Inflammatory Bowel Disease, Colon

Cancer and Chemoprevention

[염증성 장염에 의해 발생하는 대장암의 화학적 암예방]

부산대학교 약학대학

김 남 득 교수

1

Contents • IBD overview

• Inflammatory Bowel Disease

• Ulcerative colitis vs. Crohn’s disease

• Diagnosis

• Treatment

• Colon cancer

• IBD mice model

• Chemoprevention of IBD-related colon cancer

• Summary

2

Inflammatory Bowel Disease (IBD) • Def.: a group of inflammatory conditions of the

colon and small intestine. • The major types of IBD are Crohn's disease (CD) and

ulcerative colitis (UC). • Accounting for far fewer cases are other forms of

IBD, which are not always classified as typical IBD: • Collagenous colitis • Lymphocytic colitis • Ischaemic colitis • Diversion colitis • Behçet's disease • Indeterminate colitis

Micrograph showing inflammation of the large bowel in a case of inflammatory bowel disease. Colonic biopsy. H&E stain.

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Inflammatory Bowel

Disease - What Is It? • Inflammatory bowel disease (IBD)

results from the inappropriate and

ongoing activation of the mucosal

immune system, and this is driven by

the presence of normal luminal flora.

• Can be thought of as an autoimmune disease

• This unusual response is due to defects in:

1. the barrier function of the intestinal epithelium

2. the mucosal immune system

4

• Onset of IBD typically occurs in the second

and third decades of life.

The majority of patients experience relapse or

will progress to chronic disease.

• Two main forms of IBD are Crohn’s Disease and Ulcerative

Colitis (UC).

Susceptibility to IBD is inherited and genetic factors are

more important in Crohn's Disease than in Ulcerative Colitis.

Inflammatory Bowel

Disease - What Is It?

5

IBD: A comparison of incidence rates

Incidence Year

Crohn’s disease (patients/105/year)

Ulcerative colitis (patients/105/year)

North America 1993 6.9 8.3

2004 14.6 14.3

North Europe 1993 7.0 11.8

South Europe 1993 3.9 8.7

UK 1994 8.3 13.9

India 2000 NA 6.0

New Zealand 2004 7.6 16.5

South Korea 2005 1.34 3.08

Japan 1991 0.51 1.9

Hong Kong 2001 1.0 1.2

*Incidence rates (cases/100,000/year) *Ahuja and Tandon, J. Digestive Diseases 2010; 11; 134-147.

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IBD: A comparison of prevalence rates

Prevalence Year

Crohn’s disease (patients/105)

Ulcerative colitis (patients/105)

North America 2001 162 246

2004 201 238

North Europe 1987 54 161.2

South Europe 1992 40 121

UK 1996 214 122

India 2000 NA 44.3

New Zealand 2004 104 155.2

South Korea 2005 11.2 30.9

Japan 1998 13.5 -

1991 - 18.1

Hong Kong 2006 - 7.0

1994 1.3 -

*Prevalence rates (cases/100,000) *Ahuja and Tandon, J. Digestive Diseases 2010; 11; 134-147.

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Impact of Race and Ethnicity

A

Whites had stronger family history of IBD and colorectal cancer.

African Americans with CD had higher incidence of arthritis.

Disease severity similar across all groups.

pANCA served as a sensitive maker for Mexican Americans as 100% with

UC were positive compared with 40% in whites

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• Incidence of IBD in South Korea:

– Prevalence for UC in South Korea: quadrupled from

7.57/105 individuals in 1997 to 30.9/105 individuals in

2005

– Onset: 30% between 10 and 19 years of age

– Young children: <2%

– Peak age of onset: 20s & 30s, again in 60s (smaller

second peak)

– Adjusted prevalence rates of CD and UC per 100,000

individuals were 11.2 and 30.9, respectively

– The ratio of the incidence rates of UC to CD was

decreasing in Korea CD showed a trend of more

accelerated incidence rate compared to UC.

