Investigation of DNA damage in patients with colorectal cancer and their first degree relatives

Yıl 2011, Cilt: 1 Sayı: 3, 155 - 161, 31.01.2014

Ayfer Tozan-beceren Gülden Z. Omurtag Cumhur Yeğen Semra Şardaş

Öz

Objective: Studies in all over the world have reported that genetic susceptibility and environmental factors play an important role in the formation of colorectal cancer. Overall, individuals with incidence of colorectal cancer in the first-degree relatives are about twice as likely to develop colorectal cancer as those without any family history. The rapid increase about the information on molecular and biological characteristics of colorectal cancer has shed light on the pathogenesis of colorectal cancer. Biomarkers are often utilized for the determination of increased risk of cancer. The aim of our study was to investigate the potential DNA damage using by comet assay in peripheral lymphocytes of colorectal cancer patients, their first degree relatives compared with healthy subjects.
Methods: Peripheral blood samples were taken from untreated patients diagnosed with colorectal cancer (n=26), their first degree relatives (n=26) and healthy subjects (n=18) were analyzed by comet assay. A total of 50 individual cells were screened per sample (from each slide). The length of the DNA migrated in the comet tail, which is an estimate of DNA damage, was measured with image analysis system. Mean tail %DNA for each cell was calculated as 100-Head %DNA. Each participant was interviewed with a questionnaire which covered a detailed sociodemographic attributes including variables known to induce the comet frequency. 
Results: The DNA damage was found significantly higher in colorectal cancer patients and their first degree relatives (respectively 10.45±1.50, 9.83±1.40) as compared with the control group (8.59±0.76) (respectively p <0.001, p <0.01). 
Conclusion: In this study, our results demonstrate that comet assay can be used as a biomarker for detecting DNA damage in patients diagnosed with colorectal cancer and especially in to demonstrate the damage in their first degree relatives.

Key words: DNA damage, colorectal cancer, comet assay

Anahtar Kelimeler

-

Kolorektal kanser tanısı konmuş olgularda ve birinci derece yakınlarında DNA hasarının araştırılması

Öz

Amaç: Dünyanın pek çok ülkesinde yapılan araştırmalar kalıtımsal duyarlılık ve çevresel faktörlerin etkileri sonucu kolorektal kanserlerin oluştuğuna işaret etmektedir. Kolorektal kanser hastalarının birinci derece yakınlarında kansere yakalanma riskinin diğer bireylere oranla iki kat daha yüksek olduğu yapılan çalışmalar ile gösterilmiştir. Kolorektal kanserin moleküler ve biyolojik özellikleri hakkındaki bilgilerin hızla artması patogenezine ışık tutmaktadır. Artan kanser riskinin belirlenmesi için biyogöstergelerden sıklıkla yararlanılmaktadır. Bu çalışmanın amacı, genotoksisite çalışmalarında DNA hasarının gösterilmesinde oldukça başarılı bir biyogösterge olan comet tekniği ile kolorektal kanser hastaları ve birinci derece yakınlarının lenfositlerindeki muhtemel genotoksik etkilerin sağlıklı gönüllüler ile karşılaştırılmasıdır.
Yöntemler: Henüz hiç tedavi görmemiş, yeni teşhis edilmiş kolorektal kanser hastalarından (n=26), birinci derece yakınlarından (n=26) ve kontrol grubundan (n=18) kan örnekleri toplandı. Comet tekniğinde her örnek için incelenen 50 hücrenin (slayt başına) her birinde kuyruktaki DNA yüzdesi oranlarının ortalaması “mean tail DNA %” (%DNA T) görüntüleme analiz sistemi kullanılarak hesaplandı. Yapılan anket değerlendirmelerinde sosyodemografik özellikler ve DNA hasarını etkileyebilecek faktörler göz önünde bulundurulmuştur. 
Bulgular: Kolorektal kanser hastaları ve birinci derece yakın bireylerin periferal kan lenfositlerinde comet tekniği uygulanması sonucunda elde edilen ortalama %DNA T değerleri (sırasıyla 10.45±1.50 ve 9.83±1.39) olarak saptanmış olup, kontrol grubu (8.59±0.76) ile kar- şılaştırıldığında istatistiksel olarak anlamlı bir farklılık tespit edilmiştir (sırasıyla p <0.001, p <0.01). 
Sonuç: Bu çalışmada elde edilen sonuçlar kolorektal kanser teşhisi konmuş hastalarda ve özellikle de birinci derece yakınlarında comet tekniğinin, DNA hasarını belirlemede bir biyogösterge olarak kullanılabileceğini göstermiştir.

