line of defence against aerosol-based Mtb transmission;
(C) Upon internalisation, Mtb pathogen inhibits phagosome maturation, and induces M1 phenotypic differentiation via ESAT-6. This leads to the formation of primary innate granuloma, which is MTB antigen ESAT-6 dependent and T cell independent;
(D) Eventually, a pro-inflammatory environment is generated enhancing efficient recruitment of adaptive immunocompetent cells (Th1 and B cells) forming a bactericidal granuloma (non-caseous granuloma)
= PROTECTIVE anti-MTB
(E) With persistent infection for a longer period, ESAT-6 drives polarisation of M1 towards M2 phenotype, inducing foamy cells differentiation non-caseous granuloma). Th2 and Treg cells surround the granuloma. Mtb infected M2 Mφs upregulates KLF4 and C/EBPβ. KLF4 signalling inhibits phagosome/lysosome fusion preventing autophagy and thus helps in Mtb survival
= NON-PROTECTIVE pro-MTB.
Fig. 2. Cytokines and mediators involved in Anti-Mtb response & Pro-Mtb response.
The upregulated cytokines and mediators are shown with upward arrows and the downregulated ones with downward arrows during M1 and M2 polarisation.
Ep 338-11: Sasha Larsen Front Immunol provides a slightly different model on MTB-relayed immune mechanisms.
Modeling immunological mechanisms of early prevention of infection (POI) or later prevention of disease (POD).
- In early infection (upper), detection of Mtb bacilli in the pulmonary alveoli by macrophages leads to downstream activation of innate immune cells, which may include neutrophils, NK cells, and DCs. Activated APCs can then prime T cells (CD4+ or CD8+) for further Mtb-specific adaptive responses to target Mtb and Mtb-infected cells and control infection. Vaccines for prevention of infection (POI) should prophylactically activate these mechanisms, before infection.
- Later stages of Mtb infection (lower) and pathology are defined by formation of a granuloma, which contains Mtb by a surrounding composition of immune cells.
- Containment (proactive immunity) seem to correlate with IgM, robust pulmonary CD4+ and CD8+ T cells, and activated inflammatory M1 macrophages and DCs at the granuloma.
- In contrast, regulatory TH2 CD4+ T cells, abundance of IgA or IgG4, and M2 macrophages are more associated with loss of control. Higher peripheral gamma-delta T cells are associated with active TB disease in humans, but their direct role in granuloma control or bacterial dissemination requires further study.
Vaccines for prevention of disease (POD) should modulate this delicate balance towards containment.
IS THERE A SOLID CORRELATE OF PROTECTION? NOT YET, but there are several suggestions….
Ep 338-12: Jeffrey Morgan Immunol Rev 2021
The immune response against Mtb is complex and involves all aspects of the immune system,
however, the optimal protective, non-pathogenic T cell response against Mtb is still elusive.
Nevertheless: studies have identified specific classical CD4 T cell subsets important for immune responses against Mtb in various states of infection.
Characteristics of Mtb-specific classical CD4 T cells in different disease stages and potential correlates of protection (CoP).
Latent TB infection (LTBI): a higher frequency of Tcm (central memory), Th1* and CD153 *
Acute TB (ATB): higher frequency of Tem (effector memory) and Tscm (stem cell memory), HLA-DR and more differentiation
Th1 and CD153 are important for control of Mtb infection.
Other cytokines and chemokines include: IL-2, IL-10, IL-17, TNFα, and CXCL9/10/11/12/13
* CD153 (cluster of differentiation 153) also known as tumor necrosis factor ligand superfamily member 8 is is encoded by the TNFSF8 gene. It is a cytokine ligand for the TNF receptor CD30. It plays a role in the T cell dependent anti- MTB response.
Ep 388-13: Role of CD30/CD153(CD30L) in the anti-MTB T cell response.
- After mycobacterial antigens presentation to naïve CD4+CD30− T cells by dendritic cells (DC), the Th0 cells produces both IL-4 and IFN-γ and differentiate into Th1 or Th2, CD4+CD30+. After secondary antigen exposure, binding of CD153 to CD30 may complement and even replace other costimulatory signals for further differentiation and clonal expansion of Th2 and preferentially of Th1 effector and memory cells, which upon continuous activation release soluble CD30. Although there is no direct evidence it would be possible that sCD30 down regulates the production of IFN-γ.
- Evidence from Guo et al suggests that during early M. bovis BCG infection Vγ6/Vδ1 CD30+CD153+ T cells produce IL-17A, possibly involving homotypic T-T cell interactions which may play an important role in the early inflammatory response.
