Maryse Delehedde

Vascular endocan (ESM-1) is markedly overexpressed in clear cell renal cell carcinoma.

Leroy X, Aubert S, Zini L, Franquet H, Kervoaze G, Villers A, Delehedde M, Copin M‐C & Lassalle P 
(2010) Histopathology56, 180–187

Endocan in Cancers: A Lesson from a Circulating Dermatan Sulfate Proteoglycan

As most proteoglycans exert their biological activities in the pericellular region, circulating Endocan has appeared since its discovery as an atypical dermatan sulfate proteoglycan, with distinctive structural and functional properties. Endocan is naturally expressed by endothelial cells, highly regulated in presence of proinflammatory and proangiogenic molecules, binds to matrix proteins, growth factors, integrin, and cells, and may be then considered as an accurate marker of endothelial activation. Consequently, Endocan expression has been associated with a growing number of pathological conditions where endothelium gets challenged and notably in highly vascularized cancers. In this context, Endocan has indeed been rapidly emerging as a promising tissue- and blood-based marker of the vascular growth and neoangiogenesis during cancer progression. Furthermore, very recent studies have reported an expression of Endocan by the tumor cells themselves. This highlights Endocan as a multifaceted molecule with a great interest for researchers and clinicians to better understand tumor development, from the bench to the clinics. With promising perspectives of clinical applications, Endocan thus appears as an exciting model for on going and future developments of proteoglycan-based approaches in cancer diagnostics and/or therapy.

Serum proteoglycans as prognostic biomarkers of hepatocellular carcinoma in patients with alcoholic cirrhosis

Background: Proteoglycans are involved in neoangiogenesis and transduction of oncogenic signals, two hallmarks of carcinogenesis. Methods: This study sought to assess the prognostic value of serum levels of three proteoglycans (endocan, syndecan-1, and glypican-3) and VEGF in 295 patients with alcoholic cirrhosis: 170 without hepatocellular carcinoma, 58 with early hepatocellular carcinoma, and 67 with advanced hepatocellular carcinoma at inclusion. We analyzed the association between proteoglycan levels and prognosis using Kaplan–Meier and Cox methods. Results: Serum levels of the three proteoglycans and VEGF were increased in patients with advanced hepatocellular carcinoma compared with those without hepatocellular carcinoma or with early hepatocellular carcinoma. In multivariate analysis, high levels of serum endocan (>5 ng/mL) were independently associated with death [HR, 2.84; 95% confidence interval (CI,) 1.18–6.84; P = 0.02], but not with hepatocellular carcinoma occurrence, in patients without hepatocellular carcinoma at baseline. High serum endocan (>5 ng/mL) and syndecan-1 (>50 ng/mL) levels were significantly associated with greater risk of tumor recurrence (P = 0.025) in patients with early hepatocellular carcinoma treated by radiofrequency ablation. In patients with advanced hepatocellular carcinoma, high serum levels of endocan (P = 0.004) and syndecan-1 (P = 0.006) were significantly associated with less favorable overall survival. However, only a high level of serum syndecan-1 (>50 ng/mL) was independently associated with greater risk of death (HR, 6.21 95% CI, 1.90–20.30; P = 0.0025). Conclusion: Serum endocan and syndecan-1 are easily assessable prognostic serum biomarkers of overall survival in alcoholic cirrhosis with and without hepatocellular carcinoma. Impact: These new biomarkers will be useful to manage patients with hepatocellular carcinoma developed on alcoholic cirrhosis. Cancer Epidemiol Biomarkers Prev; 22(8); 1343–52. ©2013 AACR.

ESM-1 expression in stromal cells is predictive of recurrence after radiofrequency ablation in early hepatocellular carcinoma

BACKGROUND & AIMS The prognosis of hepatocellular carcinoma (HCC) treated by radiofrequency ablation (RFA) is mainly linked to tumor recurrence. So far, no tissue biomarker of recurrence has been validated in biopsy samples. We aimed at investigating the prognostic value of tissue biomarkers in HCC biopsy samples of patients treated with RFA. METHODS All consecutive naive patients from 3 university hospitals, with compensated cirrhosis, early-stage (BCLC 0/A) uninodular HCC treated with RFA, and available tumor biopsy, were included. Edmondsons grade, and the expression of cytokeratin 19, glutamine synthase, beta-catenin, epithelial cell adhesion molecule (EpCAM), and endothelial cell-specific molecule 1 (ESM-1) were assessed. Main clinical end points were overall and early recurrence. Statistical analyses were performed using Kaplan Meier, Log-rank test, and Cox models. RESULTS 150 patients were included. Recurrence, death or liver transplantation occurred in 85, 51, and 12 patients, respectively. Median follow-up was 27months. ESM-1 expression by HCC stromal endothelial cells was observed in 58 patients (40%) and was associated with higher serum AFP levels, larger tumor, and more frequent expression of EpCAM and surrogate markers of activation of the Wnt-ß-catenin pathway. The 2 independent predictive factors of overall recurrence were serum AFP (HR 1.11 [1.002; 1.22], p=0.045) and ESM-1 expression (HR 1.56 [1.004; 2.43], p=0.048). ESM-1 expression was also an independent predictive factor of early recurrence (HR 1.81 [1.02; 3.21], p=0.042). CONCLUSIONS ESM-1 expression by stromal endothelial cells, in tumor biopsy samples, has an independent predictive value of early recurrence after RFA.

