On recent advances of colorectal carcinoma, basically focusing on molecular pathogenesis

1. Recent advances on Colo-rectal
carcinoma
Dr. Samriddhi Karki
1st year Resident
Department of Pathology

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Introduction
Epidemiology
Etiology
MOLECULAR PATHOGENESIS
Clinical features
Morphology
Diagnostic modalities
Staging and grading
Spread and Metastasis
Treatment
Prognosis

3. INTRODUCTION
• Third most common type of cancer and second
most frequent cause of cancer-related death.
• Most curable form of carcinoma of the
gastrointestinal tract.
• Usually begins as a noncancerous polyp that
can, over time, become a cancerous tumor.
• Males and females are equally affected.
• Mean age : 62 year

4. EPIDEMIOLOGY
• Worldwide distribution
• Highest incidence rates
in
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United States
Canada
Australia
New Zeaand
Denmark
Sweden, and
Other developed countries

5. • Colorectal Carcinoma (CRC) among
Nepalese young adults accounts for a high
incidence (28%) of all CRC cases.
• Although right sided colonic cancer has
been increasing, rectum is
the commonest site.
Asian Pacific Journal of Cancer Prevention, Vol 13, 2012

6. ETIOLOGY
1.
2.
3.
4.
Dietary factors
Obesity
Smoking
Inflammatory bowel disease

7. 5. Polyps
6. Pelvic Irradiation
7. Genetic factors
▫ Hereditary non-polyposis colorectal cancer (HNPCC) Lynch syndrome
▫ Familial adenomatous polyposis (FAP)

9. Hereditary Non-polyposis colorectal
cancer ( HNPCC)
• Autosomal dominant disorder.
• Cancers at several sites
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Colorectum
Endometrium
Stomach
Ovary
Uterus
Brain
Small bowel
Hepatobiliary
Skin

10. Features of CRC in HNPCC
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Young age
Right –sided location
Mucinous features
Poor differentiation
Lymphocytic infiltration
Lack of necrosis

13. Familial adenomatous polyposis
(FAP)
• Autosomal dominant disorder.
• Numerous ( 100- 1000) colorectal adenomas.
• Mutation of the adenomatous polyposis coli (APC)
gene.
• In left untreated  colorectal carcinoma ( 100%) ,
before age 30.

14. MOLECULAR PATHOGENESIS
• TWO distinct genetic pathways .
1. APC / β- catenin pathway
▫
Associated with WNT signaling pathway and
the chromosomal instability pathway.
2. Microsatellite instability pathway
▫ Associated with defects in DNA mismatch
repair

15. APC / β- catenin pathway
• APC  tumor suppressor gene (5q21)
• Downregulate growth promoting signals
(β-catenin)
• Component of WNT signaling pathway.
• Catenins proteins found in complexes
with cadherin cell adhesion molecules.
• β-catenin participates in the WNT
signaling pathway as a growth promoting
signals.

16. WNT signaling pathway
• Network of proteins that passes signals from cell
surface receptors to the nucleus through
cytoplasm leading to expression of target genes
(transcription regulator genes- c MYC)
• Major role in controlling cell fate, adhesion, and
cell polarity during embryonic development.
• WNT signaling is also required for self –renewal
of the hematopoetic stem cells.

18. Chromosomal instability pathway
(CIN)
• Increased rate of chromosome
missegregation in mitosis .
• Due to
– Gain / loss of chromosome ( aneuploidy)
– Gross chromosomal rearrangements (GSM)

19. • Earliest event involved in CIN is APC gene
mutation ( 80%)
▫ K-RAS mutation
▫ P53 gene mutation
• Late event : DCC (deleted in colonic
carcinoma) gene mutation
• Advanced event : DPC4/ SMAD4 mutation
(18q21)

20. CIN forms the basis of
Adenoma -Carcinoma
Sequence

21. Adenoma-carcinoma sequence

22. MICROSATELLITE INSTABILITY pathway
(MSI )
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Satellite DNA?
Microsatellite DNA?
Why is it more liable to be unstable ?
How this instability leads to colorectal
carcinoma ?

23. Satellite DNA
• Satellite DNA is composed of
tandemly repeating DNA
( non - coding regions)
• Tandem repeats occur in DNA when a
pattern of two or more nucleotides is
repeated.
A-T-T-C-G-A-T-T-C-G-A-T-T-C-G

24. Type of DNA repeat
1. Satellite
No. of nucleotide repeat
5-200
2. Minisatellite
a. Hypervariable
b. Telomeric
3. Microsatellite
a. Monomorpic
b. Polymorphic
10-60
6
1-4

25. • Microsatellite DNA :
If the number of nucleotide repeat is 1-4
▫ Dinucleotide repeat:
 When exactly two nucleotides are repeated.
 Eg: ACACACAC
 Such regions in DNA are commonly affected in
HNPCC.

