New tricks with an old dog
1890
1907
1934
Mahdad Noursadeghi
UCL Division of Infection & Immunity
New insights from an old test
1890
1907
1934
Mahdad Noursadeghi
UCL Division of Infection & Immunity
2016
New insights from an old test
2016
Is there more to it than erythema & induration?
IL12
M
Th1
IFN
Necessary but
insufficient
Tomlinson, Eur J Immunol. 2011
Is there more to it than erythema & induration?
IL12
M
Th1
IFN
1. Reflects molecular
immunology at the site of
disease?
Is there more to it than erythema & induration?
IL12
M
Th1
IFN
1. Reflects molecular
immunology at the site of
disease?
2. Molecular composition
reveals differences in
immune responses between
people or groups of people?
Molecular assessment of human cell mediated
immune responses in vivo…
Molecular assessment of human cell mediated
immune responses in vivo…
Tomlinson, Eur J Immunol. 2011
HIV-ve patients with active TB
800
600
400
200
0
123456789
Fold increase (Log2)
50
Z score
No of genes
1000
NFkB
RELA
NFKB1
REL
30
STAT1
20IRF2
STAT3
10
0
5 10 15
Gene hits (x100)
40
Bell et al. PLoS Pathogens 2016
Independent cell & stimulus specific modules
LPS
Spn
2.0
4
4
1.5
3
3
1.0
2
2
0.5
1
1
0.0
0
0
-0.5
-1
-1
3
-1
IL
4
R
TL
2
R
TL
n
Sp
tb
M
/
-4
IL
F
TN
N
IF
ll
ce
l
hi
op
tr
eu te
N
y
oc
on
M ll
ce
K
N
ll
ce
B
T
Bell et al. PLoS Pathogens 2016
TST gene signature in active TB explains all the
variability in molecular changes at site of disease
Molecular
Distance
to Health
Kim et al. EMBO Mol Med 2010
Random modul.
0.0
0.5
r2=0.99
p=0.0003
0.0
-0.5
-0.5
1.5
2.0
2.5
MDH (x105)
Blood module
0.6
Module score
0.5
r2=0.11
p=0.594
Module score
1.0
TST module
1.0
r2=0.81
0.4 p=0.0365
0.2
0.0
-0.2
-0.4
1.5
2.0
2.5
MDH (x105)
1.5
2.0
2.5
MDH (x105)
Bell et al. PLoS Pathogens 2016
A reference gene list…
HIV-ve Active TB TST+ve
Unpublished data
Positive TST in patient with & without active TB
are not the same…
HIV-ve Active TB TST+ve
Healthy
Active TB
DEFB4A
MMP1
PI3
APOBEC3A
LCE3A
VEGFA
S100A9
FCER1G
PKM2
DSC2
FCGR2A
S100A12
LY6E
CSF3
C13orf33
FOXN2
PDIA3
C4orf7
744
968
347
TB (mdn exp log2)
HIV-ve Healthy TST+ve
18
in active TB
15
12
in healthy
9
6
r2 = 0.83
Slope = 0.97
6
9 12 15 18
Healthy (mdn exp log2)
-5
+5 Fold change (Log2)
Unpublished data
18
12
9
6
Relative expression (Log2)
15
Number of genes
Differences in gene-specific variance…
P1 0
M 1
M R1B
K A
A B4
EF
D T6B C
R 3
K P1R A
7
PP 0A
0 4
S1 FM B3
L IN
O RP
1
SE P2S B4
Y IN
C RP
5
SE ISA
SHSS1
PDX5
F
R 9 D
F 3
IR LR
POG5
T 5
A rf1
5o
C T2
O
FL IP1
K
A P3
P
D
Unpublished data
HIV+ve patients with clinically positive TSTs
HIV-ve Active TB TST+ve
515
1210
81
HIV+ve TST+ve
(mdn log2 exp)
HIV+ve Active TB TST+ve
18 r2 = 0.97
Slope = 0.99
15
12
9
6
in HIV+ve
6 9 12 15 18
HIV-ve TST+ve
(mdn log2 exp)
Bell et al. PLoS Pathogens 2016
HIV+ve patients with clinically positive TSTs
HIV-ve Active TB TST+ve
IL10 specific modules
4
0.8
515
1210
3
0.6
*
2
*
5
4
3
0.4
2
1
0.2
0
0.0
1
2
18 r = 0.97
Slope = 0.99
15
12
9
6
in HIV+ve
0
+
IV
H
IV
H
10
L.I
al
et
s
le
Te
n
10
/IL
an
os
m
Zy
0
-1
IL
HIV+ve TST+ve
(mdn log2 exp)
HIV+ve Active TB TST+ve
Module score
*
81
IFN
6 9 12 15 18
HIV-ve TST+ve
(mdn log2 exp)
Bell et al. PLoS Pathogens 2016
HIV+ve patients with TB-IRIS
HIV-ve Active TB TST+ve
158
1567
1213
HIV+ve IRIS
(mdn exp log2)
HIV+ve TST+ve IRIS
18
15
in IRIS
12
9
6
r 2 = 0.97
Slope = 0.97
6 9 12 15 18
HIV-ve TST+ve
(mdn exp log2)
Bell et al. PLoS Pathogens 2016
HIV+ve patients with TB-IRIS
HIV-ve Active TB TST+ve
158
1567
1.5
IL4 & IL13
1213
*
1.0
in IRIS
0.0
0
12
r 2 = 0.97
Slope = 0.97
6 9 12 15 18
HIV-ve TST+ve
(mdn exp log2)
TB
e
IS tiv
s
-IR ac
al
TB +ve B idu
IV T iv
H ive ind
ct y
A lth
ea
H
TB
e
IS iv
t
s
-IR ac
al
TB +ve B idu
IV T iv
H ive ind
ct y
A lth
ea
H
HIV+ve IRIS
(mdn exp log2)
1
18
6
4
2
0.5
9
IFN
3
HIV+ve TST+ve IRIS
15
5
Bell et al. PLoS Pathogens 2016
Immunostaining for IRF4 in TST biopsies
Bell et al. PLoS Pathogens 2016
Conclusions
• The TST reflects molecular features at the site of active TB
• Molecular profiling of the TST reveals differences in immune responses
between groups of people and between individuals
– Donor- donor variation  ? Clinical outcome
– Gene specific differences in the TST between active & ‘latent’ TB
– HIV-1 infected patients with positive TSTs exhibit deficient IL10
responses  ? Failure of immuneregulation
– Unmasking TB-IRIS is associated with an exaggerated Th2 response
• New window on immune correlates of protection and/or pathogenesis
– Inform risk stratification, host directed therapies & vaccine design
Gillian
Tomlinson
Rob Miller
Gabriele
Jennifer Roe Pollara
Benny Chain
Lucy Bell
Keertan
Carolin Haas Dheda
www.Innate2Adaptive.com