Annese Tiziana, Tamma Roberto, Ribatti Domenico
Department of Basic Medical Sciences, Neurosciences and Sensory Organs, Section of Human Anatomy and Histology, University of Bari Medical School, Bari, Italy.
Methods Mol Biol. 2022;2475:143-155. doi: 10.1007/978-1-0716-2217-9_10.
Different pro-angiogenic factors, such as vascular endothelial growth factor-A (VEGF-A), have been related to microvascular density, clinicopathologic factors, and poor prognosis in many tumors. VEGF-A binds its receptor 2 (VEGFR2) to induce neo-angiogenesis, a constant hallmark of tumor initiation and progression. Based on VEGF-A/VEGFR2 relevance in tumor angiogenesis, several inhibitors were developed. However, the clinical benefits of anti-angiogenic therapies are limited because tumors activate different mechanisms of drug resistance.The need for understanding tumor biology, limitation or failure of anti-angiogenic therapies, and the demand for a personalized therapeutic approach has boosted the search for robust biomarkers for patient stratification as responder or non-responder to anti-VEGF therapies.This chapter presents a detailed protocol to perform chromogenic VEGF-A mRNA detection and quantification in human tumor bioptic specimens using RNAscope technology and RNA-in situ hybridization (ISH) algorithm. RNAscope for VEGF-A detection, even for small amounts, is compatible with precious clinical samples and diagnostic laboratory workflows.
不同的促血管生成因子，如血管内皮生长因子-A（VEGF-A），已被证实与许多肿瘤的微血管密度、临床病理因素及不良预后相关。VEGF-A与其受体2（VEGFR2）结合以诱导新生血管生成，这是肿瘤发生和进展的一个持续特征。基于VEGF-A/VEGFR2在肿瘤血管生成中的相关性，人们研发了几种抑制剂。然而，抗血管生成疗法的临床益处有限，因为肿瘤会激活不同的耐药机制。理解肿瘤生物学、抗血管生成疗法的局限性或失败以及对个性化治疗方法的需求，推动了对用于患者分层的强大生物标志物的探索，以区分抗VEGF疗法的应答者和无应答者。本章介绍了一种详细方案，用于使用RNAscope技术和RNA原位杂交（ISH）算法在人肿瘤活检标本中进行VEGF-A mRNA的显色检测和定量。用于VEGF-A检测的RNAscope，即使针对少量样本，也与珍贵的临床样本和诊断实验室工作流程兼容。
