HER2受体核转运途径参与乳腺癌细胞曲妥珠单抗耐药及其机制的研究

Despite the survival gains provided by trastuzumab, patients with advanced HER2+ breast cancer frequently display primary resistance to trastuzumab-based therapy, and even if they initially respond. the majority of mechanisms of resistance described to date mainly focused on continued HER2 downstream signaling activation,such as PI3K and MAPK. However,Recent research showed that HER2 receptor could tanslocate into nuclear to activate kinase and regulate gene transcription which result in improvement of cellular growth and proliferation.And no direct evidence indicate relation between nuclear HER2 receptor and resistance to trastuzumab .To investigate relation between HER2 receptor nuclear translocation and resistance to trastuzumab in breast cancer cell with her2 over expression , the inhibitory effect of trastuzumab on breast cancer cell expressed mutated HER2 recetpors lack of nuclear location signal will be observed. Nuclear expression of her2 receptors in different cell lines will be detected and the physical interaction between Her2 receptor and target gene and protein wil be investigated; Western blotting and imunoprecipitation will also be applied to detect expression and activity of target gene and proten, hoping to elucidate the role of nuclear HER2 in trastuzumab resistance and its mechanism.

HER2过表达的乳腺癌患者在曲妥珠单抗治疗缓解后普遍出现耐药，其机制的研究主要集中在PI3K和MAPK等传统信号通路。HER2受体转运入细胞核激活目标基因和蛋白可以促进细胞增殖，生长。我们前期研究显示与曲妥珠单抗共培养的乳腺癌细胞核内的HER2受体表达增多,但核内的HER2受体是否与耐药相关目前尚无明确证据。本研究通过基因突变技术使HER2受体缺失转运入核所必须的核定位信号，形成无法转运入核的突变型HER2受体，来验证HER2受体转运入核是否能增加对曲妥珠单抗的耐受。还通过免疫共沉淀和染色体共沉淀的方法观察耐药细胞中HER2受体与核内蛋白和靶基因的结合情况，并观察靶蛋白和靶基因的表达和活性，来探讨HER2入核后增强细胞对曲妥珠单抗耐药的机制，以期阐明HER2受体核转运在曲妥珠单抗耐药中的作用和机制。

HER2过表达的乳腺癌患者在曲妥珠单抗治疗缓解后普遍出现耐药，其机制的研究主要集中在PI3K和MAPK等传统信号通路。HER2受体转运入细胞核激活目标基因和蛋白可以促进细胞增殖，生长。我们前期研究显示与曲妥珠单抗共培养的乳腺癌细胞核内的HER2受体表达增多,但核内的HER2受体是否与耐药相关目前尚无明确证据。本研究通过基因突变技术使HER2受体缺失转运入核所必须的核定位信号，形成无法转运入核的突变型HER2受体，来验证HER2受体转运入核是否能增加对曲妥珠单抗的耐受。还通过免疫共沉淀和染色体共沉淀的方法观察耐药细胞中HER2受体与核内蛋白和靶基因的结合情况，并观察靶蛋白和靶基因的表达和活性。在研究中通过体外实验发现曲妥珠单抗药物作用下乳腺癌细胞核内HER2表达增多。在成功构建HER2突变型和野生型cDNA的质粒后，成功转染乳腺癌细胞.研究显示乳腺癌细胞缺失NLS的HER2受体无法转运入细胞核。还在体外研究发现HER2缺失NLS的乳腺癌细胞对曲妥珠单抗耐受力降低。同时，动物实验体外验证细胞核内HER2能提高肿瘤对曲妥珠单抗的耐受能力。进一步的研究发现耐药细胞核内的HER2受体与RNA POL I 及stat3相结合而进一步调控其功能；耐药细胞核内的HER2受体可直接作用于COX-2的启动子。耐药细胞中HER2核转运与SRC激酶激活以及HER2二聚化可能相关。实验研究验证了细胞核内HER2的表达与乳腺癌细胞辐射耐受相关，并且进一步研究显示细胞核内HER2的表达可能与紫杉醇耐药相关。我们得出结论HER2核转运途径参与了乳腺癌细胞曲妥珠单抗获得性耐药的形成，对于其进一步研究可能会为逆转曲妥珠单抗耐药找到作用靶点。