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Presenter:Presenter: Venkat Rao Katkoori., Ph.D.Venkat Rao
Katkoori., Ph.D.
AuthorsAuthors:: Venkat R. Katkoori1, Xu Jia1, Tom Callens2,
Ludwine Messiaen2, Wen Wan3, Sreelatha Meleth3, Harvey Bumpers4 and
Upender Manne1.
Departments of Pathology1 and Genetics2 and Medicine3,
University of Alabamaat Birmingham;
Department of Surgery 4, Morehouse School of Medicine, Atlanta,
GA.
Prognostic Significance of p53 Codon 72Polymorphism Differs with
Race in Colorectal
Adenocarcinoma
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Normal
Epithelium
Abnormal
Crypts Foci
Early
Adenoma
Late
Adenoma
In situ
Carcinoma
Invasive
Carcinoma
Metastatic
Carcinoma
APC,
5q Loss
DCC/Smad4
18q Loss
p53, BAT-26
17p Loss
Many genes
8p Loss
Many genesCell
Adhesion
Many decades 2 - 5 years
2 - 5 years
A:A: Chromosomal Instability (CIN) Pathway (Gene mutations,
Chromosomal deletions)
K-Ras, β-CateninMMR (hMSH2)
B: Microsatellite Instability (MSI) Pathway(Mismatch Repair
Inactivation)
Multi-Step Molecular Pathways of Colorectal Multi-Step Molecular
Pathways of Colorectal CarcinogenesisCarcinogenesis
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Prognostic Molecular Markers of SporadicPrognostic Molecular
Markers of SporadicColorectal AdenocarcinomaColorectal
Adenocarcinoma
Promising Prognostic MarkersPromising Prognostic Markers: :
Microsatellite Instability &Microsatellite Instability &18q
deletions18q deletions
Candidate Prognostic Markers:p53, p27kip-1, Bcl-2, Bax, MUC1
& RPH3AL (Rabphillin-3A-like),
Potential Prognostic Makers: MUC4, MUC17, VEGF,EGFR, c-erb-2,
TGF-α, COX-2,TP, TS, DPD, 17p deletions17p deletions
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STUDY RATIONALESTUDY RATIONALE
PREMISEPREMISE: Several studies from ourlaboratory as well as
others have demonstratedthat prognostic value of molecular makers
differbased on tumor location, tumor stage and/orpatient
race/ethnicity in colorectal cancer(CRC).
AIMAIM: To identify race-specific prognosticmolecular markers in
CRCs, first we evaluatedmutational patterns of the p53 gene
andcorrelated with patient outcomes based onrace.
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Patient SelectionPatient Selection
CRC and corresponding normal tissues collected from373 patients
(137 African-Americans patients & 236 non-Hispanic Caucasians
patients).
Patients who underwent surgery with curative or palliativeintent
at UAB Hospital during 1984-1995.
We selected only patients who have not received anypre- or –post
surgery adjuvant therapy.
Patients with family or personal history were excluded. Patient
demographics, pathologic, and clinical
information was extracted from medical records andphysicians
charts
The follow-up information was collected from the UAB-Tumor
Registry
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Microsatellite Instability AnalysisMicrosatellite Instability
Analysis
Microsatellite instability was analyzed at BAT25, BAT26,
D2S123,D5S346 & D17S250 loci in 373 CRCs and marching normal
tissues
BAT-25 by ABI
Bat 25 InstableBat 25 Stable
Bat 25 InstableBat 25 Stable
BAT-25 by Light Cycler
BAT-26 by Light Cycler BAT-26 by ABI
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Microsatellite Instability & RaceMicrosatellite Instability
& Race
Similar incidence of MSI-H has been observed betweenSimilar
incidence of MSI-H has been observed betweenAfrican-American and
Caucasian patientsAfrican-American and Caucasian patients
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+1 Coding Region
Exons 1 to 11
5’ 3’
+ 1179+1
Analysis of Analysis of p53p53 in CRCs based on Race in CRCs
based on Race
A] Sequence Analysis exon 4 through 9
B] Frequency of p53 mutations Codon wise by race
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P53 mutations have been associated with poor survival of
African-American(p= 0.001) and Caucasian (p= 0.028) patients with
CRCs
African-American Patients Caucasian Patients
Survival Analysis based on p53 Status in CRCsby race
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Codon 72 polymorphism of p53Codon 72 polymorphism of p53
SNP at codon 72 of p53 results in thesubstitution of arginine
for proline
Susceptible for p53 mutations and Increaseddegradation of p53
protein
Increased tumorigenecity and oncogenic effect Uncontrolled
malignant process and Increased
invasiveness and metastasis Risk of cancer progression and poor
prognosis and Resistance to anticancer therapy
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Analysis of genotype status at Codon 72 of p53By RFLP and
Sequencing
M = Marker (100bp ladder)Lane 1 = G/G homozygous (Arg/Arg
phenotype)
(160 and 119bp)Lane 2 = G/C heterozygous (Arg/Pro phenotype)
(279bp, 160 and 119bp)Lane 3 = C/C homozygous (Pro/Pro
phenotype)
(279bp)
SequencingSequencing
PCR-RFLPPCR-RFLP
Genotype G/GHomozygous
Genotype G/CHeterozygous
Genotype C/CHomozygous
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Codon 72 Phenotypes of p53 and Race &Tumor
Characteristics
Pro/Pro phenotype is correlated with nodal metastasis tumor
gradeand higher incidence of p53 mutations in African American
patients
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Hypothesis: The proline/proline phenotypes areassociated with
aggressive tumor behavior.
Since higher frequency of proline/prolinephenotypes and these
exhibited higher incidenceof p53 mutations, we analyzed their
prognosticvalue in relation to other phenotypes(proline/arginine
and arginine/arginine) based onrace .
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Survival Analysis based on phenotypes codon 72polymorphism of
p53 in CRCs by race
African-American Patients Caucasian Patients
Pro/Pro phenotype of P53 at codon 72 is associated with poor
survival Only in African-American patients (p= 0.005), but not in
Caucasian ( p= 0.886)patients with CRCs
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Cox regression analysis to determine prognostic significance of
p53 codon 72 phenotypes
Pro/Pro phenotype is an independent prognostic marker Only in
AfricanAmerican patients with CRCs
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Mutational spectra of p53 in African-American and Caucasian
patients in relation to Pro/Pro phenotype
Pro/Pro phenotype of P53 at codon 72 is associated with higher
frequency of P53 mutations particularly disruptive type only in in
African-American patients[Differential mutational spectra in
relation to Pro/Pro phenotype]
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ConclusionsConclusions
p53 mutational pattern and prognostic value ofcodon 72
polymorphism differs with race incolorectal adenocarcinomas.
The incidence of Pro/Pro phenotype at codon 72of p53 is higher
in African- American patients andassociated with higher incidence
of p53mutations
Pro/Pro phenotype is associated with and poorclinical outcome of
African-American but notCaucasian patients with CRCs.
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Mentor: Upender Manne, Ph.D.Colleagues : Xu Jia, MD
Chandra Kumar , MD
Collaborators: Wen Wan, Ph.D.Sreelatha Meleth, Ph.D
Harvey Bumpers, MD,FACS. William E. Grizzle, MD, Ph.D.
Grant Support: RO1-CA98932-01 (NIH/NCI)2U54-CA118948-03
(NIH/NCI)
AcknowledgementsAcknowledgements