Immunology of Immunology of ChikungunyaChikungunya and and implications for disease implications for disease

interventionsinterventions

Lisa F.P. Ng, PhDLisa F.P. Ng, PhD

Principal Investigator, Singapore Immunology NetworkPrincipal Investigator, Singapore Immunology Network

Adjunct Assistant Professor, National University of SingaporeAdjunct Assistant Professor, National University of Singapore

Challenges and insights towards understandingChallenges and insights towards understandingthe reemergence of the reemergence of ChikungunyaChikungunya

20 Mar 2009, York Hotel, Singapore20 Mar 2009, York Hotel, Singapore

Expand and strengthen the immunology Expand and strengthen the immunology expertise in Singaporeexpertise in Singapore

www.sign.a-star.edu.sg

ChikungunyaChikungunya FeverFever-1st isolated in a 1953 outbreak in Tanzania- Mosquito-transmitted: Aedes- Re-emerged in 2005 in the Indian Ocean Islands and has since spread to SE Asia and other parts of the world

Arthralgia

- Acute infection lasts 1-10 days: sudden onset of high fever, rash, painful arthralgia > incapacitating joint disease

- Pathogen: CHIKV, an alphavirus of the Togaviridae family- Positive-strand RNA virus of 11.8 kb

CHIKV

What are the current needs?What are the current needs?

Build up a strong platform in CHIKV clinical immunology research for better translation into medical applications

How?How?

- Identification of targets, immune mechanisms, and correlates of protection

FieldField BenchBench BedBed

Global Research InitiativesGlobal Research Initiatives

To gather fundamental knowledge on the immune To gather fundamental knowledge on the immune responses mounted against CHIKV with a view to exploit responses mounted against CHIKV with a view to exploit this to develop new immunethis to develop new immune--based preventive and based preventive and treatment strategiestreatment strategies

1. Candidate biomarkers from CHIKV-infected patients?

2. Innate immune responses against CHIKV-infection?

3. B- and T-cell responses?

Important QuestionsImportant Questions

Identify BiomarkersIdentify Biomarkers

-- Diagnostics for improved detectionDiagnostics for improved detection

-- Pathology to devise appropriate treatmentsPathology to devise appropriate treatments

-- Identify novel targetsIdentify novel targets

-- Development of novel assays for drug screenDevelopment of novel assays for drug screen

11stst Singaporean Outbreak: Jan Singaporean Outbreak: Jan –– Mar 08Mar 08

Demographic and epidemiologic data on 10 patients with PCR-confirmed chikungunya infection.

Patient No. Age (years) Gender NationalityDuration of fever (days) Illness severitya Pre-morbid condition Clinical Outcomeb

1 45 M Bangladeshi 2 Not Severe None Persistent arthralgia2 32 M Bangladeshi 4 Not Severe None Complete recovery3 33 M Indian 5 Not Severe None Complete recovery4 45 M Indian 5 Not Severe None Complete recovery5 28 M Malaysian 3 Not Severe None Complete recovery

6 65 M Singapore resident 10 SevereLiver cirrhosis, hypertension, atrial fibrillation, anaemia Complete recovery

7 34 M Indian 4 Severe None Complete recovery8 39 M Indian 6 Severe None Persistent arthralgia9 37 M Indian 9 Severe None Complete recovery10 22 M Malaysian 4 Severe None Developed myalgiaa Severity was defined as having a temperature >38.5degC or pulse rate >100/min or platelet count <100x10^9/Lb Clinical outcome at 2 weeks post-illness onset

Clinical features

Sign/Symptom No. (%) of patientsFever 10 (100)Arthralgia 9 (90)Rash 5 (50)Conjunctivitis 4 (40)Gastrointestinal symptom+ 3 (30)

Headache 3 (30)Eye pain 2 (20)Back pain 2 (20)Mylagia 1 (10)Arthritis 1 (10)+ Nausea, vomiting, diarrhoea, or abdominal pain

Patients had fever, rash and arthralgia

None had neurologic involvement

CH

IKV Positives

Non-severe

Severe

Healthycontrols

Blood Mediators during Acute CHIKV InfectionBlood Mediators during Acute CHIKV Infection

IL-6

Controls CHIK +ve0

50

100

150p = 0.0007

pg /

ml

IL-8

Controls CHIK +ve0

200

400

600p = < 0.0001

pg /

ml

IL-10

Controls CHIK +ve0

20

40

60p = 0.0297

pg /

ml

IL-2R

Controls CHIK +ve0

500

1000

1500p = < 0.0001

pg /

ml

Differences in Cytokines LevelsDifferences in Cytokines Levels

IL-5

Controls CHIK +ve0

10

20

30

40

50p = 0.0159

pg /

ml

IFN-a

Controls CHIK +ve0

100

200

300p = 0.0010

pg /

ml

IL-7

Controls CHIK +ve0

100

200

300p = < 0.0001

pg /

ml

IL-15

Controls CHIK +ve0

100

200

300p = 0.0016

pg /

ml

Differences in Cytokines LevelsDifferences in Cytokines Levels

IP-10

Controls CHIK +ve0

2000

4000

6000p = < 0.0001

pg /

ml

MIG

Controls CHIK +ve0

200

400

600

800

1000p = 0.0006

pg /

ml

Eotaxin

Controls CHIK +ve0

50

100

150

200

250p = 0.0030

pg /

ml

Differences in Differences in ChemokinesChemokines LevelsLevels

EGF

Controls CHIK +ve0

100

200

300p = < 0.0001

pg /

ml

HGF

Controls CHIK +ve0

200

400

600

800

1000p = 0.0030

pg /

ml

FGF-b

Controls CHIK +ve0

50

100

150

200p = < 0.0001

pg /

ml

VEGF

Controls CHIK +ve0

100

200

300p = < 0.0001

Differences in Growth Factors LevelsDifferences in Growth Factors Levels

High levels of pro-inflammatory cytokines

How do they contribute to immuno-pathology?

IL-1B

Controls Non Severe Severe0

100

200

300

400p = 0.0026

pg /

ml

IL-6

Controls Non Severe Severe0

50

100

150p = 0.0009

pg /

ml

RANTES

Controls Non Severe Severe0

5000

10000

15000

20000 p = 0.01

pg /

ml

Disease Severity?Disease Severity?

Clinical definition of Severity: Fever > 38.5 C;Maximum pulse rate > 100 beats/min;Platelet count < 100 x 109/L

Uninfected ChikungunyaInfection

SevereChikungunya

IL-2RIL-5IL-6IL-7IL-10IL-15IFN-aIP-10MIGFGF-basicVEGF

IL-8EotaxinHGFEGF

IL-1bIL-6

RANTES

11stst Singapore CHIKF Outbreak ProfileSingapore CHIKF Outbreak Profile

PLoS ONE 4: issue 1, e4261, 21 Jan 2009