Speaker Abstract: S-12
Microarray Assessment of Cell Substrates and Stem Cells
Raj K. Puri, M.D., Ph.D., Center for Biologics Evaluation and Research/FDA
DNA microarray technology may offer a unique opportunity to make use of the entire human genome sequence for medical discoveries including characterization and quality assessment of cell substrates used to produce cellular and gene therapy and other biological products. Our goal has been to determine the quality of cell substrates and identity of ES cells by microarray technology, and in doing so, identify critical issues that will help FDA to develop a guidance document and standards for not only microarray technology but also for the characterization of complex cellular products. We used total RNA from a commercially available human embryonic kidney cell line and six approved stem cell lines for microarray analysis. High quality microarrays containing 10,000 cDNAs produced at the Advanced Technology Center, NCI or ~16,659 seventy base pair oligonucleotides produced at the CBER Genomics Program were used. The results were analyzed by GenPix software and the microarray database (mAdb) tools developed by the Center for Information Technology, NIH. We hypothesized that monitoring of gene expression profiles during various growth conditions may provide a novel approach in the assessment of the effect of confluency, transformation and culture conditions on the quality of products made by these cells. Increases in expression of genes belonging to metabolism and stress pathways were observed in over-confluent cells compared to 40% confluent cells. When ES cell derived RNA was analyzed, all six samples expressed multiple markers of the undifferentiated state and shared significant homology in gene expression. A common subset of 92 "stemness" genes was identified. Gene expression was confirmed by a variety of techniques including comparison with databases, RT-PCR, focused cDNA microarrays and immunocytochemistry. Further studies are focused on the characterization of additional cell substrates and cellular products.
2004 FDA Science Forum | FDA Chapter, Sigma Xi | CFSAN | FDA
Last updated on 2004-MAY-28 by frf