– Male predominance in the incidence of CD.

*Kim and Kim, Gut and Liver, 2010; 4(1) 1-14. 9

• Scope of disorder (United States)

– 700,000 physician visits per year

– 100,000 hospitalizations per year

– CD accounts for two thirds

• Long-term outlook

– Chronic, lifelong disease

– Acute flare-ups alternating with remission

– Complications and increased mortality

– Surgery for 50% to 80% of CD patients

Calkins BM. Digestive Diseases in the United States: Epidemiology and Impact. Bethesda, Md: National Institutes of Health; 1994.

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• CD is a condition of

– Chronic inflammation potentially involving any

location of the GIT from mouth to anus.

– It is a lifelong disease arising from an interaction

between genetic and environmental factors

• UC is an inflammatory disorder that affects the

rectum and extends proximally to affect

variable extent of the colon.

Crohn’s Disease vs. Ulcerative Colitis

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Crohn’s Disease vs. Ulcerative Colitis • Crohn’s and UC differ in the location and nature of the inflammation.

• Crohn's Disease: can affect any part of the GI tract

(most cases start in the terminal ileum)

• Ulcerative Colitis: restricted to the colon and rectum

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Comparison of the distribution patterns of Crohn’s disease and ulcerative colitis, as well as the different conformations of the ulcers and wall thickenings.

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Crohn’s Disease (크론씨 병)

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Chronic Ulcerative Colitis(만성 궤양성대장염)

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Crohn’s Disease vs. Ulcerative Colitis

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Clinical features of CD

• Abdominal pain

• Diarrhea

• Fever

• Malabsorption

• Colonic bleeding

• Intestinal obstruction

• Fistulas

• Small bower cancer

• Colorectal cancer

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Clinical features of UC • Mild colitis • Moderate colitis • Severe colitis • Extraintestinal manifestations: a. arthritis b. Uveitis c. Erythema nodosum d. Pyoderma gangrenosum e. Primary sclerosing

cholangitis • UC and colorectal cancer: colorectal epithelial dysplasia colorectal carcinoma

• CRC: UC > CD

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ETIOLOGIC HYPOTHESES

• Persistent infection

– Mycobacteria

– Helicobacter sp.

– Listeria

– Toxigenic E. coli

• Defective mucosal integrity

– Altered mucus

– Increased permeability

– Cellular starvation

– Impaired restitution

• Dysbiosis (장내미생물교란)

– Protective bacteria

– Aggressive commensals

• Dysregulated immune

response

– Loss of tolerance

– Aggressive cellular activation

– Defective apoptosis

– Antineutrophil cytoplasmic

antibodies are found in 80%

of patients with UC

• Family history of IBD – More common for CD than for UC

– CD: chromosome 16 with NOD2/CARD15 locus

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Pathogenesis of IBD

• 항염증: IL-10, TGFβ

• 염증유발: - Th1 cytokines (IL-12, IL-18, IFN-γ, TNFα, IL-2, IL-1β, TL1A)

- Th17 cytokines (IL-23, IL-17)

- Th2 cytokines (IL-4, IL-5, IL-13)

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New IBD genetics: common pathways with other diseases

Gut 2011;60:1739e1753 23

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IBD: Systemic Complications

*Higher incidence in women.

Eye

inflammation*

Liver and bile duct inflammation

Skin lesions

Arthritis and joint pains

Kidney stones

Growth failure in children

Lower

bone density*

Subfertility*

Gallstones

Ovaries

Uterus

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(말초 관절염)

(공막염) (상공막염)

(천장골염)

(강직성 척추염)

(아프타성 구내염)

(결절성홍반) (괴저성 농피증)

(포도막염)

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Diagnosis of IBD Diagnosis is generally done by:

an assessment of inflammatory markers in stool

a colonoscopy (to allow for a visual diagnosis)

a biopsy of pathological lesions

non-specific: can also use blood test (surrogate markers for

inflammation, e.g. ESR, nutritional deficiencies)

Diagnosis of non-specific IBD is only made once all other

possibilities have been ruled out, such as:

Infection

Ischemia

Physical damage

Allergies

Colon Cancer

NSAID enteropathy

(ESR, 적혈구침강속도)

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Treatment of IBD

• Goals of therapy

– Induce and maintain remission.

