The emergence of extracellular vesicles in urology: fertility, cancer, biomarkers and targeted pharmacotherapy

Figures & data

Fig. 1.  Extracellular vesicle (EV) origin: EV may originate from the endosomal compartment by exocytosis of vesicles formed within the multivesicular bodies or (a) from the cell surface by budding of plasma membrane. These shedding vesicles, sorted from the cell surface by budding of cell plasma membrane, are also named microvesicles. (b) Exocytic multivesicular bodies fuse with membrane after cell stimulation and release by exocytosis vesicles named exosomes. (b) These multivesicular bodies are created within the Golgi apparatus as a result of endosome compartmentalization. The insets are representative transmission electron microscopy of exosome generation from a multivesicular body and of vesicle generation by budding of plasma membrane (modified, in part, from Refs. (3) and 7).

Table I. Extracellular vesicle mechanisms of action with respect to fertility

Action	Mechanism
Promote motility	Fusion c spermatozoa at sperm neck → Ca^2 + signalling tools (prog receptor/cADPR/RyRs)
Immunosuppressive	Fusion c spermatozoa → CD59 (membrane attack complex inhibitory protein) delivery → Inhibits female compliment system lysis
	Chromogranin B is bactericidal
	Inhibits PMN phagocytosis
Antioxidant	NADPH inhibition → decreased superoxide anion generation by PNMs
Capacitation and acrosome reaction	Fusion c spermatozoa transfers cholesterol (preventing premature activation in lower female reproductive tract)
	Increases progesterone sensitivity (progesterone released by cumulus cells and stimulates acrosome reaction)

EV are key modulators in reproduction promoting spermatozoa motility, protecting spermatozoa from active and innate immune responses, act as an antioxidant and promote capacitation and acrosome reaction (14–18).

Fig. 2.  Tumour-derived EV and local invasion and metastasis: EV derived from primary tumour act to enhance matrix remodelling via a) matrix metalloproteinase (MMP) and urokinase-type plasminogen activator (uPA), enhance b) endothelial angiogenesis via vascular endothelial growth factor (VEGF) and c) EV released from PC3 cells activate fibroblasts sending antiapoptotic signals and growth signals (23, 25). Ultimately, EV tumour release leads to downstream enhanced tumour cell migration, adhesion and invasion.

Fig. 3.  Tumour cells release extracellular vesicles that can influence the malignant phenotype. Various examples include and are not limited to: (a) drug resistance; EV can influence the efflux of chemotherapeutic drugs via various mechanisms including ATPase and drug transporters. (b) apoptosis; through FasL and TRAIL, EV can induce apoptosis in activated antitumor T cells, abrogating T-cell-mediated apoptosis of tumour cells. (c) Local invasion and metastasis; EV can promote local invasion by activating fibroblasts and reducing fibroblast apoptosis, enhancing extracellular matrix degradation (mRNAs for MMP2 and MMP9), promoting angiogenesis (mRNA VEGF, FGF2, angiopoietin1) and increasing tumour cell adhesion. EV may enhance metastasis by promoting a pre-metastatic niche in lung tissue via upregulation of VEGFR1 expression, MMP2 in lung blood vessels and MMP9 in alveolar epithelial cells and blood vessels. (d) Immunosuppression; EV can alter monocyte differentiation into myeloid suppressive cells. This inhibits T-cell proliferation. Inhibiting T-cell responses upstream would abrogate antitumor immune potential. This figure is modified from (66).

Fig. 4.  Tumour extracellular vesicles and chemoresistance (67). Tumour EV act in 2 ways to reduce the efficacy of chemotherapy. (1) intracytoplasmic chemotherapy exportation via shedding vesicles. (2) Antibody sequestration.

Table II. Proposed therapeutic strategies to mitigate extracellular vesicle effects in carcinogenesis

Category	Target
Block tumour cell exocytosis of EV	1) Alter creation of EV in endoplasmic reticulum 2) Block membrane fusion of EV to plasma membrane to prevent exocytosis
Neutralize EV released from tumour cells	1) Antibody blockade of EV to prevent binding and fusion of EV to target cells (target neoplasm specific membrane bound receptors) 2) Selectively filter EV from circulation → Haemopurification – antibody bound hollow fibre cartridge to filter selected EV from blood
Use EV from therapeutic cell population (ideal phenotype/mesenchyme stem cells/bone marrow stem cells) to alter phenotype of malignant cell population	1) Reverse chemoresistance 2) Stop growth factor production/release 3) Prevent formation of pre-metastatic niche

Blocking tumour cell shedding or exocytosis of EV may act to mitigate the contribution of tumour EV to local tumour invasion, metastasis, immune evasion and chemoresistance. Similarly, neutralizing EV already released from tumour cells via an antibody blockade of binding/fusion with target cells or filtering tumour EV from circulation may be effective. Another strategy is to use EV from a therapeutic cell population such as mesenchymal or bone marrow stem cells to alter the phenotype of the malignant cell population to reverse chemoresistance, and metastasis (29, 53) (67–71).