26. • Microsatellites are more prone to get unstable compared
to other neutral regions of DNA
• This instability is due to any errors , most likely error is
▫ Slippage during DNA replication .
• Such error is normally repaired by Mismatch repair
enzymes which are encoded by MisMatch repair genes.

27. Defect in MMR gene
Reduced capacity of cells to repair specific types
of DNA damage
Increased rate of mutation accumulation in
microsatellite DNA

28. Mismatch repair genes
• MSH2 (2p21) , MSH3, MSH4, MSH5, MSH6
• MLH1(3p21.3), MLH2, MLH3
• PMS1, PMS2

29. • As majority of microsatellites are located in
the non-coding region, these mutations are
generally silent.
• Some microsatellites which are present in
the coding region of the gene are involved in
the regulation of cell growth , like those
encoding
▫ Type II TGF-ß receptor
▫ Proapoptotic protein BAX

30. Defect in MMR genes
• Mutation  HNPCC
• CpG island Hypermethylation in
MMR genes  Sporadic CCR

31. CpG Island Hypermethylation
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What is CpG?
Terms like : CpG site and CpG Island?
What is methylation?
What is hypermethylation ?
How hypermethylation leads to carcinoma?

32. • CpG sites:
▫ Regions of DNA where a cytosine occurs next
to a guanine.
▫ DNA methylation occurs at these sites by an
enzyme called DNA methyltransferases.
• In humans, 80 to 90% of all CpGs are
methylated.
• This methylation results in the conversion of
the cytosine to 5-methylcytosine.

33. • The remaining 10% nonmethylated CpGs are grouped
in a cluster forming CpG
island , and is usually located
in the promoter regions
towards 5’ end.
• The unique property of CpG
island is that it is unmethylated
in the germ line.
• Methylation of CpG island
within the promoters of genes
 silencing of tumor
suppressor genes  Cancer

35. Microsatellite instability

37. MSI testing
• MSI can be detected by PCR amplification of
microsatellite loci in DNA extracted from CRC
specimens .
• Newer tests: Nucleic acid flourescence labelling,
laser scanning , flourescence PCR amplification .
• To identify the risk for hereditary cancer and
predict the outcome of CRC.
• To detect MLH1 and MSH2 germline mutations .

38. The Bethesda Guidelines
MSI testing is recommended in people with any of the following
features :
1. Cancer in families that meet Amsterdam criteria.
2. Two HNPCC- related cancers
3. A first degree relative with CRC and/or HNPCC- related
extracolonic cancer and/or colorectal adenoma diagnosed
under 40yr.
4. Right-sided CRC with an undifferentiated pattern on HPE.
5. Signet-ring cell type CRC diagnosed under 45yr.
6. Adenomas diagnosed under 40 yr.

39. CLINICAL FEATURES
• Asymptomatic for years.
• Left sided colonic carcinomas
▫ occult bleeding
▫ changes in bowel habit
▫ crampy left lower quadrant discomfort
• Right sided colonic carcinomas
▫ fatigue
▫ weakness
▫ iron deficiency anemia
(Anemia in females may arise from gynecologic causes, but it is a clinical
maxim that iron deficiency anemia in an older man means gastrointestinal
cancer until proved otherwise)

40. MORPHOLOGY
• 50%  rectosigmoid area (involvement of
the proximal colon is increasing)
• Right-sided tumors more common in the
▫ elderly
▫ blacks
▫ patients with diverticular disease

41. GROSS
A. PROXIMAL COLON
• Polypoid:
▫ Bulky mass, well-defined/ rolled margins and a
sharp dividing line with the normal bowel.
• Ulcerative:
▫ Less elevated surface and is centrally ulcerated
• These tumors rarely cause obstruction

43. B. DISTAL COLON
• Annular lesions producing “napkin – ring”
constrictions and luminal narrowing .
• These tumors can cause obstruction.

