Dysregulation of Glypicans and Notum in Osteoarthritis: Plasma Levels, Bone Marrow Mesenchymal Stromal Cells and Osteoblasts.

In this study of the alterations of Glypicans 1 to 6 (GPCs) and Notum in plasma, bone marrow mesenchymal stromal cells (BM-MSCs) and osteoblasts in Osteoarthritis (OA), the levels of GPCs and Notum in the plasma of 25 patients and 24 healthy subjects were measured. In addition, BM-MSCs from eight OA patients and eight healthy donors were cultured over a period of 21 days using both a culture medium and an osteogenic medium. Protein and gene expression levels of GPCs and Notum were determined using ELISA and qPCR at 0, 7, 14 and 21 days. GPC5 and Notum levels decreased in the plasma of OA patients, while the BM-MSCs of OA patients showed downexpression of GPC6 and upregulation of Notum. A decrease in GPC5 and Notum proteins and an increase in GPC3 were found. During osteogenic differentiation, elevated GPCs 2, 4, 5, 6 and Notum mRNA levels and decreased GPC3 were observed in patients with OA. Furthermore, the protein levels of GPC2, GPC5 and Notum decreased, while the levels of GPC3 increased. Glypicans and Notum were altered in BM-MSCs and during osteogenic differentiation from patients with OA. The alterations found point to GPC5 and Notum as new candidate biomarkers of OA pathology.

Sandwich-type supersensitive electrochemical aptasensor of glypican-3 based on PrGO-Hemin-PdNP and AuNP@PoPD.

A new sandwich-type electrochemical biosensing platform was developed by gold @polyphthalenediamine nanohybrids (AuNP@PoPD) as the sensing platform and phosphorus doped reduced graphene oxide-hemin-palladium nanoparticles (PrGO-Hemin-PdNP) as the signal amplifier for phosphatidylinositol proteoglycan 3 (GPC3). AuNP@PoPD, co-electrodeposited into the screen printed electrode with high conductivity and stability, is dedicated to assembling the primary GPC3 aptamer (GPC3Apt). The second GPC3Apt immobilized on the high conductivity and large surface area of PrGO-Hemin-PdNP was utilized as an electrochemical signal reporter by hemin oxidation (PrGO-Hemin-PdNP-GPC3Apt). In the range 0.001-10.0 ng/mL, the hemin oxidation current signal of the electrochemical aptasensor increased log-linearly with the concentration of GPC3, the lowest detection limit was 0.13 pg/mL, and the sensitivity was 2.073 µA/µM/cm2. The aptasensor exhibited good sensing performance in a human serum sample with the relative error of 4.31-8.07%. The sandwich sensor showed good selectivity and stability for detection GPC3 in human serum samples, providing a new efficient and sensitive method for detecting HCC markers.

A dual-signal output electrochemical aptasensor for glypican-3 ultrasensitive detection based on reduced graphene oxide-cuprous oxide nanozyme catalytic amplification strategy.

Glypican-3 (GPC3) is an essential reference target for hepatocellular carcinoma detection, follow-up and prediction. Herein, a dual-signal electrochemical aptasensor based on reduced graphene oxide-cuprous oxide (RGO-Cu2O) nanozyme was developed for GPC3 detection. The RGO-Cu2O nanoenzyme displayed excellent electron transport effect, large specific surface area and outstanding peroxidase-like ability. The differential pulse voltammetry (DPV) signal of Cu2O oxidation fraction and the chronoamperometry (i-t) signal of H2O2 decomposition catalyzed by RGO-Cu2O nanozyme were used as dual-signal detection. Under optimal conditions, the log-linear response ranges were 0.1 to 500.0 ng/mL with the limit of detection 0.064 ng/mL for DPV technique, and 0.1-50.0 ng/mL for i-t technique (detection limit of 0.0177 ng/mL). The electrochemical aptasensor has remarkably analytical performance with wide response range, low detection limit, excellent repeatability and specificity, good recovery in human serum samples. The two output signals of one sample achieve self-calibration of the results, effectively avoiding the occurrence of possible leakage and misdiagnosis of a single detection signal, suggesting that it will be a promising method in the early biomarker detection.

Serum glypican-4 is a marker of future vascular risk and mortality in coronary angiography patients.

