Study Of Mutations And Polymorphisms Of The TIMP-3 Gene In Breast Cancer

[Back ground and Objective] Breast cancer is one of the most common malignant tumors in women, and the incidence rate is rising. It becomes a serious threat to women's health now. Early diagnosis is the key for preventing the cancers. According to the theories of modern molecular biology, the occurrence of cancer is the result of a series of genetic changes. The loss of chromosome segments harbored by TIMP-3 gene, the methylation of its promoter region, the coding sequences, loss of protein expression has all closely relevance to development and metastasis of many diseases. The expression of TIMP-3 was abserved in muscle epithelial cells, cavity epithelial cells and fibroblasts in normal breast tissue. However, the expression of TIMP-3 is correlation with the survival rate of patients, but it is not correlated with breast cancer and its metastasis in the axillary lymph node. Therefore, a study for expression and function of TIMP-3 gene in breast cancer has benefit of further understanding for breast cancer in the molecular mechanism of evolution, and finding a new indicator for early diagnosis and prognosis. By detecting the TIMP-3 gene mutation hot spots and polymorphisms in breast cancer we can explore the role of gene mutations and polymorphisms in the development of breast cancer, and kown the molecular mechanisms in breast cancer more clearly, and can find a new indicator for early diagnosis and prognosis for breast cancer. SSCP technology and DNA sequencing have used to screen the potential mutations, polymorphisms of TIMP-3 exons 1-5, splice junction between exons and introns in breast cancers , adjacent hyperplasia, benign breast hyperplasia , normal breast tissue and lymphcytoes from healthy donor's peripheral blood.[Methods] 50 cases of breast cancer tissue samples and adjacent hyperplasia tissues, 50 cases of benign hyperplasia tissues, 50 cases of normal breast tissues and lymphcytoes from 15 healthy donor's peripheral blood samples have collected. And all the cases of breast cancers and hyperplasia had been pathologically diagnosed by WHO standard. The TIMP-3 gene mutations in exon 1-5 of all samples were detected by polymerase chain reaction, SSCP analysis, and DNA direct sequencing. Then the sequencing datas were analyszed by the x~2 test.[Results] (1) We identified 2 polymorphisms in TIMP-3 :249(T>C),261(OT). We found 249 (T>C) of TIMP-3 exon3 in 40/50 cases (80%) of breast cancer, 36/50 cases (72%) of the corresponding adjacent hyperplasia, 39/ 50 cases (78%) benign hyperplasia, 37/50 Cases (74%) of healthy breast tissue and 13/15 cases (86.67%) of lymphcytoes from healthy donor's peripheral blood; And for 261(C>T) of TIMP-3 exon3 in 38/50 cases (76%) of breast cancer, 32/50 cases (64%) of the corresponding adjacent hyperplasia, 40/50 cases (80%) benign hyperplasia, 35/50 cases (70%) of healthy breast tissue and 11/15 cases (73.33%) of lymphcytoes from healthy donor's peripheral blood. There were no mutations found in Exon 1,2,4,5. (2) Statistics showed that: this two gene sequence changes in TIMP-3 exon 3 had no significant difference between the groups and the various genotypes. And the differences in genotype were not the cause of the susceptibility to breast cancer. These two changes can be found in the breast cancer samples, the corresponding adjacent samples of benign hyperplasia samples of healthy breast tissue samples and lymphcytoes from healthy donor's peripheral blood samples at the same time. According to the studies of polymorphism in TIMP-3 gene in the healthy group of white population, we can presume these two sites as polymorphic sites.[Conclusion] (1) There were no mutations in TIMP-3 exons in breast cancer in this study. This suggested that it was no significance to take the mutation detection for TIMP-3 in breast cancer for early diagnosis. (2) This two polymorphisms in TIMP-3 exon3 in breast cancer tissues, adjacent hyperplasia tissues, benign hyperplasia tissues, normal breast tissues and lymphcytoes from healthy donor's peripheral blood samples have not caused the changes of TIMP-3 protein. So we have speculated that the two sites of polymorphisms do not play an important role in the development of breast cancer and they can not be the indicators for early diagnosis in breast cancer.