(Arg)(9)-SH2 superbinder: a novel promising anticancer therapy to melanoma by blocking phosphotyrosine signaling.
(Arg)(9)-SH2 superbinder: a novel promising anticancer therapy to melanoma by blocking phosphotyrosine signaling.
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(Arg)9-SH2 superbinder：一种通过阻断磷酸酪氨酸信号传导来治疗黑色素瘤的新型有前途的抗癌疗法。
DOI：
10.1186/s13046-018-0812-5
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发表时间：
2018-07-05
期刊：
影响因子：
--
通讯作者：
Cao X
中科院分区：
文献类型：
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作者：
Liu AD;Xu H;Gao YN;Luo DN;Li ZF;Voss C;Li SSC;Cao X
黑色素瘤是一种误诊率高、预后差的恶性肿瘤。生物靶向治疗是治疗黑色素瘤的主流方法；然而，随之而来的耐药性是不可避免的。 SH2 superbinder 是 Src 同源 2 (SH2) 结构域的三重突变体，通过替换天然的含有 SH2 的蛋白质并阻断多个基于 pY 的信号通路，显示出有效的抗肿瘤能力。聚精氨酸 (Arg)9 是一种用于细胞内递送大分子的强大载体，可以跨细胞膜运输治疗剂。本研究的目的是构建(Arg)9-SH2 superbinder并研究其对黑色素瘤细胞的作用，以期为抗癌治疗提供潜在的新途径，克服单靶点抗肿瘤药物不可避免的耐药性。 (Arg)9和SH2 superbinder通过基因工程融合形成(Arg)9-SH2 superbinder。进行 Pull down 测定以确定 (Arg)9-SH2 superbinder 可以捕获多种 pY 蛋白。采用免疫荧光法检测(Arg)9-SH2 superbinder进入细胞的效率。通过MTT和集落形成实验评估增殖能力。此外，进行伤口愈合和transwell实验来评估B16F10、A375和A375/DDP细胞的迁移。此外，通过基于流式细胞术的膜联蛋白 V/PI 分析了 (Arg)9-SH2 superbinder 引起的细胞凋亡。此外，蛋白质印迹显示 (Arg)9-SH2 superbinder 影响一些 pY 相关信号通路。最后，建立B16F10异种移植模型以证实(Arg)9-SH2 superbinder是否能够抑制肿瘤的生长。我们的数据表明，(Arg)9-SH2 superbinder 能够有效进入黑色素瘤细胞，并对各种 pY 蛋白表现出很强的亲和力。此外，(Arg)9-SH2 superbinder可以通过调节PI3K/AKT、MAPK/ERK和JAK/STAT信号通路抑制黑色素瘤细胞的增殖、迁移并诱导细胞凋亡。重要的是，(Arg)9-SH2 superbinder可以显着抑制小鼠肿瘤的生长。 (Arg)9-SH2 superbinder 对 pY 蛋白表现出高亲和力，通过替换含有 SH2 的蛋白并阻断多种基于 pY 的途径，显示出有效的抗癌能力。 (Arg)9-SH2 superbinder 抑制癌细胞增殖和肿瘤生长的卓越能力可能为探索 SH2 superbinder 作为癌症治疗药物打开了大门。本文的在线版本 (10.1186/s13046-018-0812-5) 包含补充材料，可供授权用户使用。
Melanoma is a malignant tumor with high misdiagnosis rate and poor prognosis. The bio-targeted therapy is a prevailing method in the treatment of melanoma; however, the accompanying drug resistance is inevitable. SH2 superbinder, a triple-mutant of the Src Homology 2 (SH2) domain, shows potent antitumor ability by replacing natural SH2-containing proteins and blocking multiple pY-based signaling pathways. Polyarginine (Arg)9, a powerful vector for intracellular delivery of large molecules, could transport therapeutic agents across cell membrane. The purpose of this study is to construct (Arg)9-SH2 superbinder and investigate its effects on melanoma cells, expecting to provide potential new approaches for anti-cancer therapy and overcoming the unavoidable drug resistance of single-targeted antitumor agents. (Arg)9 and SH2 superbinder were fused to form (Arg)9-SH2 superbinder via genetic engineering. Pull down assay was performed to identify that (Arg)9-SH2 superbinder could capture a wide variety of pY proteins. Immunofluorescence was used to detect the efficiency of (Arg)9-SH2 superbinder entering cells. The proliferation ability was assessed by MTT and colony formation assay. In addition, wound healing and transwell assay were performed to evaluate migration of B16F10, A375 and A375/DDP cells. Moreover, apoptosis caused by (Arg)9-SH2 superbinder was analyzed by flow cytometry-based Annexin V/PI. Furthermore, western blot revealed that (Arg)9-SH2 superbinder influenced some pY-related signaling pathways. Finally, B16F10 xenograft model was established to confirm whether (Arg)9-SH2 superbinder could restrain the growth of tumor. Our data showed that (Arg)9-SH2 superbinder had the ability to enter melanoma cells effectively and displayed strong affinities for various pY proteins. Furthermore, (Arg)9-SH2 superbinder could repress proliferation, migration and induce apoptosis of melanoma cells by regulating PI3K/AKT, MAPK/ERK and JAK/STAT signaling pathways. Importantly, (Arg)9-SH2 superbinder could significantly inhibit the growth of tumor in mice. (Arg)9-SH2 superbinder exhibited high affinities for pY proteins, which showed effective anticancer ability by replacing SH2-containing proteins and blocking diverse pY-based pathways. The remarkable ability of (Arg)9-SH2 superbinder to inhibit cancer cell proliferation and tumor growth might open the door to explore the SH2 superbinder as a therapeutic agent for cancer treatment. The online version of this article (10.1186/s13046-018-0812-5) contains supplementary material, which is available to authorized users.