غربالگری ژرم لاین جهش Phe80Val ژن MLH1 در زنان مبتلا به سرطان تخمدان با تکنیک Tetra ARMS-PCR
محورهای موضوعی : نشانگر های زیستی پلاسما
1 - گروه ژنتیک، واحد تنکابن، دانشگاه آزاد اسلامی، تنکابن، ایران
کلید واژه: سرطان تخمدان, پروموتر, بیان ژن MLH1,
چکیده مقاله :
ﺳﺮﻃﺎن اﭘﯽﺗﻠﯿﺎل ﺗﺨﻤﺪان 90 درصد ﺑﺪﺧﯿﻤﯽ ﻫﺎي ﺗﺨﻤﺪان و 25 درصد ﺑﺪﺧﯿﻤﯽﻫﺎي ﺗﻨﺎﺳـﻠﯽ زﻧﺎن را ﺗﺸﮑﯿﻞ ﻣﯽدﻫﺪ و از آﻧﺠﺎ ﮐﻪ دﯾـرتشخیص داده می شود،ﺑﻪ ﻫﻤﯿﻦ دﻟﯿﻞ در ﺑﯿﻦ همه ی ﺑﺪﺧﯿﻤﯽ ﻫﺎي دﺳﺘﮕﺎه ﺗﻨﺎﺳﻠﯽ زﻧـﺎن از ﺑـﺎﻻﺗﺮﯾﻦ ﻣﻮارد ﻣﺮگ وﻣﯿﺮ ﺑﺮﺧﻮردار اﺳت. ﺗﻐﯿﯿﺮات microRNA که بیان ژن را کنترل میکند در این سرطان به چشم می خورد. تغییر در الگوی متیله شدن در ناحیه پروموتر ژن ها نیز از دیگر پیشامد های سرطان تخمدان است. در مطالعه حاضر نمونه های مورد آزمايش ازافراد بيمار و افراد سالم از آزمایشگاه جمع آوري گردید.جمعيت مورد مطالعه شامل 25فرد مبتلا به سرطان تخمدان و 25فرد سالم در سنین مختلف بوده است. نمونه خون افراد در ويال ٣سي سي حاوي EDTA جمع آوری و در دماي 20- درجه نگهداري شد.از روش کیت، برای استخراج DNA ژنومی از خون، استفاده شد. سپس ازجایگاه Phe80Val ژن MLH1 جهت بررسی پلی مورفیسم آن با روش ARMS-PCR Tetraاستفاده و همچنین یک جفت پرایمر جهت تایید نهایی ARMS-PCR Tetra طراحی گردید. از بین نمونه های مورد بررسی هیچکدام از نمونه ها موتاسیونی مشاهده نشد و دو عدد از نمونه ها تعیین توالی شدند و جایگاه نوکلئوتید بدون هیچ تغییری مشاهده شد.بطور کلی نتایج تحقیق حاضر نشان داد که هیچگونه ارتباطی بین موتاسیون ژن MLH1مربوط به Phe80Val و سرطان تخمدان در نمونه های حاضر وجود ندارد هرچند نیاز به بررسی روی نمونه های بیشتراز سایر مناطق ایران می باشد.
Epithelial ovarian cancer accounts for 90% of ovarian malignancies and 25% of female genital malignancies, and since it is diagnosed late, it has the highest mortality rate among all female genital malignancies. Changes in microRNA that controls gene expression are seen in this cancer. Changes in the methylation pattern in the promoter region of genes are also another occurrence of ovarian cancer. In the present study, the samples tested were collected from patients and healthy individuals from the laboratory. The study population included 25 individuals with ovarian cancer and 25 healthy individuals of different ages. Blood samples were collected in 3cc EDTA vials and stored at -20°C. A kit method was used to extract genomic DNA from blood. Then, rs63749990 of the MLH1 gene was used to examine its polymorphism using the ARMS-PCR Tetra method, and a pair of primers was designed for final confirmation by ARMS-PCR Tetra. No mutation was observed in any of the samples examined, and two of the samples were sent abroad for sequencing, and the nucleotide position was observed without any change. In general, the results of the present study showed that there is no association between the MLH1 gene mutation related to rs63749990 and ovarian cancer in the present samples, although there is a need to investigate more samples from other regions of Iran.
References
1. Flam, F., Einhorn, N., Sjovall Km, (1988), Symptomatology of ovarian cancer. Eur J Gynecol Obstet Reprod Bio. 27:53-7.
2. Stratton, J.F., Thompson, D., Bobrow L., et al. (1999), The genetic epidemiology of early- onset epithelial ovarian cancer: A population- based study Am. J. Hum. Genet. 65: 1725-1732.
