| Background and Objection:Worldwide, breast cancer has become the highest incidence of the malignant tumors of women. Also, the incidence of breast cancer has increased in China. The mortality rate increased by 99.99 percent over the past 30 years while its age of onset is gradually dropping. With the gradual improvement of diagnostic techniques and treatment, the survival rate was significantly longer, but still very poor prognosis of patients with advanced breast cancer. Therefore, early diagnosis of breast cancer is rather crucial for prognosis. The occurrence and development of breast cancer is a complex pathological process, which is the result of many factors such as genetic, environmental, physiological status and living habits.At present, it has been publicly known that the status of estrogen receptor, progesterone receptor, HER2, Ki-67 has particularly close ties with breast cancer. Such genetic and molecular events lead to the different characteristics of breast cancer. As for the most important predictors, ER and PR predict the effectiveness of the adjuvant neoadjuvant hormone therapy for metastatic breast cancer. NCI put forward the conception of Molecular typing in 1999. Based on the conception, breast cancer can be divided into 4 subtypes:luminal A,luminal B, HER2 over-expression and basal-like type, luminal A type which immunohistochemical detection of ER-positive or PR-positive, HER-2-negative, Ki67 of low expression is the most common molecular subtype of breast cancer. Its incidence rate is from 44.5% to 69.0%, while luminal A type of breast cancer subtypes prognosis for patients with early majority and the lower risk of recurrence. And for the treatment, luminal A type of breast cancer is sensitive to the endocrine therapy and the efficiency is up to 40%, what’s more, sensitivity to endocrine therapy and ER level are positively correlated. Luminal A type of breast cancer is not suitable for molecular targeted therapy because the level of HER-2 is negative. Studies have shown that luminal A type of differential efficacy of neoadjuvant chemotherapy in breast cancer patients is worse than other subtypes. HER-2 overexpression subtype of breast cancer is that immunohistochemistry ER, PR-negative, HER-2-positive, Ki-67 expression, mostly, 20% to 30% of primary breast cancer patients have HER-2 overexpression. HER-2 overexpressing breast cancer is characterized by ERBB2 amplification, gene expression on the 17th chromosome TRAP 100, GRB7 upregulated, RRM2, RAD5 downregulation. It is almost ineffective for endocrine treatment because that ER, PR are negative. For the Adjuvant chemotherapy, this type is more sensitive to chemotherapy regimens containing anthracyclines, and has a dose-effect relationship. HER-2 is both a prognostic indicator and the predictor of the application of HER-2 targeted drugs. For the luminal B type of breast cancer, the immunohistochemical detection of ER, PR are positive, and HER-2 is also positive. It is more common in the elderly breast cancer patients. This subtype also belong to endocrine therapy-sensitive tumors, the effect of tamoxifen is worse than luminal A type while the effect of aromatase inhibitors is much better. Basal-like type refers to the type of breast cancer which immunohistochemical of ER, PR, HER-2 are negative.RHAMM is also known as CD168 on the surface of cell. It is separated from the culture supernatant from the 3T3 mouse fibroblasts in 1987. The gene for the receptor for hyaluronan-mediated motility, RHAMM(designated hyaluronan-mediated motility receptor, HMMR(human) and Hmmr(mouse), for mapping purposes, was localized to human chromosome 5q33.2-qter by somatic cell and radiation hybrid analyses. Investigation of two interspecific backcrosses localized the mouse RHAMM(Hmmr)locus 18cM from the centromere of mouse chromosome 5q23-q35 gene. The map position of the human RHAMM gene places it in a region comparatively rich in disease-associated genes, including those for low-frequency hearing loss, dominant limb-girdle muscular dystrophy.Single nucleotide polymorphisms refers to a single nucleotide variation in the genome, including conversion, transversions, deletions, and insertions. The formation of genetic markers is of rich polymorphism. From the theoretical point of view, each SNP locus can have four different forms of variation, but