BACKGROUND AND AIMS: Glypican-4 (GPC4) is a cell surface protein, but can be released into circulation under various clinical conditions. The association of circulating GPC4 with the risk of future cardiovascular events or death is unclear. In the present study, we aimed to investigate the association between serum GPC4 and major adverse cardiovascular events (MACE), vascular mortality, and all-cause mortality in a prospective cohort study. METHODS: Our study included 760 patients undergoing coronary angiography. During a mean follow up period of 6.3 years, the incidence of MACE, vascular mortality, and all-cause mortality was recorded. Serum GPC4 levels were determined using an enzyme-linked immunosorbent assay. RESULTS: Serum GPC4 was highly significantly associated with increased age, body mass index, brain natriuretic peptide, and oxidized low density lipoprotein, as well as with decreased estimated glomerular filtration rate. During the follow-up period, 145 patients died, including 67 vascular deaths. MACE occurred in 137 patients. Serum GPC4 was significantly associated with MACE, vascular mortality, and all-cause mortality independently of traditional cardiovascular risk factors, with adjusted hazard ratios (HR) and 95% confidence intervals for one standard deviation change of serum GPC4 of 1.32 [1.10-1.58], 1.38 [1.06-1.78], and 1.53 [1.29-1.82], respectively. The best cut-off value for serum GPC4 for predicting MACE, vascular mortality, and all-cause mortality was 7.24 ng/ml for all three endpoints. Respective adjusted HRs were 1.61 [1.07-2.43], 2.85 [1.62-5.01], and 2.92 [2.00-4.27]. CONCLUSIONS: Our study indicates that elevated serum GPC4 levels are significantly associated with an increased risk of MACE, vascular mortality, and all-cause mortality.

How Reliable are Commercially Available Glypican4 ELISA Kits?

OBJECTIVE: Glypican4 is an interesting new adipokine, which seems to play an important role in developmental processes and is potentially associated with metabolic changes in obesity and type 2 diabetes mellitus. Currently, only a few studies examined glypican4 in human blood, mainly in adults. DESIGN, PATIENTS AND MEASUREMENTS: The aim of our study was to investigate glypican4 serum levels in lean, overweight, and obese children and adolescents, to unravel a possible association between glypican4 serum levels and parameters of obesity and insulin resistance. In order to determine a suitable method for investigating glypican4 serum levels, we validated two commercially available human glypican4 ELISA kits, using serum and plasma samples of an obese, insulin-resistant patient, and a healthy control subject, a human recombinant glypican4 protein fragment and glypican4-overexpressing cell lysate. RESULTS: Using ELISA kit #1 we were not able to detect values above background level, apart from standard curve values. ELISA kit #2 initially seemed suitable to measure glypican4, but further validation experiments showed non-linearity of serial dilutions, no recognition of a human recombinant glypican4 protein fragment and non-linearity in the recovery of glypican4-overexpressing cell lysate. In addition, there was a considerable decrease (approx. 68%) of measured values between two experiments, performed at different time points with aliquots of the same serum sample. Contrary to that, further experiments found sample stability not to be compromised. CONCLUSIONS: Extensive evaluation of the performance of two commercially available ELISA kits led to the conclusion that none of them is applicable for the measurement of glypican4 in human blood samples.

Glypican-4 in pregnancy and its relation to glucose metabolism, insulin resistance and gestational diabetes mellitus status.

Glypican-4 (GPC-4) is an adipokine that enhances insulin receptor signaling. Plasma concentrations were found to be elevated in patients with prediabetes but reduced in type 2 diabetes mellitus. No study on Glypican-4 in pregnancy and pregnancy-related insulin resistance has been published yet. GPC-4 levels were investigated in 59 overweight women throughout their pregnancy at the Medical University of Vienna. GPC-4 levels, fasting insulin, fasting glucose, estradiol, liver and renal parameters, and markers of bone development were assessed before the < 21st week of gestation (GW), and at GW 35-37. GPC-4 levels increased from < 21 GW (mean = 2.38 pg/ml, SD = 0.68 pg/ml) to GW 35-37 (mean = 2.96 pg/ml, SD = 0.77 pg/ml, p < 0.001). At the same time, GPC-4 levels correlated negatively with estimated glomerular filtration rate (eGFR), serum protein and serum albumin levels and were positively related to creatinine and uric acid levels at GW 35-37. Concerning glucose metabolism, GPC-4 levels were inversely related to ISSI-2, fasting insulin and HOMA-IR, however, not significantly different between women with normal glucose tolerance (NGT) and GDM (p = 0.239). In conclusion, GPC-4 levels rose significantly during pregnancy, correlated negatively with fasting insulin and HOMA-IR but might not be related to gestational diabetes mellitus status.

