scholarly journals Hepatocyte growth factor is a preferred in vitro substrate for human hepsin, a membrane-anchored serine protease implicated in prostate and ovarian cancers

2005 ◽  
Vol 390 (1) ◽  
pp. 125-136 ◽  
Author(s):  
Sylvia Herter ◽  
Derek E. Piper ◽  
Wade Aaron ◽  
Timothy Gabriele ◽  
Gene Cutler ◽  
...  
Keyword(s):  
Growth Factor ◽  
Substrate Specificity ◽  
Hepatocyte Growth Factor ◽  
Serine Protease ◽  
Cleavage Site ◽  
Biologically Active ◽  
Ovarian Cancer Cells ◽  
Single Chain ◽  
Hepatocyte Growth

Hepsin is a membrane-anchored, trypsin-like serine protease with prominent expression in the human liver and tumours of the prostate and ovaries. To better understand the biological functions of hepsin, we identified macromolecular substrates employing a tetrapeptide PS-SCL (positional scanning-synthetic combinatorial library) screen that rapidly determines the P1–P4 substrate specificity. Hepsin exhibited strong preference at the P1 position for arginine over lysine, and favoured threonine, leucine or asparagine at the P2, glutamine or lysine at the P3, and proline or lysine at the P4 position. The relative activity of hepsin toward individual AMC (7-amino-4-methylcoumarin)-tetrapeptides was generally consistent with the overall peptide profiling results derived from the PC-SCL screen. The most active tetrapeptide substrate Ac (acetyl)-KQLR-AMC matched with the activation cleavage site of the hepatocyte growth factor precursor sc-HGF (single-chain HGF), KQLR↓VVNG (where ↓ denotes the cleavage site), as identified by a database analysis of trypsin-like precursors. X-ray crystallographic studies with KQLR chloromethylketone showed that the KQLR peptide fits well into the substrate-binding cleft of hepsin. This hepsin-processed HGF induced c-Met receptor tyrosine phosphorylation in SKOV-3 ovarian cancer cells, indicating that the hepsin-cleaved HGF is biologically active. Activation cleavage site mutants of sc-HGF with predicted non-preferred sequences, DPGR↓VVNG or KQLQ↓VVNG, were not processed, illustrating that the P4–P1 residues can be important determinants for substrate specificity. In addition to finding macromolecular hepsin substrates, the extracellular inhibitors of the HGF activator, HAI-1 and HAI-2, were potent inhibitors of hepsin activity (IC50 4±0.2 nM and 12±0.5 nM respectively). Together, our findings suggest that the HGF precursor is a potential in vivo substrate for hepsin in tumours, where hepsin expression is dysregulated and may influence tumorigenesis through inappropriate activation and/or regulation of HGF receptor (c-Met) functions.

scholarly journals In vitro evidence that KLK14 regulates the components of the HGF/Met axis, pro-HGF and HGF-activator inhibitor 1A and 1B

2016 ◽  
Vol 397 (12) ◽  
pp. 1299-1305 ◽  
Author(s):  
Janet C. Reid ◽  
Nigel C. Bennett ◽  
Carson R. Stephens ◽  
Melanie L. Carroll ◽  
Viktor Magdolen ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Multiple Levels ◽  
Hgf Activator ◽  
Hepatocyte Growth ◽  
Activator Inhibitor

Abstract Kallikrein-related peptidase (KLK) 14 is a serine protease linked to several pathologies including prostate cancer. We show that KLK14 has biphasic effects in vitro on activating and inhibiting components of the prostate cancer associated hepatocyte growth factor (HGF)/Met system. At 5–10 nm, KLK14 converts pro-HGF to the two-chain heterodimer required for Met activation, while higher concentrations degrade the HGF α-chain. HGF activator-inhibitor (HAI)-1A and HAI-1B, which inhibit pro-HGF activators, are degraded by KLK14 when protease:inhibitor stoichiometry is 1:1 or the protease is in excess. When inhibitors are in excess, KLK14 generates HAI-1A and HAI-1B fragments known to inhibit pro-HGF activating serine proteases. These in vitro data suggest that increased KLK14 activity could contribute at multiple levels to HGF/Met-mediated processes in prostate and other cancers.

