Project 2 Fetuin-A in Prostate Cancer

项目概要：完整项目2 尽管前列腺癌（PCa）的生长在很大程度上是由雄激素驱动的，但一部分通常会发展为 雄激素消融（也称为去势抵抗性PCa; CRPC）难治，并有转移的可能。 我们实验室的初步数据表明，胎球蛋白A，也称为α 2-赫尔曼-施密德 在PCa细胞的生长和“摄取能力”外泌体的产生中，蛋白质是糖蛋白（AHSG）的主要成分。的 所提出的研究的目的是确定胎球蛋白-A在前列腺癌进展中的作用和意义。 我们假设PCa细胞表达异位胎球蛋白-A，其通过TLR 4分泌并被细胞摄取， 介导通过激活pAKT/pERK促进PCa生长的“可摄取”外泌体的生物发生; 此外，我们推测升高的胎球蛋白-A表达可作为前列腺癌的预后生物标志物。三 具体目标如下：目标1。为了确定胎球蛋白-A在AA PCa组织中的表达是否相对较高， 是否高胎球蛋白-A表达与高前列腺癌相关， Gleason评分（>6）和增强的pAKT和pERK。我们将分析胎球蛋白-A的mRNA表达， 使用人PCa的免疫组织化学（IHC）分析NanoString以及pAKT/pERK蛋白水平 组织中将使用多变量线性回归分析来确定胎球蛋白A、pAKT、 AA和CA患者PCa组织中的pERK和Gleason评分，以及其他进展参数， PCa的正边际和扩散。预期胎球蛋白-A、pAKT和pERK将以高水平表达， 在AA患者的PCa组织中，特别是具有高Gleason评分（>6）的那些患者的PCa水平。目标2.确定 异位胎球蛋白-A在外泌体生物发生、促进2-D和3-D生长、运动和侵袭中的作用 PCa细胞的能力。在这个目标中，我们将在两个PCa细胞系中过表达和敲除胎球蛋白-A，以确定 胎球蛋白-A是否在传递生长的“摄取能力”外泌体的生物发生中起因果作用 受体细胞中的信号。我们希望证明来自胎球蛋白-A过表达细胞的外泌体， 促进PCa细胞的2-D、3-D生长和运动以及侵袭，而来自胎球蛋白-A无效细胞的外来体则不会。 目标3：研究靶向胎球蛋白-A介导的信号转导对前列腺增生抑制的有效性， 肿瘤的发生和生长。在这个目标中，我们将利用Pten-敲除小鼠PCa模型来确定 Pten缺失是否需要完整的胎球蛋白-A基因介导其致瘤作用， Pten-/-/胎球蛋白-A-/-双突变小鼠减弱Pten-null的致瘤作用。我们希望肿瘤缩小 与Pten缺失胎球蛋白-A +/+小鼠相比，双突变小鼠的生长。重要性：迫切需要 鉴定可以区分CRPC和惰性PCa的生物标志物，该提案解决了这一需求， 评估了胎球蛋白-A增强PCa肿瘤生长的过程。

PROJECT SUMMARY: FULL PROJECT 2 Even though the growth of prostate cancer (PCa) is largely driven by androgens, a subset usually develops that is refractory to androgen ablation (also known as castration resistant PCa; CRPC) with potential for metastasis. Preliminary data from our laboratory has implicated fetuin-A, also known as alpha 2-Heremans-Schmid glycoprotein (AHSG), in the growth of PCa cells and in the production of “uptake-competent” exosomes. The objective of the proposed studies is to define the role and significance of fetuin-A in prostate cancer progression. We hypothesize that PCa cells express ectopic fetuin-A which is secreted and taken up by the cells via TLR4 to mediate the biogenesis of `uptake-competent' exosomes that promote PCa growth via activation of pAKT/pERK; moreover, we postulate that elevated fetuin-A expression serves as a prognostic biomarker for PCa. Three specific aims are proposed: Aim 1. To determine if fetuin-A expression is higher in AA PCa tissues relative to Caucasian American (CA) PCa tissues and whether high fetuin-A expression is associated with high Gleason Scores (>6) and enhanced pAKT and pERK. We will analyze mRNA expression of fetuin-A using NanoString as well as pAKT/pERK protein levels using immunohistochemistry (IHC) analysis of human PCa tissues. Multivariable linear regression analysis will be used to determine the correlation between fetuin-A, pAKT, pERK and Gleason scores in PCa tissues of AA and CA patients, as well as other progression parameters such as positive margins and spread of PCa. It is expected that fetuin-A, pAKT and pERK will be expressed at high levels in PCa tissues of AA patients particularly those with high Gleason scores (>6). Aim 2. To determine the role of ectopic fetuin-A in exosome biogenesis, promotion of 2-D and 3-D growth, motility and invasive capacity of PCa cells. In this aim, we will overexpress and knockout fetuin-A in two PCa cell lines to determine whether fetuin-A plays a causal role in the biogenesis of `uptake competent' exosomes that transmit growth signals in recipient cells. We expect to demonstrate that exosomes from fetuin-A overexpressing cells will promote 2-D, 3-D growth and motility and invasion of PCa cells while exosomes from fetuin-A null cells will not. Aim 3. To investigate the efficacy of targeting fetuin-A mediated signaling on the suppression of prostate tumor initiation and growth in mice. In this aim, we will utilize the Pten-null mouse model for PCa to determine whether Pten loss requires intact fetuin-A gene to mediate its tumorigenic role and whether loss of fetuin-A in Pten-/-/fetuin-A-/- double mutant mice attenuates the tumorigenic role of Pten-null. We expect reduced tumor growth in the double mutant mice compared to Pten-null fetuin-A+/+ mice. Significance: There is an urgent need to identify biomarkers that can differentiate CRPC from indolent PCa and this proposal addresses that need and evaluates the process by which fetuin-A enhances PCa tumor growth.