45. MICROSCOPIC
Tall columnar carcinomas looks similar.
• •All colorectalcells resembling dysplastic epithelium
as in adenomas.
• Almost all – ADENOCARCINOMAS
•secreting variable amounts of mucin.
Inflammatory infiltrations (lymphocytes, plasma cells,
eosinophils, histiocytes ) are prominent at the edge of
the tumor.
• Ranges from well-differentiated to
undifferentiated, frankly anaplastic
•masses.
Poorly differentiated tumors might form few GLANDS.
• Rarely, the tumor stroma may exhibit metaplastic
bone formation

46. Well - differentiated
Poorly differentiated

47. Mucinous
Anaplastic
Signet
Ring
Micropapillary
Oncocytic
Medullary
Serrated
Other
microscopi
c variants
Glassy
cell
Squamous
differentiation
Neuroendocrine
Trophoblastic
differentiation
Basaloid
Hepatoid

48. Mucinous adenocarcinoma
• 15 % of all CRCs
• Common in rectum.
• Microscopically :
more than 50%
extracellular mucin
• High association
with MSI
• Worse prognosis.

49. Signet ring adenocarcinoma
• Rare
• Microscopically :
more than 50%
intracellular mucin
• About one third
cases are associated
with MSI
• Worst prognosis.

50. Medullary carcinoma
Rare.
Common in proximal colon .
Occurs in elderly.
Microscopic : Sheets of malignant cells with
vesicular nuclei , prominent nucleoli,
abundant pink cytoplasm.
• Invariably associated with MSI .
• Favourable prognosis .
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51. Serrated adenocarcinoma
• 7.5% of all CRCs.
• Common in proximal colon.
• Derived from serrated
adenoma.
• Microscopic:
▫ serrated, mucinous or trabecular pattern of
growth
▫ abundant eosinophilic cytoplasm
▫ chromatin condensation
▫ preserved polarity, and
▫ no necrosis.

52. Squamous differentiation
• Common in proximal colon.
• Usually associated with glandular elements
(adenosquamous carcinoma)
• Occasionally , seen in a pure form (squamous
cell carcinoma).
• Evidence for human papilloma virus 16
involvement in the pathogenesis of some rectal
cases .

53. Trophoblastic differentiation
• Can occur focally in CRCs.
• hCG can be demonstrated
immunohistochemically in such tumor cells.
• Occasionally the entire tumor has the
appearance of a choriocarcinoma.
• This phenomenon should be distinguished from
conventional adenocarcinomas( where hCG
positivity is more common )

54. Immunohistochemical features
• Conventional adenocarcinoma of large bowel express are
MUC1 and MUC3.
• Mucinous carcinoma express MUC2.
• CRCs invariably positive for cytokeratin (CK)  positivity for
CK20 and negativity for CK7
• Positive for CEA.
• Positive for CDX2, in majority of CRCs.
• Tumor-associated glycoprotein (TAG-72) is present in 100 %
of invasive colorectal carcinoma.
• CRCs , especially poorly differentiated show loss of blood
group isoantigens and of HLA A, B, and C expression.

55. Other markers
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Villin
Cathepsin B
Neurolipin-1
SRCA2
Cadherin-17
Calretinin
Human chorionic gonadotropin (hCG)
Placental alkaline phosphatase (PLAP) ~10%
Estrogen and progesterone receptors
Racemase

56. Electron microscopic features
Prominent
microfilaments

57. Diagnostic modalities
BIOPSY
CYTOLOGY

58. Biopsy
• There is a need of POSITIVE BIOPSY before
radical surgery for CRC.
• In large lesions, several biopsies should be taken
form diverse areas.
• Biopsy from center  only granulation tissue
• Biopsy from the very periphery only
hyperplastic colonic epithelium

59. Cytology
• Its an accurate way of diagnosing CRC.
• Little practical value.
• Low-lying rectal lesions can be easily sampled.
• Brush cytology can also be performed via the
fiberoptic scope.
• It is a sensitive technique, perhaps even more so
than endoscopic biopsy, but it has not yet found
widespread acceptance.

60. Various screening modalities
• Colonoscopy
• Virtual colonography
• Sigmoidoscopy
• Fecal occult blood test
• Double contrast barium enema
• Digital rectal examination

61. Staging and Grading
• In 1937, Dukes proposed staging for rectal
carcinoma .
• In 1954, Astler and Coller proposed different
staging system.
• American Joint committee on Cancer(AJCC)
• The Union Internationale Countre Le Cancer
(UJCC)

62. Dukes’ Stage A
• The tumor involve the
wall of the bowel only.
• Treatment is surgery
to remove the tumor
and some surrounding
lymph nodes

63. Dukes’ Stage B
• The cancer extend through
the wall has not spread to the
lymph nodes.
• Colon cancer is treated with
surgery and, in some cases,
chemotherapy after surgery.
• Rectal cancer is treated with
surgery, radiation therapy,
and chemotherapy

