ALTERED GENE EXPRESSION IN CELL CULTURES

本工作说明的目的是使用基因表达阵列检查至少两个人类预侵袭细胞培养系统中的每一个中至少6种化学预防剂处理后的表达变化。 所提出的表达系统允许采用标准化软件包快速读出所提出的RNA水平。乳腺癌细胞的预侵袭性培养系统来源于乳腺癌细胞。 此外，NCI还提供了另外40个细胞样本，可以进行这种RNA分析。这些样品来自人细胞或细胞培养物。 细胞培养物中的初步毒性决定了使用的剂量。 从暴露于对照培养基和三个剂量的两种不同化学预防剂至少三天的重复细胞培养物测定RNA表达水平。 所用的特定化学预防剂为类维生素A、DFMO、抗雌激素、抗雄激素或NSAIDS。 使用的试剂将取决于所用的具体细胞系。 这些研究应确定a）测定的重现性，B）在人类侵袭前细胞暴露于不同类别的化学预防剂后，它们是否具有明显不同的RNA模式，以及（c）是否可以观察到剂量效应。在完成和分析这些初步研究之后，将进行进一步的研究。在三个不同的时间点（加入化学预防剂后4小时、24小时和96小时），采用3种不同剂量的每种试剂，一式两份测定五种不同的试剂。

The objective of this workstatement is to use gene expression arrays to examine for changes in expression following treatment with at least 6 chemopreventive agents in each of at least two human preinvasive cell culture systems. The proposed expression system allows for rapid readout of the proposed RNA levels employing a standardized software package. The preinvasive cell culture systems are derived from mammary. In addition NCI is suppling an additional 40 cell samples on which such an RNA analyses can be performed. These samples are from human cells or cell cultures. A preliminary toxicity in the cell culture determines the doses to be used. The RNA expression levels are determined from duplicate cell cultures, at each dose, exposed to control medium and three doses of two different chemopreventives for a period of at least three days. The specific chemopreventives to be employed will be retinoids, DFMO, antiestrogens , antiandrogens or NSAIDS. The agent to employed will depend on the specific cell line used. These studies should determine a) the reproducibility of the assay, b) whether their are distinctly different RNA patterns for following exposure of human preinvasive cells to different classes of chemopreventive agents, and (c) whether a dose effect can be seen. Following completion and analysis of these initial studies further studies will be undertaken. Five different agents in duplicate at three different time points (4 hours, 24 hours and 96 hours after the addition of the chemopreventive agents) employing 3 different doses of each agents is being assayed.