Laboratory of Cancer Molecular Genetics, Faculty of Medical Sciences (FCM), University of Campinas (Unicamp), Campinas, SP, Brazil.
Laboratory of Nanobiotechnology, Institute of Genetics and Biochemistry (Ingeb), Federal University of Uberlandia, Uberlandia, MG, Brazil.
Br J Cancer. 2014 Jul 29;111(3):551-8. doi: 10.1038/bjc.2014.331. Epub 2014 Jun 17.
This study aimed to identify novel biomarkers for thyroid carcinoma diagnosis and prognosis.
We have constructed a human single-chain variable fragment (scFv) antibody library that was selected against tumour thyroid cells using the BRASIL method (biopanning and rapid analysis of selective interactive ligands) and phage display technology.
One highly reactive clone, scFv-C1, with specific binding to papillary thyroid tumour proteins was confirmed by ELISA, which was further tested against a tissue microarray that comprised of 229 thyroid tissues, including: 110 carcinomas (38 papillary thyroid carcinomas (PTCs), 42 follicular carcinomas, 30 follicular variants of PTC), 18 normal thyroid tissues, 49 nodular goitres (NG) and 52 follicular adenomas. The scFv-C1 was able to distinguish carcinomas from benign lesions (P=0.0001) and reacted preferentially against T1 and T2 tumour stages (P=0.0108). We have further identified an OTU domain-containing protein 1, DUBA-7 deubiquitinating enzyme as the scFv-binding antigen using two-dimensional polyacrylamide gel electrophoresis and mass spectrometry.
The strategy of screening and identifying a cell-surface-binding antibody against thyroid tissues was highly effective and resulted in a useful biomarker that recognises malignancy among thyroid nodules and may help identify lower-risk cases that can benefit from less-aggressive management.
本研究旨在寻找甲状腺癌诊断和预后的新型生物标志物。
我们构建了一个人源单链可变片段(scFv)抗体文库，该文库使用 BRASIL 方法（生物淘选和选择性相互作用配体的快速分析）和噬菌体展示技术针对肿瘤甲状腺细胞进行了选择。
通过 ELISA 证实了一个具有高反应性的克隆 scFv-C1 与甲状腺乳头状瘤蛋白特异性结合，该克隆进一步在包含 229 个甲状腺组织的组织微阵列上进行了测试，包括：110 个癌（38 个甲状腺乳头状癌（PTC），42 个滤泡状癌，30 个 PTC 的滤泡变体），18 个正常甲状腺组织，49 个结节性甲状腺肿（NG）和 52 个滤泡性腺瘤。scFv-C1 能够区分癌与良性病变（P=0.0001），并且优先与 T1 和 T2 肿瘤阶段反应（P=0.0108）。我们进一步使用二维聚丙烯酰胺凝胶电泳和质谱鉴定了一个包含 OTU 结构域的蛋白 1，即 DUBA-7 去泛素化酶，作为 scFv 的结合抗原。
筛选和鉴定针对甲状腺组织的细胞表面结合抗体的策略非常有效，产生了一种有用的生物标志物，可识别甲状腺结节中的恶性肿瘤，并可能有助于识别低风险病例，从而受益于不那么激进的管理。