Table III. Extracellular vesicle characteristics that are favourable as a vehicle of therapy

1) Cell-type specificity
2) Predictable endocytosis/fusion with effector cells
3) Lipophilic
4) Not filtered by glomerulus
5) Diverse carrying capacity
• Cell surface receptors/proteins
• Cytosolic protein
• DNA
• mRNA
• miRNA
• Long non-coding RNA

EV are ideal vehicles of targeted pharmacotherapy because of cell-type specificity, predictable patterns of endocytosis/fusion with effector cells, lipophilic properties, and can carry a large array of cargo including cell surface receptors, cytosolic protein's, DNA, mRNA and miRNA.

Thery C, Ostrowski M, Segura E. Membrane vesicles as conveyors of immune responses. Nat Rev Immunol. 2009; 9: 581–93. [PubMed Abstract].

 Google Scholar

Arienti G, Carlini E, Saccardi C, Palmerini CA. Role of human prostasomes in the activation of spermatozoa. J Cell Mol Med. 2004; 8: 77–84. [PubMed Abstract].

 Google Scholar

Rooney IA, Atkinson JP, Krul ES, Schonfeld G, Polakoski K, Saffitz JE et al. Physiologic relevance of the membrane attack complex inhibitory protein CD59 in human seminal plasma: CD59 is present on extracellular organelles (prostasomes), binds cell membranes, and inhibits complement-mediated lysis. J Exp Med. 1993; 177: 1409–20. [PubMed Abstract].

 Google Scholar

Castellana D, Zobairi F, Martinez MC, Panaro MA, Mitolo V, Freyssinet JM et al. Membrane microvesicles as actors in the establishment of a favorable prostatic tumoral niche: a role for activated fibroblasts and CX3CL1-CX3CR1 axis. Cancer Res. 2009; 69: 785–93. [PubMed Abstract].

 Google Scholar

Abe K, Shoji M, Chen J, Bierhaus A, Danave I, Micko C et al. Regulation of vascular endothelial growth factor production and angiogenesis by the cytoplasmic tail of tissue factor. Proc Natl Acad Sci USA. 1999; 96: 8663–8. [PubMed Abstract] [PubMed CentralFull Text].

 Google Scholar

Yang C, Robbins PD. The roles of tumor-derived exosomes in cancer pathogenesis. Clin Dev Immunol. 2011; 2011: 1–11.

 Google Scholar

Safaei R, Larson BJ, Cheng TC, Gibson MA, Otani S, Naerdemann W et al. Abnormal lysosomal trafficking and enhanced exosomal export of cisplatin in drug-resistant human ovarian carcinoma cells. Mol Cancer Ther. 2005; 4: 1595–604. [PubMed Abstract].

 Google Scholar

Cocucci E, Racchetti G, Meldolesi J. Shedding microvesicles: artefacts no more. Trends Cell Biol. 2009; 19: 43–51. [PubMed Abstract].

 Google Scholar

Lötvall J, Hill AF, Hochberg F, Buzás EI, Di Vizio D, Gardiner C. Minimal experimental requirements for definition of extracellular vesicles and their functions: a position statement from the International Society for Extracellular Vesicles. J Extracell Vesicles. 2014; 3: 26913. doi: http://dx.doi.org/10.3402/jev.v3.26913.

 Google Scholar

Renzulli JF 2nd, Del Tatto M, Dooner G, Aliotta J, Goldstein L, Dooner M et al. Microvesicle induction of prostate specific gene expression in normal human bone marrow cells. J Urol. 2010; 184: 2165–71. [PubMed Abstract].

 Google Scholar

Grange C, Tapparo M, Collino F, Vitillo L, Damasco C, Deregibus MC et al. Microvesicles released from human renal cancer stem cells stimulate angiogenesis and formation of lung premetastatic niche. Cancer Res. 2011; 71: 5346–56. [PubMed Abstract].

 Google Scholar

Rini BI. Metastatic renal cell carcinoma: many treatment options, one patient. J Clin Oncol. 2009; 27: 3225–34. [PubMed Abstract].

 Google Scholar