[Frontiers In Bioscience, Elite, 10, 375-383, March 1, 2018]

Five- hit hypothesis in ATM gene: An individualized model in a breast cancer patient

Parvin Mehdipour1, Asaad Azarnezhad1

1Department of Medical genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and Methods
3.1. Design of study
3.2. Proband information and clinical data
3.3. DNA extraction, Polymerase chain reaction (PCR), and sequencing
3.4. Molecular cloning
3.5. Immunofluorescence (IF) assay
3.6. In silico analysis
4. Results
4.1. Sequencing findings
4.2. Allelic location of hits
4.3. Immunofluorescence (IF)
4.4. In silico predictions
5. Discussion
6. Acknowledgement
8. References

1. ABSTRACT

The aim of this study was to trace D1853N in a proband affected with primary BC to explore the molecular, cellular and structural aspects of ATM. Exon 37 and splicing regions were PCR-sequenced. Allelic location of the alterations was determined by molecular cloning. Possible impact of alterations was investigated through the bioinformatics and protein expression assays. Five genetic variants including IVS 36-91 AA>TT, IVS 36-8 T>C, D1853N, IVS 37+47 A>G, IVS 37+60 Del T were found in the target regions of ATM and all the alterations were occurred heterezygously. IVS 36-8 T>C and D1853N were observed in blood and tumor tissue, whilst splicing variants were only occurred in tumor tissue. Missense D1853N alteration seems to be effective on 2D and 3D structure of ATM protein and the probability of splicing found to be decreased by intronic variants. Protein expression of ATM also confirmed the occurrence and functional impact of alterations. Results reflect a five-hit hypothesis in a proband with BC that influence ATM, as a guard of genomic stability, at molecular, cellular, and structural levels.

7. REFERENCES

1. Tao Z., Shi A., Lu C., Song T., Zhang Z. and Zhao J. Breast cancer: epidemiology and etiology. Cell Biochem Biophys, 72, 333-338 (2015)
https://doi.org/10.1007/s12013-014-0459-6
PMid:25543329

2. Karami F., Mehdipour P. A comprehensive focus on global spectrum of BRCA1 and BRCA2 mutations in breast cancer. Biomed Res Int, (2013)

3. Turnbull C., Rahman N. Genetic predisposition to breast cancer: past, present, and future. Annu. Rev. Genomics Hum. Genet., 9, 321-345 (2008)

4. Carranza D., Vega A. K., Torres-Rusillo S., Montero E., Martinez L. J., Santamaría M., Santos J. L., Molina I. J. Molecular and Functional Characterization of a Cohort of Spanish Patients with Ataxia-Telangiectasia. Neuromolecular Med, 1-14 (2016)

5. Deng Q., Sheng L., Su D., Zhang L., Liu P., Lu K., Ma S. Genetic polymorphisms in ATM, ERCC1, APE1 and iASPP genes and lung cancer risk in a population of southeast China. Med Oncol, 28, 667-672 (2011)
https://doi.org/10.1007/s12032-010-9507-2
PMid:20354815

6. Ćmielová J., Havelek R., Vávrová J., Řezáčová M. Changes in the response of MCF-7 cells to ionizing radiation after the combination of ATM and DNA-PK inhibition. Med Oncol, 32, 138 (2015).

7. Kim J. H., Kim H., Lee K. Y., Choe K.-H., Ryu J.-S., Yoon H. I., Sung S. W., Yoo K.-Y., Hong Y.-C. Genetic polymorphisms of ataxia telangiectasia mutated affect lung cancer risk. Human molecular genetics, 15, 1181-1186 (2006)
https://doi.org/10.1093/hmg/ddl033
PMid:16497724

8. Thompson D., Duedal S., Kirner J., McGuffog L., Last J., Reiman A., Byrd P., Taylor M., Easton D. F. Cancer risks and mortality in heterozygous ATM mutation carriers. J Natl Cancer Inst, 97, 813-822 (2005)
https://doi.org/10.1093/jnci/dji141
PMid:15928302

9. Mehdipour P., Mahdavi M., Mohammadi-Asl J., Atri M.: Importance of ATM gene as a susceptible trait: predisposition role of D1853N polymorphism in breast cancer. Med Oncol, 28, 733-737 (2011)
https://doi.org/10.1007/s12032-010-9525-0
PMid:20396981

10. Schrauder M., Frank S., Strissel P., Lux M., Bani M., Rauh C., Sieber C., Heusinger K., Hartmann A., Schulz-Wendtland R. Single nucleotide polymorphism D1853N of the ATM gene may alter the risk for breast cancer. J Cancer Res Clin Oncol, 134, 873-882 (2008)
https://doi.org/10.1007/s00432-008-0355-9
PMid:18264724