– Ameliorate symptoms

– Improvements quality of life

– Adequate nutrition

– Prevent complication of both

the disease and medications

Adapted from: Hanauer et al, Am J Gastroenterol 2001; 96: 635 28

Drug therapy for Crohn’s disease - 2008

First line therapy

5-ASA (mesalamine)

balsalazide

budesonide

antibiotics

(metronidazole,

Cipro, rifaximin,

amoxicillin,

minocycline,

tetracycline)

Immunomodulators/

Second line therapy

corticosteroids

budesonide

azathioprine/6-MP

methotrexate

Biologic Therapy

infliximab

adalimumab

certolizumab pegol

natalizumab

Investigational

Immunomodulators

mycophenolate mofetil

leflunamide

FK 506

thioguanine

stem cell transplant

Biologics - in development

mesenchymal stem cells

abatacept

thalidomide

anti IL-12 (ABT-874)

Trichuris suis

probiotic therapy

visilizumab (anti-CD3)

Adacolumn (leukocytopharesis)

golimumab

fontalizumab

Nutritional therapy

elemental diet

TPN

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UC-약물치료(질병관리본부) 메살라민(Mesalamine) : 경증에서 중증의 궤양성 대장염을 치료하고 궤양성 대장염의

증상을 완화시키기 위해 사용. 국소적인 메살라민(Mesalamine)으로 좌약형과 관장형이

있으며 좌약형은 직장에 염증이 있을 때만 사용할 수 있고, 관장형은 하행결장이 시작

되는 부분까지만 약물이 도달하므로 하행결장 이후에 있는 병변에만 사용할 수 있음(예:

아사콜(Asacol)).

발살라자이드(Balsalazide) : 메살라민과 유사한 약효를 가진 전구약물로, 증상을 완화

시키는데 좋은 효과를 보임. 보통 활성기에서 호전된 궤양성 대장염 환자의 유지

(maintainance)요법에 사용됨.

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코르티코스테로이드 : 활성기 치료에 있어서는 메살라민보다 효과적임. 그러

나 유지요법으로 장기간 사용하는 것은, 치료효과보다 부작용이 더 많이 발

생할 수 있어서 권장되지 않음.

궤양성 대장염 환자의 약 50%는 아미노살리실레이트(Aminosalicylate)와 같

은 항염증제에 치료 효과를 보임. 활성기에는 코르티코스테로이드(예: 프레

드니손)가 중요한 치료제이나 이로 인해 체중 증가, 여드름과 같은 부작용이

생길 수 있으므로 주의해야 함. 심한 경우 병원에 입원해야 하고, 코르티코스

테로이드의 정맥 주입이 필요하다. 또한 과도한 출혈 시 수혈이 필요함

Summary of Standard Therapy Induction of

Remission

Maintenance of

Remission

Adverse Effects

Steroids Established 70-

90%

Ineffective Yes

5-ASA

(Mesalamine, 5-aminosalicylic

acid)

