miR-206与miR-132之靶基因结合位点的单核苷酸多态对乳腺癌发病风险和转移潜能的影响

Hypoxia plays an important role in the genesis and development for breast cancer. However, little has been known about the mechanism of hypoxia on breast cancer risk and metastasis. Our preliminary results revealed that hypoxia affected treatment sensitivity and metastatic potential by modifying the phenotype of breast cancer cells.Additionally, we found that single nucleotide polymorphism (SNP) in the binding sites of target genes with miR-206 was significantly correlated with the risk for breast cancer. On these premises, we postulate that breast cancer risk and metastasis might result from the joint effects of multiple SNPs in hypoxia-related miRNAs and their target sites. In order to verify this hypothesis, we aim at hypoxia-related miRNAs such as miR-206 to explore the association between SNPs in its binding sites with potential target genes and breast cancer risk. We also probe into the effect of these candidate miRNAs on breast cancer metastasis in breast cancer cell lines and their nude mice models by utilizing transfection of miRNA vectors and antisense oligonucleotides. Furthermore, we sought to identify other potential target genes of these candidate miRNAs, which offers implications for clinical-epidemiological studies into the correlation of SNPs in the binding sites of such target genes with breast cancer development.The joint effects of multiple literally independent but naturally dependent SNPs may enable a brand new vision into the mechanisms of breast cancer genesis and progression.

缺氧是乳腺癌发生发展过程中的重要环节，但目前缺氧如何影响乳腺癌发病风险和转移潜能仍知之甚少。我们前期研究发现：缺氧可通过改变乳腺癌分子表型进而影响其治疗敏感性和转移潜能；同时miR-206等缺氧相关miRNA与靶基因结合区SNP和乳腺癌发病风险显著相关。为此我们设想：乳腺癌发病风险及转移潜能可能是缺氧相关miRNA之靶基因结合区多个SNP位点变化的综合结果。为验证此假说，我们以miR-206等缺氧相关miRNA为切入点，探究其潜在靶基因结合区的SNP位点与乳腺癌发病风险的关系；采用载体/反义miRNA转染等手段，从分子、细胞及动物等多水平探讨候选miRNA对乳腺癌转移潜能的影响，并探索候选miRNA的其他可能靶基因；指导临床流行病学研究进一步探索这些靶基因结合区SNP与乳腺癌恶性程度的相关性。上述既独立又关联的SNPs综合效应对探索乳腺癌形成和进展机制或能提供崭新思路。

现有研究表明，雌激素与乳腺细胞的癌变密切相关。新近发现的基因表达调控因子miRNA在乳腺癌发生发展中所起的作用业已受到重视。本研究发现，携带SULF1基因3’-UTR区rs3802278 AA型的女性乳腺癌发病风险较GG+GA型为低，这一现象在ER阳性乳腺癌以及HR阳性乳腺癌发病中亦适用。ADRB2 rs1042713位点的等位基因型G可能是乳腺癌发病的保护性因素，而HIF1AN rs11292位点的等位基因型C可能增加乳腺癌发病风险。生存分析显示，将rs1042713与rs11292两个位点进行联合分析，同时携带rs1042713 G和rs11292 C等位基因女性的DFS优于其他患者。进一步探究miR-206与乳腺癌生物学行为的关系及其可能机制后发现，miR-206过表达后ER阳性乳腺癌细胞侵袭能力减弱，并可抑制ER阳性乳腺癌细胞系出现EMT。miR-206过表达细胞TGF-β mRNA表达水平及分泌蛋白水平均比对照细胞显著下降。过表达miR-206后，靶向抑制TGF-β通路中NRP1、SMAD2的表达，并抑制ANXA2、STC2表达。外源性TGF-β1刺激后，ER阳性乳腺癌细胞miR-206表达反馈性升高。添加外源性TGF-β刺激后，miR-206过表达细胞侵袭能力恢复。miR-206过表达导致下游细胞迁移相关基因PLD1 mRNA表达下降。另外，以乳腺癌患者及对照人群血清样本中游离miRNA为研究对象，运用高通量芯片表达分析筛选手段检测同一患者手术前后有明显改变的microRNA。研究发现血清miR-130b-5p、miR-151a-5p、miR-206、miR-222-3p具有潜在的乳腺癌诊断价值，且通过计算ROC曲线下面积后发现将4个miRNA联合起来的诊断价值更好。因此，本研究对探索乳腺癌形成和进展机制或能提供崭新思路，在后续成果转化中可能具有一定的社会效益和经济效益。