Microsatellite alteration and metylation of RASSF1 gene in renal cell carcinoma

  • H. V. Dubrovska
  • K. V. Onyshchenko
  • L. V. Pereta
  • O. V. Kashparova
  • V. M. Grygorenko
  • I. Ya. Skrypkina
DOI: https://doi.org/10.7124/FEEO.v23.1013

Abstract

Aim. It becomes more and more relevant to study as the promising diagnostic and prognostic markers the epigenetic and genetic changes that determine the malignant transformation of cells and biological behavior of tumors. The purpose of our research was to study the loss of heterozygosity of the RASSF1 gene and the status of methylation of the locus of the RASSF1A gene in malignant tumors of the kidneys and to assess the potential for its use as a marker in clinical practice. Methods. The determination of the allele imbalance in the RASSF1 gene locus was performed by PCR using highly polymorphic markers D3S966, D3S1568 for renal cancer (RC) in paired samples of tumor and conditionally normal tissue and the PCR products detection in polyacrylamide gels. The epigenetic variability analysis of the RASSF1A gene was performed by using methyl-specific PCR, which was preceded by bisulfite DNA treatment. The statistical significance of the differences between the study groups was analyzed by using F-test and the U-test. Results. Our studies showed the predictive value of inactivation of RASSF1A in RC. The methylation level of the genomic DNA of the RASSF1A loci of the RASSF gene was 72 %. The loss of the heterozygosity of the RASSF1 gene in this type of oncological disease was about 35.4 %. Conclusions. The results may indicate that the gene RASSF1can become a candidate for inclusion in the prognostic system of clinical course of the disease.

Keywords: kidney cancer, epigenetic changes, methylation, loss of heterozygosity, RASSF1.

References

Cohen H.T., McGovern F.J. Renal-cell carcinoma. N Engl. J. Med. 2005. Vol. 353, No. 23. P. 2477–2490. doi: 10.1056/NEJMra043172.

Lam J.S., Leppert J.T., Belldegrun A.S., Figlin R.A. Novel approaches in the therapy of metastatic renal cell carcinoma. World J. Urol. 2005. Vol. 23, No. 3. P. 202–212. doi: 10.1007/s00345-004-0466-0.

National Health Service (England) NHS. URL: http://www.nhs.uk/conditions/Cancer-of-the-kidney/Pages/Introduction.aspx (last accessed: 22.05.2017).

Lujambio A., Calin G.A., Villanueva A., Ropero S., Sánchez-Céspedes M., Blanco D., Montuenga L. M., Rossi S., Nicoloso M.S., Faller W.J., Gallagher W.M., Eccles S.A., Croce C.M., Estellera M.A microRNA DNA methylation signature for human cancer metastasis. Proc. Natl. Acad. Sci. 2008. Vol. 105, No. 36. P. 13556–13561. doi: 10.1073/pnas.0803055105.

Sandeep N. Shah, Suzanne E. Hile Kristin A. Eckert. Defective mismatch repair, microsatellite mutation bias, and variability in clinical cancer phenotypes. Cancer Res. 2010. Vol. 70, No. 2. P. 431–435. doi: 10.1158/0008-5472.CAN-09-3049.

Kawakami K., Minamoto T. DNA methylation and cancer metastasis. Gan. to Kagaku Ryoho. Cancer & Chemotherapy. 2010. Vol. 37 (11). P. 2042–2046. doi: 10.1186/bcr2121.

Kristensen L.S., Hansen L.L. PCR-based methods for detecting single-locus DNA methylation biomarkers in cancer diagnostics, prognostics, and response to treatment. Clin Chem. 2009. Vol. 55 (8). P. 1471–1483. doi: 10.1373/clinchem.2008.121962.

Zambrano N.R., Lubensky I.A., Merino M.J., Linehan W.M., McClellan M.W. Histopathology and molecular genetics of renal tumors: toward unification of a classification system. J. Urol. 1999. No. 163. P. 1246–1258. doi: 10.1016/S0022-5347(05)68259-6.

Rini B.I., Campbell S.C., Escudier B. Renal cell carcinoma. Lancet. 2009. Vol. 373, No. 9669. P. 111–1132. doi: 10.1016/S0140-6736(09)60229-4.

Katz M.E., McCormick F. Signal transduction from multiple Ras effects. Current Opinion in Genetics & Development. 1997. Vol. 7, No. 1. P. 75–79. doi: 10.1016/S0959-437X(97)80112-8.

Liu L., Tommasi S., Lee D.H., Dammann R., Pfeifer G.P. Control of microtubule stability by the RASSF1A tumor suppressor. Oncogene. 2003. Vol. 22, No. 50. P. 8125–8136. doi: 10.1038/sj.onc.1206984.

Dammann R., Li C., Yoon J.H., Chin P.L., Bates S., Pfeifer G.P. Epigenetic inactivation of RAS association domain family protein from the lung tumor suppressor locus 3p21.3. Nat Genet. 2000. Vol. 25, No. 3. P. 315–319. doi: 10.1038/77083.

Yamamoto N., Kuroiwa T., Katakura A., Shibahara T., Choudhury C. Loss of Heterozygosity (LOH) on Chromosome 2q, 3p and 21q in Indian Oral Squamous Cell Carcinoma. Bull Tokyo Dent Coll. 2007. Vol. 48, No. 3. P. 109–117. doi: 10.2209/tdcpublication.48.109.

Trimeche M., Braham H., Ziadi S., Amara K., Hachana M., Korbi S. Investigation of allelic imbalances on chromosome 3p in nasopharyngeal carcinoma in Tunisia: High frequency of microsatellite instability in patients with early – onset of the disease. Oral Oncology. 2008. Vol. 44. P. 755–783. doi: 10.1016/j.oraloncology.2007.10.001.