Introduction

Objective

Methods

Results
Baseline characteristics

● A 10-fold greater reduction from Baseline was seen for tralokinumab versus placebo in the full population 
 (ratio=0.09; P<0.0001) at Week 16 (Figure 4)
● Use of rescue therapy did not impact the results

Neutralizing interleukin-13 increases skin microbial diversity: results from a Phase 3, randomized, 
double-blind, placebo-controlled trial of tralokinumab in adult patients with atopic dermatitis

Thomas Bieber,1 Lisa A Beck,2 Andrew Pink,3 Hidehisa Saeki,4 Lawrence Eichenfield,5 Thomas Werfel,6 Anders Rosholm,7 Mads Røpke,7 Amy Paller8
1Department of Dermatology and Allergy, University Hospital, Bonn, Germany; 2Department of Dermatology, Medicine and Pathology, University of Rochester Medical Center, Rochester, NY, USA; 3St. John's Institute of Dermatology, Guy’s and St. Thomas’ Hospitals, London, UK; 4Department of Dermatology, Nippon Medical School, Tokyo, Japan; 

5Departments of Dermatology and Pediatrics, University of California San Diego School of Medicine, San Diego, CA, USA; 6Department of Dermatology and Allergy, Hannover Medical School, Hannover, Germany; 7LEO Pharma A/S, Ballerup, Denmark; 8Department of Dermatology, Feinberg School of Medicine, Northwestern University, Chicago IL, USA

● A healthy skin barrier supports the growth of commensal bacteria that protect the host from 
 pathogenic bacteria and their virulence factors
● In atopic dermatitis (AD), recent studies have pointed to a lack of microbial diversity in lesional 
 and non-lesional skin
● People with AD have high levels of Staphylococcus aureus colonizing both lesional and 
 non-lesional skin1 
● Dysregulation of the skin microbiome in AD is believed to be influenced by epidermal barrier 
 disruption and Th2-driven inflammation, in which the IL-13 cytokine plays a major role 
● Tralokinumab is a fully human, high-affinity monoclonal antibody that neutralizes the IL-13 
 cytokine, and has been shown to improve signs and symptoms in adults with  
 moderate-to-severe AD2,3 

● To examine the impact of tralokinumab treatment on microbial diversity in lesional skin of 
 adults with moderate-to-severe AD from the Phase 3 ECZTRA 1 trial (NCT03131648)

Study design (Figure 1)
● Bacteria were collected from areas (5 x 10 cm) of lesional skin
● S. aureus and overall bacterial abundance was assessed in subjects in ECZTRA 1 (n=780) at 
 Baseline and Week 16 using qPCR of the femA gene and the 16S rRNA using the Ba04230906 
 and Ba04230899 assays from Thermo Fischer, respectively 
● Microbiome profiling was done in 84 subjects (59 on tralokinumab and 25 on placebo) at 
 selected sites from ECZTRA 1 at Baseline, Week 8, and Week 16
● Relative microbial abundance and Shannon diversity were assessed based on DNA sequencing 
 of 16S ribosomal RNA V3-V4 regions 
● A total of 30,276 amplicon sequence variants (ASVs) representing known taxa were identified 
 from 205 samples. After filtering for ASVs found in more than one sample, a total of 9,130 ASVs 
 were used  for analysis representing 21 phyla, 468 genera and 791 species 
● Serum biomarkers were also measured (IL-13 and IL-22 in Singulex Erenna Array; CCL17 
 by ELISA)

Conclusions

Figure 2. S. aureus abundance correlates with select biomarkers at Baseline

10 -1 10 0 10 1 10 2
10 0
10 1
10 2
10 3
10 4
10 5
10 6

IL-13

IL-13 (pg/mL)

S
.a

u
re

u
s

(g
e

n
e

c
o

p
ie

s/
c

m
2
)

p<0.0001
r=0.3545

10 0 10 1 10 2 10 3 10 4
10 0
10 1
10 2
10 3
10 4
10 5
10 6

IL-22

IL-22 (pg/mL)

S
.a

u
re

u
s

(g
e

n
e

c
o

p
ie

s/
c

m
2
)

p<0.0001
r=0.4340

10 2 10 3 10 4 10 5
10 0
10 1
10 2
10 3
10 4
10 5
10 6

CCL17/TARC

CCL17 (pg/mL)