Anahtar Kelimeler : DNA hasarı, kolorektal kanser, comet tekniği

Anahtar Kelimeler

DNA hasarı, kolorektal kanser, comet tekniği

Kaynakça

• Yardım N, Mollahaliloğlu S, Bora Başara B. Türkiyede Kanser Durumu ve Uluslararası Göstergeler İle Uyumun Değerlendirilmesi. İçinde: Türkiye’de Kanser Kontrolü. Eds. Tuncer AM, Özgül N, Olcayto E, Gültekin M. T.C. Sağlık Bakanlığı, Kanserle Savaş Dairesi Başkanlığı, Koza Matbaacılık, Ankara, 2009. p. 51-63.
• Potter JD., Hunter D. Colorectal Cancer: Epidemiology. In: Genetics of Colorectal Cancer. Eds: Potter JD, Lindor NM, Springer, USA, 2009. p.5-25.
• Kılıç S, Kömürcü Ş, Rzayev M, Özet A, Kır T, Arpacı F, Açıkel CH, Öztürk B, Oğur R, Ataergin S, Kuzhan O, Hadse M. Gata Tıbbi Onkoloji Bilim Dalı’nda izlenen hastaların bazı sosyodemografik özellikleri ve tanıları. Gülhane Tıp Derg. 2004;46(2):115-124.
• Foulkes WD, Bolduc N, Lambert D, Ginsburg O, Olien L, Yandell DW, Tonin PN, Narod SA. Increased incidence of cancer in first degree relatives of women with double primary carcinomas of the breast and colon. J Med Genet. 1996;33:534-539.
• Taşçıoğlu N, Taheri S, Saatçi Ç, Özkul Y. Gastrointestinal sistem kanserlerinde metilentetrahidrofolat redüktaz geni 677C→T polimorfizminin incelenmesi. Sağlık Bilimleri Dergisi. 2006;15(1): 41-45.
• Eraslan E, Turkay C. Kolorektal kanser etyolojisi ve predispozan faktörler. Güncel Gastroenteroloji. 2007;11:19-26.
• Polat MH, Caner M. Kolon kanserli hastalarda dermatoglifik bulgular. Ege Tıp Dergisi. 2000;39:39-44.
• Lichtenstein P, Holm NV, Verkasalo PK, Iliadou A, Kaprio J, Koskenvuo M, Pukkala E, Skytthe A, Hemminki K. Environmental and heritable factors in the causation of cancer. N Engl J Med. 2000;343(2): 78-85.
• Slattery ML, Kerber RA. Family history of cancer and colon cancer risk: the Utah Population Database. J Natl Cancer Inst. 1994;86(21):1618- 1626.
• Libutti SK, Saltz LB, Teper JE. Colon Cancer. In: Devita, Hellman & Rosenberg’s Cancer: Principles and Practice of Oncology. Eds: DeVita VT, Lawrence TS, Rosenberg SA, 8th ed, Lippincott Williams & Wilkins, Philadelphia, 2008. p.1233-1286.
• Akay C. Biyomarkörlerin Toksikolojide Kullanımı. Gülhane Tıp Dergisi. 2004;46(1):73-83.
• Au WW. Usefulness of biomarkers in population studies: From exposure to susceptibility and to prediction of cancer. Int J Hyg Environ Health. 2007;210:239-246.
• Irvine T, Scott M, Clark CI. A small rise in CEA is sensitive for recurrence after surgery for colorectal cancer. Tumori. 2007;9:527-531.
• Bonassi S, Abbondandolo A, Camurri L, Dal Pra A, De Ferrari M, Degrassi F, Forni A, Lamberti L, Lando C, Padovani P, Sbrana I, Vecchio D, Puntoni R. Are chromosome aberrations in circulating lymphocytes predictive of a future cancer onset in humans? Preliminary results of an Italian cohort study. Cancer Genet Cytogenet. 1995;79:133-135.
• Bonassi S, Hagmar L, Stromberg U, Montagud AH, Tinnerberg H, Forni A, Heikkila P, Wanders S, Wilhardt P, Hansteen IL, Knudsen LE, Norppa H. Chromosomal aberrations in lymphocytes predict human cancer independently of exposure to carcinogens. European Study Group on Cytogenetic Biomarkers and Health. Cancer Res. 2000;60:1619- 1625.
• Norppa H, Bonassi S, Hansteen IL, Hagmar L, Stromberg U, Rossner P, Boffetta P, Lindholm C, Gundy S, Lazutka J, Cebulska-Wasilewska A, Fabia-nova E, Sram RJ, Knudsen LE, Barale R, Fucic A. Chromosomal aberrations and SCEs as biomarkers of cancer risk. Mutat Res. 2006;600:37-45.
