Institute of Experimental Medicine and Biotechnology, Samara State Medical University, Chapaevskaya St., 89, 443099 Samara, Russia.
Biomolecules. 2020 Nov 17;10(11):1563. doi: 10.3390/biom10111563.
In the context of modern drug discovery, there is an obvious advantage to designing phenotypic bioassays based on human disease-relevant cells that express disease-relevant markers. The specific aim of the study was to develop a convenient and reliable method for screening compounds with Tumor Necrosis Factor-alpha (TNF-α) inhibitory activity. This assay was developed using cryopreserved ready-to-use cartilage-derived cells isolated from juvenile donors diagnosed with polydactyly. It has been demonstrated that all donor (10 donors) cells were able to respond to TNF-α treatment by increased secretion of pro-inflammatory cytokine IL-6 into subcultural medium. Inhibition of TNF-α using commercially available TNF-α inhibitor etanercept resulted in a dose-dependent decrease in IL-6 production which was measured by Enzyme-Linked Immunosorbent Assay (ELISA). TNF-α dependent IL-6 production was detected in the cells after both their prolonged cultivation in vitro (≥20 passages) and cryopreservation. This phenotypic bioassay based on ready-to-use primary human cells was developed for detection of novel TNF-α inhibitory compounds and profiling of biosimilar drugs.
在现代药物发现的背景下，基于表达与疾病相关标志物的人类疾病相关细胞来设计表型生物测定具有明显的优势。本研究的具体目的是开发一种方便可靠的方法，用于筛选具有肿瘤坏死因子-α（TNF-α）抑制活性的化合物。该测定法是使用从患有多指畸形的青少年供体中分离出的冷冻即用型软骨衍生细胞来开发的。已经证明，所有供体（10 个供体）细胞都能够通过增加促炎细胞因子 IL-6 分泌到亚培养物中来响应 TNF-α 处理。使用市售的 TNF-α抑制剂依那西普抑制 TNF-α 会导致 IL-6 产生的剂量依赖性降低，这通过酶联免疫吸附测定（ELISA）进行测量。在体外（≥20 代）和冷冻保存后延长培养后，均可检测到细胞中 TNF-α 依赖性的 IL-6 产生。已经开发了这种基于即用型原代人细胞的表型生物测定法，用于检测新型 TNF-α 抑制化合物和生物类似药的特性。
