Human bacterial challenge experiments as an alternative

November 2013 MRF Annual Conference, London

Experimental Human Infection with Pneumococcus

Stephen Gordon

2© The Liverpool School of Tropical Medicine

Pathogenesis of pneumococcal disease


Outline

•Previous studies of experimental pneumococcal carriage–Jeff Weiser early work–Our first experiments

•How we do our EHPC model–Method, safety issues

•Results–Challenge / re-challenge–Humoral response–Cellular response

•Where next?–Vaccine testing–Vaccine discovery–Host immunity–Microbial ecology


Previous studies of experimental carriage

•Jeff Weiser (U Penn, Philadelphia)

•JEM 2002 and I&I 2003

•Type 23F 6/14 colonised

•Type 6B 6/8 colonised

•Association noted with IgG to 22kD protein in serum

•Did not particularly comment on dose dependency

•3 washes prior to inoculation

•F/up twice weekly and then weekly

Type cfu/naris colonised

23F 5000 0 (4)

23F 7000 3 (6)

23F 17000 3 (4)

6B 22000-39000

6 (8)

J Expt Med 2002;195:359-362 and I&I 2003;71:5724-32


Experimental Human Pneumococcal Carriage Early Studies

Study number Description Dates n Carriers

1 23F Bronchoscopy Nov 09-Feb 10 9 1 =11%

2 23F No Bronchoscopy Mar 10-Apr 10 6 0

3 6B No Bronchoscopy Oct 2010 7 5= 71%

4 6B Bronchoscopy Jan 11-Mar 11 9 0

5 6B No Bronchoscopy Mar 11-Aug 11 9 2 =22%

6 6B Re-challenge Jun 11-Sep 11 3 0

7 6B No Bronchoscopy-inocula test Sep 2011 5 1=20%

8 23F No Bronchoscopy-inocula test Sep 2011 4 2 =50%

9 Gates EHPC –dose ranging 6B

Nov 11-Feb 12 50 21=42%

10 Gates EHPC –dose ranging 23F

Feb 12- 60 3=6.0%

11 Gates EHPC - 6B re-challenge Mar 12-May 12 7 0


Outline






Method

Watch the video


Experimental Human Pneumococcal Carriage


Outline



•Results–Dose ranging and challenge / re-challenge–Humoral response–Cellular response



Dose ranging study

2. Reproducibility study: 42% carriage

1. Dose ranging studies: 6B and 23F

(Gritzfeld, J) in preparation

Over 200 people inoculated with pneumococcus, no SAE


Re-challenge: Carriage protects against acquisition of carriage

Subjects PS Proteins Neither

s1 ++ +s2 =s3 +s4 =s5 ++s6 + ++s7 +s8 ++s9 + +

s10 +

Ferreira et al, AJRCCM 2013


Response to carriage- cellular

Note IL-17 response following colonisation

Responses can be compared in non-carriage vs carriage


12.5ng/ml 125ng/ml 625ng/ml

Increased uptake compared to

vehicle

Decreased uptake compared to

vehicle

p = 0.013

p = 0.004

26% ↑ 37% ↑-Overall % increase in uptake

rhIL-17 dose

Macrophages activated with IL17 kill pneumococci

Percentage change in cfu recovered compared to media


Outline






Carriage model to test vaccine efficacy (2010)

N=58

Vaccine 29

Placebo 29

*mock challenge

8

challenge21

*mock challenge

8

challenge21

*Mock challenge to control for natural acquisition

95% 83%

1 1

12 10

T=0 T=4T=1


Vaccine (PCV vs Hep A): protection against carriage (2013)


Vaccine discovery – Me Vaccine

MRC/FAPESP Bilateral AgreementLSTM/ Butantan Institute

Peptide Arrays: linear epitopes

Fragments (conformational epitopes)

Phage display: protective anti-PspA monoclonal antibodies

- IgG - CD4+ Th17

Learning from acquired immunity (EHPC) how to designbetter vaccines – Cross-reactive epitopes of PspA

Me Vaccine to confer serotype-independent protection


Vaccine discovery - AmiC


Host immunity – correlates of protection


Microbial ecology - nasotypes


Summary of Expt Human Pneumo Carriage project

•Safe inoculation of over 200 volunteers–Strain differences in carriage

•First vaccine study ongoing–Start with PCV–Plan phase 2a in 2014

•Where next?–Vaccine testing – in risk populations–Vaccine discovery – by antigen knockout–Host immunity – role of adjuvant–Microbial ecology – effect of eradication


Thank YOU!!Thank YOU!!


Home Team• Daniela M. Ferreira, Adam Wright, Sarah

Glennie, Toni Banyard• Andrea Collins, Sherouk El Batrawy, Jamie

Rylance, Dan Wootton, Ben Morton• Jenna Gritzfeld, Mathieu Bangert, Kondwani

Jambo, Shaun Pennington• Angela Wright, Carole Hancock, Dave

Shaw, Lorna Roche, Jane ArdreyCollaborators• Eddie Ades (CDC)• Jerry Brown (UCL, London)• Geoff Ginsberg (Duke)• David Goldblatt (ICH, London)• Peter Hermans, Amelieke Cremers, Aldert

Zomer (Nijmegen)• Rob Heyderman (Malawi)• Aras Kadioglu (Liverpool)• Keith Klugman, Josh Shak (Emory)• Rick Malley, Mark Lipsitch (Harvard)• Eliane Miyaji, Adriane Moreno (Butantan)Advisors• Jeff Weiser (U Penn), Mark Alderson (PATH)Safety panel• Rob Read, David Lalloo, Brian Faragher...and our willing volunteers

Royal Liverpool and Broad Green University Hospitals NHS Trust

AcknowledgementsAcknowledgements


Summary of Expt Human Pneumo Carriage project

•Safe inoculation of over 200 volunteers–Strain differences in carriage

•First vaccine study ongoing–Start with PCV–Plan phase 2a in 2014

•Where next?–Vaccine testing – in risk populations–Vaccine discovery – by antigen knockout–Host immunity – role of adjuvant–Microbial ecology – effect of eradication

Key QuestionsCan we test the model in the elderly? HIV infected?Can we use GMO in the model? Adjuvant?


Response to carriage - humoral

P=0.01*

PLoS Pathog. 2012 Apr;8(4):e1002622.


Pre-challenge samples

Post-challenge samples (2 weeks post inoculation)

* ***

*

* * ** **

*** ***** *

Anti-protein IgG - serum

Ab increased 20/27proteins

* P≤0.05 comparing pre vs post samples

Serum response to challenge by MSD


* *** *

** * * * *

** *

* *** * * * ** * *

***

Pre-challenge samples

Post-challenge samples(2 weeks post inoculation)

*P≤0.05 comparing pre vs post samples




Serum response to challenge by MSD

Mock challenge No carriage found Carriage found

Mock challenge shows lower post inoculation serum levels; carriage alters response to many proteins

Ferreira et al. 2012 Unpub


Response to carriage- cellular

There are no naive mice in human studies of carriage

We can detect Ag-specific responses in BAL but not NW

Wright et al. 2012 Unpub.


Colonization density- 6B culture

A higher dose does not equal higher colonization density

Relatively low dose recovery follows inoculation


A simple model of defence vs pneumococcal disease

Health & Medicine

Human bacterial challenge experiments as an alternative