Ep 338-14: Todia P. Setiabudiawan Cell 2022 questions the role of TH1 and multifunctional CD4 T cells as correlate and argues in favor of central memory T cells (TCM, “trained immunity” as well as specific IgA and IgG
This author also argues that the most used outcome in trials, IGRA (Interferon-Gamma Release Assay) is probably not a 100 % reliable parameter of TB infection, as it does not measure infection directly, but the IFN production as a result of MTB infection. However:
- False positive: people with positive IGRA may have self-cleared the infection but still have a Th1 immune response,
- False negative: people with a negative IGRA might be infected with Mtb but only have IFN-g independent responses
The proposed better alternative is the Mycobacterial growth inhibition assays (MGIA) it measures the growth of mycobacteria in whole blood or PBMCs ex vivo, assessing the combined, complex interaction between T cell, B cell, and innate immunity that affects mycobacterial growth.
Inhibition of mycobacterial growth has been associated with multifunctional CD4+ T cell, B cell, and IgG1 responses and markers of trained immunity
Ep 388-16: Brennan Clin Vaccine Immunology 2017 MGIA review
Par 3 Good old Bacille Calmette-Guérin (BCG) vaccine
Ep 338-14: Todia P. Setiabudiawan Med Cell 2022 History of discovery, evolution and distribution of BCG.
In 1921, Bacillus Calmette-Guerin (BCG) was used for the first time in an infant.
1921-1930 BCG use increased in Europe
1929-30 until the Lubeck disaster caused a downturn in public and expert opinion regarding the safety of the vaccine.
Nevertheless, BCG use increased again because of the resurgence of TB after World War II, and it continues to be used worldwide.
The Lübeck disaster: During the second half of 1929, the medical personnel in Lübeck requested a supply of BCG from Calmette for a vaccination program targeting infants born into tuberculosis-endemic environments. After delivery, the vaccine was unfortunately stored and processed in a hospital laboratory where virulent M tuberculosis was also stored. During the spring of 1930, BCG vaccination was offered to all 412 newborn babies in Lübeck: 72 infants died.
An in-depth assessment showed:
- no evidence that BCG had reverted to pathogenicity;
- rather the consequence of negligent contamination of BCG with virulent MTB in the lab
Potential factors influencing the beneficial effect of BCG
Ep 388-17 Lange Lancet Infect Dis 2022 and Ep 388-18 Mangtani CID 2014 investigate the evidebce of some of these factor in meta-analysis.
BCG provides long-lasting strong protection against miliary and meningeal tuberculosis in children,
but it is less effective for the prevention of pulmonary tuberculosis, especially in adults
Excluding tuberculine-reactive individuals = to exclude already (latently) infected individuals
RR= relative risk or rate ratio of getting pulmonary TB after BCG versus controls
A clear effect of age and latitude; but not really the BCG strain used.
- Latitude presumed to be linked with higher exposure to environmental mycobacteria closer to equator.
- Studies that exclude tuberculin-reactive children in efficacy analysis show larger vaccine efficacy.
Effect on meningeal and/or miliary (generalized) TB shows
- Large confidence intervals and some trials show paradoxical effect (Georgia/Alabama 1950).
- Clear effect of age: neonatal immunization provides most pronounced protection
- Clear effect of previous sensitization (stringency): more pronounced protection by BCG if tuberculin-positive (= already infected) children are excluded.
Ep 388-19: Roy in BMJ 2014 is a metanalysis of BCG effect on MTB infection versus active TB disease in children.
Overall protection against infection (estimated by IGRA), based on 14 studies and 3855 participants: 19 %
Restriction to 6 studies (n = 1745) with info on clinical progression to active TB:
- Protection against infection of 27% (risk ratio 0.73, 0.61 to 0.87)
- Protection against active TB = 71% ( RR 0.29, 0.15 to 0.58)
- Among those infected, protection against progression to disease was 58% ( RR 0.42, 0.23 to 0.77).
Ep 388-20: WHO statement 2023 on BCG vaccination
- has a documented protective effect against meningitis and disseminated TB in children
- does not prevent primary infection in adults and, more importantly,
- does not prevent reactivation of latent pulmonary infection in adults = principal source of bacillary spread.
The impact of BCG vaccination on transmission of Mtb is therefore limited
Ep 388-5: Kaufmann Essential Tuberculosis 2021: Tuberculosis Vaccine Development Pipeline
Whilst BCG immunization can protect against extrapulmonary TB in neonates, it fails to eliminate Mtb, hence does not diminish risk of pulmonary TB in all age groups.
