New pharmacological approach for inflammatory bowel disease

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N

EW

PHARMACOLOGICAL

APPROACH

FOR

I

NFLAMMATORY

B

OWEL

D

ISEASE

2019

Inês Silva

1,2

_{, Rui Pinto}

2,3

_{, Vanessa Mateus}

1,2

1. H&TRC – Health and Technology Research Center, Portugal

2. iMed.ULisboa, Faculdade de Farmácia, Universidade de Lisboa, Portugal

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I

NTRODUCTION

I

NFLAMMATORY

B

OWEL

D

ISEASE

(IBD):

Include Crohn’s disease and Ulcerative colitis

IBD affects 7–10% of people worldwide, mainly of Caucasian descent.

2

1. Pithadia A, Jain S. Treatment of inflammatory bowel disease (IBD). Pharmacological Reports 2011;63:629-42 2. Spiegel B. The burden of IBS: looking at metrics. Current Gastroenterology Reports 2009;11:265–9 3. Mowat C, et al. Guidelines for the management of inflammatory bowel disease in adults - On behalf of the IBD Section of the British Society of Gastroenterology. Gut 2011 https://www.jhmicall.org/GDL_Disease.aspx?CurrentUDV=31&GDL_Disease_ID=2A4995B2-DFA5-4954-B770-F1F5BAFED033&GDL_DC_ID=D03119D7-57A3-4890-A717-CF1E7426C8BA

IBD manifests into several intestinal and extra-intestinal symptoms,

mainly related to oxidative stress, inflammation and autoimmune reaction.

3 Chronic inflammatory disease of the

gastrointestinal (GI) tract characterized by

recurrent ulceration.

1

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I

NTRODUCTION

Valatas V, et al.. The value of experimental models of colitis in predicting efficacy of biological therapies for inflammatory bowel diseases. Am J Physiol Gastrointest Liver Physiol. 2013;305:763–785 Sartor B. Mechanisms of Disease: pathogenesis of Crohn's disease and ulcerative colitis. Nature Clinical Practice Gastroenterology & Hepatology . 2006;3:390-407

Chron Disease - Th

₁

and Th

₁₇

response

Ulcerative colitis - Th

₂

response

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I

NTRODUCTION

C

URRENT

PHARMACOLOGICAL

APPROACHES

ON

IBD

1. Pithadia A, Jain S. Treatment of inflammatory bowel disease (IBD). Pharmacological Reports 2011;63:629-42 Aloi M, Nuti F, Stronati L, Cucchiara S. Advances in the medical management of paediatric IBD. Nature Reviews Gastroenterology & Hepatology 2014;11:99-108

Induce and maintain the patient in remission

however no modify or reverse

the underlying pathogenic mechanism.

1 Nonclinical studies

for

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I

NTRODUCTION

A

NIMAL

MODELS

M

ODEL

S

PECIES

M

ETHOD

OF

INDUCTION

T

IME

COURSE

D

ISEASE

LOCATION

T

YPE

OF

COLITIS

I. C

HEMICALLY

INDUCED

MODELS

TNBS

Rats, mice and

rabbits

TNBS enema

(20-30mg in 30-50% EtOH)

3 days – 8 weeks

Small intestine or

colon

Acute and chronic

DSS

Hamsters, mice and

rats

2 - 10% DSS feeding

5 days – 15 weeks/

Colon

Acute and chronic

Acetic acid

Rats

1 – 10% acetic acid enema

1 day – 3 weeks

Colon

Acute

Carrageenan

Rats, guinea, pigs

and rabbits

Variable oral dosing

1 – 4 weeks

Cecum and colon

Acute and chronic

Indomethacin

Rats

Oral or SC once or twice

< 1 - 8 days

Small intestine

Acute

(6)

I

NTRODUCTION

5. Padua, D. et al. The Role of Neuropeptides in Mouse Models of Colitis. J Mol Neurosci 2016, 59(2), 203–210. 6. Wirtz, S. et al. Chemically induced mouse models of acute and chronic intestinal inflammation. Nature Protocols 2017, 12(7), 1295–1309. 7. Sartor R. Microbial Influences in Inflammatory Bowel Diseases. Gastroenterology. 2008;134(2): 577–94

E

THANOL

is proposed to elicit a transient increase in intestinal permeability.

5 TNBS

reaches the subepithelial space and haptenate tissue and microbial proteins.

6 Mechanism of action of TNBS / Ethanol

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I

NTRODUCTION

Erythropoietin

(EPO)

is a potent stimulator of erythroid progenitor cells.

Furthermore, EPO inhibits NF-kB activation and proinflammatory gene expression in the lamina propria.

5,6

5 Cuzzocrea S, Mazzon E, Paola R, et al. Erythropoietin Reduces the Development of Experimental Inflammatory Bowel Disease. J Pharma Exp Therap 2004;311:1272–80. 6. Nairz M,Schroll A,Moschen A. Erythropoietin Contrastingly Affects Bacterial Infection and Experimental Colitis by Inhibiting Nuclear Factor-kB-Inducible Immune Pathways. Immunity. 2011;34:61–74. http://www.hammernutrition.com/knowledge/diet-for-increasing-your-natural-epo.280.html?sect=endurance-library-section

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Evaluate the influence of a new pharmacological approach with erythropoietin

in the establishment and development of inflammation associated with IBD,

through of an experimental colitis model in rodents.