Galanin and glypican-4 levels depending on metabolic and cardiovascular risk factors in patients with polycystic ovary syndrome

ABSTRACT Objective: Galanin is a neuropeptide which has effects not only on metabolic syndrome but also on reproduction. Glypican-4 is an adipokine associated with insulin sensitivity by interacting directly with the insulin receptor. This study evaluated serum concentrations of galanin and glypican-4 in relation with the hormonal profile as well as metabolic and cardiovascular risk factors in patients with and without polycystic ovary syndrome (PCOS). Subjects and methods: A total of 44 women with PCOS and 44 age-matched controls were eligible. Hirsutism scores, hormonal profile, metabolic and cardiovascular risk factors as well as galanin and glypican-4 levels were evaluated in each subject. Results: Women with PCOS exhibited lower levels of galanin (20.2 pg/mL versus 26.4 pg/mL, p = 0.002) and higher concentrations of glypican-4 (3.1 ng/mL versus 2.6 ng/mL, p < 0.001) than controls. Both adipokines were correlated positively with body mass index (BMI), insulin, triglyceride and Homeostasis Model Assessment (HOMA) index; glypican-4 also showed positive correlations with fasting blood glucose, free testosterone, modified Ferriman-Gallwey scores (p < 0.05). Multiple Linear Regression analyses showed that PCOS and BMI were the best predictors affecting galanin levels with a decreasing and increasing effect respectively; however BMI was the best predictor affecting glypican-4 levels with an increasing effect (p < 0.001). Conclusion: Galanin levels were lower and glypican-4 levels were higher in women with PCOS than controls. Further studies are needed to determine whether these adipokines could be used as additional markers for insulin sensitivity and lipid profile and whether they might play a role in the pathogenesis of PCOS, in which metabolic cardiovascular risks are increased.

Shed antigen-induced blocking effect on CAR-T cells targeting Glypican-3 in Hepatocellular Carcinoma.

BACKGROUND: Glypican-3 (GPC3), a cell surface glycoprotein that is pathologically highly expressed in hepatocellular carcinoma (HCC), is an attractive target for immunotherapies, including chimeric antigen receptor (CAR) T cells. The serum GPC3 is frequently elevated in HCC patients due to the shedding effect of cell surface GPC3. The shed GPC3 (sGPC3) is reported to block the function of cell-surface GPC3 as a negative regulator. Therefore, it would be worth investigating the potential influence of antigen shedding in anti-GPC3 CAR-T therapy for HCC. METHODS: In this study, we constructed two types of CAR-T cells targeting distinct epitopes of GPC3 to examine how sGPC3 influences the activation and cytotoxicity of CAR-T cells in vitro and in vivo by introducing sGPC3 positive patient serum or recombinant sGPC3 proteins into HCC cells or by using sGPC3-overexpressing HCC cell lines. RESULTS: Both humanized YP7 CAR-T cells and 32A9 CAR-T cells showed GPC3-specific antitumor functions in vitro and in vivo. The existence of sGPC3 significantly inhibited the release of cytokines and the cytotoxicity of anti-GPC3 CAR-T cells in vitro. In animal models, mice carrying Hep3B xenograft tumors expressing sGPC3 exhibited a worse response to the treatment with CAR-T cells under both a low and high tumor burden. sGPC3 bound to CAR-T cells but failed to induce the effective activation of CAR-T cells. Therefore, sGPC3 acted as dominant negative regulators when competed with cell surface GPC3 to bind anti-GPC3 CAR-T cells, leading to an inhibitory effect on CAR-T cells in HCC. CONCLUSIONS: We provide a proof-of-concept study demonstrating that GPC3 shedding might cause worse response to CAR-T cell treatment by competing with cell surface GPC3 for CAR-T cell binding, which revealed a new mechanism of tumor immune escape in HCC, providing a novel biomarker for patient enrolment in future clinical trials and/or treatments with GPC3-targeted CAR-T cells.

Galanin and glypican-4 levels depending on metabolic and cardiovascular risk factors in patients with polycystic ovary syndrome.

OBJECTIVE: Galanin is a neuropeptide which has effects not only on metabolic syndrome but also on reproduction. Glypican-4 is an adipokine associated with insulin sensitivity by interacting directly with the insulin receptor. This study evaluated serum concentrations of galanin and glypican-4 in relation with the hormonal profile as well as metabolic and cardiovascular risk factors in patients with and without polycystic ovary syndrome (PCOS). METHODS: A total of 44 women with PCOS and 44 age-matched controls were eligible. Hirsutism scores, hormonal profile, metabolic and cardiovascular risk factors as well as galanin and glypican-4 levels were evaluated in each subject. RESULTS: Women with PCOS exhibited lower levels of galanin (20.2 pg/mL versus 26.4 pg/mL, p = 0.002) and higher concentrations of glypican-4 (3.1 ng/mL versus 2.6 ng/mL, p < 0.001) than controls. Both adipokines were correlated positively with body mass index (BMI), insulin, triglyceride and Homeostasis Model Assessment (HOMA) index; glypican-4 also showed positive correlations with fasting blood glucose, free testosterone, modified Ferriman-Gallwey scores (p < 0.05). Multiple Linear Regression analyses showed that PCOS and BMI were the best predictors affecting galanin levels with a decreasing and increasing effect respectively; however BMI was the best predictor affecting glypican-4 levels with an increasing effect (p < 0.001). CONCLUSION: Galanin levels were lower and glypican-4 levels were higher in women with PCOS than controls. Further studies are needed to determine whether these adipokines could be used as additional markers for insulin sensitivity and lipid profile and whether they might play a role in the pathogenesis of PCOS, in which metabolic cardiovascular risks are increased.