in fact only two occurred:transitions and transversions, the ratio of which is 2:1. Generally, SNP refers to a single nucleotide polymorphism mutation frequency greater than 1%. In the human genome, there is a SNP in about 1000 bases, and there are about 3×10E6 SNPs in the human genome. Therefore, SNP has became the third generation of genetic markers, many human phenotypic differences, drug or disease susceptibility, etc., may be related to the SNP and SNP research is an huge step towards the human genome project applications. This is mainly because the SNP will provide a powerful tool for the discovery of high-risk groups, to identify the disease-related genes, drug testing as well as basic research, such as biology. SNP in the genome is fairly well distributed, studies show that the SNP appears once in every 300 base pairs in the human genome. The existence of a large number of SNP loci create the opportunity to discover the variety of diseases, including cancer-related mutations in the genome; from the experimental operation, the detection of disease-related genes by SNP mutation is easier than through the family study; otherwise, some SNP does not directly lead to expression of disease genes, but because it is adjacent to genes associated with certain diseases, it also has become an important marker. SNP has also played a great role in basic research. Through the analysis of the SNP on the Y chromosome, scientists have made a series of important results in the field of evolution and migration of human populations.Methods:1. Select samples. A total of 473 breast cancer cases and 506 healthy controls were identified from the outdoor and indoor patients at hospitals in China during October 2010 to December 2013. The whole subjects were unrelated Chinese females of Han ancestry. The healthy controls were female examinees of the corresponding period with no history of breast related disease or family history of breast cancer. All breast cancer cases were diagnosed as invasive breast cancer using haematoxylin and eosin(H&E)staining by at least two pathologists. Patients with personal or family history of breast cancer were excluded. Patients’characteristics including age and histological information were collected. Including estrogen receptor(ER), progesterone receptor(PR), human epidermal growth factor receptor 2(HER-2)and Ki-67 status.2. Select SNPs. In total, we selected 16 SNPs which have been previously reported in GWAS and all of them were in HMMR gene. These SNPs were rs213505, rs299290, rs299318, rs427056, rs2069345, rs2069347, rs2303077, rs4869089, rs6556674, rs10035535, rs10056017, rs13183712, rs17535909, rs17601374, rs74805168,rs193028482.3. Mutation detection. Primers were designed using Sequenom MassARRAY(?)Assay Design 3.1.All SNPs were genotyped using the SEQUENOM MassARRAY platform(Sequenom, San Diego, California, USA). The call rate for each assay was set at>90%. Two or more technicians analyze polymorphisms image by automatic way and manual way.4.Statistical analysis. The Hardy-Weinberg equilibrium(HWE)test was carried out to compare the genotype and allele distributions between case and control subjects. Further stratifying analysis was conducted based on histological information including ER, PR, HER2, and Ki-67 status. The Chi square test was carried out to evaluating the statistical significance of differences the genotype and allele distributions between case and control groups. The unconditional logistic regression was used to assess the allelic odds ratio(OR) and 95% confidence intervals (95%CIs) after age-adjustment in case-control study. Finally, the association between the detected SNPs and breast cancer susceptibility can be evaluated. A P value less than 0.05 was considered statistically significant. All statistical analyses were carried out with IBM SPSS version 18.0 and the web-based toolSNPstats(http://bioinfo.iconcologia.net/SNPstats).Result:1. Basal-line characteristics of breast cancer patients were shown in Table 3-1. The most common histological type was invasive ductal carcinoma, accounting for 92.36% of all cases. In terms of ER and PR status,the proportion of positive cases were 60.5% and 57.1%, respectively. In addition, patients with luminal A breast cancer represented more than half of all cases. Though we excluded patients with family history of