Loss of Endocan tumorigenic properties after alternative splicing of exon 2.

BackgroundEndocan was originally described as a dermatan sulfate proteoglycan found freely circulating in the blood. Endocan expression confers tumorigenic properties to epithelial cell lines or accelerate the growth of already tumorigenic cells. This molecule is the product of a single gene composed of 3 exons. Previous data showed that endocan mRNA is subject to alternative splicing with possible generation of two protein products. In the present study we identified, and functionally characterized, the alternative spliced product of the endocan gene: the exon 2-deleted endocan, called endocanΔ2.MethodsStable, endocanΔ2-overexpressing cell lines were generated to investigate the biological activities of this new alternatively spliced product of endocan gene. Tumorigenesis was studied by inoculating endocan and endocanΔ2 expressing cell lines subcutaneously in SCID mice. Biochemical properties of endocan and endocanΔ2 were studied after production of recombinant proteins in various cell lines of human and murine origin.ResultsOur results showed that the exon 2 deletion impairs synthesis of the glycan chain, known to be involved in the pro-tumoral effect of endocan. EndocanΔ2 did not promote tumor formation by 293 cells implanted in the skin of severe combined immunodeficient (SCID) mice.ConclusionOur results emphasize the key role of the polypeptide sequence encoded by the exon 2 of endocan gene in tumorigenesis, and suggest that this sequence could be a target for future therapies against cancer.

Efficient long-term and high-yielded production of a recombinant proteoglycan in eukaryotic HEK293 cells using a membrane-based bioreactor

Standard culture systems of eukaryotic cells generally failed to deliver sufficient amounts of recombinant proteins without increasing the costs of production. We here showed that membrane-based bioreactors, initially developed for the production of monoclonal antibodies, can be very useful for the production using engineered HEK293 cells, of a recombinant proteoglycan called endocan, with achievement of high level expression and efficient long-term production. When compared to standard procedures, the growth in suspension and at high density of these cells in one bioreactor promoted a 60-fold increase of the concentration of the soluble recombinant endocan. These culture conditions did not affect cell viability, stable expression, recognition by specific monoclonal antibodies or electrophoretic profile of the recombinant endocan. Such an easy to scale up system to produce recombinant protein should open soon new opportunities to study structure and functions of endocan or any other glycosylated cell products newly investigated.

Endocan (endothelial cell-specific molecule-1) as a pertinent biomarker of endothelial dysfunction in sepsis

One of the main players in the severity of sepsis is the endothelium integrity. Endocan, also called endothelial cell-specific molecule-1 (ESM-1), was shown to be preferentially expressed in lung vasculature. Structurally, endocan/ESM-1 is a 50 kDa proteoglycan that can interact with ICAM-1 and LFA-1 integrins and consequently prevents inflammatory events. In an experimental rat endotoxemic shock model, we previously showed that a decrease in the leukocyte-endothelial cell contacts (induced by drugs) is clearly linked to an increase of blood endocan levels. Blood levels of endocan/ESM-1 were also shown to be associated with the severity and evolution of septic states in preliminary studies.

Identification of cathepsin G in the generation of elastase-resistant fragment of vascular endocan: involvement in the regulation of LFA-1-dependent cascade

The migration of polymorphonuclear neutrophils (PMN) into inflamed tissue requires fine interactions with the endothelial cell surface. The PMN serine proteases cathepsin G (CG), neutrophil elastase (NE) and proteinase 3 (PR3) were originally thought to play a role by the cleavage of endothelial cell proteins that control the PMN firm adhesion and the transendothelial cell migration. However, how these proteases participate in leukocyte adhesion and transmigration remains controversial. Vascular endocan, also called esm1, is a restricted endothelial cell-secreted proteoglycan constituted by a protein core of 20 kDa and by a unique glycosaminoglycan chain of dermatan sulphate (DS). Endocan is preferentially expressed in lung and kidney tissues. Endocan binds to its leukocytic receptor, the LFA-1 integrin, with an affinity of 18 nM, and inhibits the LFA-1-ICAM-1 interactions. Its expression is upregulated by the proinflammatory mediators TNF, IL-1, and lipopolysaccharide (LPS). In human sepsis, the serum endocan increases from fivefold to 30-fold the normal value and correlates with bad prognosis. Here, we examined the role of PMN-derived proteases in the degradation of endocan.

Differential kinetics of endothelial cell activation biomarkers E-selectin and endocan during nonlethal endotoxemia in 129Sv mice: a role for PMN-derived serine proteases in the transient decrease of circulating endocan levels

Severe septic syndrome remains one of the most frequent causes of death in ICUs. One of the main players in this pathology is the endothelium integrity. Our laboratory has demonstrated in preliminary clinical studies among the various biomarkers of endothelial dysfunction that blood levels of endocan (ESM-1), a pulmonary vascular endothelial cell-specific molecule participating in the control of endothelial-leukocyte interactions, are associated with the severity and evolution of septic states. On the other hand, we showed in vivo that antiprotease therapy is associated with a decrease of the leukocyte rolling and firm leukocyte adhesion to endothelium in sepsis. This decrease in the leukocyte-endothelial cell contacts was associated with an increase of blood endocan levels, suggesting a linkage between leukocyte proteases, endocan, and inflammation during sepsis.