genomic signatures in colorectal cancer mirco menigatti md, phd institute of molecular cancer...
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Genomic signatures in colorectal cancer
Mirco Menigatti MD, PhD
Institute of Molecular Cancer Research
University of Zurich (Switzerland)
“Highlights in Metastatic Colorectal Cancer”
Roma, 4 marzo 2011
adapted from Kemp Z et al. Hum Mol Genet. 2004 Oct 1;13 Spec No 2:R177-85.
Familial
Sporadic
Hereditay
Colorectal cancers
The colorectal tumor progression
Grady WM, Carethers JM. Gastroenterology. 2008 Oct;135(4):1079-99
(HNPCC)
Genomic instability
Alterations in mitotic-spindle checkpoint and sister-chromatid separation pathways.
Chromosomal instability (85%)
Loss of function of the DNA Mismatch Repair system (HNPCC, epigenetic silencing of MLH1).
Microsatellite instability (15%)
A genetic model for colorectal tumorigenesis.
Fearon ER, Vogelstein B. Cell. 1990 Jun 1;61(5):759-67
The genomic landscape of human colorectal cancers
Wood LD et al. Science. 2007 Nov 16;318(5853):1108-13.
analysis of exons representing 20,857 transcripts from 18,191 genes
Somatic mutations
• Drivers: causally involved in the
neoplastic process and positively selected for during tumorigenesis.
• Passengers: no positive or negative
selective advantage to the tumor.
Colorectal cancers a median 76 genes altered by point mutations.Only 14 can be considered as drivers.
Copy number changes
Colorectal cancers have a median 9 genes altered by a major copy number change
Homozygous deletions(i.e. PTEN, TP53, MAP2K4, SMAD2)
Amplifications(i.e. MYC, EPPB9, EGFR)
Leary RJ et al. Proc Natl Acad Sci U S A. 2008 Oct 21;105(42):16224-9
Alterations of signaling pathways
• WNT
• TGFβ / SMAD
• Notch
• Hedgehog
• RAS
•EGFR
• PI3K/Akt
Cancer phenotype
Genetic and Epigenetic alterations
Epigenetics
Historically, the word “epigenetics” was used to describe events that could not be explained by genetic principles.
Waddington, C.H. (1942). Endeavour 1, 18–20.
microRNAsLong non coding
RNAs
Goldberg AD et al Cell. 2007 Feb 23;128(4):635-8.
EPIGENETICSThe study of any potentially stable and inheritable change in gene expression or cellular phenotype that occurs in the absence of changes in Watson-Crick base-pairing of DNA.
DNA methylation
histone modifications
In mammals, nearly all DNA methylation occurs on cytosine bases that are located 5' to a guanosine in a CpG dinucleotide (CpG sites)
DNA methylation
DNA methylation
It is the best characterized chemical modification of chromatin.
It plays a role in many cellular processes :
• Silencing of repetitive and centromeric sequences
• X chromosome inactivation in female mammals
• Mammalian imprinting
ALTERATED LEVELS OF DNA METHYLATION IN TUMORS
HYPOMETHYLATION within the coding regions of genes are present CpG sites with low density. Most of them are normally methylated
Hypomethylation,during carcinogenesis, may lead to chromosomal instability
HYPERMETHYLATION de novo methylation of CpG islands present in ~60% of human gene promoters which are normally found unmethylated
Silencing of gene expression (depending on the density of promoter methylation)
Cell cycle control → p16
Repair of DNA damage → MLH1
Apoptosis → Dap kinase
Tumor-cell invasion → TIMP3
Growth-factor response → ER
Transcription
Silencing
HYPERMETHYLATION
MGMT
CpG unmet
CpG met
Menigatti et al Oncogene 2009
adapted from Sparmann A, van Lohuizen M. Nat Rev Cancer. 2006
Histone changes
Iorio, M. V. et al. J Clin Oncol; 27:5848-5856 2009
microRNAs
Iorio, M. V. et al. J Clin Oncol; 2009
Mechanisms of microRNA (miR) regulation
GENETIC ALTERATIONS CAUSING EPIGENETIC CHANGES IN CANCER
PML-RAR fusion protein in acute promyelocytic leukemias induces RARß2 gene promoter hypermetylation and silencing
by recruiting DNA methyltransferases to its promoter (Di Croce et al. Science, 2002)
EPIGENETIC CHANGES CAUSING GENETIC ALTERATIONS IN CANCER
MLH1 promoter hypermethylation
Mismatch repair deficiency
Mutation in genes with repetitive sequences
(BAX, TGFß RII, etc.)
MGMT promoter hypermethylation
No removal of G alkyl adducts
G to A mutations in oncogenes(KRAS) and tumor suppressor genes (p53)
32 normal mucosal samples 32 adenomas
level of expression (blue, low; red, high)
Microarray transcription profiling
NormalMucosa(no=32)
Polypoidadenomas
(n=32)
Colorectalcancers(n=25)
Colon cancercell lines(n=18)
Nor
mal
ized
inte
nsi
ty
PTPRR mRNA levels
PTPRR
• Encodes the classical transmembrane protein-tyrosine phosphatase (PTP) known as PTP, receptor type, R.
• Reversible tyrosine-specific phosphorylation of cellular proteins is a key signalling mechanism used to evoke essential cell decisions such as proliferation and differentiation and its proper regulation depends on the balanced activities of PTPs and protein tyrosine kinases (PTKs).
Re-activation of PTPRR expression
Colo205 (PTPRR silenced) SK-N-SH (expressing PTPRR)
Inpu
t %Histone code
H3K9ac H3K27me3 H3K9me3
in progress
Stem cells
miRNAs
CRC stem-like cell lines CD133+Angelo Vescovi, University of Milano, Italy
Expressed only in CRC stem-like cell lines CD133+
Silenced only in CRC stem-like cell lines CD133+
Translational Epigenetics
• Early detection
• Response to therapeutics
• Epigenetic therapy
Early detection
Aberrant methylation of some gene promoters is more common and easier to detect than mutations
Non invasive tests: serum, bronchoalveolar lavage , urine and stool
Stool-based analyses of a combination of DNA methylation markers achieving at least 85% sensitivity for cancer, 50% sensitivity for pre-cancer with 90% specificity.
sensitivity/specificity: 92% / 86%
Response to therapeutics
Patients with MGMT methylation median survival of 21.7 months (15.3 months without temozolomide therapy).
Patients without MGMT methylation median survival of 12.7 months (11.8 months without temozolomide therapy).
Epigenetic therapy
Taby R, Issa JP. CA Cancer J Clin. 2010 Nov-Dec;60(6):376-92.
Readings
Jones PA, Baylin SB. The epigenomics of cancer.Cell 2007 Feb 23;128(4):683-92.
Esteller M. Epigenetics in cancer. N Engl J Med. 2008 Mar 13;358(11):1148-59
Brena RM, Costello JF. Genome-epigenome interactions in cancer.Hum Mol Genet. 2007 Apr 15;16 Spec No 1:R96-105.
Jiricny J, Menigatti M. DNA Cytosine demethylation: are we getting close?Cell. 2008 Dec 26;135(7):1167-9.
LINKS
The Epigenome Network: www.epigenome-noe.net
Epigenetics society: www.dnamethsoc.com
DNA Methylation in Cancer: www.mdanderson.org/departments/methylation
EMBOSS CpGPlot : www.ebi.ac.uk/emboss/cpgplot
GRAZIE ROMA