• Wogan GN. Molecular epidemiology in cancer risk assessment and prevention: recent progress and avenues for future research. Environ Health Perspect. 1992;98:167-178.
• McKenna DJ, McKeown SR, McKelvey-Martin V. Potential use of the comet assay in the clinical management of cancer. Mutagenesis. 2008;23: 183-190.
• Pirc-Danoewinata H, Bull JP, Okamoto I, Karner J, Breiteneder S, Liebhardt A, Budinsky A, Marosi C. Cytogenetic findings in colorectal cancer mirror multistep evolution of colorectal cancer, Wien Klin Wochenschr. 1996;108:752-758.
• Gebhart E, Romahn R, Schneider A, Hoffmann M, Rau D, Tittelbachl H. Cytogenetic studies in lymphocytes of patients with rectal cancer. Environ Health Perspect. 1993;101(3):169-175.
• Baltacı V, Sardas S, Aytac B, Cakar S, Karakaya AE. Assessment of cytogenetic aberrations and comet assay in colorectal adenocarcinomas. Tumori. 2003;89:305-310.
• Bardi G, Johansson B, Pandis N, Bak-Jensen E, Orndal C, Heim S, Mandahl N, Andrén-Sandberg A, Mitelman F. Cytogenetic aberrations in colorectal adenocarcinomas and their correlation with clinicopathologic features. Cancer. 1993;71:306-314.
• Bardi G, Sukhikh T, Pandis N, Fenger C, Kronborg O, Heim S. Karyotypic characterization of colorectal adenocarcinomas. Genes Chrosomes Cancer. 1995;12(2):97-109.
• Singh NP, McCoy MT, Tice RR, Schneider EL. A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res. 1988;175(1):184-191.
• Dhawan A, Bajpayee M, Parmar D. Detection of DNA Damage in Drosophila and Mouse. In: The Comet Assay in Toxicology. Eds: Dhawan A, Anderson D, The Royal Society of Chemistry Publishing, UK, 2009. p. 151-170.
• Collins AR. The Comet Assay. Principles, applications, and limitations. Methods Mol Biol. 2002;203:163-177.
• Moller P. Assessment of reference values for DNA damage detected by the comet assay in human blood cell DNA. Mutat Res. 2006;612:84- 104.
• Kumar V, Abbas AK, Fausto N, Aster JC. Small intestine & colon, Adenocarcinoma. Chapter 7, In: Robbins and Cotran, Pathologic Basis of Disease. 8th ed. Elsevier Saunders, Philadelphia PA. 2010.
• Johns LE, Houlston RS. A systematic review and meta-analysis of familial colorectal cancer risk. Am J Gastroenterol. 2001;96(10):2992- 3003.
• Butterworth AS, Higgins JPT, Pharoah P. Relative and absolute risk of colorectal cancer for individuals with a family history: A meta- analysis. Eur J Cancer. 2006;42:216-227.
• Migliore L, Migheli F, Spisni R, Coppede F. Genetics, cytogenetics, and epigenetics of colorectal cancer. J Biomed Biotechnol. 2011;2011:1- 19.
• Ahsan H, Neugut AI, Garbowski GC, Jacobson JS, Forde KA, Treat MR, Waye JD. Family history of colorectal adenomatous polyps and increased risk for colorectal cancer. Ann Intern Med. 1998;128(11):900- 905.
• Norppa H. Genetic susceptibility, biomarker response, and cancer. Mutat Res. 2003;544:339-349.
• Colleu-Durel S, Guitton N, Nourgalieva K, Legue F, Lévêque J, Danic B, Chenal C. Alkaline single-cell gel electrophoresis (comet assay): a simple technique to show genomic instability in sporadic breast cancer. Eur J Cancer. 2004;40(3):445-451.
• Lou J, He J, Zheng Z, Jin L, Chen Z, Chen S, Lin Y, Xu S. Investigating the genetic instability in the peripheral lymphocytes of 36 untreated lung cancer patients with comet assay and micronucleus assay. Mutat Res. 2007;617(1-2):104-110.
• McKelvey-Martin VJ, Green MH, Schmezer P, Pool-Zobel BL, De Meo MP, Collins A. The Single Cell Gel Electrophoresis Assay (Comet Assay): A European Review, Mutat Res. 1993;288:47-63.