ALCAM-mediated cell adhesion and extracellular vesicle biogenesis in bladder cancer

膀胱癌中 ALCAM 介导的细胞粘附和细胞外囊泡生物发生

• 批准号: 10066187
• 负责人: Ariana Kathryn von Lersner
• 金额: $ 3.02万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10066187
• 关键词: 3-Dimensional; ALCAM gene; Activated-Leukocyte Cell Adhesion Molecule; Adaptor Signaling Protein; Adhesions; Adhesives; Affect; Alternative Splicing; Binding; Biogenesis; Biological; Biology; Birds; Bladder; Bladder papilloma; Cancer Patient; Cancer Relapse; Carcinoma; Cell Adhesion; Cell Communication; Cell Line; Cell membrane; Cells; Chemotaxis; Cytometry; Cytoplasmic Tail; DLG4 gene; Data; Detection; Development; Disease; Disease Progression; Electron Microscopy; Embryo; Environment; Event; Goals; Immunoblotting; Immunoglobulin Domain; Immunoglobulins; In Vitro; Intercellular Junctions; Intervention; Laboratories; Length; Leukocyte Adhesion Molecules; Link; Lipid Bilayers; Liquid substance; Literature; Malignant Neoplasms; Malignant neoplasm of urinary bladder; Mass Spectrum Analysis; Mediating; Mediator of activation protein; Membrane; Modeling; Molecular; Monitor; Multivesicular Body; Neoplasm Metastasis; Pathologic; Pathway interactions; Physiology; Plasma; Production; Property; Protein Analysis; Protein Isoforms; Proteins; Publishing; Regulation; Research; Risk; Scaffolding Protein; Site; Techniques; Testing; Tight Junctions; Tumor Cell Migration; Tumor-Derived; Urine; Vesicle; Western Blotting; Work; base; cancer cell; cell motility; clinically relevant; exosome; experimental study; extracellular; extracellular vesicles; in vivo Model; insight; liquid biopsy; member; microvesicles; migration; molecular marker; mortality; nanoparticle; neoplastic cell; novel; particle; prevent; response; synergism; targeted treatment; treatment response; treatment strategy; tumor progression

PROJECT SUMMARY Extracellular vesicles (EVs) are a heterogenous group of lipid bilayer enclosed particles secreted from cells and are found in most, if not all, biological fluids. The term EV encompasses a wide range of vesicle classes, including exosomes (50-150 nm) and microvesicles (150 nm - 1000+ µm), derived from the endosomal pathway after fusion of multivesicular bodies with the plasma membrane or from direct budding from the plasma membrane, respectively. EVs are further characterized by membrane and luminal cargoes that give rise to distinct functional properties. An emerging body of literature suggests that EVs are major contributors in cell-cell communication in normal physiology and pathological events, such as cancer. EVs have been shown to affect cancer cell chemotaxis and metastatic organotropism. Notably, a specific class of EV is dependent upon syntenin-1, an adaptor protein that contains two PSD95/Dlg/zonula occludens 1 (PDZ) domains, for biogenesis and cargo loading through the endosomal pathway. This suggests that upstream effectors which bind to syntenin-1 could modulate its availability, and in turn, regulate exosome biogenesis and cargo loading. Activated Leukocyte Cell Adhesion Molecule (ALCAM) is an adhesion protein in the immunoglobulin superfamily found at sites of cell-cell junctions. ALCAM is a dynamic regulator of cell adhesion through differential shedding of its ectodomain. Our laboratory demonstrated that the full-length ALCAM Isoform (Iso1) resists shedding, promotes adhesion and limits metastasis. Conversely, the alternatively spliced isoform 2 (Iso2) is susceptible to shedding, facilitates motility and metastasis. ALCAM shedding and elevated expression of ALCAM Iso2 correlates with bladder cancer (BCa) disease progression. ALCAM can be linked to EV biogenesis through its association with the cytoplasmic scaffolding protein syntenin-1. Sequestering syntenin-1 via the cytoplasmic tail of ALCAM abrogates motility and metastasis, further underscoring the functional relevance of the regulatory mechanism. Additional preliminary studies revealed that the expression of Iso1 suppresses large EV biogenesis while Iso2-facilitates the biogenesis of large EVs. Based on these findings we hypothesize that ALCAM shedding controls syntenin-dependent EV biogenesis in tumor cells. In addition, we hypothesize that ALCAM-mediated adhesion can control the pro-migratory function of EVs by regulating their cargo. To test this hypothesis, I will utilize cell lines expressing different isoforms of ALCAM and BCa patient-derived plasma to determine the mechanism in which ALCAM-mediated cell adhesion affects EV biology. Experiments proposed in Aim 1 will investigate how modulation of a cells' adhesive state through ALCAM expression alters EV biogenesis and function. EV biogenesis will be assessed through complementary techniques of nanoparticle tracking analysis, western blotting, and microflow cytometry. EV function will be determined with the avian embryo model of metastasis and a 3D organotypic bladder model. Aim 2 will characterize the changes in EV cargos upon ALCAM shedding and disease progression in BCa through palmitoylated-mass spectrometry. Collectively, these data will define a relationship between cell adhesion EV biology, both mechanistically and functionally. Additionally, these findings will identify informative molecular markers to aid in the monitoring of disease state through a non-invasive means.

项目总结