Transient receptor potential ion-channel subfamily V member 4: a potential target for cancer treatment

The transient receptor potential ion-channel superfamily consists of nonselective cation channels located mostly on the plasma membranes of numerous animal cell types, which are closely related to sensory information transmission (e.g., vision, pain, and temperature perception), as well as regulation of intracellular Ca2+ balance and physiological activities of growth and development. Transient receptor potential ion channel subfamily V (TRPV) is one of the largest and most diverse subfamilies, including TRPV1–TRPV6 involved in the regulation of a variety of cellular functions. TRPV4 can be activated by various physical and chemical stimuli, such as heat, mechanical force, and phorbol ester derivatives participating in the maintenance of normal cellular functions. In recent years, the roles of TRPV4 in cell proliferation, differentiation, apoptosis, and migration have been extensively studied. Its abnormal expression has also been closely related to the onset and progression of multiple tumors, so TRPV4 may be a target for cancer diagnosis and treatment. In this review, we focused on the latest studies concerning the role of TRPV4 in tumorigenesis and the therapeutic potential. As evidenced by the effects on cancerogenesis, TRPV4 is a potential target for anticancer therapy.


Introduction
Malignant tumors are still the primary health problems that plague people 1 , which are clinically treated mainly through chemotherapy, biological therapy, radiotherapy, and surgical resection 2 . However, chemotherapy usually causes severe side effects and proneness to drug resistance 2 . Therefore, researchers have endeavored to find molecular targeted drugs which are less toxic and more efficient. Targeted therapy using drugs which designed based on antitumor targets interferes with or even blocks the physiological activities of tumors 3 . Drugs that target cancer cell membrane receptors have the advantages of high affinity and recognition efficiency 4 . For instance, drugs targeting epidermal growth factor receptors and some other cell membrane receptors have been clinically used 5 . Several important cellular functions are related to transmembrane potentials and lie under the control of ion channels. The role of ion channels play in connecting the intracellular to the extracellular environment and controlling almost any cellular function makes ion channels ideal potential therapeutic targets. Nowadays, increasing interest has been given to ion channels as potential drug targets in many cancer conditions 6,7 . Many observations found that some cancer cell lines display unusual ion channel expression, and the altered ion transport may play an important role in human cancer progression.
When ion channels open, certain ion species can pass through which affects such basic cellular characteristics, including membrane potential, cell volume, and the state of intracellular signaling pathways. Thus, aberrant expression or function changes of ion channels may produce global or local, spatially restricted changes in these characteristics and driving the transformation of normal cells into malignant derivatives [8][9][10][11] . Some previous research also found that regulating of certain ion channels can inhibit the occurrence and development of tumor [12][13][14][15][16] . Beyond that, the expression level of transient receptor potential ion channel, subfamily V, member 4 (TRPV4) vary in different tumors and play important role in multiple processes of tumor progression. TRPV4 is a calcium-permeable nonselective cation channel of TRP family, which biologically involved in osmotic sensitivity and mechanosensitivity 17,18 . TRPV4 participates in many physiological processes, such as liver 19 , intestinal 20 , renal 21 and bladder 22 functions, growth and structural integrity of the skeleton 23 , together with systemic osmotic pressure induced by the brain 24 . Until now, TRPV4 has been reported functionally related to cell proliferation 25 , differentiation 26 , apoptosis 27 , migration 28 , and many other physiological processes. In addition, TRPV4 has also been significantly correlated with tumor angiogenesis 29 . Targeting TRPV4 may have inhibitory effects on tumor onset and progression, so it is a potential prognostic index and therapeutic target for malignant tumors. We herein reviewed the research focus on the relationship between TRPV4 and cancer, also explored the mechanism of TRPV4-mediated oncogenesis and the strategy that target TRPV4 for tumor metastasis. We hope this finding provide valuable reference for further research and clinical application.

