MET, encoded by Met proto-oncogene located on chromosome 7q31, is a tyrosine kinase-type specific receptor of HGF, which forms disulfide-inked heterodimer consisting of an extracellular alpha chain and single-pass transmembrane beta chain [3,13,14,15]. As shown in Figure 1, the intracellular domain of the beta chain comprises a juxtamembrane domain and catalytic kinase domain containing an activation loop and carboxy-terminal multifunctional docking site. The juxtamembrane domain downregulates the kinase activity by phosphorylation of Ser975, while the catalytic kinase domain upregulates the activity by phosphorylation of Tyr1234 and Tyr1235. The multifunctional docking sites contain Tyr1349 and Tyr1356, which lead to downstream signaling through several intracellular adaptor proteins [3,13,14,15,16]. Increased expression of MET with worse prognosis has been reported in various cancer cells, and phosphorylation (activation) potently promotes invasion and metastasis [16,17,18,19]. Activation of HGF/MET signaling axis in cancer cells also plays a significant role in proliferation, angiogenesis, epithelial-mesenchymal transition (EMT), and drug resistance [3,13,14]. Activation is introduced by: 1) ligand (HGF)-dependent activation, 2) reciprocal activation by overexpression-induced MET oligomerization, 3) activating point mutation of tyrosine kinase domain, and 4) transactivation by heterodimerization with another receptor tyrosine kinase [3,13,14]. In the ligand-dependent activation, proteolytic activation of pro-HGF is necessary. As mentioned above, two major activating protease families were reported: 1) a serum serine protease, HGFA; and 2) type II transmembrane serine proteases (TTSPs) such as matriptase, hepsin, and transmembrane protease/serine (TMPRSS) 2 [3,10,11,12]. Although these pro-HGF activating proteases are tightly regulated by two transmembrane serine protease inhibitors, HAI-1 and HAI-2, downregulation of HAIs has been observed in several cancers and has been shown to induce progression [11,12].

The TTSP family in humans consists of 17 serine proteases [3,10,17]. The structures are specified as a single-pass hydrophobic transmembrane domain near the N-terminus with a short intracellular domain and a large extracellular portion including a carboxy-terminal serine protease domain [3,10,17]. All TTSPs are divided into the four subfamilies of hepsin, matriptase, human airway trypsin-like protease (HAT) and corin (Table 1) [3,10,17]. All TTSPs belong to the S1 peptidase family (noted in MEROPS as clan PA, family S1), and a catalytic triad consists of serine, aspartate, and histidine residues, as shown in Figure 2 [20,21]. Hepsin, matriptase and TMPRSS2 shows a strong cleavage preference for substrate with arginine in the P1 position [20,21]. In urogenital cancers, the expression of matriptase, hepsin and TMPRSS2 has been reported (Figure 2). Therefore, we focused on these TTSPs in this review.

Matriptase (MT-ST1, ST14) gene is located on human chromosome 11q24-25, and 855 amino acids are encoded in the gene [3,10,17,18]. The molecular weight of matriptase is 80–90-kda. Matriptase was first discovered in breast cancer cell line (T-47D) and purified from human milk [3,10,17,18]. It is expressed in human epithelial cells of various organs to maintain the formation of epithelial barrier formation [3,10,11,12,19]. In addition, the major enzymatic functions are reported as follows: 1) activation of hepatocyte growth factor zymogen (pro-HGF), pro-platelet-derived growth factor (PDGF)-C, -D, and pro-macrophage stimulating protein (MSP); 2) activation of protease-activated receptor (PAR)-2; 3) activation of urokinase-type plasminogen activator; 4) degradation of extracellular matrix; and 5) activation of prostasin, which is a glycosylphosphatidylinositol (GPI)-anchored protease known to activate epithelial sodium channel (ENaC) [3,10,17,18]. Among TTSPs, matriptase has been reported as the most efficient activator of pro-HGF [3,10,11,12]. HAIs are major regulators of matriptase, and deregulation of matriptase activity facilitates cancer progression [3,10,11,12]. Indeed, matriptase expression is reported to be upregulated in various cancers (breast, ovarian, uterine, colon, cervical, skin, pancreatic, esophageal, head and neck, prostate, bladder and renal cell carcinoma: RCC) with poor prognosis [3,10,11,12,22].