Minor effect Conflicting

evidence

Yes

Antibiotics No No

Immune

Suppresants

Established

55%

Established Yes

Methotrexate Established Not

demonstrated

Yes -

teratogenic

Biologicals Established Established Yes

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The pipeline of IBD

Pre-clinical

Phase I Phase II Phase III Pre-reg. Launched

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Biologicals

Small molecules

Natalizumab/

Elan/Biogen

STA5236/Syntha

Kappaproct/

Index/Serono

OCP 6535/

Otsuka

RDP58/

Genzyme/SangStat

Lecithin/

Dr. Stremmel

Cytokine/chemokine

Adhesion molecules

Transcription factors

Anti TNF

Mucosal barrier

Phosphodiesterase IV inhibi

tion

Cell homing Cytokine release

Onercept/Serono

MLN-2/Millenium

EGF/

Hitachi-Nippon

Fontolizumab/

PDL

Basilixmab/Novartis

ABT-874/J695/

Abbott-Wyeth

Visilizumab/

PDL

CNI1493/

Pharma Science

Early pipelines not empty but specific IBD i

nformation is lacking on the plethora of ant

i-inflammatory approaches. Most such com

pounds are patented for IBD which does no

t infer IBD development intent. Adalimumab/

Abbott

Sargamostim/

Schering-Berlex

CDP-870

Celltech-UCB

Alicaforsen/

Isis

Oprelvekin/

Wyeth

Infliximab/

Centocor/Schering- Pl

ough

ADDITIONAL OTHER INDICATIONS:

Oprelvekin thrombocytopenia post-chemo (launched)

CDP-870 rheumatoid arthritis (Ph III)

Adalimumab rheumatoid arthritis (launched)

Sargamostim neutropenias post-chemo (launched)

Natalizumab multiple sclerosis (pre-reg)

Infliximab rheumatoid arthritis (launched)

psoriasis/psoriatic arthritis (pre-reg/Ph III)

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<성별 10대암 조발생률, 2009년> (2012년 2월 현재)(단위: 명/10만명)

위(80.2)

대장(60.6)

폐(56.4)

간(47.9)

전립선(29.6)

갑상선(20.8)

방광(10.3)

췌장(9.8)

담낭 및 기타 담도(9.6)

신장(9.4)

갑상선(108.2)

유방(54.1)

대장(40.0)

위(39.4)

폐(22.8)

간(16.2)

자궁경부(15.1)

담낭 및 기타 담도(9.8)

췌[장(8.0)

난소(7.2)

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2010년 암종별 사망자수 : 남녀전체 한국의 사망원인(2010)

1. Cancer (25.6%) 2. 심혈관질환

폐암 15,623

간암 11,205

위암 10,032

대장암 7,701

췌장암 4,306

담낭 및 기타 담도암 3,502

유방암 1,868

백혈병 1,618

비호지킨 림프종 1,430

식도암 1,352

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Colon Cancer is the third most common

cause of cancer-related death in the USA

36

Stages of colon carcinogenesis ~50% of US population

have adenoma(s) by age 70 years

37

Schematic of the morphologic and molecular changes in the adenoma-carcinoma sequence. It is

postulated that loss of one normal copy of the tumor suppressor gatekeeper gene APC occurs early.

Indeed, individuals may be born with one mutant allele of APC, rendering them extremely likely to

develop colon cancer. This is the "first hit," according to Knudson's hypothesis. The loss of the

normal copy of the APC gene follows ("second hit"). Mutations of the oncogene K-RAS seem to

occur next. Additional mutations or losses of heterozygosity inactivate the tumor suppressor gene

p53 (on chromosome 17p) and SMAD2 and SMAD4 on chromosome 18q, leading finally to the

emergence of carcinoma, in which additional mutations occur. It is important to note that while there

seems to be a temporal sequence of changes, as shown, the accumulation of mutations, rather than

their occurrence in a specific order, is more important.

대장 – 결장과 직장의 선암 (선종-선암종 과정)

38

대장 - 선종-선암종

39

• As many as 4 million people (including one million Americans, 23,000 Australians, and 250,000 Canadians) worldwide suffer from a form IBD.

• The cost of lost productivity to U.S. businesses due to IBD is estimated to be as much as $.8 billion a year.

• The risk of colon cancer for people with IBD increases by 0.5% to 1% yearly approximately 8 to 10 years after diagnosis.

• Also people with IBD are five times more likely to develop colon cancer than the general public.

Inflammatory Bowel Diseases (IBD, e.g., Crohn’s Disease, Ulcerative Colitis)

40

Dysplasia-Carcinoma Sequence

Adenoma Normal

Dysplasia Colitis Cancer

Cancer

41

Sporadic Colon Cancer vs.