TRP superfamily
Similar to voltage-dependent cation channel, TRP channel also has six-transmembrane (S1-S6) domains, and both the N-terminus and C-terminus are intracellular. The segment between S5 and S6 of the TRP channel is embedded to form an ion-passage channel 30 . The S4 fragment lacks positively charged amino acid residues of that in normal voltage-dependent cation channel, i.e., the TRP channel is nonvoltage dependent. Besides, there are several ankyrin (ANK) repeat domains at the N-terminus of many TRP subtypes 31 . According to the amino acid sequence homology, more than 30 mammalian TRP channels have been classified into seven subfamilies: TRPC, TRPV, TRPM, TRPA, TRPN, TRPML, and TRPP 32 . The TRP channel is a nonselective cation channel on the cell membrane through which calcium and sodium ions mainly pass 33 . However, the ratios of selectivity of calcium ion to that of sodium ion (PCa/PNa, P stand for permeability) are different, ranging from higher than 100:1 to lower than 0.05:1 34,35 . In addition, many divalent ions, such as magnesium (Mg 2+ ), zinc (Zn 2+ ), manganese (Mn 2+ ), and cobalt (Co 2+ ), can also pass through TRPM6 and TRPM7 channels [36][37][38] .
As a crucial sensor of cells, TRP channels can transmit information between intracellular and extracellular, also being regulated by changes in messenger molecules, compounds, temperature, and osmotic pressure 39 . Different TRP channels have various regulatory mechanisms, many of which are modulated by a variety of stimulations. Although the activation mechanisms of different TRP channels vary, they can be roughly classified into receptor activation, ligand activation, and temperature-change activation. G protein-coupled receptors or tyrosine kinase receptors can activate TRP channels by activating phospholipase C (PLC) 40,41 . Some exogenous small molecules, compounds, inorganic ions, and endogenous substances can also activate these channels 42 . Temperature changes directly activate multiple TRP channels, and the activation thresholds of different temperaturesensitive TRP channels vary 43 . TRPV4 is activated by moderate heat (>24-27°C), while TRPA1 is activated by noxious cold at 17°C and below. TRP channel is widely distributed in the peripheral nervous system, skin, cardiovascular system, respiratory system, gastrointestinal system, genitourinary system, and immune systems in addition to the central nervous system [44][45][46][47][48] . Unlike the classical voltage-dependent cation channel that participates in specific functions, TRP channel has a variety of functions, mainly including temperature and pain perception 49 , gustation 50 , feeling of mechanical force 51 , mediation of PLC-dependent calcium influx 52 , maintenance of cell ion homeostasis 35 , as well as regulation of cell growth 53 , neurotransmitter release, and hormone secretion 54,55 .
Recently, several members of the TRP superfamily have been confirmed to play certain roles in tumor progression. Accumulating evidence has indicated that TRP members were expressed abundantly in cancer cells and closely related to tumor progression. Kim et al. reported that TRPM7 played a key role in the growth and survival of gastric cancer cells 56 . Elevated expression of TRPC5 protein is associated with the drug resistance of breast cancer cells 57 , and promotes colon cancer metastasis via the hypoxia-inducible factor 1-alpha/Twist (HIF-1α/ Twist) signaling pathway 58 . Meanwhile, TRPV4 is aberrantly expressed in many tumors and strongly corelated with the prognosis of cancer. The different members of the TRP family and their main characteristics are summarized in Table 1, including the physiological functions, activation conditions, and their effects on cancer.

