“what are the current knowledge · challenge route and host defence humoral immunity may provide...

“What are the current knowledge

status and gaps towards

establishing correlates of

protection against plague ?”

Dr. Simon Funnell

PHE Porton 23rd April 2018

Current know approaches

Live “attenuated” Yersinia sp.

Live attenuated alternative recombinant bacteria

Live attenuated recombinant virus

Recombinant purified proteins (eg rF1+rV, rF1-V, Yops, etc)

Yersinia OMV

Recombinant OMV from alternative sp. expressing protein (F1+rV)

2 Plague correlates discussion






Yersinia OMV



What does a vaccine need to do?

Rapidly induce lasting immunological protection by;

Inducing the production of neutralising antibodies

Inducing the production of protective cellular immunity

Immune response protects by;

1. Killing Y. pestis

• Antibody directed complement mediated lysis

• Opsonophagocytosis

2. Antibody directed or specific cellular immunity

3. Blocking Yops being delivered via the TTSS to host cells

4. Recognising cells infected with Y. pestis and killing them


Challenge route and host defence

Humoral immunity

May provide “front-line” protection and

help prevent septicaemia

Mucosal immunity

May help against pneumonic

and ingestional routes of infection

Cellular immunity

May provide critical protection against

intracellular pathogens


Bubonic

Flea (or insect) bite or

Infected cut/graze

Pneumonic

Inhalation from another patient

or an aerosol

Ingestional

Eating, drinking, mucosal entry

Does the vaccine protect humans?

Human clinical trials – not possible without existing disease burden

Endemic country trials?

Response to an emergency?

Controlled trials are very difficult

Ethics of unvaccinated

“Emergency response” may change dynamics

“Animal Rule” – one or more suitably qualified models of infection

How do we know if animals and NHPs accurately predict protection?

Need correlates of protection to help in the assessment of vaccine


Does the vaccine protect humans? Human clinical trials


Challenges Possibilities Pro Cons

Not ethical to

infect humans

Trial in endemic

country

Should have

good uptake

rate

Difficult to separate pre-

existing immunity

How to show efficacy

Wait for an

outbreak to start

Should help

the power of

the study

Timing, supplies and Ops

How to show efficacy

Surrogate model

“Animal rule”

Can control

all aspects

Can’t be certain that human

protection correlates

Use Phase 1

sera in passive

protection

Has been

published

FDA approve

T cell immunity probably also

needed

Humoral and T

cell assays

Likely to

correlate

Not always possible in

remote locations

Overview of some assay approaches


Author ELISA Flow

Bacterial

surface

labelling

Fluorescent

Microscopy

Cytokine +

chemokine

assays

Lymphocyte

stimulation

Murine

passive

protection

Murine

active

protection

NHP active

protection

Bashaw

et al 2007 (M)

Williamson

et al 2007 Comp.

(& M)

Graham

et al 2014

Eisele

et al 2011

Xiao

et al 2010

Szaba

et al 2014

Zhang

et al 2014

Derbise

et al 2015 (spleen)

Spinner

et al 2016

Tao

et al 2017 (+rat)

Other vaccines Assay used as correlates in other fields

N. meningitidis - Serum Bactericidal assay (Agar)

Opsonophagocytosis (Flow)

Complement binding (Flow)

B. anthracis - Toxin neutralisation test (macrophages)

B. pertussis - Serum Bactericidal assay (Agar)

Opsonophagocytosis (Flow)

Complement binding (Flow)


Serum bactericidal assay


Yersinia pestis CO92

Functional assays


Functional assessment of Antibody

Biological activity of the humoral response;

• Inhibition of host-cell invasion, cytotoxicity, translocation of virulence factors

or toxins, giant cell formation and plaque formation

• Bactericidal activity

• Flow cytometric measurement of Ab mediated complement binding

• Passive immune therapy

How do we know which of these are good correlates?

Unless we can rely on NHP data, only a human trial will tell us.


Stimulation of PBMCs


Nithichanon et al., (2018) Immune Control of Burkholderia pseudomallei––Common, High-Frequency T-Cell Responses to a Broad

Repertoire of Immunoprevalent Epitopes. Front. Immunol. 9:484. doi: 10.3389/fimmu.2018.00484

A diverse range of T cell epitopes?


Gaps

Relevance – Assays must be relevant to protecting humans

High throughput – Should be able to analyse at least 50 samples a day?