11. Gao L.-B., Pan X.-M., Sun H., Wang X., Rao L., Li L.-J., Liang W.-B., Lv M.-L., Yang W.-Z., Zhang L. The association between ATM D1853N polymorphism and breast cancer susceptibility: a meta-analysis. J Exp Clin Cancer Res, 29, 117 (2010)

12. Mehdipour P., Habibi L., Mohammadi-Asl J., Kamalian N., Azin M. M. Three-hit hypothesis in astrocytoma: tracing the polymorphism D1853N in ATM gene through a pedigree of the proband affected with primary brain tumor. J Cancer Res Clin Oncol, 134, 1173-1180 (2008)
https://doi.org/10.1007/s00432-008-0404-4
PMid:18465141

13. Azarnezhad A., Sharifi Z., Seyedabadi R., Hosseini A., Johari B., Fard M. S. Cloning and Expression of Soluble Recombinant HIV-1 CRF35 Protease-HP Thioredoxin Fusion Protein. Avicenna J Med Biotechnol, 8, 175 (2016)

14. Mehdipour P., Pirouzpanah S., Sarafnejad A., Atri M., Shahrestani S. T., Haidari M. Prognostic implication of CDC25A and cyclin E expression on primary breast cancer patients. Cell Biol Int, 33, 1050-1056 (2009)
https://doi.org/10.1016/j.cellbi.2009.06.016
PMid:19555767

15. Ahmed M., Rahman N.: ATM and breast cancer susceptibility. Oncogene, 25, 5906-5911 (2006)
https://doi.org/10.1038/sj.onc.1209873
PMid:16998505

16. Milne R. L. Variants in the ATM gene and breast cancer susceptibility. Genome Med, 1, 12 (2009)

17. Hall J. The Ataxia-telangiectasia mutated gene and breast cancer: gene expression profiles and sequence variants. Cancer lett, 227, 105-114 (2005)

18. Keller G. M. In vitro differentiation of embryonic stem cells. Curr Opin Cell Biol, 7, 862-869 (1995)

19. Surani M. A., Hayashi K., Hajkova P. Genetic and epigenetic regulators of pluripotency. Cell, 128, 747-762 (2007)
https://doi.org/10.1016/j.cell.2007.02.010
PMid:17320511

20. Ly S. Embryonic Differentiation in Animals. Embryo Project Encyclopedia (2012)

21. Black D. L.: Mechanisms of alternative pre-messenger RNA splicing. Annu Rev Biochem, 72, 291-336 (2003)
https://doi.org/10.1146/annurev.biochem.72.121801.161720
PMid:12626338

22. Wang G.-S., Cooper T. A. Splicing in disease: disruption of the splicing code and the decoding machinery. Nature Reviews Genetics, 8, 749-761 (2007)
https://doi.org/10.1038/nrg2164
PMid:17726481

23. Douglas A. G., Wood M. J. RNA splicing: disease and therapy. Briefings in functional genomics, 10, 151-164 (2011)
https://doi.org/10.1093/bfgp/elr020
PMid:21628314

24. Jian X., Boerwinkle E., Liu X. In silico prediction of splice-altering single nucleotide variants in the human genome. Nucleic Acids Res, 42, 13534-13544 (2014)
https://doi.org/10.1093/nar/gku1206
PMid:25416802 PMCid:PMC4267638

25. Tazi J., Bakkour N., Stamm S. Alternative splicing and disease. BBA MOL BASIS DIS, 1792, 14-26 (2009)
https://doi.org/10.1016/j.bbadis.2008.09.017
PMid:18992329

26. Ward L. D., Kellis M. Interpreting noncoding genetic variation in complex traits and human disease. Nat Biotechnol, 30, 1095-1106 (2012)
https://doi.org/10.1038/nbt.2422
PMid:23138309 PMCid:PMC3703467

Abbreviations: BC: Breast cancer; ATM: Ataxia telangiectasia-mutated; Family History (FH); Human Splice Finder (HSF); 2D: Secondary structure; 3D: Tertiary structure; IF: Immunofluorescence; PE: Protein expression;

Key Words: Ataxia Telangiectasia Mutated, Polymorphism, Breast cancer, hit- hypothesis, Evolution

Send correspondence to: Parvin Mehdipour, Department of Medical Genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran, Tel: 0098218830100, Fax: 0098218830100, E-mail: mehdipor@tums.ac.ir