Hepsin (HPN, TMPRSS1) gene is located on human chromosome 19q13.11, and 417 amino acids are encoded [3,10,17,18]. The molecular weight of hepsin protein is 45-kda. Although mRNA is highly expressed in liver and kidney, ubiquitous expression of the protein is reported [3,10,23]. The functions are reported as follows: 1) activation of pro-HGF; 2) activation of pro-MSP; 3) activation of pro-urokinase-type plasminogen activator; and 4) cleavage of laminin-332 [24]. Similar to matriptase, the catalytic activities of hepsin are regulated by HAI-1 and HAI-2 [3,10,11,12]. In cancer, overexpression of hepsin mRNA is reported in prostate, ovary, kidney, and breast [3,10,17,18]. Increased expression of the protein is also reported in prostate, ovarian, breast, and endometrial cancer [3,10,17,18].

HAI-1 (SPINT-1) gene is located at 15q 15.1 and HAI-2 (SPINT-2) is located at 19q 13.2 [3,11,12]. Both proteins were initially identified in conditioned medium of human gastric cancer cell line MKN45 [3,11,12,25,26]. HAI-2 was also purified as placental bikunin from placenta [3,11,12,27]. The proteins have two specific extracellular Kunitz-type serine protease inhibitor domains, (KD)-1 and KD-2, except for a splicing variant of HAI-2 (isoform B has single KD) (Figure 3), which can inhibit several trypsin-like serine proteases, including all pro-HGF-activating enzymes [3,11,12,28,29]. Whereas, HAIs were initially discovered as HGFA inhibitors, they also inhibit matriptase and hepsin [3,11,12]. In addition, HAIs are required for intracellular transport and cell surface localization of matriptase in several types of cells [3,10,11,12]. HAI-1 is reported to express in the majority of normal epithelial cells [3,11,12,30]. In physiological condition, HAI-1 maintains epithelial integrity through regulation of matriptase activity [3,11,12,30]. HAI-1 is also required for placental differentiation, embryonic development and postnatal survival [11,12,31]. However, it has been reported that insufficient expression revealed dysregulation of pro-HGF activating enzymes in various cancers leading to progression [11,12]. Indeed, decreased expression of HAI-1 induced carcinogenesis (skin, intestine) and progression with worse prognosis (gastrointestinal, breast, ovarian, endometrial cancers and RCC) [11,12,32,33,34,35,36,37,38,39,40,41,42]. In addition, HAI-1 is also known as a suppressor of epithelial mesenchymal transition (EMT) [43].

HAI-2 is ubiquitously expressed in normal cells, including epithelial, mesenchymal, blood cells and trophoblasts [12]. HAI-2 is reported to maintain the integrity of intestinal epithelium through regulation of matriptase-induced epithelial cell adhesion molecule (EpCAM) cleavage [44]. Downregulation by hypermethylation of SPINT2 gene has been reported in several cancers, including hepatocellular carcinoma, RCC, melanoma, gastric carcinoma, and esophageal squamous cell carcinoma [45,46,47,48]. Expression of HAI-2 is also decreased in PC. However, no apparent SPINT2 promoter methylation has been observed in either clinical samples or cell lines [49]. In this report, the authors suggest that posttranslational regulation of HAI-2 expression is essential in prostate cancer. The regulatory role of HAI-2 in the activation of pro-HGF by inhibiting the activating proteases (including matriptase), which induces HGF/MET signaling axis, has been considered a major suppressive function in cancer progression [11,12]. Additionally, an alternative function such as the activation of caspase 3 in esophageal squamous cell carcinoma leading to the promotion of apoptosis and inhibition of proliferation was also reported [47,50]. However, HAI-2 has also been reported to be required for invasive growth in oral squamous cell carcinoma, which suggests that the role of HAI-2 may be tissue or cell-type specific and dependent on targeting TTSPs [51].

