DIAGNOSTIC AND FOLLOW UP OF FAP, HNPCC AND SPORADIC COLORECTAL CANCER

4. A GENETIC MODEL OF CARCINOGENESIS

5. GENETIC PATHWAYS OF CANCER PATHOGENESIS Loss of heterozygosity (LOH) A complete loss of function of two corresponding genes (alleles) – deletion. Most frequent areas are 17p and 18 where tumor suppressor genes are located Defect in mismatch repair (MMR) genes The changes in short seguences of bases that are repeated and are found in the entire genome of cancer DNA, wich is damaged in microsatellite regions – microsatelitte instability (MSI) Formation of new alleles at microsatellite loci in tumor DNA

6. MSI COLORECTAL CANCER Somatic mutation MMR genes – sporadic MSI colorectal cancer Germ line mutation MMR genes – hereditary MSI colorectal cancer (Hany at al, 2001) 90% of hereditary cancers are MSI, and 10% of sporadic cancers are MSI Classification of MSI: MSI-H (2 locus), MSI- L (1 locus) and MSI-S (no MSI) RER+: defective MMR genes in at least 2 loci

7. MOLECULAR SCREENING Using 5 microsatellite markers: BAT26, BAT 40, DS 123 and two from literatute MSI causes an expresion of tranformated tumor growth factor on the cell surface (Markowitz at al, 1995) Bethesda criteria: cancers that meet Amsterdam criteria should be tested on MSI

8. CLASSIFICATION OF CANCERS ACCORDING TO THE HEREDITARY INFLUENCE Sporadic colorectal cancer (60%) Familial colorectal cancer (30%) Hereditary colorectal cancer (10%) The American Cancer Society : averige, moderate and high risk for development of colorecatl cancer

9. INHERETED POLYPOSIS SYNDROMES The first report in 1864 and 1873 Adenomatous and hamartomatous polyposis syndromes Lockart-Mummery (1925)- prophylactic examination of family members at risk and colectomy in affected individuals redused the frequency of colorectal cancer The St. Mark’s Hospital Polyposis Registry was founded on this bases- the first polyposis regester in the world

11. GENETIC TESTS Linkage testing, in vitro protein synthesis (IVPS) testing and identification of the specific mutation DNA is obtaned from white cell of peripheral blood Direct DNA sequencing-to detrmine the precise mutation Germline mutation of APC-carriers-surveillance by regular endoscopies, and non-carriers-exclude from a surveillance programme Correlation between the site of mutation in the APC gene and severity of intestinal polyposis Mutation at codon 1309-develop adenomas more frequently and at earlier age

13. MANAGEMENT OF A NEW PATIENTSt. Mark's Hospital protocol (2008.) Confirm diagnosis by histopathology, colonoscopy with dye spray, detailed family pedigree through Registry-individuals at risk Obtain consent for DNA testing, draw blood, monitor processing for use with at risk individuals, arrange consultation with Geneticist The early appearance of clinical features of FAP-recommendations for screening beginning at puberty-medical benefit for 50% of children who will be found not to be mutation carriers-can be spared to necessity of unpleasent and costly annual sigmoidoscopy

14. TREATMENT OF THE LARGE BOWEL Colectomy with ileorectal anastomosis (IRA) Indications: young patients (15-25 years), low density polyposis phenotype (< 1000 polyps overall), low density genotype<5 rectal polyps, concurent desmoid disease or paliative operation Folow up after treatment: 6 monthly flexible sigmoidoscopy and remove polyps >5 mm or if confluent, and consider chemoprevention with Sulindac 150 mg BD or celecobix, a selective cyclooxigenase-2 inhibitor, and consider pouch conversion Patients with IRA are at risk for rectal cancer

15. TREATMENT OF THE LARGE BOWEL Restorative proctocolectomy Older new patients (over 25 year old), conversion of ileorectal anastomosis in patients over 35 years, high density polyposis phenotype >1000 polyps overall, regardless of age, high density genotype and some patients with rectal cancer The main premise: a pouch procedure minimizes the risk of rectal cancer Folow up after treatment: annual flexible examination of pouch. Pouch biopsy, blood tests (particularly B12 an serum folic acid

16. TRETMENT OF THE LARGE BOWEL A proctocolectomy with end ileostomy A permanent stoma should not be considered unless there is low rectal cancer, sometimes when there is desmoid disease or when the anal sphincter is weak

17. MANAGEMENT OF AT RISK RELATIVES A detailed history of family Relatives at risk inheriting FAP should be identified, informed of their risk and offered screening They should understand the implications and application of advanced predictive DNA testing To obtain consent for DNA testing and to impart the results If predictive DNA test is negative-a full explanation must be given before the patient can be discharged. If the causative mutation in the family is unknown-a clinical screening programme should be advised Yearly flexible sigmoidoscopy at ages 14,15,16,17,18 and 19 Five yearly colonoscopy with dye spray at age 20 with flexibile sigmoidoscopy in the intervening years

18. OUR STUDY University Clinical Center Tuzla University School of Medicine,LJubljana Genetic testing for loss of heterozygousity (LOH)- intragenic marcers for tumor supresor genes NM23, p53, APC, RB1, DCC1 i DCC2 microsatellite instability (MSI) – mononucletide and dinucleotide marcers – BAT25, BAT40, BAT26, TP53 i DS123 direct seguencing Colonoscopy and chromoendoscopy

19. FAP-studies Hamamatsu University School of Medicine-Yapan ( Baba et al., 1994) 28 families, 50 patients 6 first degree relatives FAP patients germ line mutations APC gene, codon 1309 (17%), codon 1061 (8,5%) University clinical center Tuzla 10 families, 17 FAP patients, LOH APC and RB1; MSI on BAT 25 and BAT 40 1 first degree relative FAP patient germ line mutation APC gene, codon 1309