64. Dukes’ Stage C
• The cancer has spread to
the regional lymph nodes
(lymph nodes near the
colon and rectum)
▫ C1: regional L.N
▫ C2: mesenteric B.V.
ligature
• Colon cancersurgery and
chemotherapy
• Rectalcancersurgery,
radiation therapy, and
chemotherapy

65. Dukes’ Stage D
• Spread outside of the
colon or rectum to other
areas of the body
• Treatment : chemotherapy.
• Surgery to remove the
colon or rectal tumor may
or may not be done
• Additional surgery to
remove metastases may
also be done in carefully
selected patients

66. Astler and Coller Staging
System
• Stage A
▫ Limited to mucosa
• Stage B1
▫ Involving the muscularis externa but not penetrating it
• Stage B2
▫ Penetrating through the muscularis externa
• Stage C1
▫ Confined to the bowel wall but with nodal metastasis
• Stage C2
▫ Penetrating through the wall and with nodal metastsis

67. •
TNM Staging of Colon Cancer
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Tumor (T)
T0 = none evident
Tis = in situ (limited to mucosa)
T1 = invasion of lamina propria or submucosa
T2 = invasion of muscularis propria
T3 = invasion through muscularis propria into subserosa or nonperitonealized perimuscular tissue
T4 = invasion of other organs or structures
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Lymph Nodes (N)
0 = none evident
1 = 1 to 3 positive pericolic nodes
2 = 4 or more positive pericolic nodes
3 = any positive node along a named blood vessel
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Distant Metastases (M)
0 = none evident
1 = any distant metastasis
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5-Year Survival Rates
T1 = 97%
T2 = 90%
T3 = 78%
T4 = 63%
Any T; N1; M0 = 66%
Any T; N2; M0 = 37%
Any T; N3; M0 = data not available
Any M1 = 4%

68. Microscopically, colorectal carcinoma
can be graded into
▫ I – well differentiated
▫ II- moderately differentiated
▫ III- poorly differentiated

69. Spread and Metastasis
•Common sites
▫Regional lymph nodes
▫Liver

70. Lymph node metastasis
• More common in the tumors showing
▫ poorly differentiated areas
▫ highly infiltrative pattern of growth.
• Minimum number of nodes recovered from a
surgical specimen of colorectal carcinoma
should be 14 or 15.

71. Liver metastases
• More common in the tumors showing
evidence of blood vessel invasion.

72. • Other relatively common
metastatic sites include
▫ peritoneum
▫ lung
▫ ovaries.

73. TREATMENT
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Chemotherapy
Radiotherapy
Photodynamic therapy
Radical surgery
Gene therapy

74. PROGNOSIS
• The 5-year survival rate after curative resection
for CRC ranges between 40% and 60% .
• Local recurrence and/or regional lymph node
metastases occur in over 90% of the failure cases.
• Over two-thirds of the recurrences are evident
within the first 2 years and 91% by 5 years.
• The prognosis of colorectal carcinoma is related
to a number of clinical and pathologic parameters.

75. • Category I
▫ Well supported by the literature, generally used in
patient management and of sufficient importance to
modify TNM stage groups.
• Category IIA
▫ Extensively studied biologically and/or clinically.
Prognostic value for therapy, sufficient to be noted in
pathology report
• Category IIB
▫ Well studied but not sufficiently established for
Category I or IIA
• Category III
▫ Not yet established to meet criteria for Category I or II
• Category IV
▫ Studied and shows no consistent prognostic
significance

76. Age
P
Perforation
P
Sex
M-P
Category I
CEA
seum
levels P
Obstruction
P
Prognosis
Tumor
edge
NP-P
Category III
Tumor
location
+/-
Tumor
size +/Local
extent P
Tumor
multiplicity
(S)

77. III
Presence of
neuroendocrine
cells +/-
Tumor
margins/
inflm rxn
G
Category IIA
Tumor
budding
P
Acinar
morphology
P
Vascular
invasion
P
PROGNOSIS
Microscopic
tumor type
(Mu/S/A -P,
Me -G)
Category IIB
Tumor
thickness
+/-
Pericolonic
tumor
deposits
P
Surgical
Margins
(R - P)
Perineurial
invasion
P
Category IIA

78. Claudin-1
Loss- P
Tumor
angiogenesis
P
III
Mucin
related
antigens
P
Cell
proliferation
+/-
Fascin
P
PROGNOSIS
DNA
ploidy
+/-
HLA-DR
expression
G
BCL2
protein
expression
G
IIB
hCG
expression
+/-
pRB and
P16
P

79. Category IIB
Category IIA
Category I

80. THANK YOU