Transcription-dependent and -independent signaling of RSK2 in cancer metastasis

描述(申请人提供)：转移是导致大多数癌症死亡的原因，包括乳腺癌、肺癌或头颈癌。然而，促进肿瘤转移的细胞变化背后的分子过程仍然知之甚少。我们发现，P90核糖体S6激酶2(RSK2)在包括乳腺癌、肺癌和头颈癌在内的转移性人类癌症中，通常在促进肿瘤转移的多个步骤中起重要作用，如抵抗失巢诱导、细胞迁移和侵袭以及肿瘤转移。通过结合磷蛋白组学和基于基因组学的研究，我们证明了RSK2通过磷酸化一系列下游因子来介导抗失巢凋亡、促进侵袭和促进转移的信号。特别是，我们确定了RSK2的新底物，包括凋亡信号调节激酶1(ASK1)和IRS1，分别提供抗失巢凋亡保护和促进癌细胞的侵袭和转移潜能。此外，我们还发现RSK2-CREB信号通路调节一组参与细胞死亡调控的蛋白质的基因表达，包括促凋亡的ING3以提供抗失巢凋亡保护，以及促进上皮-间充质转化(EMT)的蛋白质，包括Vitronectin(VTN)。这些因素相互交织形成一个信号网络，该网络介导RSK2信号，以转录依赖和非依赖的方式为人类癌症提供促进生存和促进转移的优势。此外，我们的初步研究表明，用一种新型的、生物可用的RSK特异性抑制剂FMK-MEA治疗可以显著减少癌细胞的侵袭和肿瘤转移。因此，我们的中心假设是RSK2信号通路在转移性癌症中普遍被激活，它以转录依赖和非转录依赖的方式提供抗失巢凋亡保护，促进癌细胞侵袭和肿瘤转移。RSK2信号通路在肿瘤治疗中是一种很有吸引力的抗转移治疗方法。我们提出了三个具体的目标：(1)确定RSK2是否通过抑制新发现的磷酸化靶标ASK1和下调CREB依赖的促凋亡蛋白ING3的基因表达来为癌细胞提供抗失巢凋亡保护；(2)确定RSK2是否通过磷酸化促转移蛋白IRS1和上调作为RSK2-CREB转录靶标的EMT促进效应因子VTN的基因表达来介导癌细胞的促侵袭和促迁移信号；(3)验证肿瘤标本中RSK2及其信号转导效应分子作为治疗靶点，并将新型RSK抑制剂FMK-MEA与包括顺铂在内的抗癌药物联合使用，在体内外治疗人转移癌。

DESCRIPTION (provided by applicant): Metastasis is responsible for most cancer deaths, including breast, lung or head and neck cancers. However, the molecular processes underlying cellular changes that promote tumor metastasis are still poorly understood. We found that p90 Ribosomal S6 kinase 2 (RSK2) is commonly important to promote multiple steps that comprise tumor metastasis, such as resistance to anoikis induction, cell migration and invasion, and tumor metastasis in metastatic human cancers including breast, lung, and head and neck cancers. Through incorporated phosphor-proteomics and genomics based studies, we demonstrated that RSK2 mediates anti-anoikis, pro-invasive and pro-metastatic signals by phosphorylating a spectrum of downstream factors. In particular, we identified novel substrates of RSK2, including apoptosis signal-regulating kinase 1 (ASK1) and IRS1, to provide anti-anoikis protection and promote pro-invasive and pro-metastatic potentials in cancer cells, respectively. Moreover, we demonstrated that RSK2-CREB signaling pathway regulate gene expression of a group of proteins that are involved in cell death regulation including pro-apoptotic ING3 to provide anti-anoikis protection, as well as proteins that promote epithelial-mesenchymal transition (EMT) including vitronectin (VTN). These factors intertwine with each other to form a signaling network, which mediates RSK2 signals to provide a pro-survival and pro-metastatic advantage to human cancers in both transcription-dependent and -independent manners. Furthermore, our preliminary studies showed that treatment with a novel, bio-available RSK specific inhibitor, FMK-MEA significantly attenuates cancer cell invasion and tumor metastasis. Thus, our central hypothesis is that RSK2 signaling pathway is commonly activated in metastatic cancers, which provides anti-anoikis protection and promotes cancer cell invasion and tumor metastasis in both transcription-dependent and -independent manners. RSK2 signaling represents an attractive anti-metastasis therapy in cancer treatment. Three Specific Aims are proposed: (1) To determine whether RSK2 provides anti-anoikis protection to cancer cells by inhibiting a newly identified phosphorylation target ASK1 and downregulating CREB-dependent gene expression of pro-apoptotic protein ING3; (2) To determine whether RSK2 mediates pro-invasive and pro-migratory signals in cancer cells by phosphorylating pro-metastatic protein IRS1 and upregulating gene expression of EMT promoting effector VTN that is a RSK2-CREB transcription target; (3) To validate RSK2 and its signaling effectors in tumor specimens as therapeutic targets and treat human metastatic cancers in vitro and in vivo using a novel RSK inhibitor FMK-MEA in combination with anti-cancer agents including cisplatin.