S
.a

u
re

u
s

(g
e

n
e

c
o

p
ie

s/
c

m
2
)

p<0.0001
r=0.3521

Figure 3. Correlation of change in S. aureus abundance with change in EASI score

-100 -50 0 50 100
10 -6
10 -5
10 -4
10 -3
10 -2
10 -1
10 0
10 1
10 2
10 3

Tralokinumab

Change in EASI from Baseline to Week 16 (%)

C
h

a
n

g
e

in
S

.a
u

re
u

s
a

b
u

n
d

a
n

c
e

(W
e

e
k

16
/B

a
se

lin
e

)

p<0.0001
r=0.284

-100 -50 0 50 100
10 -5
10 -4
10 -3
10 -2
10 -1
10 0
10 1
10 2
10 3
10 4

Placebo

Change in EASI from Baseline to Week 16 (%)

C
h

a
n

g
e

in
S

.a
u

re
u

s
a

b
u

n
d

a
n

c
e

(W
e

e
k

16
/B

a
se

lin
e

)

p<0.0001
r=0.430

Figure 4. Treatment with tralokinumab led to a greater reduction in S. aureus abundance in 
lesional skin relative to placebo at Week 16

Placebo Tralokinumab

Baseline Week 16 Baseline Week 16 

10 0

10 1

10 2

10 3

10 4

10 5

10 6

S
.a

u
re

u
s

(g
e

n
e

c
o

p
ie

s
/c

m
2 )

p<0.0001

p<0.0001 p=0.182

Figure 7. Relative abundance of the most dominant Staphylococcus species over time

0.0

0.1

0.2

0.3

0.4

0.5

0.6

Staphylococcus species relative to all bacteria

A
ve

ra
g

e
 o

f 
sp

e
c

ie
s 

p
e

r 
sa

m
p

le
 g

ro
u

p

0.0

0.2

0.4

0.6

0.8

1.0

Staphylococcus species relative to the genus

A
ve

ra
g

e
 o

f 
sp

e
c

ie
s 

p
e

r 
sa

m
p

le
 g

ro
u

p

S. aureus

S. caprae (CoNS)
S. capitis (CoNS)

S. epidermidis (CoNS)
S. haemolyticus (CoNS)
S. warneri (CoNS)
S. lugdunensis (CoNS)

S. argenteus

S. schleiferi (CoNS)

S. saprophyticus (CoNS)
S. pettenkoferi (CoNS)
S. cohnii (CoNS)

Other species

Placebo Tralokinumab Placebo Tralokinumab 

Baseline Week 16 Week 8 Baseline Week 16 Week 8 Baseline Week 16 Week 8 Baseline Week 16 Week 8 

Figure 5. Treatment with tralokinumab led to increased Shannon diversity

0

1

2

3

4

5

S
h

a
n

n
o

n
 D

iv
e

rs
it

y

Week 8 Baseline 

*    p<0.05 
**  p<0.01 
*** p<0.001 

Placebo 

Tralokinumab 

Figure 6. Relative abundance of the most dominant phyla and genera over time

0.0

0.2

0.4

0.6

0.8

1.0

A
ve

ra
g

e
p

ro
p

o
rt

io
n

o
f

m
a

jo
r

p
h

yl
a

a
n

d
g

e
n

u
s

Staphylococcus
Streptococcus
Lactobacillus
Brochothrix
Other Firmicutes
Corynebacterium
Micrococcus
Dermacoccus
Kocuria
Other Actinobacteriota
Moraxella
Acinetobacter
Paracoccus
Haemophilus
Photobacterium
Brucella
Other Proteobacteria
Chryseobacterium
Prevotella
Porphyromonas
Bacteroides
Other Bacteroidota
Other phyla

Bacteriodota

Proteobacteria

Actinobacterota

Firmicutes

Placebo Tralokinumab 

Baseline Week 16 Week 8 Baseline Week 16 Week 8 

Figure 1. ECZTRA 1 trial design and microbiome sample collection.