Colitis-associated Colon Cancer

42

Contributive role of chronic inflammation in colon

carcinogenesis

43

Inflammation Dysplasia Carcinoma

Inflammatory Bowel Disease

44

Risk of CRC in IBD: Factors that Increase Risk

• Duration >8-10 years

• Extent of colitis:

• Extensive disease

• Backwash ileitis

• Family history of colon cancer

• Primary sclerosing cholangitis (1차성 강직성 담관염)

• Early age at onset of colitis

• Histologic activity

• Dysplasia at surveillance

45

Risk of CRC in IBD: Factors that Decrease Risk

• Prophylactic colectomy

• Surveillance colonoscopy

• Regular doctor visits

• Chemoprevention

• 5-ASA ?

• UDCA Yes (PSC patients)

• Folate ?

• Thiopurines No

46

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Cell death

Activated Carcinogens

Procarcinogens

Detoxification & Secretion

Metabolic activation

Healthy Cell Damaged Cell (precancerous)

Cancer Cell

Repeated damage

Chemotherapy

C h e m o p r e v e n t i o n

Initiation Promotion Progression

(1-2 days) ( >10 years) ( >1 years )

Initiated Cell

48

Animal Models of Inflammatory Bowel Disease

Human IBD

IBD represents an important chronic

disease affecting the GI tract of man

and domesticated animal species.

The 2 IBD entities in humans are Crohn’s

disease (CD) and Ulcerative colitis

(UC).

Immune mediated - responds to

immunomodulatory drugs

The pathogenesis of IBD involves: 1. Failure of immune regulation

2. Genetic susceptibility.

3. Environmental triggers (microbial flora).

4. Disruption of the mucosal barrier.

Mouse models of IBD

DSS (Dextran sulphate) colitis: Oral administration of this

sulphated polysaccharide to mice induces a self limiting

colitis

TNBS (Trinitrobenzene sufonic acid) colitis: Colonic

inflammation is induced by intrarectal administration of

TNBS dissolved in 50% ethanol

Canine IBD

Small intestine: Lymphocytic-plasmacytic enteritis (LPE),

eosinophilic enteritis and eosinophilic gastro-enteritis

(EGE)

Large intestine; Lymphocytic-plasmacytic colitis,

eosinophilic colitis, histiocytic ulcerative colitis (HUC)

(mainly PAS-positive macrophages), and regional

granulomatous colitis (mainly PAS-negative macrophages)

Organ site Species Carcinogen Endpoint measured

Mammary Gland

Rat MNU adenocarcinoma

Rat DMBA adenoma, adenocarcinoma

Lung

Mice

B[a]P adenoma

NNK adenoma

Vinyl carbamate adenoma, adenocarcinoma

Colon

Rat AOM adenoma, adenocarcinoma

Mice AOM adenoma, adenocarcinoma

Mice DMH adenocarcinoma

Mice MAMAc adenocarcinoma

Mice

AOM + DSS (inflammation)

adenoma adenocarcinoma

Carcinogenesis Models:

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50

51

New therapeutic aspects of flavones: The anticancer properties of Scutellaria and its main active constituents Wogonin,

Baicalein and Baicalin. Cancer Treatment Reviews 35 (2009) 57–68

기원

Scutellaria baicalenesis (속썩은풀)

생약명 : 황금 (黃芩)

Scutellariae Radix

속썩은풀의 주피를 벗긴 뿌리가 약용으로 사용됨

성미

청열조습 (淸熱燥濕): 열을 내리고 땀조절

사화해독 (瀉火解毒): 항염증

Effect of baicalein on AOM/DSS-

induced colon cancer model in mice

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김동환 박사과정

In vitro study

(HCT116 Human Colon Cancer Cell Lines p53 wild type, HCT116 p53-/-)