scholarly journals The Serine Protease Matriptase Acts As a Tumour Suppressor in Multiple Myeloma

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 14-14
Author(s):  
Ida Steiro ◽  
Pegah Abdollahi ◽  
Magne Børset ◽  
Tobias S. Slørdahl
Keyword(s):  
Multiple Myeloma ◽  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Serine Protease ◽  
Cell Lines ◽  
Myeloma Cell ◽  
Tumour Suppressor ◽  
Hepatocyte Growth

Both in newly diagnosed multiple myeloma (MM) and during progression of the disease, malignant plasma cells are found circulating in peripheral blood as well as in the bone marrow (BM). The disseminated nature of MM is strongly dependent on the interplay between the cancer cells and the BM microenvironment, promoting myeloma cell migration in the BM. Matriptase (ST14), a type-II transmembrane serine protease primarily found in epithelial tissues, is overexpressed in a variety of human malignancies and is sufficient to induce tumour formation in mice. Frequently, a concomitant reduction in the levels of its cognate inhibitor hepatocyte growth factor activator inhibitor (HAI)-1 (SPINT1) is observed in carcinomas, while expression and function of the related inhibitor HAI-2 (SPINT2) is yet to be clarified. Dysregulated expression causing increased matriptase proteolytic activity has been associated with cancer growth, survival and metastasis. Here, we show for the first time a role of matriptase as a possible tumour suppressor in myeloma pathogenesis. Gene expression analysis of primary cells from MM patients (n=24) and human myeloma cell lines (n=8) revealed highly variable levels of matriptase, HAI-1 and HAI-2. This observation prompted us to investigate the functional role of matriptase in vitro. We showed that stable overexpression of matriptase in INA-6, a MM cell line with no endogenous ST14 expression, reduced migration by more than 50% in response to the combination of the pro-migratory cytokines stromal cell-derived factor-1 alpha (SDF-1α) and hepatocyte growth factor (HGF, Fig. 1A). Conversely, stable knockdown of matriptase in two MM cell lines with high endogenous matriptase expression (RPMI-8226 and JJN-3) significantly enhanced migration in vitro. Mechanistically, matriptase overexpression blocked activation of Src kinase (Fig. 1B), well-known as a critical player in metastasis formation promoting cancer cell motility, invasiveness and angiogenesis. In agreement with our result, previous studies have demonstrated the activation of Src family kinases (SFK) downstream SDF-1/CXCR4-signaling. Finally, we performed survival analyses in the public available MMRF CoMMpass trial database (release version IA14). Low ST14 expression was associated with significant worse overall survival (P=0.05, Fig. 1C) and progression-free survival (P=0.02, Fig. 1D). Altogether, our data are in marked contrast to the role ascribed to matriptase in epithelial and certain non-epithelial tumours as an oncogenic protein and an unfavourable prognostic marker. In conclusion, these findings suggest a novel role of matriptase as a tumour suppressor in MM pathogenesis. Disclosures Slørdahl: Celgene: Consultancy; Janssen and Celgene: Honoraria.

scholarly journals Targeting hepatocyte growth factor in epithelial–stromal interactions in an in vitro experimental model of human periodontitis

Odontology ◽  
2021 ◽  
Author(s):  
Yoko Yamaguchi ◽  
Akira Saito ◽  
Masafumi Horie ◽  
Akira Aoki ◽  
Patrick Micke ◽  
...  
Keyword(s):  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Experimental Studies ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Neutralizing Antibody ◽  
Chronic Inflammatory Disease ◽  
Collagen Degradation ◽  
Hepatocyte Growth