ECZTRA 1: Phase 3, randomized, double-blind, placebo-controlled trial 

Tralokinumab 300 mg Q2W

Placebo Q2W

Tralokinumab 300 mg Q2W

Tralokinumab 300 mg Q4W
Alternating with placebo

Placebo Q2W

Placebo Q2W

Tralokinumab 300 mg Q2W
Optional TCS and optional home use

Initial treatmentScreening Maintenance treatment
Patients with clinical response IGA-0/1 or EASI-75

ytefaS
follow-up

Open-label treatment

ECZTRA 1 (n=603)

ECZTRA 1 (n=199)

3:1 
randomization

Patients not achieving IGA=0/1 or EASI-75 at 16 weeks
Patients transferred from

temeraairetirccificepsfitnemtaertecnanetniam

Up to 6 weeks washout 
of AD medication
(2 weeks for TCS)

laitiniretfaW2Qgm003
 loading dose (600 mg)

2:2:1 randomization

16 weeks0–6 weeks 52 weeks 66 weeks4 8 28 weeks

AD, atopic dermatitis; EASI, Eczema Area and Severity Index; IGA, Investigator’s Global Assessment; Q2W, every 2 
weeks; Q4W, every 4 weeks; TCS, topical corticosteroid.  

  Characteristic
All randomized 

(N=802)

Skin swab (microbiome) subgroup 
(N=84)

Tralokinumab Q2W 
(n=59)

Placebo 
(n=25)

Age 

Mean (SD) 38.8 (14.1) 39.9 (13.8) 36.8 (13.1)

Sex, n (%)

Male 474 (59.1) 37 (62.7) 16 (64.0)

Female 328 (40.9) 22 (37.3)  9 (36.0)

Race, n (%)

White 564 (70.3) 48 (81.4) 22 (88.0)

Black  59 (7.4)  2 (3.4)  1 (4.0)

Asian 160 (20.0)  8 (13.6)  2 (8.0%)

IGA, n (%)

Moderate Disease 391 (48.8) 23 (39.0) 14 (56.0)

Severe Disease 407 (50.7) 36 (61.0) 11 (44.0)

EASI

Mean (SD) 32.4 (13.8)
a

35.6 (14.7) 32.8 (13.1)

SCORAD 

Mean (SD) 70.6 (12.9)
a

74.2 (13.1) 72.1 (10.5)

DLQI

Mean (SD) 16.9 (7.0)
b

17.7 (6.6)
d

18.3 (6.6)

Worst Daily Pruritus NRS (weekly average) 

Mean (SD) 7.7 (1.4)
c

8.1 (1.4) 8.1 (1.2)

Table 1. Baseline demographics and clinical characteristics for randomized subjects in parent 
study (ECZTRA 1) and in the skin swab subgroup.

S. aureus abundance
● At Baseline, S. aureus abundance was moderately correlated with IL-13, IL-22, and 
 CCL17/TARC  serum levels based on a non-parametric Spearman correlation (Figure 2)

● Patients with the greatest reduction in S. aureus abundance from Baseline to Week 16 also had 
 the greatest improvement in EASI score (Figure 3)

Microbiome diversity
● The tralokinumab group showed a significant increase in microbial diversity over time and 
 relative to the placebo group at Week 8 and Week 16 (Figure 5) 
● The results are presented as Shannon diversity index, which is a quantitative measure of how 
 many bacterial species are present on the skin and also accounts for the phylogenetic 
 relations between the different species 

● Relative abundance of major phyla and genera remained stable for patients receiving 
 placebo, while the relative abundance of Staphylococcus was reduced 47.5% from Baseline 
 for the tralokinumab group (Figure 6)

● At the species level, the overall decrease in the relative abundance of Staphylococcus was 
 primarily due to decreased relative abundance of S. aureus, from comprising almost 32% of all 
 bacteria at Baseline to less than 8% of all bacteria at Week 16 (Figure 7) 
● Relative abundance also decreased for S. argenteus, a pathogenic hemolysin-producing 
 species associated with S. aureus, from 5% of Staphylococcus at Baseline to 2% at Week 16
● In contrast, the relative abundance of commensal coagulase-negative staphylococci (CoNS), 
 such as S. epidermidis and S. capitis, were moderately increased