 

 

• Janetka, W.J.; Benson, M.R. Extracellular Targeting of Cell Signaling in Cancer; Strategies Directed at MET and RON Receptor Tyrosine Kinase Pathways; Wiley: Hoboken, NJ, USA, 2018; pp. 1–154. [Google Scholar]
• Kataoka, H.; Miyata, S.; Uchinokura, S.; Itoh, H. Roles of hepatocyte growth factor (HGF) activator and HGF activator inhibitor in the pericellular activation of HGF/scatter factor. Cancer Metastasis Rev. 2003, 22, 223–236. [Google Scholar] [CrossRef]
• Miyazawa, K.; Shimomura, T.; Kitamura, N. Activation of hepatocyte growth factor in the injured tissues is mediated by hepatocyte growth factor activator. J. Biol. Chem. 1996, 16, 3615–3618. [Google Scholar] [CrossRef] [PubMed]
• Shimomura, T.; Kondo, J.; Ochiai, M.; Naka, D.; Miyazawa, K.; Morimoto, Y.; Kitamura, N. Activation of the zymogen of hepatocyte growth factor activator by thrombin. J. Biol. Chem. 1993, 25, 22927–22932. [Google Scholar]
• Kataoka, H.; Kawaguchi, M. Hepatocyte growth factor activator (HGFA): pathophysiological functions in vivo. FEBS J. 2010, 277, 2230–2237. [Google Scholar] [CrossRef]
• Kirchhofer, D.; Peek, M.; Lipari, M.T.; Billeci, K.; Fan, B.; Moran, P. Hepsin activates pro-hepatocyte growth factor and is inhibited by hepatocyte growth factor activator inhibitor-1B (HAI-1B) and HAI-2. FEBS Lett. 2005, 28, 1945–1950. [Google Scholar] [CrossRef]
• Owen, K.A.; Qiu, D.; Alves, J.; Schumacher, A.M.; Kilpatrick, L.M.; Li, J.; Harris, J.L.; Ellis, V. Pericellular activation of hepatocyte growth factor by the transmembrane serine proteases matriptase and hepsin, but not by the membrane-associated protease uPA. Biochem. J. 2010, 426, 219–228. [Google Scholar] [CrossRef]
• Martin, C.E.; List, K. Cell surface-anchored serine proteases in cancer progression and metastasis. Cancer Metastasis Rev. 2019, 38, 357–387. [Google Scholar] [CrossRef]
• Kawaguchi, M.; Kataoka, H. Mechanisms of hepatocyte growth factor activation in cancer tissues. Cancers (Basel) 2014, 29, 1890–1904. [Google Scholar] [CrossRef]
• Kataoka, H.; Kawaguchi, M.; Fukushima, T.; Shimomura, T. Hepatocyte growth factor activator inhibitors (HAI-1 and HAI-2): Emerging key players in epithelial integrity and cancer. Pathol. Int. 2018, 68, 145–158. [Google Scholar] [CrossRef] [PubMed]
• Gentile, A.; Trusolino, L.; Comoglio, P.M. The Met tyrosine kinase receptor in development and cancer. Cancer Metastasis Rev. 2008, 27, 85–94. [Google Scholar] [CrossRef] [PubMed]
• Trusolino, L.; Bertotti, A.; Comoglio, P.M. MET signalling: Principles and functions in development, organ regeneration and cancer. Nat. Rev. Mol. Cell Biol. 2010, 11, 834–848. [Google Scholar] [CrossRef] [PubMed]
• Ma, P.C.; Maulik, G.; Christensen, J.; Salgia, R. c-Met: structure, functions and potential for therapeutic inhibition. Cancer Metastasis Rev. 2003, 22, 309–325. [Google Scholar] [CrossRef] [PubMed]
• Tovar, E.A.; Graveel, C.R. MET in human cancer: germline and somatic mutations. Ann. Transl. Med. 2017, 5, 205. [Google Scholar] [CrossRef]
• Tanabe, L.M.; List, K. The role of type II transmembrane serine protease-mediated signaling in cancer. FEBS J. 2017, 284, 1421–1436. [Google Scholar] [CrossRef]