AbstractPeriodontitis is a chronic inflammatory disease leading to progressive connective tissue degradation and loss of the tooth-supporting bone. Clinical and experimental studies suggest that hepatocyte growth factor (HGF) is involved in the dysregulated fibroblast–epithelial cell interactions in periodontitis. The aim of this study was to explore effects of HGF to impact fibroblast-induced collagen degradation. A patient-derived experimental cell culture model of periodontitis was applied. Primary human epithelial cells and fibroblasts isolated from periodontitis-affected gingiva were co-cultured in a three-dimensional collagen gel. The effects of HGF neutralizing antibody on collagen gel degradation were tested and transcriptome analyses were performed. HGF neutralizing antibody attenuated collagen degradation and elicited expression changes of genes related to extracellular matrix (ECM) and cell adhesion, indicating that HGF signaling inhibition leads to extensive impact on cell–cell and cell–ECM interactions. Our study highlights a potential role of HGF in periodontitis. Antagonizing HGF signaling by a neutralizing antibody may represent a novel approach for periodontitis treatment.

Abstract 1340: Disruption of a Hepatocyte Growth Factor/c-Met Receptor Autocrine Loop Severely Impairs the Potency of Pluripotent Adipose Stromal Cells

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Liying Cai ◽  
Brian H Johnstone ◽  
Zhong Liang ◽  
Dmitry Traktuev ◽  
Todd G Cook ◽  
...  
Keyword(s):  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Rank Order ◽  
Residual Activity ◽  
Autocrine Loop ◽  
Major Mode ◽  
Factor C ◽  
Hepatocyte Growth

Background Paracrine stimulation of endogenous repair, rather than direct tissue regeneration, is increasingly accepted as a major mode of therapeutic stem and progenitor cell action; yet, this principle has not been fully established in vivo . Adipose-derived stem cells (ASCs) secrete many factors and promote reperfusion and tissue repair in ischemia models. RNA interference was used to silence the expression of the abundant protein, hepatocyte growth factor (HGF), to determine its contribution to ASC potency in vivo . Methods and Results Dual-cassette lentiviral vectors, expressing GFP and either a small hairpin RNA (shRNA) specific for HGF mRNA (shHGF) or a control sequence (shCtrl), were used to stably transduce ASCs (ASC-shHGF or ASC-shCtrl). ASC-shHGF secreted 5-fold less HGF, which resulted in a reduced ability of these cells to promote survival, proliferation and migration of mature and progenitor endothelial cells in vitro ( p <0.01). HGF knockdown also severely impaired the ability of ASCs to promote reperfusion in a mouse hindlimb ischemia model. Perfusion of the ischemic leg at 15 d in mice treated with ASC-Ctrl was 84±4%, compared to only 69±5% for ASC-shHGF ( p <0.05). Even so, ASC-shHGF retained residual activity as indicated by greater reperfusion ( p <0.05) than with saline treatment (58±6%). Capillary densities in ischemic tissues from each group followed a similar rank order (ASC-Ctrl>ASC-shHGF>saline) ( p <0.05 between each group). While there was no difference in total GFP + cells in ischemic limbs at 5 d after infusion, indicating similar homing potentials, 3-fold fewer ASC-shHGF were present in ischemic tissues at 15 d compared to ASC-shCtrl ( p <0.01). This was accompanied by an increase in TUNEL-positive ASC-shHGF cells (61 ± 0.1%) compared to ASC-Ctrl (41% ± 3.2%) in ischemic tissues at 5 d ( p <0.01); suggesting that attenuated potency of ASC-shHGF was related to reduced survival in ischemic tissues. Conclusions These results indicate that secretion of HGF is critically important for ASC potency. In addition to promoting endogenous repair, the data suggest that an important effect of HGF is autocrine promotion of ASC survival in ischemic tissue. Enhanced donor cell survival is an important goal for increasing the efficacy of cell therapy.

scholarly journals First-in-human phase I trial of anti-hepatocyte growth factor antibody (YYB101) in refractory solid tumor patients

