Discovering how oncogenes remodel the surfaceome of cells

发现癌基因如何重塑细胞表面组

• 批准号: 10212408
• 负责人: JAMES A WELLS
• 金额: $ 53.49万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10212408
• 关键词: Amines; Antibodies; Antineoplastic Agents; BRAF gene; Bacteriophages; Bar Codes; Biological Markers; Cell Line; Cell Surface Proteins; Cell surface; Cells; Collaborations; DNA; Data Set; Drug Targeting; Epidermal Growth Factor Receptor; Event; Foundations; Goals; Human; KRAS2 gene; Lead; Logic; MEKs; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Membrane; Membrane Proteins; Methods; Molecular; Mutation; N-terminal; Oncogenes; Oncogenic; Patients; Phage Display; Polysaccharides; Population; Proteins; Proteome; Proteomics; Recombinant Antibody; Regulation; Sampling; Signal Transduction; Surface; Technology; Tissues; antibody detection; base; cancer cell; metaplastic cell transformation; new technology; new therapeutic target; next generation sequencing; particle

Title: Discovering how oncogenes remodel the surfaceome of cells Abstract: The cell surface is the primary hub for cells to communicate with the outside world. Cancer cells have numerous challenges for survival and we hypothesize many of these start at the cell surface made up of some 3000-4000 proteins. Our primary goal is to systematically understand how cancer cells remodel their membrane proteomes (surfaceomes) during oncogenic transformation to survive. We propose to develop enabling technologies for surfaceomics at the population, single cell, and tissue level. We will generate foundational data sets to understand the molecular logic and signaling mechanisms behind the coordinated remodeling events that drive cellular transformation. (Fig 1). We will focus our studies on mutationally activated KRAS and five other highly proliferative oncogenes (EGFR, BRAF, MEK, Akt and PI3K) that together are found in nearly half of all human cancers. We will study how these oncogenes induce coordinate changes in the surfaceome compared to isogenic cells of the same type, in patient derived cancer cells and tissues. We propose new mass spectrometry-based methods to allow for quantification of membrane proteins using two complementary enrichment methods that target surface glycans or surface N-terminal α-amines. We will also develop a new, highly sensitive and multiplexed technology (phage-antibody next generation sequencing, or PhaNGS) to simultaneously detect 100s of surface proteins in very small Figure 1. (A) To understand how oncogenes such as KRAS induce massive cellular changes we will (B) develop new technologies to measure surfaceome changes to understand coordinate regulation, and identify new drug targets. samples or even single cells. The Antibiome Center, which I direct, in collaboration with the Recombinant Antibody Network, has produced recombinant antibodies to 100s of cell surface proteins using Fab-phage display. Each Fab-phage is essentially a DNA barcoded antibody because it has a functional Fab displayed from the phage particle with the DNA encoding it inside. PhaNGS has tremendous potential for highly multiplexed, inexpensive and ultrasensitive means of probing 1000s of cells or cell lines for changes in their surfaceomes. I am excited that these studies will reveal how coordinate remodeling of membrane protein teams contributes to cell state changes. I believe we will also discover new biomarkers and cancer drug targets.

标题：发现癌基因如何重塑细胞表面组 摘要：细胞表面是细胞与外界沟通的主要枢纽。癌细胞 生存面临许多挑战，我们假设其中许多 它们从细胞表面开始，由大约 3000-4000 种蛋白质组成。 我们的主要目标是系统地了解癌细胞如何 重塑其膜蛋白质组（表面组） 致癌转化以求生存。我们建议开发 群体、单细胞、 和组织水平。我们将生成基础数据集 了解背后的分子逻辑和信号机制 驱动细胞转化的协调重塑事件。 （图1）。我们将重点研究突变激活的 KRAS 和其他五种高度增殖的癌基因（EGFR、BRAF、MEK、 Akt 和 PI3K）共同存在于近一半的人类体内 癌症。我们将研究这些癌基因如何诱导协调 与相同的同基因细胞相比，表面组发生变化 类型，在患者来源的癌细胞和组织中。我们提出新的 基于质谱的方法可以量化 使用两种互补富集方法的膜蛋白 靶向表面聚糖或表面 N 末端 α-胺。我们将 还开发了一种新的、高灵敏度的多重技术 （噬菌体抗体下一代测序，或 PhaNGS） 同时检测非常小的数百个表面蛋白 图 1.(A) 了解如何 KRAS 等致癌基因可诱导 我们将发生巨大的细胞变化 (B) 开发新技术 测量表面组变化 了解协调调节， 并确定新的药物靶点。 样品甚至单个细胞。我领导的抗体组中心与重组中心合作 Antibody Network 使用 Fab 噬菌体生产了针对数百种细胞表面蛋白的重组抗体 展示。每个 Fab 噬菌体本质上都是 DNA 条形码抗体，因为它显示有功能性 Fab 来自内部编码 DNA 的噬菌体颗粒。 PhaNGS 具有巨大的潜力 多重、廉价且超灵敏的方法可探测数千个细胞或细胞系的变化 表面组。我很高兴这些研究将揭示膜蛋白如何协调重塑 团队有助于细胞状态的改变。我相信我们还会发现新的生物标志物和抗癌药物 目标。