Ep 388-21: Marco Antonio HUMAN VACCINES & IMMUNOTHERAPEUTICS 2020: Multiple faces of BCG
- efficacy against disseminated and meningeal tuberculosis.
- significant protective effect against leprosy.
Trained immunity: beneficial heterologous (nonspecific) effects on mortality in some developing countries.
- considered the gold standard adjuvant treatment for non-muscle-invasive bladder cancer.
- has also been tested as treatment for diabetes and multiple sclerosis: with questionable results.
In Kawasaki erythema of the BCG site is recognized as a clinical clue in Kawasaki disease.
BCGitis and BCGosis: local or disseminated disease in the immunodeficient patient.
BCG administration has been associated with the development of autoimmunity ?
Ep 338-17: Lange Lancet Infect Dis 2022 explains the concept of trained immunity by BCG
Trained immunity induced by BCG results in
- In monocytes-macrophages: improved cytokine (TNF, IL-1, IL-6) responses
- Neutrophils more effective release of reactive oxygen species, antimicrobial proteases, and enhanced pathogen killing by neutrophils.
To achieve long-term effects, functional changes are induced by BCG at the level of bone marrow progenitors of myeloid cells biasing towards myelopoiesis: can last for months and even years.
The molecular processes responsible are changes in chromatin accessibility due to chemical processes at the DNA (methylation) and histone (methylation, acetylation) level, leading to more effective transcription of genes important for host defense (figure 3).
Nevertheless, it remains a somewhat vague concept that is often invoked to explain unexpected clinical phenomena without much evidence. See the claim that lesser pathogenicity of SARS-CoV-2 in Africa was due to “trained immunity”, while, in fact, more obvious explanations are present, such as the younger population.
Par 4: Novel vaccine concepts
It is evident that BCG has limitations and it can have serious adverse effects in immunocompromised subjects
To be distinguished:
- Prevention of infection (POI) or pre-exposure vaccines versus prevention of disease (POD) or post-exposure prophylaxis.
- POI can be evaluated with prevention of conversion of TST or IGRA (tuberculin skin test or interferon-gamma release assays)
- POD requires more complicated clinical parameters.
- There are several concepts under development:
Ep 338-22: Shruti Srivastava Vaccines MDPI 2023: TB vaccine where are we now?
- Protein subunit vaccine: antigens, selected for their protective effect against TB in animal models, with strong innovative adjuvants, which promote a Th1 and polyfunctional CD4 T cell response;
(b) Recombinant BCG vaccines: BCG with extra MTB antigens, associated with protective immune responses in animal models
c) Viral vector vaccines; contains well known attenuated, replication-deficient non-pathogenic viruses, which express potentially protective MTB antigens.
- The vectors include: Modified vaccina Ankara (MVA), Human or Rhesus Adenoviruses, attenuated Influenza, Rhesus Cytomegalovirus (RhCMV) or Lymphocytic Chorio-Meningitis virus
- Attenuation or Inactivation of whole-cell-based vaccine using related mycobacteria;
Ep 338-11: Sasha Larsen Front Immunol 2022: Multifaceted immunity to MTB
Par 5 Some examples of early success with novel concepts
5.1. Boost with H4:IC32 subunit vaccine as compared to BCG boost to prevent infection (POI)
Ep 338-23: Nemes NEJM 2018 provides evidence that boost with BCG or subunit vaccine H4:IC31 reduces MTB infection in HIV-negative adolescents who received BCG as neonates, were not TB infected (negative IGRA) and were at high risk for MTB infection in South-Africa
H4:IC31: H4 = recombinant subunit of Ag85B + TB10.4 with IC31 adjuvant which is TLR-9 activator
Evidence of infection = sustained positive result in IGRA
QFT sustained conversion
after 3 and 6 months
Reduction sustained conversion compared to placebo
Clearly, this result is not based on direct proof of presence or absence MTB infection, but on T cell response against MTB antigens as measured by IGRA with:
- Conversion indicates infection,
- Sustained conversion = established infection
- Reversion indicates clearing of infection.
Note: The paper provides no information on clinical active TB
5.2. M72/AS01 subunit vaccine: acting in latent TB, hence prevention of disease (POD)
Ep 338-24: van der Meeren Phase 2B M72/AS01 NEJM 2018
About 3300 HIV-negative adults with proven latent TB (by IFN-gamma release assay or IGRA) in Kenya, Zambia and South-Africa followed after 2 doses of M72/AS01 for 2.3 years.