Structure and function of TRPV4
The TRPV family consists of six members (TRPV1-TRPV6), which all function as tetramers. Among them, TRPV1-TRPV4 have moderate permeabilities for calcium ions, with the PCa/PNa ratios of 1-10. With the ratios of over 100, TRPV5-TRPV6 are highly permeable for calcium ions 34 . The TRPV4 ion channel was described in detail dating back to 2000, and characterized as a volume-regulated channel due to osmotic sensitivity and cell volume regulation 59 . TRPV4 gene encodes TRPV4 ion channel protein, which was initially referred to as "vanilloid-receptor related osmotically activated channel" and "OSM9-like transient receptor potential channel, member 4" 60 , as a member of the vanilloid subfamily in the TRP superfamily 61 . TRPV4 channel protein consisting of 871 amino acids has a homodimeric tetramer structure which is similar to those of other transient potential receptor proteins, with six transmembrane spanning α-helices (S1-S6) per monomer 62 . The structure of TRPV4 is shown in Fig. 1. In addition to the transmembrane region, the remaining part of this protein is located in the cytoplasm. Similar to other TRPVs, it has six ANK repeats at the N-terminus, which are essential for the regular functioning of ion channels and protein-protein interactions. As a nonselective cation channel (Ca 2+ or Mg 2+ as the permeating extracellular cation), TRPV4 is characterized with a moderate high Ca 2+ permeability ratio (PCa/PNa= 6-10, PMg/PNa = 2-3) 63,64 . The pore-forming loop that allows the ionic flow is located between S5 and S6 domains of TRPV4 65 . Some molecules, such as phosphatidylinositol 4,5bisphosphate, can bind firmly to the ANK repeats end of TRPV4, thereby inhibiting the effects of TRPV4 66 . Temperature, mechanical force, hypotonia, phorbol ester derivatives, and other physical and chemical stimuli can activate TRPV4, allowing calcium-based cations to rapidly enter the cytoplasm to maintain osmotic pressure stability and signal transmission 66 . The representative agonists and antagonists of TRPV4 are organized in Table 2. TRPV4 is widely expressed in the nervous system 67 , immune system 68 , eye 69 , ear 70 , cardiovascular system 71 , respiratory system 72 , urinary system 73 , and digestive system 74 . Moreover, TRPV4 maintains osmotic pressure homeostasis by activating, rapidly and efficiently causing the influx of calcium-based cations, and maintaining cell morphology 75 . When skin tissue is physically and chemically stimulated, opening of the TRPV4 promotes the mechanical responses of subcutaneous fibroblasts and endothelial cells, manifested as vasodilation and skeletal muscle relaxation 76 . Different physical and chemical stimuli include heat, mechanical force, and endogenous substances, such as arachidonic acid and its cytochrome P450-derived metabolites (epoxyeicosatrienoic acids), endocannabinoids (anandamide and 2-arachidonoylglycerol), as well as synthetic a-phorbol derivatives can activate TRPV4. TRPV4 integrates multiple stimuli, then transmitting calcium signals and inducing a series of stress responses, such as promotion of release of nitric oxide, prostaglandin I2, and endothelial-derived enoic acid in the vascular endothelial system, relaxation of vascular smooth muscles, production of inflammatory factors (e.g., interleukin-6 (IL-6)) in lung tissue, and development of inflammatory responses 77,78 . At the early stage of vascular and neuronal development, activation of the TRPV4 channel of capillary endothelial cells and neurons activates downstream phosphatidylinositol 3kinase (PI3K) and induces the activation of α-integrin protein, thereby facilitating the localization and remodeling of neurons and endothelial cells 79 . In adipocytes, TRPV4 is involved in fatty acid metabolism. Activating TRPV4 not only increases fatty acid synthesis by regulating RAC-alpha serine/threonine-protein kinase (AKT) phosphorylation but also attenuates fatty acid oxidation to reduce heat production 80 .