2020 ◽  
Vol 12 ◽  
pp. 175883592092679
Author(s):  
Seung Tae Kim ◽  
Jung Yong Hong ◽  
Se Hoon Park ◽  
Joon Oh Park ◽  
Young Whan Park ◽  
...  
Keyword(s):  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Phase I ◽  
Cancer Patients ◽  
Phase I Trial ◽  
Maximum Tolerated Dose ◽  
Safety And Efficacy ◽  
Hepatocyte Growth

Background: YYB101, a humanized monoclonal antibody against hepatocyte growth factor (HGF), has shown safety and efficacy in vitro and in vivo. This is a first-in-human trial of this antibody. Materials and Methods: YYB101 was administered intravenously to refractory cancer patients once every 4 weeks for 1 month, and then once every 2 weeks until disease progression or intolerable toxicity, at doses of 0.3, 1, 3, 5, 10, 20, 30 mg/kg, according to a 3+3 dose escalation design. Maximum tolerated dose, safety, pharmacokinetics, and pharmacodynamics were studied. HGF, MET, PD-L1, and ERK expression was evaluated for 9 of 17 patients of the expansion cohort (20 mg/kg). Results: In 39 patients enrolled, no dose-limiting toxicity was observed at 0.3 mg/kg, and the most commonly detected toxicity was generalized edema ( n = 7, 18.9%) followed by pruritis and nausea ( n = 5, 13.5%, each), fatigue, anemia, and decreased appetite ( n = 4, 10.8%, each). No patient discontinued treatment because of adverse events. YYB101 showed dose-proportional pharmacokinetics up to 30 mg/kg. Partial response in 1 (2.5%) and stable disease in 17 (43.5%) were observed. HGF, MET, PD-L1, and ERK proteins were not significant predictors for treatment response. However, serum HGF level was significantly lowered in responders upon drug administration. RNA sequencing revealed a mesenchymal signature in two long-term responders. Conclusion: YYB101 showed favorable safety and efficacy in patients with refractory solid tumors. Based on this phase I trial, a phase II study on the YYB101 + irinotecan combination in refractory metastatic colorectal cancer patients is planned. Conclusion: ClinicalTrials.gov Identifier: NCT02499224

Protamine enhances the proliferative activity of hepatocyte growth factor in rats

1998 ◽  
Vol 274 (1) ◽  
pp. G21-G28 ◽  
Author(s):  
Ke-Xin Liu ◽  
Yukio Kato ◽  
Tai-Ichi Kaku ◽  
Kunio Matsumoto ◽  
Toshikazu Nakamura ◽  
...  
Keyword(s):  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Proliferative Activity ◽  
Plasma Clearance ◽  
Mitogenic Response ◽  
Enhancing Effect ◽  
A Cell ◽  
Hepatocyte Growth

The effect of protamine on the proliferative activity of hepatocyte growth factor (HGF) was examined in α-naphthyl isothiocyanate-intoxicated rats. Protamine preinjection increased the hepatocyte labeling index induced by HGF four- to fivefold. A similar effect was also observed in partially hepatectomized rats. Because a cell surface heparin-like substance can bind to HGF and protamine has an affinity for heparin, protamine may affect HGF pharmacokinetics. In fact, protamine injection caused a transient increase in plasma HGF concentrations after administration of HGF and, in vitro, protamine eluted HGF prebound to heparin-Sepharose. Protamine also reduced the plasma clearance of HGF and increased 2.5-fold the exposure of hepatocytes to HGF in vivo. The enhancing effect of protamine on the mitogenic response of hepatocytes to HGF was also observed in vitro (∼2-fold after protamine pretreatment compared with HGF alone), suggesting that the enhancing effect of protamine on HGF-induced liver regeneration results from dual effects exerted by protamine 1) lowering the overall elimination of HGF and 2) directly stimulating hepatocyte mitosis induced by HGF.

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