Colorectal cancer (CRC)Colorectal cancer (CRC)

Professor Yaron NivProfessor Yaron Niv

Rabin Medical CenterRabin Medical CenterTel Aviv UniversityTel Aviv University

CRC CRC Colon anatomyColon anatomy The problemThe problem Clinical pictureClinical picture DiagnosisDiagnosis Staging and prognosisStaging and prognosis Etiology/pathogenesisEtiology/pathogenesis Genetic pathwaysGenetic pathways Genetic applicationsGenetic applications

CRC – the problemCRC – the problem

150,000 new cases every year in 150,000 new cases every year in USAUSA

lifetime probability is 6% lifetime probability is 6% (death = 3%)(death = 3%)

Dukes D 5-y-survival = 6-12%Dukes D 5-y-survival = 6-12% Colorectal cancer kills 55,000 Colorectal cancer kills 55,000

Americans every yearAmericans every year 33000 new cases every year in 000 new cases every year in

Israel, 1Israel, 15500 deaths00 deaths

CRC – clinical presentationCRC – clinical presentation

Men:women = 1.2:1Men:women = 1.2:1 Sporadic CRC related to age, rare < 40ySporadic CRC related to age, rare < 40y Genetic syndromes – youngerGenetic syndromes – younger Early stages – asymptomaticEarly stages – asymptomatic Average risk vs. high risk populationsAverage risk vs. high risk populations Symptoms/signs – advanced stages:Symptoms/signs – advanced stages:Abdominal pain, rectal bleeding, weight loss, iron Abdominal pain, rectal bleeding, weight loss, iron

deficiency anemia (>R), change in bowel habits deficiency anemia (>R), change in bowel habits (>L)(>L)

CRC – diagnosis (sensitivity/specificity)

CRC – diagnosis (sensitivity/specificity)

Physical examinationPhysical examination Rectal examinationRectal examination Iron deficiency anemia Iron deficiency anemia Elevated CEAElevated CEA FOBTFOBT BAE/virtual colonoscopyBAE/virtual colonoscopy CT/USCT/US Colonoscopy/sigmoidoscopyColonoscopy/sigmoidoscopy

Dukes’ StagingDukes’ Staging

Dukes 5-year-survivalDukes 5-year-survival

A 95%A 95%

B 80-85%B 80-85%

C 50-70%C 50-70%

D 6-12%D 6-12%

TNM System - 1958TNM System - 1958

T = tumor, N = nodes, T = tumor, N = nodes,

M = metastasisM = metastasis

Stage I = T1,N0,M0Stage I = T1,N0,M0

Stage IV = T1,N1,M1Stage IV = T1,N1,M1 SurvivalSurvival TreatmentTreatment Clinical trialsClinical trials Accurate communicationAccurate communication Uniform reportingUniform reporting

Grade:Grade: Well, Moderately

well, Poorly differentiated,

Mucinous adenocarcinoma

Multi-step carcinogenesisproliferation, differentiation,

apoptosis, angiogenesis

Multi-step carcinogenesisproliferation, differentiation,

apoptosis, angiogenesis

Series of mutational events Series of mutational events that release the cell from that release the cell from some level of growth some level of growth restraint, followed by waves restraint, followed by waves of clonal expansion of these of clonal expansion of these cells.cells.

Vogelgram

Genetic definitions - colorectal cancer

Genetic definitions - colorectal cancer

OncogenesOncogenes Tumor Tumor

suppressor genessuppressor genes DNA repair DNA repair

genesgenes Epigenetic Epigenetic

processesprocesses

OncogenesOncogenes Transformed proto-oncogenesTransformed proto-oncogenes Signal trasduction and regulation of gene Signal trasduction and regulation of gene

expressionexpression SRC, RAS, MYC, FOS, JUNSRC, RAS, MYC, FOS, JUN Mutation increases the activity of the encoded Mutation increases the activity of the encoded

proteins - proliferationproteins - proliferation Dominantly actingDominantly acting JCV, DNA virus encodes for T Ag, JCV, DNA virus encodes for T Ag,

transform cells of GI tracttransform cells of GI tract

Tumor suppressor genesTumor suppressor genes

Act in normal tissues to restrain Act in normal tissues to restrain growth and maintain the growth and maintain the differentiated phenotype.differentiated phenotype.

Recessively acting: one copy Recessively acting: one copy undergoes an inactivating mutation undergoes an inactivating mutation (heterozygosity - silent), the second (heterozygosity - silent), the second copy is deleted (loss of heterozygosity copy is deleted (loss of heterozygosity or LOH - no more protection).or LOH - no more protection).

Tumor specificTumor specific 5q(APC), 17p(P53), 18q(DCC,SMAD)5q(APC), 17p(P53), 18q(DCC,SMAD)

Gene silencing prevents translation and expression and may start tumorigenesisGene silencing prevents translation and expression and may start tumorigenesis

Genetic:Genetic: Mutation, deletion, insertion Mutation, deletion, insertion TSG (LOH, dominant - 2 hits) TSG (LOH, dominant - 2 hits) Oncogene (recessive – 1 hit)Oncogene (recessive – 1 hit) Epigenetic:Epigenetic: Promoter (CpG island) Promoter (CpG island)

methylation leads to loss of imprinting (LOI) methylation leads to loss of imprinting (LOI) When both alleles (maternal and paternal) When both alleles (maternal and paternal)

are methylated, imprinting patterns are lost – are methylated, imprinting patterns are lost – carcinogenesiscarcinogenesis

DNA methylation in CRC DNA methylation in CRC

Activated by DNA-methyltransferase (uses S-Activated by DNA-methyltransferase (uses S-adenosyl methionine) adds CH3 to Cytosine adenosyl methionine) adds CH3 to Cytosine followed by Guanine (CpG island)followed by Guanine (CpG island)

