Targeting Glucose Transporters Using Rapafucins

药物发现中的一个主要挑战是鉴定用于治疗癌症的小分子抑制剂。 具有挑战性的目标，包括蛋白质-蛋白质相互作用和多通道 跨膜蛋白大自然已经为这个问题进化出了一个巧妙的解决方案 如大环天然产物的亲免素配体家族所例示的， 包括雷帕霉素和FK 506除了他们自己的更大的尺寸， 与蛋白质更广泛的相互作用，这些天然产物也招募 FKBP-家族的伴侣蛋白形成更大的二聚体复合物， 与它们各自的靶点mTOR和钙调神经磷酸酶相关联。受此启发 独特的作用模式，我们产生了45，000个雷帕霉素样化合物库， 通过融合雷帕霉素的FKBP结合结构域， 与组合四肽库。筛选该文库导致了 溶质载体（SLC）转运蛋白超家族的多个成员的发现， 包括葡萄糖转运蛋白（GLUT）家族的成员。两种不同的抑制剂 鉴定了葡萄糖转运蛋白（GLUT）的两个亚型：雷帕凝集素A（rapaglutin A，RgA）和雷帕凝集素E（rapaglutin E，RgE）。而RgE RgA对GLUT 1具有高度特异性，RgA似乎抑制多种GLUT亚型。RGA 已经显示在乳腺癌的异种移植模型中是有效的，而RgE 被发现通过阻断DNA- NFAT的结合活性。我们将进一步表征RgA的抗肿瘤活性 通过系统评估其对细胞代谢谱的影响， AMPK-mTOR信号通路。在初步研究中，我们发现RgA 在抑制三阴性乳腺癌细胞方面具有协同作用 （TNBC）。我们将进一步阐明协同机制和 用于抑制体内TNBC肿瘤生长的RgA-苯福明组合。那个RgE 抑制钙刺激的DNA结合活性的NFAT表明一个潜在的作用 GLUT 1在调节细胞内T细胞受体信号传导中的作用。我们将阐明 RgE的免疫抑制活性的机制，并研究 RgE作为体内免疫抑制剂的潜力。最后，使用新的 开发的微阵列平台，我们将筛选rapafucin库对每一个 GLUT的14种亚型，以寻找不同亚型的特异性抑制剂， GLUT可以成为研究GLUT生物学的新化学工具。

A major challenge in drug discovery is to identify small molecule inhibitors for the challenging targets including protein-protein interactions and multi-pass transmembrane proteins. Nature has evolved an ingenious solution to this problem as exemplified by the immunophilin ligand family of macrocyclic natural products, including rapamycin and FK506. Aside from their own larger sizes that enable more extensive interactions with proteins, these natural products also recruit the FKBP-family of chaperones to form much larger dimeric complexes before associating with their respective targets, mTOR and calcineurin. Inspired by this unique mode of action, we generated a 45,000-compound library of rapamycin-like macrocycles, named rapafucins, by fusing the FKBP-binding domain of rapamycin with a combinatorial tetrapeptide library. Screening of this library has led to the discovery of multiple members of the solute carrier (SLC) transporter superfamily, including members of the glucose transporter (GLUT) family. Two distinct inhibitors of GLUT were identified, rapaglutin A (RgA) and rapaglutin E (RgE). Whereas RgE is highly specific for GLUT1, RgA seems to inhibit multiple GLUT isoforms. RgA has been shown to be efficacious in a xenograft model of breast cancer while RgE was found to inhibit intracellular T cell receptor signaling by blocking the DNA- binding activity of NFAT. We will further characterize the antitumor activity of RgA by systematically assessing its effect on cellular metabolomic profiles and the AMPK-mTOR signaling pathway. In preliminary studies, we have found that RgA and phenformin have synergy in inhibiting triple negative breast cancer cells (TNBC). We will further delineate the mechanism of synergy and the effect of the RgA-phenfomin combination for inhibiting TNBC tumor growth in vivo. That RgE inhibited calcium stimulated DNA-binding activity of NFAT suggests a potential role of GLUT1 in regulating intracellular T cell receptor signaling. We will elucidate the mechanism underlying the immunsuppressive activity of RgE and investigate the potential of RgE as an immunosuppressant in vivo. Lastly, using a newly developed microarray platform, we will screen rapafucin libraries against each of the 14 isoforms of GLUTs in search of specific inhibitors of different isoforms of GLUT, which can become new chemical tools for studying the biology of GLUT.