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MLH1 and HNPCC. March 29, 2005 Tammy Jernigan. http://www.macgn.org/cc_hnpcc1.html. Type I Site Specific Colorectal Cancer. Type 2 Colorectal Cancer Extracolonic Cancers: -Endometrial (80%) -Ovarian -Stomach, Urologic.
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MLH1 and HNPCC March 29, 2005 Tammy Jernigan http://www.macgn.org/cc_hnpcc1.html
Type I Site Specific Colorectal Cancer Type 2 Colorectal Cancer Extracolonic Cancers: -Endometrial (80%) -Ovarian -Stomach, Urologic MLH1 and HNPCCHereditary Non-Polyposis Colon CancerLynch I SyndromeSite SpecificColorectal CancerMLH1 and HNPCCHereditary Non-Polyposis Colon CancerLynch I SyndromeSite SpecificColorectal CancerHNPCCHereditary Non-Polyposis Colon Cancer Lynch Syndrome http://my.webmd.com/hw/colorectal_cancer/nord953.asp
~5% Colorectal Cancers Autosomal, dominant inheritance (follows Tumor-Suppressor 2-Hit rule) 80-90% Penetrance 40% of these are related to the gene MLH1, a DNA Mismatch Repair gene 90% of the tumors related to MLH1 deficiency show microsatellite instability (MSI or MIN) HNPCCHereditary Non-Polyposis Colon Cancer http://www.healthsystem.virginia.edu/uvahealth/hub_cancer/hnpcc.cfm http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=120436
Treatment for HNPCC Colectomy • Removal of entire colon and reconnection of small intestine to the remaining rectum • Followed by the Health Maintenance Guideline http://www.kimmelcancercenter.org/kcc/HereditaryCancer/HNPCC/hnpcc.htm
Health Maintenance Guideline http://www.clevelandclinic.org/registries/inherited/hnpcc.htm
MLH1 Location • Chromosome 3 • On short arm (p) at location 21.3 (3p21.3) US National Library of Medicine http://ghr.nlm.nih.gov/gene=mlh1#name
Mutations in MLH1 • Over 300 have been characterized Common Mutations include: • Entire deletion of the exon (Codons 578-672 on short arm of chromosome 3) • Frameshift mutation (new stop codon); truncation • Insertion of nucleotides http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?CMD=search&DB=omim
MLH1 Function • One of the two most common genes related to HNPCC • One of five genes involved in DNA Mismatch Repair during cell division (MSH2, MLH1, MSH6, PMS1, PMS2) • MLH1 protein is part of a DNA repair complex that includes PMS2, which is part of the MMR system. • MLH1 deficiency results in increased spontaneous mutations throughout the genome due to dysfunction of the MMR system http://www.ohsu.edu/tech-transfer/technologies/wct_389.shtml http://www.exactsciences.com/pregen26/professionals/about_hnpcc/_index.htm
MLH1: A Mismatch Repair Gene Role of Mismatch Repair System: - Eliminate base-base mismatches - due to replication errors or - deamination (cytosine to thymidine) - Eliminate insertion-deletion loops - due to DNA polymerase pausing and template slippage Peltomaki, Human Molecular Genetics, 2001. Vol. 10, No. 7 735-740 http://saturn.roswellpark.org/cmb/huberman/DNA_Repair/mmr.html
Base-Base Mismatches • MSH2 and MSH6 bind to mispaired DNA segment. • This triggers the binding of the MLH1 endonuclease complex (includes PMS2). DNA helicase unwinds helix, daughter stand is cut, exonuclease removes many nucleotides including mismatched base. • Gap filled by DNA polymerase and sealed by DNA ligase. - E. coli studied in depth for this mechanism (methylation) Lodish et al, Figure 23-28 http://saturn.roswellpark.org/cmb/huberman/DNA_Repair/mmr.html