20. Follow up after treatment according to the St. Mark´s hospital protocol Colectomy with ileorectal anastomosis Endoscopy with dye spray every 6 months Adenomas tubulare were developed in 12 patients with low grade displasia, size < 10 mm Polypectomy were performed 2 patients preapered for surgery – Restorative proctocolectomy

22. GENETICS OF HNPCC SYNDROME Genetic disorder is common for both variant of HNPCC syndrome. Autosomal dominant inheritance, in around 50% of cases there is a chance of passing the mutated gene to each children Carriers of genetic mutations have an 80% chance of developing cancer There is genetic linkage between cancer occurrence and chromosome 2p and 3p where are located MMR genes Mutations in these genes appear to account for 90% of all known HNPCC families

23. THE REVISED BETHESDA GUIDELINES Those who meet the Amsterdam criteria should be tested on MSI 90% of HNPCC are MSI – the result of inactivating of MMR gene alleles. One mutation is inherited and MSI folows the mutation of the second alleles. If positive, a search for constitutional mutations of MMR genes should be recomended by the method of direct sequencing The benefit of these Guidelines and HNPCC testing should be directed at the relatives Approximatelly 50% HNPCC families cannot be genetically determined – to continue reaserching

24. DIAGNOSIS OF HNPCC SYNDROME Families meeting Amsterdam criteria (I, II or revised) are very likely to have HNPCC syndrome: at least three members of the femily have colorectal cancer, and two of them are first degree relatives; colorectal cancer has developed in more than one generation of the HNPCC family; at least one family member with colorectal cancer was diagnosed at age 50 years or younger; when two of the following cancers are diagnosed in the family: endometrium, ovary, stomach, hepatobiliary tract, smoll bowel, ureter, or two or more primary colorectal cancers.

25. HNPCC HNPCC is not polyposis disorder But, affected individuals do form colonis adenomas There is circumstential evidence that these adenomas are precursor lesions to malignancy But, adenoma-carcinoma sequence in HNPCC may be as short as 2 to 3 years, even 6 months There are interval cancers -gene carriers develop "de novo“ cancers

26. CANCER SCREENING GUIDELINES FOR HNPCC For individulas with HNPCC: every 1 to 3 years make a colonoscopy beginning ages 20-25, alternatively, consider surgical removal of the colon Women-have ana annual pelvic examination beginning at age 18 or with onset of sexual activity. To have ana annual endometrial biopsy beggining at age 25-35 Have an urine test yearly to check for the presence of blood, regular urine cytology tests to look for cancer cell in the urine Guidelines for screening individuals at risk (those who are close relatives of individuals with HNPCC, but have not themselves had a cancer)- the same as for individuals with HNPCC

27. SURGICAL TREATMENT If colorectal cancer occur-surgical resection must be no less than a subtotal colectomy-entire colonic mucosa is at risk for malignant transformation Prophylactic surgery- individuals who have germline mutations are given the option of prophylactic subtotal colectomy or lifetime colonoscopy screening

28. CANCER SCREENING vs PROPHYLACTIC SURGERY Gene carriers may not have confidence in the efficacy of colonoscopy-consider prophylactic surgery when the patient‘s quality of life is worsened by a severe cancer phobia

29. GENETIC COUNCELING Historically, the first example of genetic counceling in HNPCC took place in 1895-Aldred Warthin explored the family history of an patient who died of cancer Genetic counceling must be medically oriented, the cancelour will make use of all availiable diagnostic, preventive, and therapeutic regimens avalilable. Must be psychodinamically oriented The cancelour must appraise the full extent of emotional factors, and must be sympathetic to them, providing an empathetic listening ear, and must provide accurate genetic information when the patient is emotionally capable of grasping its significance

30. HNPCC study University Clinical Center Tuzla, 2000.-2008. 480 colorectal cancers, 50 colorectal cancers-genetic testing LOH APC 84,2% MSI-H (BAT40-84%) for the chromosome 2p hMSH2 HNPCC n=7- 15% , Amsterdam criteria – 23 HNPCC families, 28 patients 48 members (10%) Ponz, 1742 -17%; Aoltonen, 509-12% Boland CR.Decoding HNPCC.N Engl J Med.2006,354.2817-7 There are many chalenges, mostly technical-genetic testing, that limit the ability to make diagnosis.

31. The greatest number of patients with sporadic colorectal cancer and HNPCC displayed Astler-Coller B2 and C1 spread levels By diagnosing HNPCC it was able to do review of surgical treatment and To test and follow up relatives as the persons with high risk for development of colorectal cancer

32. SPORADIC COLORECTAL CANCER Genetic tests LOH – APC 63%, NM23 60,6%, p53 u 57,5%, dcc2 41% and DCC1 38,2% MSI - high for 2-3 locuses ( BAT25 – for microsatellite locuses on chromosome 17Q21.3, BAT26 ( chromosome 2) and BAT40 (chromosome 1p13.1) 5-year surveillance rate: 92 -26,3%

33. CONCLUSIONS The latest genetic informations could be clinically significant in the future in the prevention of colorectal cancer, pre-symptomatic cancer diagnosis, selection of patients for the most suitable treatment and evaluation of the malign potential (Baba, 1997) Colorectal cancer screening program for population with everige and high risk Develop colorectal cancer, HNPCC and FAP family Registries

34. CONCLUSIONS Identification of population at high risk of developing colorectal cancer – carriers of germ line mutations Colonoscopy should be used as screening procedure Genetic counseling should be an integral component of the management of hereditary cancer (Lynch at al, 1996) Genetic therapy will probably be used in the future as the moste optimal type of the therapeutic treatment