TRPV4 is involved in tumor onset and progression
Abnormal expression of TRPV4 is closely related to tumor formation and metastasis, which is higher in gastric cancer, lung cancer, and colorectal cancer cells, but lower in esophageal cancer and prostate cancer cells than in normal tissue cells according to the researches on TRPV4 ( Table 3). The expression data of TRPV4 obtained from oncomine (https://www.oncomine.org/resource/main.html) show similar results (Fig. 2). Since TRPV4 can be activated at the condition of body temperature, its high expression in these cancer cells may led the intracellular

Role of TRPV4 in cell proliferation and differentiation
Proliferation and differentiation are two basic cell biological events. It is well-accepted that cancer cells require unlimited replicative potential to generate macroscopic tumors 81 . This capability is different from the behaviors of most normal cell lineages which have limited number of successive growth-and-division cycles 82 . Most cancer cells have been considered immortal after transformation from normal cells, as the starting point of tumorigenesis. TRPV4 may dominate in the process of proliferation, and it's upregulation is strongly linked to the proliferation of hepatic stellate cells (HSCs). Song et al. reported that the TRPV4 mRNA and protein expression levels of rat HSC-T6 cell line after treatment with transforming growth factor β1 significantly exceeded those of control group. However, TGF-β1-induced HSC-T6 cell proliferation was inhibited by Ruthenium Red (a nonspecific inhibitor of TRPV4) or synthetic siRNA targeting TRPV4 83 . When TRPV4 activated, HSCs can be transformed into myofibroblasts which in turn secrete a large amount of collagen, leading to liver fibrosis, damage to liver tissue structure, cirrhosis, and eventually liver cancer 84,85 . Accordingly, increased expression of TRPV4 can indirectly induce liver cancer by stimulating the proliferation of HSCs. Functionally expressed in oligodendrocyte precursor cells showed that the overactivation of TRPV4 promoted the proliferation and/or migration of esophageal squamous cell carcinoma 87 . In addition, TRPV4 activation may selectively inhibits tumor endothelial cell proliferation via inhibition of ERK1/2 phosphorylation 25 . Studies have also shown that calcium-sensing receptor (CaSR) and TRPV4 were colocalized in gastric cancer cells, and CaSR activation evoked TRPV4-mediated Ca 2+ entry promote gastric cancer cells proliferation 88 .
If a gene cannot be specifically expressed during maturation, the differentiation process is inhibited, which is primarily responsible for tumorigenesis. The growth and differentiation of keratinocytes are affected by intracellular and extracellular Ca 2+ concentrations. When the extracellular Ca 2+ concentration is low, primary keratinocytes remain undifferentiated. Conversely, cell proliferation is suppressed and differentiation is thus facilitated in the presence of high-concentration Ca 2+89 . TRPV4 is highly expressed in healthy or inflamed skin, whereas lowly or even not expressed in precancerous lesions and nonmelanoma skin cancer. In human keratinocytes, activation of TRPV4 stimulates the release of IL-8, which in turn downregulates TRPV4 expression 90,91 . Hypothetically, low expression of TRPV4 in skin cancer affects the release of ATP and autocrine communication between keratinocytes by regulating Ca 2+ homeostasis, then decreasing extracellular Ca 2+ concentration, keeping cells intact and ultimately inducing tumor formation 91 . In Fig. 1 Structure of TRPV4. Similar to other transient potential receptor proteins, TRPV4 is consisted of 871 amino acids, has a homodimeric tetramer structure with six transmembrane spanning α-helices (S1-S6) per TRPV4 monomer   192,197,198 ND not determined summary, TRPV4 plays an important role in the proliferation and differentiation of cells, which further affects the progression of cancer.