3. Goodman, M.T., Howe, H.L., Tung, K.H. et al, (2003), incidence of ovarian cancer by race and ethnicity in the united states, 1992- 1997. Cancer. 97: 26-76.
4. Whitney A.S., Anil, K.S., Robert, L., Coleman,(2008), Angiogenesis as a strategic target for ovarian cancer therapy. Nat clin prac Oncol. 5(4): 194-204.
5. Oscoe Klinck A.B. Lyna Inkel. Genevie`veDufresne-Martin, Julien Gervais-Bird, et al. (2008), Multiple Alternative Splicing Markers for Ovarian Cancer. 68(3): 7-11.
6. Pal, T., Permuth- wey, J., Betts, J.A., et al. (2005), BRCAI and BRCA2 mutations account for a large proportion of ovarian carcinoma cases. Cancer, 15(104), 12. 2807- 2816.
7. Hamna, L., Adams, M, (2006), Prevention of ovarian cancer best practice and research clinical obstetrics and gynecology. 2: 339-362.
8. Dowty, J.G., Win, A.K., Buchanan, D.D., et al, (2013), Cancer risks for MLH1 and MSH2 mutation carriers. Hum Mutat. 34(3):490-7.
9. Holschneider, C.H., Berek, J.S, (2000), Ovarian cancer: epidemiology, biology, and prognostic factors. Semin Surg Oncol. 19(1): 3-10.
10. Chuchu, Zhao., Saisai, Li., Menghuang, Zhao., Haiyan Zhu and Xueqiong Zhu, (2018), Prognostic values of DNA mismatch repair genes in ovarian cancer patients treated with platinum-based chemotherapy. Arch Gynecol Obstet. 297(1): 153–159.
11. Lin, KM., et al, (1998), Colorectal and extracolonic cancer variations in MLH1/MSH2 hereditary nonpolyposis colorectal cancer kindreds and the general population. Dis Colon Rectum. 41:428-33.
12.Valérie Bonadona, MD, PhD; Bernard Bonaïti, MSc; Sylviane Olschwang, MD, PhD; et al, (2011), Cancer Risks Associated With Germline Mutations in MLH1, MSH2, and MSH6 Genes in Lynch Syndrome Article Information JAMA. 305(22):2304-2310.
13. Aquilina G, Ceccotti S, Martinelli S, Soddu S, Crescenzi M, Branch P, Karran P, Bignami M. (2000). Mismatch repair and p53 independently affect sensitivity to N-(2-chloroethyl)-N′-cyclohexyl-N-nitrosourea. Clin Cancer Res. 6:671–680
14. Scartozzi M, De Nictolis M, Galizia E, Carassai P, Bianchi F, Berardi R, Gesuita R, Piga A, Cellerino R, Porfiri E. (2003). Loss of hMLH1 expression correlates with improved survival in stage III–IV ovarian cancer patients. Eur J Cancer. 39:1144–1149.
15. Hawn MT, Umar A, Carethers JM, Marra G, Kunkel TA, Boland CR, Koi M. (1995). Evidence for a connection between the mismatch repair system and the G2 cell cycle checkpoint. Cancer Res. 55:3721–3725.
16. Helleman J, van Staveren IL, Dinjens WN, van Kuijk PF, Ritstier K, Ewing PC, van der Burg ME, Stoter G, Berns EM. (2006). Mismatch repair and treatment resistance in ovarian cancer. BMC Cancer.6:201-208.
17. Ian H, Barry R, Harvey A. Risch, KS, Mario E. Beiner,JM, Ping S, Steven A. (2008). Narod Ovarian cancer risk is associated with a common variant in the promoter sequence of the mismatch repair gene MLH1. 109(3): 384–387
18. Aquilina G, Ceccotti S, Martinelli S, Soddu S, Crescenzi M, Branch P, Karran P, Bignami M. (2000). Mismatch repair and p53 independently affect sensitivity to N-(2-chloroethyl)-N′-cyclohexyl-N-nitrosourea. Clin Cancer Res.6:671–680
19. Scartozzi M, De Nictolis M, Galizia E, Carassai P, Bianchi F, Berardi R, Gesuita R, Piga A, Cellerino R, Porfiri E. (2003). Loss of hMLH1 expression correlates with improved survival in stage III–IV ovarian cancer patients. Eur J Cancer;39:1144–1149.
20. Hawn MT, Umar A, Carethers JM, Marra G, Kunkel TA, Boland CR, Koi M. (1995). Evidence for a connection between the mismatch repair system and the G2 cell cycle checkpoint. Cancer Res.55:3721–3725