Insertion-deletion loops • Due to DNA polymerase pausing and template slippage during DNA replication • Slippage can occur in regions of tandemly repeated units (such as (CA)12). These regions of repeated units are known as microsatellites. The gain or loss of these units due to slippage is known as microsatellite instability (MSI or MIN). • DNA melts and reanneals incorrectly and creates a loop extending off of one strand. • Repair follows similar mechanism of base-base mismatches http://www.virtuallaboratory.net/Biofundamentals/lectureNotes/AllGraphics/slippage.jpg Peltomaki, Human Molecular Genetics, 2001. Vol. 10, No. 7 735-740 http://saturn.roswellpark.org/cmb/huberman/DNA_Repair/mmr.html
Model Organisms • Mouse • E. coli • S. cerevisiae • Drosophila
Knockout Mouse: MLH1 and PMS2 • Phenotype: - normal gestation - >80% developed tumors after one year - 7 of 8 tumors did not express APC (Adenomatus polyposis coli) - Frameshift and base substitution mutations increased dramatically in all examined tissues • Double knockout of MLH1 and PMS2 revealed similar results, supporting the idea that MLH1 nullizygosity is sufficient to inactivate MMR completely http://www.mdc-berlin.de/~gfactor/maus.jpg http://www.ohsu.edu/tech-transfer/technologies/wct_389.shtml http://www.pdg.cnb.uam.es/UniPub/iHOP/gs/122799.html
S. cerevisiae • Missense Codons from human HNPCC correlated with MLH1 and MSH2 introduced into yeast to note effects: - complete loss of function - silent polymorphisms - efficiency polymorphisms • Connection between clinical human data and yeast results • Supported feasibility of constructing genes that encode functional hybrid human-yeast MLH1 proteins http://www.bio.unc.edu/courses/2005Spring/Biol169/ • Ellison AR, Lofing J, Bitter GA.
Cancer Treatment Summary • Colectomy (presently used) • Hybrid human-yeast MLH1 gene introduction • Drug development that would target MLH1 -/- cells and cause cell death
Sources • http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=120436 • http://www.md.ucl.ac.be/entites/mint/intr/hainaut/dossierprojet/dossierdocsem/canccolrectgen/canccolrectgen.html • http://search.cancer.org/search?client=amcancer&site=amcancer&output=xml_no_dtd&proxystylesheet=amcancer&restrict=cancer&q=HNPCC • http://www.clevelandclinic.org/registries/inherited/hnpcc.htm • http://www.virtuallaboratory.net/Biofundamentals/lectureNotes/AllGraphics/slippage.jpg • http://www.ohsu.edu/tech-transfer/technologies/wct_389.shtml • http://www.pdg.cnb.uam.es/UniPub/iHOP/gs/122799.html • http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?CMD=search&DB=omim • http://saturn.roswellpark.org/cmb/huberman/DNA_Repair/mmr.html • http://www.exactsciences.com/pregen26/professionals/about_hnpcc/_index.htm • http://www.healthsystem.virginia.edu/uvahealth/hub_cancer/hnpcc.cfm • http://www.kimmelcancercenter.org/kcc/HereditaryCancer/HNPCC/hnpcc.htm • http://ghr.nlm.nih.gov/gene=mlh1#name • http://www.mdc-berlin.de/~gfactor/maus.jpg • http://www.bio.unc.edu/courses/2005Spring/Biol169/ • http://my.webmd.com/hw/colorectal_cancer/nord953.asp • Ellison AR, Lofing J, Bitter GA., Functional analysis of human MLH1 and MSH2 missense variants and hybrid human-yeast MLH1 proteins in Saccharomyces cerevisiae. Hum Mol Genet. 2001 Sep 1;10(18):1889-900. • Lodish et al. Molecular Cell Biology. Ed. 5, Figure 23-28. W.H. Freeman Co, NY, 2004 • Peltomaki, Paivi. Deficient DNA mismatch repair: a common etiologic factor for colon cancer. Hum Mol Genet. 2001, Vol. 10 No. 7 735-740 • Satya Narayan and Deodutta. Roy,Role of APC and DNA mismatch repair genes in the development of colorectal cancers. Molecular Cancer 2003, 2:41