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Transcript of 1 Activation of the CREB-dependent pathway distinguishes M.tuberculosis-mediated unmasking immune...
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Activation of the CREB-dependent pathway distinguishes M.tuberculosis-mediated
unmasking immune reconstitution syndrome (IRIS) from chronic HIV and HIV-MTB.
Dawany N., Kossenkov A., Chang C., Conradie F., Ive, P. Sanne I., Montaner L.J., Showe L., Azzoni L.
ART
RIPE (2 mo)u-IRIS IE (4 mo)
Blood draw
ART
ART
RIPE (1-2 mo) IE (4 mo)
Unm
aski
ng
TB-IR
ISCo
ntro
lG
roup
1: n
o M
TBCo
ntro
lgr
oup
2: M
TB
1-3 months
1-3 months
1-3 months
ART
ART + RIPE
Study design
Blood draw
Blood draw
1. Control HIV N= 58
2. MTB + HIV N= 31
3. Unmasking MTB IRIS
N= 2318 confirmed or probable TB IRIS• 11 pulmonary• 7 extra-pulmonary
Unmasking MTB IRIS diagnostic criteria: ACTG
Enrollment:
ARV regimen: 106 of 112 subjects: D4t + 3Tc + EFV, median time 45 days (IQR= [28-61]. 2 IRIS exceptions, ART = 111 and 140 days)
No significant differences in:• ART duration and regimen• Baseline CD4 count and VL• Pre-ART CD4
Study participant characteristics
IL-8
pg/m
l
C-reactive protein
ng/m
l x 1
0-3
25
20
15
10
5
0
Β2-μglobulin
pg/m
l
IL-6
pg/m
l
HGF
pg/m
l
EGF
pg/m
l
CREB-regulated cytokines and acute-phase reactants in unmasking M. tuberculosis-associated IRIS
Benjamini Hochberg FDR < 0.10
G-protein-coupled
receptor(s)
Adenylate cyclase
cAMP
PKACREB
CyclooxygenasePTGS1/2
Prostaglandins
Aspirin
NSAID
COX-2 inhibitors
Hypothesis for unmasking MTB-IRIS pathogenesis
MyD88M. tuberculosis
HIV-1Other pathogens?
Innate effectorsCellular receptors
( e .g.: TLR, Mannose receptors, DC-SIGN, others)
PI3K, AktGSK3 MAPK
Inflammatory response
Pro-inflammatory genes
NF-kB
Adaptive effectors(e.g. TCR)
Gene expression patterns highlightsThreshold= 1.3 fold background expression; ANOVA p value cutoff= 0.05
Pattern 1 (total n= 80): Caspase 6, PKA1*, CD226, Heat Shock Protein B1, cMER
HIV TB u-IRIS
Pattern 2 (total n= 61): CDK7, BCL-10, MAPK-7, ERB-B2HIV TB u-IRIS
•Pattern 3 (total n=122): Adenylate cyclase 3*, prostaglanin E synthase, TNF superfamily, E2F4
HIV TB u-IRIS
Pattern 4 (total n= 157): CXCR4, IL-16, CD44, CD160, MAPK9 HIV TB u-IRIS
unmasking MTB IRIS
MTB (independent of IRIS)
*Upstream of CREB*Downstream of CREB
IL-16: A, Baier M, Serfling E, Kurth R. Proc Natl Acad Sci U S A. 1999 Feb 16;96(4):1541-6.CXCR4: Cristillo AD, Highbarger HC, Dewar RL, Dimitrov DS, Golding H, Bierer BE. FASEB J. 2002 Mar;16(3):354-64.HSP: Billack B, Heck DE, Mariano TM, Gardner CR, Sur R, Laskin DL, Laskin JD. Am J Physiol Cell Physiol. 2002 Oct;283(4):C1267-77.CD44: Rothman BL, Kennure N, Kelley KA, Katz M, Aune TM. J Immunol. 1993 Dec 1;151(11):6036-42.TNF: Matt T. Acta Med Austriaca. 2002;29(3):77-9.; Spriggs DR, Deutsch S, Kufe DW, Immunol Ser. 1992;56:3-34..
CREB family gene expression: heat map
* *
*
*
Gen
e E
xpre
ssio
n
HIV TB IRIS
3
4
5
6x 10
4 CREB1
HIV TB IRIS
0.5
1
1.5
2
x 104 CREB1
HIV TB IRIS
1000
2000
3000
4000
CREB5
HIV TB IRIS1200
1600
2000
2400CREB3L2
HIV TB IRIS200
300
400
500
CREBL1
HIV TB IRIS
400
500
600
CREBL2
HIV TB IRIS
300
400
500
CREBBP
HIV TB IRIS
3000
3500
4000
CREBBP
HIV TB IRIS
500
1000
1500
CREBZF
CREB family gene expression summary
Kruskall Wallis FDR < 0.10 Wilcoxon < 0.05 Wilcoxon < 0.10
TLR family gene expression: heat map
Gen
e E
xpre
ssio
n
HIV TB IRIS
200
600
1000
TLR10
HIV TB IRIS
1000
2000
3000
TLR8
HIV TB IRIS
500
1000
1500
TLR7
HIV TB IRIS500
1000
1500
2000
TLR6
HIV TB IRIS
400
800
1200
TLR5
HIV TB IRIS
500
1000
1500
TLR4
HIV TB IRIS
200
300
400
TLR2
HIV TB IRIS
1000
2000
3000
TLR1
*
*
*
TLR family gene expression summary
Kruskall Wallis FDR < 0.10 Wilcoxon < 0.05 Wilcoxon < 0.10
Conclusions• Unmasking MTB-IRIS is characterized by:
• Increased expression of cytokines and acute-phase reactants regulated by the cAMP/CREB pathway
• Increased gene expression of CREBL2>CREB5 (transcription activators)• Decreased gene expression of CREBZF (transcription repressor)
• These result support a role for an activation of the CREB pathway(s) in the pathogenesis of IRIS
• Differential expression of TLR genes (e.g. increased TLR5) suggests another potential activation pathway.
Limitations• Small number of subjects• Lack of CREB and TLR protein expression and activation assessments• Lack of a comparative paradoxical MTB IRIS group• Possible confounding effect of treatment in the MTB control group
Planned studies• Validation of gene array results in larger longitudinal cohorts• Assessment of CREB and TLR protein levels and activity
Acknowledgements
Funding:NIH/NIAID grant RO1 AI069996 to L.A. The Philadelphia Foundation The Stengel-Miller familyAIDS funds from the Commonwealth of Pennsylvania and from the Commonwealth Universal Research Enhancement Program, Pennsylvania Department of HealthCancer Center Grant (P30 CA10815)Penn Center for AIDS Research (CFAR).
• The Wistar Institute, Philadelphia, PA, USA:• Noor Dawany • Andrew Kossenkov • Celia Chang • Louise Showe • Luis J. Montaner
• Department of Molecular Medicine and Haematology, University of the Witwatersrand, Johannesburg, South Africa:• Denise Lawrie• Deborah Glencross• Wendy Stevens
• Clinical HIV Research Unit, Department of Medicine, University of the Witwatersrand/Themba Lethu Clinic, Johannesburg, South Africa:• Study participants• Katerina Roussos• Prudence Ive • Francesca Conradie• Ian Sanne