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M

ATERIALS

AND

M

ETHODS

Male CD-1 mice, 30-40g in weight and 6-10 weeks of age, were housed in standard

polypropylene cages with ad libitum access to food and water in the Faculty of Pharmacy

Central Animal Facility in the University of Lisbon.

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M

ATERIALS

AND

M

ETHODS

According to Morris (1989)

9 _:



Mice were left unfed during 24h



Mice were anesthetized with Ketamine + Xilazine IP



100µl of TNBS (2,5% TNBS in 50% ethanol) was

administered through a catheter inserted into the rectum



Mice were kept for 1 min in a Tredelenburg position to

avoid reflux



EPO 500 – 1000 mg/Kg were daily administrated IP

9 Morris G, et al.Hapten-induced model of chronic inflammation and ulceration in the rat colon. Gastroenterology 1989;96:795–803

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R

ESULTS

C

HANGE

OF

BODY

WEIGHT

(%)

C

OLON

LENGTH

(cm)

0

1

2

3

4 - 2 0

- 1 0

0 1 0

2 0

3 0

4 0

T im e (d a y s )

C

h

a

n

g

e

o

f

b

o

d

y

w

e

ig

h

t

(

%

)

T N B S

T N B S + E P O 5 0 0

T N B S + E P O 1 0 0 0

E P O 1 0 0 0

E th a n o l

S h a m

p<0.0001

0

1

2

9

10

11

12 TNBS

TNBS +

EPO500

TNBS +

EPO1000

Ethanol

Sham

C

o

lo

n

l

e

n

g

th

(

c

m

)

***

_***

***

Mateus V, et al. Basic and Cinical Pharmacology and Toxicology 2016. Submitted

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R

ESULTS

F

AECAL

HAEMOGLOBIN

(µmol Hg/g feces)

A

LP

(UI/L)

EPO decreased the intestinal bleeding

EPO seems to decrease the colonic damage

****

0

2

4

6

8

10

12

14 TNBS

TNBS +

EPO500

TNBS +

EPO1000

Ethanol

Sham

F

e

c

a

l

h

e

m

o

g

lo

b

in

(



m

o

l

H

g

/g

f

e

c

e

s

)

0

10

20

30

40

50

60

70

80 TNBS

TNBS +

EPO500

TNBS +

EPO1000

Ethanol

Sham

A

L

P

(

IU/

L

)

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0 5 0 1 0 0 1 5 0 2 0 0 IL -1 0 ( p g /m l) T N B S T N B S + E P O 5 0 0 T N B S + E P O 1 0 0 0 E P O 1 0 0 0 E t h a n o l S h a m 0 1 0 2 0 3 0 4 0 5 0 M P O ( n g /m l) T N B S T N B S + E P O 5 0 0 T N B S + E P O 1 0 0 0 E P O 1 0 0 0 E t h a n o l S h a m

R

ESULTS

IL-10

(pg/mL)

MPO

(ng/mL)

****

0

100

200

300

400 TNF-



IL-1



TNBS

TNBS+

EPO500

TNBS+

EPO1000

Ethanol

Sham

P

ro

-i

n

fl

a

m

a

to

ry

c

y

to

k

in

e

s

(

p

g

/m

l)

TNF-α

and

IL-1β

(pg/mL)

EPO

presented an anti-inflammatory effect

****

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TNBS

TNBS+EPO500

TNBS+EPO1000

EPO1000

Ethanol

Sham

R

ESULTS

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0

5

10

15

20

25

30

35

40 TNBS

TNBS +

EPO500

TNBS +

EPO1000

Ethanol

Sham

A

L

T

(

IU

/L

)

R

ESULTS

ALT

(UI/L)

C

REATININE

(mg/dL)

U

REA

(mg/dL)

**

****

***

****

0

10

30

40

50

60

70 TNBS

TNBS +

EPO500

TNBS +

EPO1000

Ethanol

Sham

Ur

e

a

(

m

g

/d

l)

0.00

0.05

0.10

0.15

0.20

0.25

0.30 TNBS

TNBS +

EPO500

TNBS +

EPO1000

Ethanol

Sham

C

re

a

ti

n

in

e

(

m

g

/d

l)

EPO

decreased the extra-intestinal markers

0 2 4 0 4 2 4 4 4 6 4 8 5 0

H

e

m

a

to

c

ri

t

(%

)

T N B S + E P O 5 0 0 T N B S + E P O 1 0 0 0 E P O 1 0 0 0 S h a m

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D

ISCUSSION

/ C

ONCLUSION

EPO promoted a beneficial effect on:

body weight, faecal haemoglobin, ALP, MPO, TNF-α, IL-1β, IL-10, urea, creatinine, ALT and histopathology

• Dose-dependent effect was observed;

• Hematocrit level was normal.

http://www.hammernutrition.com/knowledge/diet-for-increasing-your-natural-epo.280.html?sect=endurance-library-section