Breast Cancer Biomarkers in the KLF4 Signaling pathway

KLF4是一种锌指转录因子，属于癌基因和肿瘤抑制基因家族的一部分。项目3表明KLF4在大多数乳腺癌中表达上调，并显示出癌基因的特性，这与最近的研究表明它在多能干细胞表型中的作用是一致的。我们证明KLF4通过调节Notch1的转录和功能而发挥癌基因的作用 (Ntc1)途径蛋白。此外，KLF4诱导维甲酸受体的表达，其功能类似于乳腺癌中的肿瘤抑制基因。我们的遗传学和药理学研究表明，这两个受KLF4调控的通路是KLF4功能的关键决定因素。使用y-分泌酶抑制剂抑制Ntc1或给予RXR选择性维甲酸有效地阻断了转化活性 KLF4在体外或体内的表达。这些效应是KFL4特有的，在对照癌基因中没有观察到。 我们已经证明KLF4和Ntc1是在人类乳腺肿瘤中一致表达的预后因子。我们使用这两种蛋白的抗体作为乳腺癌的联合预后因素。 这些研究表明，KLF4-Ntc1途径活性增加的肿瘤具有临床侵袭性，KLF4和Ntc1抗体可用于评估乳腺癌的预后(目标1)。 而Ntc1通过经典途径(即CSL和MAML1)诱导转化，而KLF4抑制CSL并将Ntc1分流到另一条途径。当KLF4或Ntc1在人乳腺MCF10A细胞中表达时，在3D培养中诱导阻断腺泡成熟。KLF4阻断了通常在MCF10A细胞接种到Matrigel层后第6天发生的上皮化。为了改进 小分子的临床前试验我们将使用MCF10A细胞开发一个模型，在该模型中细胞通过Ntc1替代途径转化(目标2)。 抑制Ntc1或促进维甲酸受体信号转导的小分子可阻断KLF4的转化。 使用体外乳腺上皮模型、人乳腺癌异种移植模型和我们开发的ErbB2诱导的乳腺癌快速模型，这些小分子将作为乳腺癌中KLF4效应的抑制剂单独和联合进行测试(目标3)。在临床试验中，我们将探讨维甲酸受体信号在KLF4-Ntc1通路中的作用，并评估γ-分泌酶抑制剂在转移性乳腺癌中的抗肿瘤效果(AIM 4)。在疗效试验中，KLF4-Ntc1通路组件将相互关联 通过肿瘤反应或耐药性来确定哪些患者最有可能受益。

KLF4 is a zinc finger transcription factor and part of a family that includes oncogenes and tumor suppressor genes. Project 3 has shown that KLF4 is upregulated in a majority of breast cancers and shows properties of an oncogene, consistent with recent studies showing its role in the pluripotent stem cell phenotype. We demonstrate that KLF4 functions as an oncogene by regulating the transcription and function of Notchl (Ntc1) pathway proteins. In addition, KLF4 induces expression of retinoid receptors that function similarly to tumor suppressor genes in breast cancer. Our genetic and pharmacologic studies show that these two KLF4-regulated pathways are critical determinants of KLF4 function. Either inhibition of Ntc1 using y-secretase inhibitors or administration of an RXR-selective retinoid efficiently blocked the transforming activity of KLF4 in vitro or in vivo. These effects were KFL4-specific and were not observed for control oncogenes. We have shown that KLF4 and Ntc1 are prognostic factors that are concordantly expressed in human breast tumors. We used antibodies to these two proteins as combination prognostic factors in breast cancer. These studies suggest that tumors with increased KLF4-Ntc1 pathway activity are clinically aggressive, and that KLF4 and Ntc1 antibodies can be use to assess prognosis in breast cancer (Aim 1). Whereas Ntc1 induces transformation through the Classical Pathway (i.e., CSL and MAML1), KLF4 suppresses CSL and shunts Ntc1 to an Alternate Pathway. When expressed in human mammary MCF10A cells, KLF4 or Ntc1 induced a block to acinar maturation in 3D cultures. KLF4 blocked the epithelialization that normally occurs by day 6 following inoculation of MCF10A cells onto a Matrigel layer. For improved preclinical testing of small molecules we will use MCF10A cells to develop a model in which cells are transformed by the Ntc1 Alternate Pathway (Aim 2). Small molecules that inhibit Ntc1 or promote retinoid receptor signaling blocked transformation by KLF4. Using in vitro breast epithelial models, human breast cancer xenografts, and a rapid model of ErbB2-induced breast cancer that we developed, these small molecules will be tested alone and in combination as inhibitors of KLF4 effects in breast cancer (Aim 3). In clinical trials, we will explore the effects of retinoid receptor signaling on the KLF4-Ntc1 pathway and estimate the anti-tumor efficacy of y-secretase inhibitors in metastatic breast cancer (Aim 4). In the efficacy trial, KLF4-Ntc1 pathway components will be correlated with tumor response or resistance to identify which patients are most likely to benefit.