Microbiome 
dysbiosis 

Skin barrier 
dysfunction 

Neutralizing IL-13 cytokine 
with tralokinumab 

modulates type 2 immunity4 

Tralokinumab treatment 
shifts both inflammatory 

mediators and skin barrier 
markers towards a non-

lesional profile5 

Tralokinumab 
treatment 

increased skin 
microbial 
diversity 

IL-13↑↑

Immune
dysregulation

References
1. Ogonowska P et al. Front. Microbiol. 2021; 11:567090; 2. Wollenberg A et al. Br J Dermatol. 2020 Sep 30. doi: 10.1111/bjd.19574. Online 
ahead of print; 3. Silverberg et al. Br J Dermatol. 2020 Sep 30. doi: 10.1111/bjd.1957 3. Online ahead of print; 4. Guttman-Yassky E, et 
al. Poster presentation at AAD VMX 2021; 5. Guttman-Yassky E, et al. Late-breaking presentation at AAD VMX 2021. 

Disclosures
Thomas Bieber is an advisor/speaker/researcher for AbbVie, Allmiral, AnaptysBio, Arena, Asana Biosciences, Astellas, Bayer, 
BioVersys, Boehringer Ingelheim, Celgene, Daiichi Sankyo, Dermavant/Roivant, Dermtreat, Domain Therapeutics, DS Biopharma, 
RAPT Therapeutics (FLX Bio), Galapagos/MorphoSys, Galderma, Glenmark, GlaxoSmithKline, Incyte, Kymab, LEO Pharma, Lilly, 
L´Oréal, Menlo Therapeutics, Novartis, Pfizer, Pierre Fabre, Sanofi/Regeneron, and UCB. Lisa A Beck is an investigator and consultant 
for Abbvie, Astra-Zeneca, LEO Pharma, Pfizer, Regeneron and Sanofi, and reports consulting fees and/or honoraria from Benevolent 
AIBio, Dermtech, Incyte, Janssen, Lilly, Novartis, Principia Biopharma, Rapt Therapeutics, Regeneron, Sanofi/Genzyme and Sanofi-
Aventis. She is an investigator for Kiniksa. Andrew Pink reports personal fees and nonfinancial support from LEO Pharma, Novartis, 
and UCB; and personal fees from AbbVie, Almirall, Janssen, La Roche Posay Lilly, and Sanofi. Hidehisa Saeki is an advisor to LEO 
Pharma. Lawrence Eichenfield has served as an advisor/speaker/researcher for AbbVie; Almirall, Arcutis,  Arena, Dermira, Forte, 
Galderma Glenmark/Ichnos,,Incyte, LEO Pharma, Lilly, Novartis, Pfizer, Ortho Dermatology  Regeneron,and Sanofi Genzyme. 
Thomas Werfel has received lecture or consultancy fees from AbbVie, Almirall, Astellas, Galderma, Janssen/Johnson & Johnson, 
LEO Pharma, Lilly, Novartis, Pfizer, and Regeneron/Sanofi. Anders Rosholm was an employee of LEO Pharma. Mads Røpke is an 
employee of LEO Pharma. Amy Paller has served as an investigator for AbbVie, Anaptysbio, Incyte, Janssen, LEO Pharma, Lilly, 
Lēnus, Novartis, Regeneron, and UCB and received honorarium for consultancy from AbbVie, Abeona, Almirall, Asana Biosciences, 
Boehringer Ingelheim, Bridgebio, Dermavant, Dermira, Exicure, Forté Pharma, Galderma, Incyte, InMed, Janssen, LEO Pharma, Lilly, 
LifeMax, Novartis, Pfizer, RAPT Therapeutics, Regeneron, Sanofi Genzyme, Sol-Gel, and UCB

Acknowledgements
The ECZTRA 1 clinical trial was sponsored by LEO Pharma

Originally presented at American Academy of Dermatology (AAD) VMX, April 23-25, 2021.

● Tralokinumab treatment was associated with decreased abundance of 
 S. aureus and increased microbial diversity in lesional skin
	 - The results support that neutralization of the IL-13 cytokine contributes to improving 
  the hallmarks of AD by shifting the microbial diversity on AD lesional skin towards 
  commensal flora 

an=798; bn=785; cn=793; dn=57