• List, K.; Bugge, T.H.; Szabo, R. Matriptase: potent proteolysis on the cell surface. Mol. Med. 2006, 12, 1–7. [Google Scholar] [CrossRef]
• Wu, C.J.; Feng, X.; Lu, M.; Morimura, S.; Udey, M.C. Matriptase-mediated cleavage of EpCAM destabilizes claudins and dysregulates intestinal epithelial homeostasis. J. Clin. Investig. 2017, 127, 623–634. [Google Scholar] [CrossRef]
• Szabo, R.; Bugge, T.H. Membrane-anchored serine proteases in vertebrate cell and developmental biology. Annu. Rev. Cell Dev. Biol. 2011, 27, 213–235. [Google Scholar] [CrossRef]
• Rawlings, N.D.; Barrett, A.J.; Thomas, P.D.; Huang, X.; Bateman, A.; Finn, R.D. The MEROPS database of proteolytic enzymes, their substrates and inhibitors in 2017 and a comparison with peptidases in the PANTHER database. Nucleic Acids Res. 2018, 46, D624–D632. [Google Scholar] [CrossRef]
• Cheng, M.F.; Huang, M.S.; Lin, C.S.; Lin, L.H.; Lee, H.S.; Jiang, J.C.; Hsia, K.T. Expression of matriptase correlates with tumour progression and clinical prognosis in oral squamous cell carcinoma. Histopathology 2014, 65, 24–34. [Google Scholar] [PubMed]
• Tsuji, A.; Torres-Rosado, A.; Arai, T.; Le Beau, M.M.; Lemons, R.S.; Chou, S.H.; Kurachi, K. Hepsin, a cell membrane-associated protease. Characterization, tissue distribution, and gene localization. J. Biol. Chem. 1991, 5, 16948–16953. [Google Scholar]
• Tripathi, M.; Nandana, S.; Yamashita, H.; Ganesan, R.; Kirchhofer, D.; Quaranta, V. Laminin-332 is a substrate for hepsin, a protease associated with prostate cancer progression. J. Biol. Chem. 2008, 7, 30576–30584. [Google Scholar]
• Shimomura, T.; Denda, K.; Kitamura, A.; Kawaguchi, T.; Kito, M.; Kondo, J.; Kagaya, S.; Qin, L.; Takata, H.; Miyazawa, K.; et al. Hepatocyte growth factor activator inhibitor, a novel Kunitz-type serine protease inhibitor. J. Biol. Chem. 1997, 272, 6370–6376. [Google Scholar]
• Kawaguchi, T.; Qin, L.; Shimomura, T.; Kondo, J.; Matsumoto, K.; Denda, K.; Kitamura, N. Purification and cloning of hepatocyte growth factor activator inhibitor type 2, a Kunitz-type serine protease inhibitor. J. Biol. Chem. 1997, 272, 27558–27564. [Google Scholar] [PubMed]
• Marlor, C.W.; Delaria, K.A.; Davis, G.; Muller, D.K.; Greve, J.M.; Tamburini, P.P. Identification and cloning of human placental bikunin, a novel serine protease inhibitor containing two Kunitz domains. J. Biol. Chem. 1997, 272, 12202–12208. [Google Scholar]
• Kataoka, H.; Itoh, H.; Nuki, Y.; Hamasuna, R.; Naganuma, S.; Kitamura, N.; Shimomura, T. Mouse hepatocyte growth factor (HGF) activator inhibitor type 2 lacking the first Kunitz domain potently inhibits the HGF activator. Biochem. Biophy. Res. Commun. 2002, 290, 1096–1100. [Google Scholar]
• Kataoka, H.; Shimomura, T.; Kawaguchi, T.; Hamasuna, R.; Itoh, H.; Kitamura, N.; Miyazawa, K.; Koono, M. Hepatocyte growth factor activator inhibitor type 1 is a specific cell surface binding protein of hepatocyte growth factor activator (HGFA) and regulates HGFA activity in the pericellular microenvironment. J. Biol. Chem. 2000, 275, 40453–40462. [Google Scholar]