Ideally need assays that can cope with 1000 samples a day

Ease of use – Need assays which do not require “green fingers”

Ideally able to Tech transfer to “in the field” scenarios

Ideally assay protocol should be reproducible on transfer

Ideally - Low cost equipment requirement

Ideally needs qualification and validation and tech transfer

Humoral assays – ELISA not good enough – SBA? Neutralisation assays?

T cell immunity assays – should be able to demonstrate a T-cell response

This might be possible using RNAseq in the future. The interim? MinION?


Gaps – some suggestions

A centralised resource for assay reagents including;

Stocks of reference strains of Y. pestis (restricted use)

Stocks of reference human convalescent antisera (restricted use)

Develop and agree; Standard in vitro functional humoral assay (SBA)

Develop and agree: Standard in vivo functional humoral assay

(Passive therapy? Would in vivo verification do)

Develop and agree; Standard T-cell assay (transferability)

Investigate; Development of RNAseq biomarker sets to detect and differentiate

from disease convalescence and a protective immune response after

vaccination. This may facilitate field testing in disposable units (eg minION?)

Develop: field use systems of all above if possible

Qualify and Validate to comply with regulatory requirements (fit for purpose)


References J. Bashaw, S. Norris, S. Weeks, S. Trevino, J. J. Adamovicz, and S. Welkos (2007) Development of In Vitro Correlate Assays of Immunity to

Infection with Yersinia pestis. CLINICAL AND VACCINE IMMUNOLOGY, May 2007, p. 605–616 Vol. 14, No. 5

Derbise et al (2015) Complete Protection against Pneumonic and Bubonic Plague after a Single Oral Vaccination. PLoS Negl Trop Dis 9(10):

e0004162. doi:10.1371/journal.pntd.0004162

NA Eisele, H Lee-Lewis, C Besch-Williford, CR Brown, and DM Anderson (2011) Chemokine Receptor CXCR2 Mediates Bacterial Clearance

Rather Than Neutrophil Recruitment in a Murine Model of Pneumonic Plague. The American Journal of Pathology, Vol. 178, No. 3, March 2011

VA Graham, GJ Hatch, KR Bewley, K Steeds, A Lansley, SR Bate, and SGP Funnell (2014) Efficacy of Primate Humoral Passive Transfer in a

Murine Model of Pneumonic Plague Is Mouse Strain-Dependent Journal of Immunology Research Volume 2014, Article ID 807564, 11 pages.

http://dx.doi.org/10.1155/2014/807564

JL. Spinner, AM. Hasenkrug, JG. Shannon, D. Kobayashi, BJ Hinnebusch (2016) Role of the Yersinia YopJ protein in suppressing interleukin-8

secretion by human polymorphonuclear leukocytes. Microbes and Infection 18 (2016) 21e29

Szaba FM, Kummer LW, Duso DK, Koroleva EP, Tumanov AV, et al. (2014) TNFa and IFNc but Not Perforin Are Critical for CD8 T Cell-

Mediated Protection against Pulmonary Yersinia pestis Infection. PLoS Pathog 10(5): e1004142. doi:10.1371/journal.ppat.1004142

Tao P, Mahalingam M, Zhu J, Moayeri M, Kirtley ML, Fitts EC, Andersson JA, Lawrence WS, Leppla SH, Chopra AK and Rao VB (2017) A

Bivalent Anthrax–Plague Vaccine That Can Protect against Two Tier-1 Bioterror Pathogens, Bacillus anthracis and Yersinia pestis. Front.

Immunol. 8:687. doi: 10.3389/fimmu.2017.00687

E.D. Williamson, H.C. Flick-Smith, E. Waters, J. Miller, I. Hodgson, C.S. Le Butt, J. Hill (2007) Immunogenicity of the rF1+rV vaccine for plague

with identification of potential immune correlates. Microbial Pathogenesis 42 (2007) 11–21

Xiao PLoS One. 2010; 5(10): e13047. Published online 2010 Oct 13. doi: 10.1371/journal.pone.0013047

Zhang et al “Yersinia pestis biovar Microtus strain 201, an avirulent strain to humans, provides protection against bubonic plague in rhesus

macaques” Human Vaccines & Immunotherapeutics 10:2, 368–377; February 2014; © 2014 Landes Bioscience


“what are the current knowledge · challenge route and host defence humoral immunity may provide...

Documents