NRP-1介导PI3K/AKT信号通路调控乳腺癌淋巴管生成和转移的机制研究

Neuropilin-1(NRP-1) is regarded as a promising target for cancer therapy.It is a non-tyrosine kinase receptor for semaphorin 3A(Sema 3A) and vascular endothelial growth factor(VEGF) involved in tumor angiogenesis, growth and metastasis.In preparatory study，the anti-NRP-1mAb was obtained and the affinity of antibody was identified. It can inhibits the adhesion of breast cancer cell MCF7 to fibronectin by suppressing FAK/p130cas signaling pathway. In this study，we aim to investigated the effects of NRP-1 to tumor cells proliferation, migration and adhesion in breast cancer MDA-MB-231 after treatment with anti-NRP-1mAb or NRP-1 RNAi, respectively. To asssay differention of the migration by boyden chamber and canaliculization by culture with MDA-MB-231 cells. To observe the tumor growth, metastasis and lymphangiogenesis in nude mice, Also to detect the expression of NRP-1, PI3K, AKT, VEGFR-3 and integrin αvβ3. Finally, to illuminate the regulation mechanism of lymphangiogenesis and metastasis in breast Cancer by NRP-1 through PI3K/AKT signaling pathway which may be a importent theory basis that the Anti-NRP-1mAb is likely to be a prospective antibody drug candidate for tumor treatment.

Neuropilin-1（NRP-1）被认为是一个很有前景的癌症治疗靶点，它是一个多功能细胞表面受体。课题组前期成功制备鉴定Anti-NRP-1mAb，初步探讨其抑制乳腺癌细胞增殖、迁移和粘附机理。但关于NRP-1在乳腺癌新生淋巴管形成和淋巴结转移中的作用和机制，目前尚不清楚。本项目拟以转移性乳腺癌MDA-MB-231为研究对象，通过干扰NRP-1表达或抑制NRP-1功能，观察NRP-1在调控乳腺癌新生淋巴管形成和转移过程中的作用。同时通过分析NRP-1与PI3K、AKT、VEGFR-3及integrinαvβ3的蛋白和mRNA表达的相关性，阐明NRP-1通过PI3K/AKT信号通路调控乳腺癌淋巴管生成和淋巴结转移的作用机制，为基于NRP-1靶点的肿瘤靶向治疗提供新策略和理论依据。

Neuropilin-1（NRP-1）被认为是一个很有前景的癌症治疗靶点，它是一个多功能细胞表面受体。课题组前期成功制备鉴定Anti-NRP-1mAb，初步探讨其抑制乳腺癌细胞增殖、迁移和粘附机理。但关于NRP-1在乳腺癌新生淋巴管形成和淋巴结转移中的作用和机制，目前尚不清楚。本项目拟以转移性乳腺癌MDA-MB-231为研究对象，通过干扰NRP-1表达或抑制NRP-1功能，观察NRP-1在调控乳腺癌新生淋巴管形成和转移过程中的作用。同时通过分析NRP-1与PI3K、AKT、VEGFR-3及integrin的蛋白和mRNA表达的相关性。项目成功制备了纯度较高、亲和力较强的NRP-1mAb，构建2株NRP-1敲低乳腺癌细胞模型，结果表明无论采用NRP-1mAb阻断NRP-1还是NRP-1表达敲低均可有效抑制乳腺癌细胞的增殖、迁移和侵袭，且抑制程度与抗体浓度有明显剂量依赖关系。研究还发现采用NRP-1mAb阻断或者NRP-1干扰敲低后的乳腺癌细胞培养上清作为条件培养基，培养人淋巴管内皮细胞，可显著抑制淋巴管内皮细胞的增殖、迁移、侵袭和小管形成能力。体内实验表明NRP-1mAb可显著抑制荷瘤裸鼠的生长，且NRP-1干扰敲低乳腺癌细胞接种裸鼠后，其成瘤能力和肿瘤生长速度较对照组显著减慢。进一步研究发现NRP-1通过调控GIPC蛋白调控AKT磷酸化，进而减少IGFBP3表达和分泌，从而影响淋巴管内皮细胞的增殖、迁移、侵袭和小管形成。本研究初步阐明NRP-1调控乳腺癌淋巴管生成和淋巴结转移的作用机制，为基于NRP-1靶点的肿瘤靶向治疗提供新策略和理论依据。