Role of TRPV4 in cell apoptosis
In multicellular organisms, the total number of cells is delicately balanced by the cell-generating effects of mitosis and cell death induced through apoptosis, which, when disrupted, results in cancer progression 92 . As a selfmonitoring mechanism of organisms for hyperproliferative cells, apoptosis is also an effective means of tumor treatment. TRPV4 can inhibit the expressions of apoptotic proteins via a variety of (direct and indirect) pathways, and plays an indispensable role in suppressing apoptosis. Knockdown of TRPV4 expression by siRNA or pharmacological inhibition of TRPV4 can attenuate neuronal apoptosis 18 . Nonetheless, overactivation of TRPV4 can induce apoptosis. Activation of TRPV4 by GSK1016790A induces apoptosis by downregulating PI3K/Akt and upregulating p38 MAPK signaling pathways 93 . TRPV4 is a nonselective cation channel that works mainly based on channel switches. Zhan et al. treated HSC-T6 cells with 4α-phorbol 12,13-didecanoate (a TRPV4 activator) which then suppressed apoptosis and enhanced autophagy 94 . There are existing insights suggested substantial cytotoxicity of TRPV4 overactivation. GSK1016790A can cause strong calcium-overload and cellular disarrangement, increased the rate of apoptosis, and strongly inhibited human melanoma cell lines (A375, SK-MEL-28, MKTBR) proliferation/survival, similarly in HaCaT keratinocytes 90 , also in breast cancer cell line MDA-MB-468 pharmacological activation of TRPV4 produced pronounced cell death through apoptosis and oncosis 27 . It can be seen that activation of TRPV4 can induce the occurrence of apoptosis from above results. Classical calcium signaling pathway has manifested that calcium influx can regulate apoptosis by regulating caspase signaling pathway which was regulated by calpain. This may also be the main mechanism of TRPV4 in apoptosis.

Role of TRPV4 in tumor metastasis
It is well-documented that the expression level of TRPV4 in tumor cells was positively correlated with their metastatic ability. Lee et al. performed high-throughput sequencing for isogenic breast cancer cell lines, and found that the expression levels of TRPV4 in 4T07 and 4T1 cells which were prone to exudation and spread during metastasis were abnormally elevated 95 . Fusi et al. detected the expression of TRPV4 by immunohistochemical assay and found that the TRPV4 in weakly metastatic squamous cell carcinoma and basal cell epithelial cancer were significantly lower than that of normal skin tissue, whereas the expression in strongly metastatic malignant melanoma was higher than that of normal tissue 89 . Mrkonjić et al. transfected HEK293 cells with TRPV4. As a result, the cell migration ability was significantly enhanced compared with that of the blank control group. After transfecting with TRPV4 lacking a phosphoinositidebinding site, TRPV4 of HEK293 cell opens continuously which allowing further augmentation of cell motility and directionality 96 . Tumor-related death has mainly been attributed to metastasis 97 . Taken together, TRPV4 is abnormally highly expressed in many tumors which significantly correlated with their metastasis potential. Hence, it may be a therapeutic target for inhibiting tumor metastasis.

Influence of TRPV4 on tumor metastasis and possible mechanism
Regardless of significant advances achieved in research, diagnosis, and treatment of cancers, the vast majority of advanced metastatic cases, with rare exception, cannot be cured by current regimens. Being closely related to the prognosis of tumor patients, TRPV4 is probably one of the main participants in tumor metastasis. Therefore, it is of great significance to clarify the mechanism of TRPV4 in tumor metastasis.

TRPV4 promotes epithelial-mesenchymal transition (EMT)
EMT confers on tumor cells properties that are critical to invasion and metastatic dissemination, notably increased motility, invasiveness, and degradation of components of the extracellular matrix. The cells are transformed into spindle-shaped fibroblast-like ones, accompanied by enhanced motility 98 . TRPV4 can mediate the occurrence of EMT. E-cadherin is a key inhibitor of neonatal epithelial tumor cell movement and invasion, thus the decreased expression of E-cadherin is considered to be a significant marker of EMT 99 . TRPV4 is highly expressed in bladder cancer cells, which, when suppressed, can significantly downregulate the expression of E-cadherin. Meanwhile, inhibiting TRPV4 can induce the activation of AKT and FAK to further alter the expression level of E-cadherin 100 . Cell motility enhancement is The results of lung cancer and colorectal cancer in this table and Fig. 2 are obtained from the same research typified by cytoskeletal protein remodeling, also as a crucial feature of EMT 101,102 . In mammal cells, TRPV4 and cytoskeletal protein actin have extensive colocalized expression. Breast cancer cells in which TRPV4 is activated undergo cytoskeletal remodeling. As an upstream "signal emitter", TRPV4 regulates not only microtubule-microfilament polymerization but also the dynamic changes of microvilli, filopodia, and slab pseudopods, thereby affecting