Overall: clear effect, but low prevalence of active TB (0.3 per 100 person/years in vaccine group versus 0.6 in placebo)
Some remarkable confounders:
Clear effect of age (> 25 better) sex (male better). However, more young men than women.
Effect of country and BCG less clear.
Ep 338-25: Tait NEJM 2019: Final analysis after 3 years: vaccine efficacy 49.7 %
Ep 338-26: Zhenhua Ji Front Immunol 2019 Meta-analysis of 7 studies with M72 vaccine (4590 participants)
Vaccine efficacy was 57.0% (but most studies with follow-up of 6-12 months, except the van der Meeren-Tait NEJM).
Side effects = acceptable and limited to local (redness, swelling), fatigue and malaise
Immune correlates of vaccination: mainly M72-specific polyfunctional Th1 CD4 T cells and antibodies
SMD (standardized mean difference between vaccinated and placebo) > 0 indicates that vaccine can effectively stimulate the growth of polyfunctional CD4+ T-cells.
RR (relative risk) > 1 indicates association with vaccination
Interestingly, there was NO association between vaccination and either CD8 T cell and Th17 CD4 responses
Ep 338-27: E Thacher AIDS: M72 AS01 was safe and immunogenic in HIV (+) subjects on antiretroviral therapy
M72-specific CD4 T-cell responses following vaccination with M72/AS01in HIV(+) subjects
Data are reported as the percentages of M72-specific CD4+ T cells expressing (after in-vitro stimulation) at least two immune markers among IFN-g, IL-2, TNF-a and CD40L of all CD4+ T cells
These results are similar as previously shown in HIV(-) subjects (Ep 338-28 I Leroux-Roels Vaccine 2013)
(also shown here is superiority of AS01 over AS02 adjuvant)
5.3. First experimental mRNA vaccine for TB
Ep 338-29 A and B: Sasha Larsen Vaccine 2022 and IJID 2023: RNA vaccine for TB ?
(A) ID91 fusion protein ID91 comprising 4 MTB antigens Rv3619, Rv2389, Rv3478, Rv1886.
Formulated with glucopyranosyl lipid adjuvant formulated in a stable emulsion (GLA-SE)= TLR-4 agonist
(B) Replicating-RNA (repRNA) = Alphavirus backbone expressing nonstructural proteins (nsP) and ID91 fusion antigen
Formulated in nonstructured lipid nanoparticle (NLC)
Experimental design: C57BL/6 mice were vaccinated two times three weeks apart with
- ID91 fusion protein +TLR4 agonist GLA-SE)as a positive control for CD4+ T-cell responses and prophylactic efficacy
- The novel candidate ID91-repRNA + NLC
Moderate protection, most pronounced with combo (= 2 X protein + RNA)
- ID91-specific IgG and IgA responses most pronounced with repeated protein vaccination
- ID91 specific CD4+ T cell responses most pronounced after repeated combo vaccination
SOME GENERAL CONCLUSIONS
- MTB itself induces partly protective immune responses, since many immune-competent infected individuals either eliminate or control the infection life-long.
- The close association between untreated HIV infection and TB reactivation shows that CD4 T cells have an important role in containing TB infection.
- The TB antigens and the type of immune response involved in protection have only partly been elucidated. There are arguments that, besides Th1 CD4 T cells, other immune mechanisms are also involved, but the role of antibodies is limited. Environmental effects (through other non-pathogenic mycobacteria?) have also a role.
- Neonatal BCG induces mainly POD (prevention of disease) immunity in children. The POI effect is limited and too many people still acquire and get ill from MTB infection in endemic areas, with generalized neonatal BCG vaccination.
- Nevertheless, according to Nemes NEJM 2018, a boost of BCG could reduce new infection in adolescents by almost half.
- BCG has pleiotropic effects on the adaptive and innate immune system, may be responsible for “trained immunity”, has been used in some cancer therapies etc.
- Novel vaccine concepts are trying to focus the immune system on (combinations of) particular MTB antigens, in combination with potent adjuvants and come in various formats, including subunit proteins themselves or expressed via recombinant viral vectors, (myco-)bacteria or self-replicating RNA.
- There are some successful clinical trials with various novel vaccines, both in the field of POI and POD, but an efficacy of about 50 % is the maximum which has been reached until now.
Developing a TB vaccine with 90 % efficacy will take more time….
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