IDENTIFICATION OF NEW RETINOID REGULATED BIOMARKERS IN BREAST AND OVARIAN CELLS

乳腺和卵巢细胞中新的视黄醇调节生物标志物的鉴定

• 批准号: 6357023
• 负责人: POWELL H BROWN
• 金额: $ 19.33万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-30 至 2001-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6357023
• 关键词: animal tissue; biomarker; breast neoplasms; cell growth regulation; chemoprevention; gene expression; genetic regulation; genetically modified animals; human genetic material tag; human tissue; neoplastic cell; neoplastic growth; northern blottings; nucleic acid probes; ovary neoplasms; polymerase chain reaction; retinoids; statistics /biometry; subtraction hybridization; tissue /cell culture

Retinoids are promising chemopreventive agents in animals and in humans. However, because currently available retinoids are relatively toxic, they are not generally used for cancer prevention. Therefore, receptor-selective retinoids are being tested to develop agents which might prevent cancer with reduced toxicity. The long-term goal of our studies is to identify genes which are regulated by these receptor-selective retinoids and to determine whether the protein products of these genes can serve as surrogate-endpoint biomarkers for clinical chemoprevention trials. Our preliminary studies demonstrated that RCR-selective retinoids including LGD1069 can inhibit the proliferation of normal and malignant breast cells and suppress mammary tumor development in a transgenic mouse model, perhaps with less toxicity than naturally occurring broad- spectrum retinoids. We now propose to identify a set of genes which are modulated by RXR- selective retinoids in breast cells and which regulate growth and invasive properties of these cells, and to determine whether changes in the expression of these genes are associated with successful chemoprevention by retinoids. First, we will determine the set of genes which are up- or down-regulated in human breast and ovarian cells upon treatment by the RCR-selective ligand LDG1069. We will use cDNA expression arrays to determine the genes which are up- or down-regulated panels of normal, premalignant, and malignant normal breast and ovarian cells treated with LGD1069. Second, we will determine whether these specific RXR- regulated genes are involved in controlling the growth and invasion of breast and ovarian cells. Genes identified in Specific Aim 1 which are involved in signal transduction pathways, or are related to proteins which regulate growth or invasion, will have expression and antisense constructs of these genes, and transfect cells to determine how expression or suppression prepared. Transfected cells will show how expression or suppression of these genes affects growth or invasiveness, or affects the actions of LGD1069 on the cells. Third, we will determine whether LGD1069 induces changes in the expression of retinoid-regulated genes in vivo in mammary glands. Using stored tissues representing stages of mammary tumor formation from an SV40 Tag transgenic mouse model treated with LGD1069, we will investigate whether LGD1069 modulated the in vivo expression of markers identified above in normal, and dysplastic, and in malignant tissue. markers tested will also include the proliferation marker Histone H3 (the phosphorylated form), as well as ER, ErbB2, and the retinoid receptors (RARalpha, beta, gamma). Through these studies we will identify promising markers regulated by RXR-selective retinoids which could serve as surrogate endpoint biomarkers in chemoprevention trials. Potential biomarkers altered by LGD1060 will then be tested directly in Projects 1 and 2 using human breast and ovarian tissues.

类维生素A是动物和人类有前途的化学预防剂。然而，由于目前可用的类视黄醇毒性较大，因此通常不用于预防癌症。因此，正在测试受体选择性类维生素A，以开发可以预防癌症并降低毒性的药物。我们研究的长期目标是鉴定受这些受体选择性类维生素A调节的基因，并确定这些基因的蛋白质产物是否可以作为临床化学预防试验的替代终点生物标志物。我们的初步研究表明，包括 LGD1069 在内的 RCR 选择性类视黄醇可以抑制正常和恶性乳腺细胞的增殖，并抑制转基因小鼠模型中乳腺肿瘤的发展，可能比天然存在的广谱类视黄醇具有更低的毒性。我们现在建议鉴定乳腺细胞中由 RXR 选择性类视黄醇调节的一组基因，这些基因调节这些细胞的生长和侵袭特性，并确定这些基因表达的变化是否与类视黄醇的成功化学预防相关。首先，我们将确定在 RCR 选择性配体 LDG1069 处理后人类乳腺和卵巢细胞中上调或下调的一组基因。我们将使用 cDNA 表达阵列来确定经 LGD1069 处理的正常、癌前和恶性正常乳腺和卵巢细胞中上调或下调的基因。其次，我们将确定这些特定的 RXR 调节基因是否参与控制乳腺和卵巢细胞的生长和侵袭。具体目标 1 中鉴定的参与信号转导途径或与调节生长或侵袭的蛋白质相关的基因将具有这些基因的表达和反义构建体，并转染细胞以确定如何准备表达或抑制。转染细胞将显示这些基因的表达或抑制如何影响生长或侵袭性，或如何影响 LGD1069 对细胞的作用。第三，我们将确定 LGD1069 是否会诱导体内乳腺中视黄醇调节基因表达的变化。使用代表来自经 LGD1069 处理的 SV40 标签转基因小鼠模型的乳腺肿瘤形成阶段的储存组织，我们将研究 LGD1069 是否调节上述在正常组织、发育不良组织和恶性组织中识别的标记物的体内表达。测试的标记物还包括增殖标记物组蛋白 H3（磷酸化形式）以及 ER、ErbB2 和类视黄醇受体（RARα、β、γ）。通过这些研究，我们将确定受 RXR 选择性视黄醇调节的有前途的标记物，这些标记物可以作为化学预防试验中的替代终点生物标记物。然后，将在项目 1 和 2 中使用人类乳腺和卵巢组织直接测试 LGD1060 改变的潜在生物标志物。