• Kawaguchi, M.; Takeda, N.; Hoshiko, S.; Yorita, K.; Baba, T.; Sawaguchi, A.; Nezu, Y.; Yoshikawa, T.; Fukushima, T.; Kataoka, H. Membrane-bound serine protease inhibitor HAI-1 is required for maintenance of intestinal epithelial integrity. Am. J. Pathol. 2011, 179, 1815–1826. [Google Scholar]
• Tanaka, H.; Nagaike, K.; Takeda, N.; Itoh, H.; Kohama, K.; Fukushima, T.; Miyata, S.; Uchiyama, S.; Uchinokura, S.; Shimomura, T.; et al. Hepatocyte growth factor activator inhibitor type 1 (HAI-1) is required for branching morphogenesis in the chorioallantoic placenta. Mol. Cell Biol. 2005, 25, 5687–5698. [Google Scholar]
• Hoshiko, S.; Kawaguchi, M.; Fukushima, T.; Haruyama, Y.; Yorita, K.; Tanaka, H.; Seiki, M.; Inatsu, H.; Kitamura, K.; Kataoka, H. Hepatocyte growth factor activator inhibitor type 1 is a suppressor of intestinal tumorigenesis. Cancer Res. 2013, 73, 2659–2670. [Google Scholar] [CrossRef] [PubMed]
• Baba, T.; Kawaguchi, M.; Fukushima, T.; Sato, Y.; Orikawa, H.; Yorita, K.; Tanaka, H.; Lin, C.Y.; Sakoda, S.; Kataoka, H. Loss of membrane-bound serine protease inhibitor HAI-1 induces oral squamous cell carcinoma cells’ invasiveness. J. Pathol. 2012, 228, 181–192. [Google Scholar] [PubMed]
• Ning, T.; Zhang, H.; Wang, X.; Li, S.; Zhang, L.; Deng, T.; Zhou, L.; Wang, X.; Liu, R.; Bai, M.; et al. miR-221 and miR-222 synergistically regulate hepatocyte growth factor activator inhibitor type 1 to promote cell proliferation and migration in gastric cancer. Tumour. Biol. 2017, 39, 1010428317701636. [Google Scholar] [CrossRef] [PubMed]
• Ye, J.; Kawaguchi, M.; Haruyama, Y.; Kanemaru, A.; Fukushima, T.; Yamamoto, K.; Lin, C.Y.; Kataoka, H. Loss of hepatocyte growth factor activator inhibitor type 1 participates in metastatic spreading of human pancreatic cancer cells in a mouse orthotopic transplantation model. Cancer Sci. 2014, 105, 44–51. [Google Scholar] [CrossRef] [PubMed]
• Oberst, M.D.; Johnson, M.D.; Dickson, R.B.; Lin, C.Y.; Singh, B.; Stewart, M.; Williams, A.; al-Nafussi, A.; Smyth, J.F.; Gabra, H.; et al. Expression of the serine protease matriptase and its inhibitor HAI-1 in epithelial ovarian cancer: Correlation with clinical outcome and tumor clinicopathological parameters. Clin. Cancer Res. 2002, 8, 1101–1107. [Google Scholar] [PubMed]
• Nakamura, K.; Abarzua, F.; Hongo, A.; Kodama, J.; Nasu, Y.; Kumon, H.; Hiramatsu, Y. The role of hepatocyte growth factor activator inhibitor-1 (HAI-1) as a prognostic indicator in cervical cancer. Int. J. Oncol. 2009, 35, 239–248. [Google Scholar] [CrossRef]
• Nakamura, K.; Hongo, A.; Kodama, J.; Hiramatsu, Y. The role of hepatocyte growth factor activator inhibitor (HAI)-1 and HAI-2 in endometrial cancer. Int. J. Cancer 2011, 128, 2613–2624. [Google Scholar]
• Hamasuna, R.; Kataoka, H.; Meng, J.Y.; Itoh, H.; Moriyama, T.; Wakisaka, S.; Koono, M. Reduced expression of hepatocyte growth factor activator inhibitor type-2/placental bikunin (HAI-2/PB) in human glioblastomas: Implication for anti-invasive role of HAI-2/PB in glioblastoma cells. Int. J. Cancer 2001, 93, 339–345. [Google Scholar] [CrossRef]
• Li, W.; Wang, B.E.; Moran, P.; Lipari, T.; Ganesan, R.; Corpuz, R.; Ludlam, M.J.; Gogineni, A.; Koeppen, H.; Bunting, S.; et al. Pegylated Kunitz domain inhibitor suppresses hepsin-mediated invasive tumor growth and metastasis. Cancer Res. 2009, 69, 8395–8402. [Google Scholar] [CrossRef]
