DNA methylation analysis as novel tool for quality control in regenerative medicine.

Stephen Rapko, Udo Baron, Ulrich Hoffmüller, Fabian Model, Leslie Wolfe, Sven Olek
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引用次数: 9

Abstract

Cell-based regenerative medicine, including tissue engineering, is a novel approach to reconstituting tissues that do not spontaneously heal, such as damaged cartilage, and to curing diseases caused by malfunctioning cells. Typically, manufacturing processes to generate cartilage for replacement therapies involve isolation and expansion of cells from cartilage biopsies. A challenge in the field is potential contamination by other cell types (e.g., fibroblast-like cells), which can overgrow the desired cells during culturing and may ultimately compromise clinical efficacy. No standard analytical system has been absolutely effective in ensuring the identity of these cell-based products. Therefore, we tested deoxyribonucleic acid methylation analysis as a quality assessment tool, applying it to Genzyme's Carticel product, a chondrocyte implant that the Food and Drug Administration has approved. We identified 7 potent discriminators by assaying candidate genomic regions derived from methylation discovery approaches and literature searches regarding a functional role of genes in chondrocyte biology. Using a support vector machine, we trained an optimal cell type classifier that was absolutely effective in discriminating chondrocytes from synovial membrane derived cells, the major potential contaminant of chondrocyte cultures. The abundant marker availability and high quality of this assay format also suggest it as a potential quality control test for other cell types grown or manipulated in vitro.

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DNA甲基化分析作为再生医学质量控制的新工具。
以细胞为基础的再生医学,包括组织工程,是一种重建不能自发愈合的组织的新方法,比如受损的软骨,以及治疗由功能失调的细胞引起的疾病。通常情况下,生产用于替代疗法的软骨的制造过程包括从软骨活检中分离和扩增细胞。该领域的一个挑战是其他细胞类型(如成纤维细胞样细胞)的潜在污染,这些细胞在培养过程中可能过度生长,最终可能损害临床疗效。没有标准的分析系统能绝对有效地确保这些细胞基产品的鉴别。因此,我们将脱氧核糖核酸甲基化分析作为一种质量评估工具进行测试,并将其应用于Genzyme的Carticel产品,这是一种获得美国食品和药物管理局批准的软骨细胞植入物。我们通过分析甲基化发现方法获得的候选基因组区域和关于基因在软骨细胞生物学中的功能作用的文献检索,确定了7个有效的鉴别器。使用支持向量机,我们训练了一个最优的细胞类型分类器,它绝对有效地区分软骨细胞和滑膜来源的细胞,软骨细胞培养的主要潜在污染物。丰富的标记可用性和高质量的分析格式也表明,它作为一个潜在的质量控制测试其他类型的细胞生长或操纵体外。
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Tissue engineering
Tissue engineering CELL & TISSUE ENGINEERING-BIOTECHNOLOGY & APPLIED MICROBIOLOGY
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