cell motility 103 . The release of many factors in the tumor microenvironment, such as epidermal growth factor (EGF), tumor necrosis factor-α (TNF-α), and signal transducer and activator of transcription 3 (STAT3), can induce the occurrence of tumor EMT, and these factors are regulated by intracellular calcium signaling 104 . Calcium ion chelators can inhibit the activation of EGF transduction signals and STAT3 in breast cancer cells 105 . Furthermore, the levels of calcium ions in tumor epithelioid cells and mesenchymal cells are different 62 . Raising calcium concentration can induce EMT of breast cancer MDA-MB-468 cells, probably by mediating their morphological changes 105 . Using the specific antagonist HC-067047 to inhibit TRPV4 in hepatocellular carcinoma cells suppressed cell proliferation, induced apoptosis, and decreased the migration capability by attenuating the EMT process in vitro and intraperitoneal injection of HC-067047 could obviously suppress tumor growth in NOD-SCID mouse xenograft models 106 . Since TRPV4 is an essential ion channelregulating cell morphology, it may predominantly regulate EMT. TRPV4-mediated calcium signaling participates in the EMT process via multiple signaling pathways, including the Wnt/β-catenin and PI3K/AKT pathway. As : series GSE19188 (title: Fibroblast growth factor 9 has oncogenic activity and is a downstream target of Wnt signaling in ovarian endometrioid adenocarcinomas), with p-value of 0.067, and 45 patients samples were used for this analysis; PRAD (prostate adenocarcinoma): series GSE21034 (title: Whole-transcript and exon-level expression data for human primary and metastatic prostate cancer samples and control normal adjacent benign prostate), with p-value of 3.71E-12, and 160 patients samples were used for this analysis; STES (esophageal carcinoma): series GSE13898 (title: Robust prognostic biomarkers for EAC identified by systems-level characterization of tumor transcriptome), with p-value of 6.58E-9, and 40 patients samples were used for this analysis. We queried oncomine for each of those specific cancers to compare the fold change of TRPV4 in cancer tissue cells with normal tissue cells. All p-values represent a Student's t test to gastric cancer metastasis, TRPV4 can enhance PI3K/ AKT activity upon activation, induce β-catenin to enter the nucleus, and activate EMT by interacting with nuclear transcription factor T lymphocyte factor/lymphoid enhancer 88 .

TRPV4 promotes expressions of tumor metastasisassociated proteins
The promotive effects of TRPV4 on the EMT process have been elucidated above. When TRPV4 is continuously activated, the expressions of genes that facilitate tumor metastasis increase, but those of genes that inhibit metastasis reduce. Lee et al. found that the expressions of cell adhesion-associated tumor suppressor genes in mouse breast cancer cell line 4T07, such as Fn1, Clu, Tubb2c, and Spp1, decreased after administration with TRPV4 agonist 4α-PDD. At the same time, the expression of tumor metastasis-promoting gene Talin secreted by exosomes increased. The Kaplan-Meier survival analysis of 4142 patients with breast cancer revealed a significant positive correlation between Talin gene expression and lymphatic metastasis 95 . In the meantime, tumor cells released metastasis-promoting factors through exosomes to mediate tumor metastasis 28 . As the main histological barrier of tumor metastasis, ECM comprises collagen, glycoprotein, proteoglycan, and other components 107 . TRPV4 seems to be colocalized with F-actin in highly dynamic membrane structures, such as filopodia, microvilli, and lamellipodia edges. The interaction between TRPV4 and F-actin may be necessary for the activation of TRPV4 by hypotonic cell swelling, and disrupting the Factin structure abolishes TRPV4-actin co-localization and leads to the loss of hypotonicity-induced Ca 2+ influx and RVD 108 . Moreover, matrix metalloproteinases (MMPs) are important enzymes for the degradation of ECM 109 . Villalta et al. evaluated the effects of TRPV4 on mouse brain edema, and found that the expression of MMP2/ MMP9 in the hippocampus was significantly inhibited by using HC-067046, a TRPV4 antagonist 110 . Furthermore, MMP2/MMP9 is associated with the metastasis of many types of cancers such as lung cancer 111 . Microtubuleassociated protein 7 (MAP7) interacts with residues 798-809 at the C-terminus of TRPV4, possibly by increasing the expression of TRPV4 in the plasma membrane and linking the channel to cytoskeletal microtubules, forming a mechanosensitive molecular complex 112 . VPAC1, a member of the G protein-coupled receptor (GPCR) superfamily, is mainly activated by vasoactive intestinal peptide (VIP). Previous studies reveal that VPAC1 activation promotes migration and invasion of GC cells through TRPV4 channel-dependent Ca 2+ entry, which in turn augments VIP expression. VIP significantly increased the lung metastasis of gastric cancer cells in vivo through the VPAC1/TRPV4/Ca 2+ signaling axis 113 . In vivo angiogenesis assay demonstrated that TRPV4 regulates tumor vessel integrity by maintaining VE-cadherin expression at cell-cell contacts 114 .