• Fukushima, T.; Kawaguchi, M.; Yamasaki, M.; Tanaka, H.; Yorita, K.; Kataoka, H. Hepatocyte growth factor activator inhibitor type 1 suppresses metastatic pulmonary colonization of pancreatic carcinoma cells. Cancer Sci. 2011, 102, 407–413. [Google Scholar] [CrossRef]
• Betsunoh, H.; Mukai, S.; Akiyama, Y.; Fukushima, T.; Minamiguchi, N.; Hasui, Y.; Osada, Y.; Kataoka, H. Clinical relevance of hepsin and hepatocyte growth factor activator inhibitor type 2 expression in renal cell carcinoma. Cancer Sci. 2007, 98, 491–498. [Google Scholar] [CrossRef] [PubMed]
• Cheng, H.; Fukushima, T.; Takahashi, N.; Tanaka, H.; Kataoka, H. Hepatocyte growth factor activator inhibitor type 1 regulates epithelial to mesenchymal transition through membrane-bound serine proteinases. Cancer Res. 2009, 69, 1828–1835. [Google Scholar] [CrossRef] [PubMed]
• Kawaguchi, M.; Yamamoto, K.; Takeda, N.; Fukushima, T.; Yamashita, F.; Sato, K.; Kitamura, K.; Hippo, Y.; Janetka, J.W.; Kataoka, H. Hepatocyte growth factor activator inhibitor-2 stabilizes Epcam and maintains epithelial organization in the mouse intestine. Commun. Biol. 2019, 2, 11. [Google Scholar] [CrossRef] [PubMed]
• Morris, M.R.; Gentle, D.; Abdulrahman, M.; Maina, E.N.; Gupta, K.; Banks, R.E.; Wiesener, M.S.; Kishida, T.; Yao, M.; The, B.; et al. Tumor suppressor activity and epigenetic inactivation of hepatocyte growth factor activator inhibitor type 2/SPINT2 in papillary and clear cell renal cell carcinoma. Cancer Res. 2005, 65, 4598–4606. [Google Scholar] [CrossRef]
• Hwang, S.; Kim, H.E.; Min, M.; Raghunathan, R.; Panova, I.P.; Munshi, R.; Ryu, B. Epigenetic silencing of SPINT2 promotes cancer cell motility via HGF-MET pathway activation in Melanoma. J. Investig. Dermatol. 2015, 135, 2283–2291. [Google Scholar] [CrossRef]
• Yue, D.; Fan, Q.; Chen, X.; Li, F.; Wang, L.; Huang, L.; Dong, W.; Chen, X.; Zhang, Z.; Liu, J.; et al. Epigenetic inactivation of SPINT2 is associated with tumor suppressive function in esophageal squamous cell carcinoma. Exp. Cell Res. 2014, 322, 149–158. [Google Scholar] [CrossRef]
• Dong, W.; Chen, X.; Xie, J.; Sun, P.; Wu, Y. Epigenetic inactivation and tumor suppressor activity of HAI-2/SPINT2 in gastric cancer. Int. J. Cancer 2010, 127, 1526–1534. [Google Scholar] [CrossRef]
• Tsai, C.H.; Teng, C.H.; Tu, Y.T.; Cheng, T.S.; Wu, S.R.; Ko, C.J.; Shyu, H.Y.; Lan, S.W.; Huang, H.P.; Tzeng, S.F.; et al. HAI-2 suppresses the invasive growth and metastasis of prostate cancer through regulation of matriptase. Oncogene 2014, 18, 4643–4652. [Google Scholar] [CrossRef]
• Roversi, F.M.; Olalla Saad, S.T.; Machado-Neto, J.A. Serine peptidase inhibitor Kunitz type 2 (SPINT2) in cancer development and progression. Biomed. Pharmacother. 2018, 101, 278–286. [Google Scholar] [CrossRef]
• Yamamoto, K.; Kawaguchi, M.; Shimomura, T.; Izumi, A.; Konari, K.; Honda, A.; Lin, C.Y.; Johnson, M.D.; Yamashita, Y.; Fukushima, T.; et al. Hepatocyte growth factor activator inhibitor-2 (HAI-2)/SPINT2 contributes to invasive growth of oral squamous cell carcinoma cells. Oncotarget 2018, 8, 11691–11706. [Google Scholar] [CrossRef]