Mutagenesis Screen for Prostate Cancer

描述(申请人提供)：前列腺癌是男性癌症相关死亡的第二大常见原因，但对疾病进展的分子机制知之甚少。识别参与这一过程的基因对于识别新的治疗靶点很重要。在这里，我们建议建立一种新的逆转录病毒穿梭载体突变技术来识别推动前列腺癌进展的基因。比较基因组杂交(CGH)、转录组深度测序和蛋白质组学已经成为前列腺癌基因发现的新技术。然而，从整个癌症基因组/转录组/蛋白质组中区分原因突变仍然是一个挑战。我们开发了一种新的突变载体，可以高效、高通量地识别候选的失调基因。在这种方法中，整合的慢病毒穿梭载体通过失调附近基因的表达来诱变靶细胞。载体前病毒：染色体连接在细菌中以质粒的形式被挽救，这些质粒被测序以确定每个载体前病毒的染色体位置。这确定了附近促进癌症生长和/或进展的候选失调基因。我们的穿梭载体技术克服了以前逆转录病毒突变筛选的严重技术限制，即基于聚合酶链式反应的检测效率低下。我们将利用已建立的LNCaP异种移植前列腺癌模型来建立我们的新技术。使用这项技术，我们希望在体内快速识别驱动前列腺癌的新基因。这些基因可以在未来的研究中作为抑制前列腺癌的靶点，或者用作生物标志物。在未来的研究中，我们的方法可能会被用于其他类型的癌症，以加快癌症研究的步伐，具有广泛的潜在影响。

DESCRIPTION (provided by applicant): Prostate cancer is the second most common cause of cancer related deaths in men, yet relatively little is known about the molecular mechanisms of disease progression. Identifying genes that are involved in this process is important to identify new therapeutic targets. Here we propose to establish a novel retroviral shuttle vector mutagenesis technology to identify genes that drive prostate cancer progression. Comparative genome hybridization (CGH), transcriptome deep sequencing, and proteomics have emerged as technologies for prostate cancer gene discovery. However, distinguishing the causal mutations from the entire cancer genome/transcriptome/proteome remains a challenge. We have developed a novel mutagenesis vector that allows efficient high-throughput identification of candidate dysregulated genes. In this approach, integrated lentiviral shuttle vectors mutagenize target cells by dysregulating the expression of nearby genes. Vector provirus:chromosome junctions are rescued as plasmids in bacteria, and these plasmids are sequenced to identify the chromosomal location of each vector provirus. This identifies nearby candidate dysregulated genes that promote cancer growth and/or progression. Our shuttle vector technology overcomes a severe technical limitation of previous retroviral mutagenesis screens, inefficient PCR-based detection. We will take advantage of an established LNCaP xenograft prostate cancer model to establish our novel technology. Using this technology we expect to rapidly identify novel genes that drive prostate cancer in vivo. These genes can be targeted in future studies to inhibit prostate cancer or used as biomarkers. In future studies our approach may be used for other types of cancer to accelerate the pace of cancer research with broad potential impact.