Regulatory effects of TRPV4 on tumor angiogenesis
Access to the host vascular system and generation of tumor blood supply are important processes in tumor progression. A large number of new blood vessels not only provide sufficient nutrients for promoting the continuous growth of tumors but also drive tumor spread and metastasis 115 . TRPV4 is critical for tumor angiogenesis, but it may affect various tumors differently. Under normal conditions, activation of TRPV4-mediated Ca 2+ entry is translated into a pro-angiogenic signal by several decoders, such as the Ca 2+ -dependent nuclear factor of activated T cells, cytoplasmic 1 (NFATc1), myocyte enhancer factor 2C (MEF2C), and Kv channel interacting protein 3, calsenilin (KCNIP3/CSEN/DREAM), which drive endothelial cell proliferation, β-integrin, and PI3K, which promote endothelial cell motility 116 . When TRPV4 expression is subjected to interference with shRNA or an inhibitor is used, retinal vascular cells fail to form tubules, indicating that TRPV4 significantly participates in retinal vascular endothelial cell migration and tubule formation 117 . The functions of TRPV4 protein from breast cancer-derived endothelial cells are similar to those of normal endothelial cells. Activating TRPV4 by using arachidonic acid can significantly promote the migration of endothelial cells derived from human breast cancer, but not that of normal human microvascular endothelial cells, which may be ascribed to the significantly higher expression of TRPV4 in breast cancer. As a result, intracellular calcium increased to promote endothelial cell migration. In contrast, loss of TRPV4 expression inhibits arachidonic acid-induced breast cancer cell migration 118 . For lung cancer and prostate cancer, the expression levels of TRPV4 in tumor-derived endothelial cells are lower than those of normal endothelial cells, thus elevating the sensitivity of endothelial cells to extracellular matrix stiffness and being conducive to the formation of abnormal blood vessels 29 . In TRPV4 knockout mice, the density and diameter of new tumor blood vessels enlarge, and the coverage of surrounding tumor capillary endothelial cells shrinks. Therefore, the formation of blood vessels is "abnormalized" to further promote the progression of lung cancer. On the contrary, activating TRPV4 with GSK1016790A can "normalize" the vascular endothelium, enhance the permeability of chemotherapeutic drugs, and ultimately reduce the exudation of cancer cells and block tumor growth 119 . With the study mentioned before, these results demonstrate the tumor angiogenesis regulatory role of TRPV4. In short, inhibition of tumor angiogenesis by targeting TRPV4 should be based on a thorough understanding of the organ specificity of the tumor microenvironment.
Nevertheless, the underlying mechanisms require further in-depth studies.

