Mechanisms Driving Cortical Cytoskeleton Dynamics in Cancer Cell Invasion

癌细胞侵袭中皮质细胞骨架动力学的驱动机制

• 批准号: 8677778
• 负责人: KATHRYN M EISENMANN
• 金额: $ 29.25万
• 依托单位: UNIVERSITY OF TOLEDO HEALTH SCI CAMPUS
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8677778
• 关键词: Actins; Adenocarcinoma Cell; Attention; Automobile Driving; Axon; Behavior; Biological Assay; Breast; Breast Cancer Cell; Bulla; Cancer cell line; Cell Adhesion; Cell Line; Cells; Cellular Membrane; Cellular Morphology; Cellular biology; Chimeric Proteins; Clinic; Complex; Coupled; Cytoskeleton; DNA Sequence Rearrangement; Detection; Development; Diagnosis; Disease; Embryo; Energy Transfer; Exhibits; Family; Fatty acid glycerol esters; Fibroblasts; Focal Adhesions; Goals; Growth Cones; Growth Factor; Guanosine Triphosphate Phosphohydrolases; Human; Image; In Vitro; Knowledge; Life; Light; Link; MDA MB 231; Malignant Neoplasms; Mammary Neoplasms; Mammary gland; Matrix Metalloproteinase Inhibitor; Matrix Metalloproteinases; Mediating; Membrane; Mesenchymal; MicroRNAs; Microfilaments; Microscopy; Mission; Molecular; Natural regeneration; Neoplasm Metastasis; Normal Cell; Peptide Hydrolases; Physiological Processes; Play; Proteins; Regulation; Research; Role; Signal Transduction; Testing; Tetanus Helper Peptide; Time; Total Internal Reflection Fluorescent; Tumor Cell Invasion; ameboid movement; base; cancer cell; cell motility; cell type; disability; formin-2; in vivo; insight; migration; mouse model; mutant; neoplastic cell; novel; optical imaging; research study; rho; therapeutic target; tumor; tumor growth; tumorigenesis; whole animal imaging

DESCRIPTION (provided by applicant): Much attention has focused upon the Rho-family of GTPases, which play fundamental roles in actin remodeling and often exhibit enhanced expression and/or activation in human cancers. However, there is a fundamental gap in understanding how key downstream effecter proteins propagate Rho signaling in cancer cells. The long-term goal of these studies is understand the molecular and cellular mechanisms driving rearrangements of the cortical actin cytoskeleton at the leading edge of invasive cancer cells. The objective of this application is to define at the basic molecular and cellular level the role of the Rho effecter mammalian Diaphanous-related formin (mDia2) and its regulator Diaphanous-interacting protein (DIP) in breast cancer cell migration in three-dimensional (3D) matrices both in vitro and in vivo. Perturbation of mDia2 activity via DIP induces a rounded cellular morphology and membrane blebbing. Membrane blebbing is a physiological process that promotes amoeboid cell motility (ACM). ACM is distinct from the mesenchymal-type of cell motility involving focal adhesions and matrix metalloproteinases (MMPs); ACM is a specialized mode of cancer cell migration and is proposed to play an essential role in metastasis. The central hypothesis is that DIP and mDia2 control changes in cortical actin assembly associated with an amoeboid transition during 3D cancer cell migration. The rationale for the proposed research is that therapeutics targeting mesenchymal-type cell migration have largely failed in the clinic for treating breast and other cancers, suggesting that cells can also utilize protease-independent mechanisms for in vivo migration; therefore, multiple modes of cell migration must be targeted to effectively block metastasis. The proposed research is relevant to NIH's mission pertaining to developing fundamental knowledge to potentially help reduce the burdens of human disability. We will pursue three specific aims: 1. To determine the spatial and temporal regulation of mDia2-associated proteins in blebbing cancer cells in 2D matrices; 2; To evaluate the requirement for the DIP/mDia2 node for driving tumor cell adhesion, migration and invasion in vitro; and 3. To determine the functional requirement for mDia2 in breast tumor growth, invasion and metastasis in vivo. To achieve this, Tet-inducible MDA-MB-231 cells, a highly invasive adenocarcinoma cell line, will be developed expressing wild-type (wt) or mutant DIP and/or mDia2 fluorescent fusion proteins, or DIP- or mDia2-directed miRNA. In Aims 1 and 2, quantitative live cell confocal, FRET and TIRF imaging will test the requirement for and spatial/temporal regulation of mDia2 and DIP in migrating breast cancer cells in 2- and 3D matrices. In Aim 3, a mammary fat pad mouse model will assess tumorigenesis/metastasis by multiple platform analyses utilizing histological detection and 3D optical imaging of tumors by whole animal imaging. The proposed research is significant as understanding the molecular basis of amoeboid motility will lend novel insight into mechanisms controlling cancer cell migration and may highlight critically needed alternative therapeutic targets for metastatic disease.