MLH1 hypermethylation is responsible for up MLH1 hypermethylation is responsible for up to 80% of MSI tumorsto 80% of MSI tumors

Therapeutic potential:Therapeutic potential: 1. Inhibition by 5-deoxy-azacytidine1. Inhibition by 5-deoxy-azacytidine 2. MSI-H tumors are resistant to 2. MSI-H tumors are resistant to 5FU-based chemotherapy5FU-based chemotherapy

Microsatellite instability (MSI – H) Microsatellite instability (MSI – H)

Stretches of DNA – a short motif (1-5 Stretches of DNA – a short motif (1-5 nucleotides) is repeated several timesnucleotides) is repeated several times

Mutation (germ-line or somatic) or Mutation (germ-line or somatic) or hypermethylation (somatic, epigenetic) of DNA hypermethylation (somatic, epigenetic) of DNA repair gene (MLH1)repair gene (MLH1)

Diagnosis: genotyping fluorescently labeled Diagnosis: genotyping fluorescently labeled PCR products with the use of an automated PCR products with the use of an automated sequencersequencer

A panel of 5 microsatellite markers – MSI-H = A panel of 5 microsatellite markers – MSI-H = 2 or more of them is a positive result2 or more of them is a positive result

Genomic instability in CRCGenomic instability in CRC

50% - Suppressor pathway - 50% - Suppressor pathway - chromosomal instability (CIN, chromosomal instability (CIN, LOH)LOH)

15% - Mutator pathway15% - Mutator pathway - - microsatellite instability (MSI)microsatellite instability (MSI) 35% - CpG island methylation 35% - CpG island methylation

phenotype of TSG (CIMP)phenotype of TSG (CIMP)

Colorectal cancer

*Chromosomal instability (CIN)

LOH, aneuploidyPredominantly Lt. sidedHighly differentiatedLittle lymphocytic infiltrationRarely mucinousWorse prognosis

50% 15%

MSI-H

DiploidyPredominantly Rt. SidedPoorly differentiatedLymphocytic infiltrationOften mucinousBetter prognosis

HNPCC Epigenetic silencing of MLH1

5% 10%*CpG island methylator phenotype (CIMP) – 35%Serrated adenoma pathway

Rt, women, old, BRAF+

35%

Genetic applicationsGenetic applications

Diagnosis of genetic (germ line) Diagnosis of genetic (germ line) syndromessyndromes

Diagnosis (screening) of sporadic Diagnosis (screening) of sporadic CRCCRC

Prognosis of sporadic CRCPrognosis of sporadic CRC

Distribution of CRC (%)Distribution of CRC (%)

Sporadic

HamartomatouspolyposisFAP

HNPCC

High risk

Any risk

FAPFAP

Hundreds to thousands of adenomas Hundreds to thousands of adenomas at the age of 16, CRC risk = 100% at the age of 16, CRC risk = 100% (39y)(39y)

Gardner, AAPC, TurcotGardner, AAPC, Turcot Annual sigmoidoscopy start at age Annual sigmoidoscopy start at age

10-12y10-12y

FAP, Extra-Colonic Cancers, Lifetime risk (%)

FAP, Extra-Colonic Cancers, Lifetime risk (%)

00.5

11.5

22.5

33.5

44.5

5

Periampulary

Pancreatic

Thyroid

Gastric

CNSHepatoblastom

a

Hereditary NonPolyposis Colorcetal Cancer

Hereditary NonPolyposis Colorcetal Cancer

Amsterdam II criteria: 3 1st degree relatives with CRC Amsterdam II criteria: 3 1st degree relatives with CRC (female genitalia, other GI), 2 generations, (female genitalia, other GI), 2 generations,

1 case < 50 y1 case < 50 y LOH of DNA mismatch repair genesLOH of DNA mismatch repair genes Microsatellite instabilityMicrosatellite instability Proximal colon, fast growing adenomas, better Proximal colon, fast growing adenomas, better

prognosis, relatively resistant to chemotherapyprognosis, relatively resistant to chemotherapy CRC risk 80%CRC risk 80% Colonoscopy every 2y, start age 25y or 10 years younger Colonoscopy every 2y, start age 25y or 10 years younger

the earliest casethe earliest case

HNPCC, Extra-Colonic Cancers - Lifetime Risk (%)

HNPCC, Extra-Colonic Cancers - Lifetime Risk (%)

05

10152025303540

Endometrial

Ovarian

Gastric

Urinary

Renal

Biliary

CNSSB

Stool: DNA quantitationLoktinov, Clin Caner Res 98;4:337Stool: DNA quantitationLoktinov, Clin Caner Res 98;4:337

Normal cells - apoptosis - Normal cells - apoptosis - short DNAshort DNA

Cancer cells - necrosis - long Cancer cells - necrosis - long DNADNA

17 CRC - 213317 CRC - 2133407 ng/ml407 ng/ml

28 healthy - 46928 healthy - 469 65 ng/ml 65 ng/ml

p=0.0005p=0.0005

Tumor MSI status – predictor of benefit from 5-FU-based adjuvant chemotherapy for CRC

Ribic, N Engl J Med 2003;349:247

Tumor MSI status – predictor of benefit from 5-FU-based adjuvant chemotherapy for CRC

Ribic, N Engl J Med 2003;349:247

570 patients – CRC tissue 570 patients – CRC tissue specimensspecimens

MSI-H MSSMSI-H MSS

N 95 475N 95 475

5-y-s5-y-s 78% 72% 78% 72%

No adjuvant 88% 68%No adjuvant 88% 68%

Adjuvant 71% 76%Adjuvant 71% 76%