TRPV4 interacts with tumor microenvironment to promote tumor metastasis
The tumor microenvironment is characterized by hypoxia, low pH, and high tissue pressure, often accompanied by inflammatory reactions, which preferentially selects more invasive and aggressive tumor cells and also impedes the tumor-killing action of immune cells 120 . Proinflammatory cytokines in the inflammatory microenvironment recruit and hijack immune cells to aid the immune escape of tumor cells 121 . These inflammatory factors also stimulate blood vessels and lymphatic vessels to change their permeability to increase protein and cell leakage, ultimately contributing to tumor metastasis 122 . TRPV4 is crucial throughout inflammation. At the initial stage of inflammation, cathepsin produced by the microenvironment of tumor inflammation can simultaneously activate TRPV4 and proteinase 2 which then synergize to continue the development of inflammatory response 123 . Meanwhile, TRPV4 is an essential effector protein that mediates inflammation and signal transduction. Inflammatory factors released in the tumor microenvironment, such as IL-1, IL-8, arachidonic acid, and phosphatase A2, as well as interstitial plasticity changes can promote opening of the tumor TRPV4, at which point the inflammatory factor release is reinforced by cascade amplification. Walter et al. reported that the expressions of inflammatory factors IL-1β, IL-6, and IL-8 decreased when intervertebral disc cells were inhibited by TRPV4 124 . Besides, Kim et al. found that inflammatory factors, such as TNF-α and IL-6, promoted the metastasis of Lewis lung cancer by increasing toll-like receptor 2 (TLR2) and its community toll-like receptor 6 (TLR6) 125 . When inflammatory response occurs, inflammatory factors recruit immune cells, and activation of the TRPV4 channel therein enables tumor cells to escape from the immune system. Scheraga et al. verified that activating the TRPV4 in bone marrow-derived macrophages was beneficial to the release of inflammatory factors, such as IL-1β and IL-10. Furthermore, activation of the TRPV4 of T cells at the site of inflammation facilitates the release of interferon-ɣ which is also an important mediator of tumor immune escape 126 . Collectively, targeting TRPV4 can simultaneously act on tumor cells and their "hijacked" immune cells, reduce the production of inflammatory factors, and finally ameliorate the inflammatory microenvironment for tumor metastasis. Fig. 3 The specific process of TRPV4 acting on tumor occurrence and development and its possible mechanism. TRPV4 is closely related with the proliferation, differentiation, apoptosis, and migration of tumor cell by regulation of Ca 2+ and production of isoforms, and finally participates in tumor onset and progression

Conclusion
In summary, this review clarified the roles of TRPV4 in tumor onset, progression, and metastasis together with the mechanisms, verifying its potential antitumor effects. The specific process of TRPV4 acting on tumor occurrence and development and its possible mechanism is shown in Fig. 3. It is explicitly that the expression and function of TRPV4 are closely related to the occurrence and development of tumors. Hence, TRPV4 is worthy of extensive research for the diagnosis, treatment, and prognosis of tumors. In recent years, the remarkable effects of calcium signal and relative channel protein on the progression of tumors have gradually been recognized. Cancer therapy can be improved by evaluating channel protein expression feature of different tumors then rationally selecting specific antagonists or agonists. Although TRPV4 inhibitors have been employed to treat various diseases such as pulmonary edema and heart failure, there are no commercially available low-toxic and efficient drugs hitherto. Furthermore, the functions of TRPV4 in tumors varied depending on the original tissue type. TRPV4 regulates cellular function by modulating calcium signaling, and finally participates in tumor onset and progression. As summarized above, TRPV4 is closely related with proliferation, differentiation, apoptosis, and migration of tumor cell by regulation of Ca 2+ and its downstream, then finally participates in tumor onset and progression. Consequently, TRPV4 could be a potential therapeutic target of cancer treatment, thereby providing a new direction that develop drugs for cancer to reach the clinic. Regardless of burgeoning studies on the roles of TRPV4 in tumors, its differential expressions in different tumor tissues and the underlying mechanisms are still largely unknown. Therefore, developing antitumor drug targeting TRPV4 remains rather challenging. In addition, targeting TRPV4 channels may also affect other stromal cells, so it is imperative to assess the overall biological effects before possible clinical use.