描述（由申请人提供）：非常关注的是GTPases的Rho-family，它们在肌动蛋白重塑中起着基本作用，并且经常在人类癌症中表现出增强的表达和/或激活。但是，了解关键下游效应蛋白如何在癌细胞中传播RHO信号的基本差距。这些研究的长期目标是了解驱动侵入性癌细胞前缘皮质肌动蛋白细胞骨架的重排的分子和细胞机制。该应用的目的是在基本的分子和细胞水平上定义Rho效应子哺乳动物驱动式驱动式甲素（MDIA2）及其调节剂dipanous diaphanous-dipanos-Contractracting蛋白（DIP）在三维（3DD）中的乳腺癌细胞迁移（3DD） ）体外和体内矩阵。 MDIA2活性通过DIP的扰动会诱导圆形的细胞形态和膜出现。膜释放是一种生理过程，可促进变形虫细胞运动性（ACM）。 ACM与涉及局灶性粘连和基质金属蛋白酶（MMP）的细胞运动性的间充质类型不同。 ACM是一种癌细胞迁移的专业模式，并被提议在转移中起着至关重要的作用。中心假设是在3D癌细胞迁移期间，浸入和MDIA2控制与动型转变相关的皮质肌动蛋白组装的变化。拟议的研究的基本原理是，针对间充质型细胞迁移的治疗剂在临床上已经在治疗乳腺癌和其他癌症的临床上几乎失败了，这表明细胞还可以利用与蛋白酶无关的机制来用于体内迁移。因此，必须针对多种细胞迁移模式以有效阻断转移。拟议的研究与NIH的使命有关，涉及发展基本知识，以帮助减轻人类残疾的负担。我们将追求三个具体目标：1。确定2D矩阵中流向癌细胞中与MDIA2相关蛋白的空间和时间调节； 2;评估对驱动肿瘤细胞粘附，迁移和体外侵袭的DIP/MDIA2节点的需求； 3。确定MDIA2在乳腺肿瘤生长，侵袭和体内转移中的功能需求。为此，将开发出表达野生型（WT）或突变体浸入和/或MDIA2荧光融合蛋白，或DIP-或MDIA2-DIA2-DIACIA2-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-DIP-或MIRNA，将开发出TET诱导的MDA-MB-231细胞，高度侵入性的腺癌细胞系。 。在目标1和2中，定量活细胞共焦，FRET和TIRF成像将测试2和3D矩阵中迁移的乳腺癌细胞中MDIA2和MDIA2和DIP的空间/时间调节。在AIM 3中，乳腺脂肪垫小鼠模型将通过整个动物成像对肿瘤进行组织学检测和3D光学成像进行多个平台分析来评估肿瘤发生/转移。拟议的研究非常重要，因为了解变形虫运动的分子基础将为控制癌细胞迁移的机制提供新的见解，并可能突出迫切需要的转移性疾病的替代治疗靶标。

项目成果

SMIFH2-mediated mDia formin functional inhibition potentiates chemotherapeutic targeting of human ovarian cancer spheroids.

SMIFH2 介导的 mDia 福明功能抑制增强了对人卵巢癌球体的化疗靶向。

• DOI: 10.1016/j.bbrc.2016.02.049
• 发表时间: 2016
• 期刊: Biochemical and biophysical research communications
• 影响因子: 3.1
• 作者: Ziske,MeganA;Pettee,KristaM;Khaing,MaNada;Rubinic,Kaitlin;Eisenmann,KathrynM
• 通讯作者: Eisenmann,KathrynM

An mDia2/ROCK signaling axis regulates invasive egress from epithelial ovarian cancer spheroids.

• DOI: 10.1371/journal.pone.0090371
• 发表时间: 2014
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Pettee KM;Dvorak KM;Nestor-Kalinoski AL;Eisenmann KM
• 通讯作者: Eisenmann KM

Carcinoma associated fibroblasts (CAFs) promote breast cancer motility by suppressing mammalian Diaphanous-related formin-2 (mDia2).

• DOI: 10.1371/journal.pone.0195278
• 发表时间: 2018
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Dvorak KM;Pettee KM;Rubinic-Minotti K;Su R;Nestor-Kalinoski A;Eisenmann KM
• 通讯作者: Eisenmann KM

Dia-interacting protein (DIP) imposes migratory plasticity in mDia2-dependent tumor cells in three-dimensional matrices.

Dia 相互作用蛋白 (DIP) 在三维基质中对 mDia2 依赖性肿瘤细胞施加迁移可塑性。

• DOI: 10.1371/journal.pone.0045085
• 发表时间: 2012
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Wyse,MeghanM;Lei,Jun;Nestor-Kalinoski,AndreaL;Eisenmann,KathrynM
• 通讯作者: Eisenmann,KathrynM

mDia2 and CXCL12/CXCR4 chemokine signaling intersect to drive tumor cell amoeboid morphological transitions.

mDia2 和 CXCL12/CXCR4 趋化因子信号交叉驱动肿瘤细胞变形虫形态转变。

• DOI: 10.1016/j.bbrc.2017.01.087
• 发表时间: 2017
• 期刊: Biochemical and biophysical research communications
• 影响因子: 3.1
• 作者: Wyse,MeghanM;Goicoechea,Silvia;Garcia-Mata,Rafael;Nestor-Kalinoski,AndreaL;Eisenmann,KathrynM
• 通讯作者: Eisenmann,KathrynM