The interplay of ion transporters and cytoskeleton in breast cancer migration and metastasis

离子转运蛋白和细胞骨架在乳腺癌迁移和转移中的相互作用

• 批准号: 10381200
• 负责人: Konstantinos Konstantopoulos
• 金额: $ 5.06万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-21 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10381200
• 关键词: 3-Dimensional; Actins; Actomyosin; Affect; Automobile Driving; Biological Models; Biomedical Engineering; Breast Cancer Cell; Breast Cancer cell line; Breast cancer metastasis; CD29 Antigen; Cancer Biology; Cancer Patient; Cancerous; Cell Cycle; Cell Volumes; Cell membrane; Cell surface; Cells; Cellular biology; Cessation of life; Clinical; Complement; Confined Spaces; Cytoplasm; Cytoskeleton; Data; Devices; Distant; Elements; Event; Extracellular Matrix; Fatty acid glycerol esters; Fiber; Gel; Goals; Growth; Image; Imaging Device; In Vitro; Individual; Intracellular Transport; Ion Channel; Ions; Light; Liquid substance; Mediating; Metastatic breast cancer; Microtubules; Modeling; Molecular; Molecular Biology; Mus; Myosin Type II; NHE1; Neoplasm Metastasis; Optics; Organ; Organoids; Patients; Phase; Primary Neoplasm; Regulation; Research Personnel; Role; Solid Neoplasm; Swelling; Testing; Tissues; Transplantation; Travel; Tumor Cell Migration; Water; Work; Zebrafish; cancer cell; cell motility; combat; experimental study; fluid flow; in vivo; in vivo Model; innovation; interdisciplinary approach; interstitial; intravital microscopy; live cell imaging; malignant breast neoplasm; mammary; mathematical model; migration; mouse model; multiphoton microscopy; neoplastic cell; new therapeutic target; novel; optogenetics; patient derived xenograft model; targeted treatment; therapeutic target; tool; triple-negative invasive breast carcinoma; tumor; tumorigenic; water channel

Summary- The inability to clinically treat tumor metastasis is responsible for the majority of cancer patient deaths. Cell migration is a pivotal step in the metastatic dissemination of cancer cells from a primary tumor to distant organs in the body. Cell motility is governed by cell-matrix interactions, the actomyosin cytoskeleton, and cell volume regulation via the involvement of ion transporters, such as the Na[+] /H[+] exchanger 1 (NHE1), as explained by the Osmotic Engine Model (OEM). The roles of cytoskeleton and ion transporters in cell locomotion have been typically studied in isolation. The overarching goal of this project is to employ a multidisciplinary approach involving state-of-the-art bioengineering and imaging tools, quantitative analysis and in vivo models to define the relative roles and potential crosstalk between ion transporters and the cytoskeleton in breast cancer cell migration and metastasis in vivo. This application will test the hypothesis, supported by intriguing preliminary data, that the coordinated action of local isosmotic swelling at the leading edge and shrinkage at the trailing edge mediated by NHE1 and SWELL1, respectively, supports migration in confinement. We further hypothesize that NHE1 and SWELL1 act in concert with cell cytoskeleton to mediate efficient migration and metastasis. Given the lack of targeted therapies for triple negative breast cancer (TNBC), we will prioritize TNBC cell lines and patient-derived xenograft (PDX) tumor cells as models. In Aim 1, we will establish the functional roles of NHE1 and SWELL1 in cell migration inside confining µ-channels of different stiffnesses, in 3D gels and in cell dissemination from 3D breast cancer cell organoids. We will also elucidate the mechanism responsible for the polarized distribution of NHE1 and SWELL1 at the cell front and rear, respectively, and use novel optogenetic tools to alter their spatial polarization and test how these alterations affect the direction and efficiency of cell migration. In parallel, we will develop an innovative mathematical model to identify the key variables that enable OEM-mediated cell motility. In Aim 2, we will delineate the interplay between OEM and the various cytoskeletal constituents, including Beta1 integrins, myosin II, actin and microtubules. Importantly, we will define the intracellular transport mechanisms responsible for NHE1 and SWELL1 shuttling along the longitudinal cell surface. We will also introduce a comprehensive mathematical model to decipher the crosstalk of OEM and cytoskeletal components in regulating migration efficiency. In Aim 3, we will demonstrate the effects of NHE1 and SWELL1 silencing on cell migration in natural mammary tissue tracks in vivo and examine their roles in breast cancer growth and metastasis, using TNBC cell lines and PDXs orthotopically transplanted to the 4th mammary fat pad of mice. We will complement mouse studies with experiments in zebrafish, which enables us to image its entire vasculature at exceptional optical clarity, in order to delineate the roles of ion transporters in different steps of the metastatic cascade. This application brings together a team of investigators with expertise in bioengineering, imaging, cell & molecular biology, quantitative analysis, PDXs, in vivo studies and breast cancer biology.

总结-无法临床治疗肿瘤转移是大多数癌症患者死亡的原因。细胞迁移是癌细胞从原发性肿瘤转移到体内远端器官的关键步骤。细胞运动性由细胞-基质相互作用、肌动球蛋白细胞骨架和通过离子转运蛋白（如Na[+] /H[+]交换器1（NHE 1））参与的细胞体积调节控制，如渗透引擎模型（OEM）所解释。细胞骨架和离子转运蛋白在细胞运动中的作用通常是孤立研究的。该项目的总体目标是采用多学科方法，包括最先进的生物工程和成像工具，定量分析和体内模型，以确定离子转运蛋白和细胞骨架在体内乳腺癌细胞迁移和转移中的相对作用和潜在串扰。该应用程序将测试的假设，支持有趣的初步数据，即协调行动的局部等渗膨胀在前缘和收缩在后缘介导的NHE 1和SWELL 1，分别支持迁移的限制。我们进一步假设NHE 1和SWELL 1与细胞骨架协同作用，介导有效的迁移和转移。鉴于三阴性乳腺癌（TNBC）缺乏靶向治疗，我们将优先考虑TNBC细胞系和患者来源的异种移植（PDX）肿瘤细胞作为模型。在目标1中，我们将确定NHE 1和SWELL 1在不同刚度的限制性μ通道内的细胞迁移中，在3D凝胶中以及在3D乳腺癌细胞类器官的细胞传播中的功能作用。我们还将阐明NHE 1和SWELL 1分别在细胞前部和后部的极化分布的机制，并使用新的光遗传学工具来改变它们的空间极化，并测试这些改变如何影响细胞迁移的方向和效率。同时，我们将开发一种创新的数学模型，以确定使OEM介导的细胞运动的关键变量。在目标2中，我们将描述OEM和各种细胞骨架成分之间的相互作用，包括β 1整合素，肌球蛋白II，肌动蛋白和微管。重要的是，我们将确定负责NHE 1和SWELL 1穿梭沿着纵向细胞表面的细胞内运输机制。我们还将引入一个全面的数学模型来解释OEM和细胞骨架成分在调节迁移效率中的串扰。在目标3中，我们将使用原位移植到小鼠第4乳腺脂肪垫的TNBC细胞系和PDX，证明NHE 1和SWELL 1沉默对体内天然乳腺组织轨道中细胞迁移的影响，并检查它们在乳腺癌生长和转移中的作用。我们将用斑马鱼实验补充小鼠研究，这使我们能够以特殊的光学清晰度对其整个血管系统进行成像，以描绘离子转运蛋白在转移级联的不同步骤中的作用。该应用程序汇集了一个研究人员团队，他们在生物工程，成像，细胞和分子生物学，定量分析，PDX，体内研究和乳腺癌生物学方面具有专业知识。