Baicalein induced cell growth inhibition

(*p<0.05, **p<0.01, ***p<0.001 vs. untreated control)

*** ***

***

* ***

***

25 C 50 100 (μM)

p53

p73

MDM2

β-actin

Wild type

Baicalein (24 h)

p21

p53

p73

p53 null type

Baicalein (24 h)

25 C 50 100 (μM)

MDM2

p21

β-actin

Baicalein induced cell growth inhibition

Con 25 50 100

p53

+/+

p53

-/-

Baicalein induced cell growth inhibition

Con 25 50 100

Annexin V

PI

p53+/+

p53-/-

14.02% 1.81%

83.11% 1.06%

18.09% 1.44%

79.97% 0.5%

18.47% 1.67%

78.88% 0.98%

19.42% 1.79%

77.77% 1.02%

9.80% 1.37%

87.91% 0.92%

4.59% 9.45%

82.87% 3.09%

8.18% 10.5%

74.79% 6.53%

15.7% 9.12%

72.93% 2.25%

Baicalein induced cell growth inhibition

Baicalein induced apoptosis through the

extrinsic pathway

25 C 50 100 (μM)

Wild type

Baicalein (24 h)

PARP

Cleavage

β-actin

25 C 50 100 (μM)

p53 null type

Baicalein (24 h)

PARP

Cleavage

β-actin

Death Receptor Pathway

Mitochondrial Pathway

Apoptotic Insults

Mitochondria

Cytochrome c

Apaf1

Pro-caspases 9

Active Caspase 9

Caspase 8 Activation

Caspase 3, 6, 7

Death Substrates

Cell Death

TNF, TL1A

TNF-R1, DR3, DR6

TRADD

FADD

Pro-caspase 8

FasL, TRAIL

Fas, DR4, DR5

FADD

Pro-caspase 8

tBID

Apoptosis: Death Receptor Pathway (Extrinsic Pathway) and

Mitochondrial Pathway (Intrinsic Pathway)

25 C 50 100 (μM)

pro-caspase-3

β-actin

p53 null type

Baicalein (24 h)

pro-caspase-8

pro-caspase-9

25 C 50 100 (μM)

pro-caspase-3

Wild type

Baicalein (24 h)

pro-caspase-8

pro-caspase-9

β-actin

DR5

TRAIL

DR5

TRAIL

Baicalein induced apoptosis through the

extrinsic pathway

25 C 50 100 (μM)

PPAR γ

Wild type

Baicalein (24 h)

p50 (NE)

p65 (NE)

iNOS

β-actin

PPAR γ

p53 null type

Baicalein (24 h)

25 C 50 100 (μM)

β-actin

iNOS

Baicalein suppressed the activation of NF-kB

through PPARγ activation

Effect of baicalein on AOM/DSS-

induced colon cancer model in mice

62

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Summary • The chronic IBDs, Crohn’s disease and ulcerative colitis have seen notable

successes culminating in the discovery of 99 published susceptibility

loci/genes (71 Crohn’s disease; 47 ulcerative colitis) to date.

Approximately one-third of loci described confer susceptibility to both

Crohn’s disease and ulcerative colitis.

• There is substantial overlap in susceptibility loci/genes between IBD and

other diseases. Presently approximately 51 IBD loci show overlap between

up to 23 different diseases.

• Need long term management with primary goal to induce then maintain

remission and prevent complications of both the disease and drugs.

• Chemoprevention uses specific natural or synthetic compounds in an attempt

to prevent, halt or reverse the process of carcinogenesis, and is an

attractive way to fight CRC development.

• Further studies are required to explore the chemopreventive effects of

diosgenin, baicalein, astaxanthin, ginseng on colon carcinogenesis in

human clinical studies.

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Further studies Molecular action mechanisms of cell cycle regulation and

inflammatory related change by baicalein

Investigate the apoptosis mechanisms both p53 wild type and

null type

PCNA and TUNNEL staining to check the inhibition of cell

proliferation and apoptosis in vivo