breast cancer, there was still 9.3% of the included patients having family history of other cancer.2. Quality control for all SNPs with Hardy-Weinberg Equilibrium (HWE) P>0.05 in cases and controls was conducted. Seven SNPs including rs299318, rs4869089, rs6556674, rs13183712, rs17601374, rs74805168, rs193028482 were left to analyze afterwards.Subsequent analyses revealed that SNP rs193028482 was non-polymorphic in our study population. Thus, only 6 of the 16 SNPs were evaluable in the present study.3. Then we assessed the distributions of the 6 evaluable SNPs in breast cancer and control groups (Table3-2). We found that the genotype distribution of the rs13183712 SNP in the breast cancer group was 57.3% GG,37.6% GT and 5.1% TT, in the control group, it was 47.5% GG,45% GT and 7.4% TT, P=0.012. These two resuts are definitely different. Compared with GG, genotype GT appeared to decrease breast cancer risk under the codominant(OR=0.69,95%CI=0.52-0.91, p=0.013), and overdominant models(OR=0.74,95%CI=0.56-0.96,p=0.026). However, the other 5 SNPs were not associated with an increased or decreased incidence in our population. The data thus highlights the critical influence of the SNP rs13183712 in HMMR gene on breast cancer.4. Moreover, the associations of the 6 evaluable SNPs with ER status(Table 3) were assessed. As shown in Table 3, compared to the GG genotype, the G/T-T/T genotypes of rs13183712 showed a significant association with both negative ER status(OR=0.66,95%CI=0.46-0.95, P=0.024)and positive ER status(OR=0.68, 95%CI=0.50-0.94, P=0.018). In addition, compared to the AA genotype, the AT genotype of rs17601374 showed a significant association with negative ER status under codominant(OR=0.62,95%CI=0.37-1.06, P=0.036).5. Further stratification based on PR status identified three significant SNPs(Table 3-4), including rs4869089, rs13183712 and rs17601374.Compared to the GG-GA genotypes, the AA of rs4869089 showed a significant association with negative PR status under recessive model(OR=1.99,95%CI=1.08-3.67, P=0.03).In contrast with the GG genotype, the GT of rs13183712 showed a significant association with negative PR status under codominant(OR=0.68,95%CI=0.48-0.95, P=0.04) and G/T-T/T genotype has a negitave association with both negative PR status and positive PR status. Besides, comparing with the AA genotype, the AT genotype of rs17601374 showed a significant association with negative PR status under both codominant(OR=0.62,95%CI=0.37-1.04, P=0.027).6. In subtype-specific analyses, as shown in Table 5,three SNPs were identified to be associated with luminal B breast cancer,including rs4869089, rs17601374 and rs13183712. In contrast with the GG-GA genotypes, the AA of rs4869089 showed a significant association with luminal B tumors(OR=2.23,95%CI=1.17-4.25, P=0.019). Likewise, comparing with AA,genotype AT of the rs17601374 SNP appeared to decrease luminal B tumors under the codominant(OR=0.50, 95%CI=0.27-0.92, p=0.015) and overdominant models(OR=0.51,95%CI=0.27-0.94, p=0.021). Different from the previous 2 SNPs, rs13183712 was found to associated with both luminal B and HER2+breast cancer. Compared with GG,genotype GT of the rs 13183712 appeared to decrease luminal B tumors under the codominant(OR=0.57,95%CI=0.38-0.86, p=0.011) and overdominant models(OR=0.62,95%CI=0.41-0.92, p=0.017). Besides, comparing with GG, genotype GT of the rs13183712 appeared to decrease HER2+tumors under the codominant(OR=0.46,95%CI=0.27-0.79, p=0.0075) and overdominant models(OR=0.51,95%CI=0.30-0.86, p=0.0093). Lastly, in contrast with the CC genotype, the CT of rs74805168 showed a significant association with basal-like tumors under both dominant(OR=0.24,95%CI=0.06-1.02, P=0.017).Conclusion:The present study showed that associations of 16 SNPs HMMR gene and breast cancer risk according to ER status, PR status and molecular subtypes.Among the 6 evaluable SNPs, rs13183712 was associated with a significantly decreased frequency of breast cancer risk.Additionally,7 associations were found as for hormone receptors, and 5 for molecular subtypes.Thus,we indicate those 4 SNPs(rs4869089, rs13183712, rs17601374 and rs74805168)in HMMR gene to be potential makers and help to develop more individualized therapeutic strategies. However, We still need more studies to confirm all of our new findings. |
PDF Full Text Request