Serum glypican4 and glycosylphosphatidylinositol-specific phospholipase D levels are associated with adipose tissue insulin resistance in obese subjects with different glucose metabolism status.

OBJECTIVES: Glypican4 (GPC4) is a novel adipokine associated with obesity and insulin resistance. GPC4 was cleaved by the glycosylphosphatidylinositol-specific phospholipase D (GPLD1) in an anchored site of the glycosylphosphatidylinositol, and then was released into the extracellular environment. Herein, we investigated the changes of serum GPC4 and GPLD1 levels in obese subjects with different glucose metabolism status and their relationship with adipose tissue insulin resistance index (Adipo-IR) in Chinese north populations. METHODS: A total of 221 obese subjects and 37 normal controls (NC) were recruited in this study. Obese subjects were divided into normal insulin (NI) group, hyperinsulinemia (HI) group, impaired glucose tolerance (IGT) group, and type 2 diabetes mellitus (DM) group. Serum GPC4, GPLD1, and adiponectin were determined by commercially available ELISA kits. RESULTS: Serum GPC4 levels in the HI, IGT, and DM groups were significantly higher than those in the NC and NI groups (2.27 ± 0.58 ng/mL, 2.21 ± 0.60 ng/mL, 2.49 ± 0.67 ng/mL vs. 1.70 ± 0.33 ng/mL, 1.93 ± 0.34 ng/mL, P < 0.05). GPC4 was positively correlated with GPLD1, which was the most important influencing factor of GPC4. Adipo-IR was independently and positively associated with serum GPC4 and GPLD1. For GPC4, after adjustment for confounders, the risk of adipose tissue insulin resistance in subjects with the highest tertile was 2.974-fold that of those with the lowest tertile (OR = 2.974, P = 0.013). For GPLD1, before adjustment for lipids, the increased probability still existed (Model 2, OR = 3.568, P = 0.003). CONCLUSION: GPC4 is an adipokine associated with adipose tissue insulin resistance, and its activity may be regulated by GPLD1. GPC4 may be a marker for adipose tissue insulin resistance in Chinese north obese populations.

Can Glypican-6 Level Predict Ejection Fraction Decline After Myocardial Infarction?

The main goals in the treatment of acute coronary syndrome are to prevent myocardial ischemia, damage, and possible complications. Accordingly, we evaluated the predictive value of glypican-6 (GPC6) for cardiac remodeling after myocardial infarction (MI). Baseline plasma GPC6 levels were measured in patients who underwent primary percutaneous coronary intervention (PCI) for acute MI. Left ventricular ejection fraction (LVEF) was measured at baseline and at 6 months with transthoracic echocardiography. Reduced LVEF persisted in 89 out of 276 patients after 6 months. The majority of the patients were male (n = 198, 72%) and the mean age was 57.8 ± 10.8 years. Glypican-6, N-terminal pro-brain natriuretic peptide (NT-proBNP), and high-sensitive troponin levels were significantly lower in the improved LVEF group compared with the low LVEF group (10.54 ± 4.46 vs 6.98 ± 3.34 ng/mL, P < .001; 500 pg/mL [range, 300-600 pg/mL] vs 350 pg/mL [range, 200-550 pg/mL], P = .008; 396 pg/mL [range, 159-579 pg/mL] vs 300 pg/mL [range, 100-500 pg/mL], P = .016, respectively). Logistic regression analysis revealed the SYNTAX Score 2, GPC6, and NT-proBNP as significant independent predictors of low LVEF (hazard ratio [HR]: 1.064, P = .041; HR: 1.215, P < .001; HR: 1.179, P < .001). Glypican-6 may prove to be useful for the detection of low LVEF development in patients undergoing PCI following MI.

Highly sensitive electrochemical aptasensor for Glypican-3 based on reduced graphene oxide-hemin nanocomposites modified on screen-printed electrode surface.

Glypican-3 (GPC3) is a highly specific tumor marker for hepatocellular carcinoma (HCC), and plays an important role in reflecting the existence, therapeutic evaluation, monitoring and prognosis of HCC. Herein, an electrochemical aptasensor was designed for GPC3 detection with the reduced graphene oxide-hemin nanocomposites (RGO-Hemin) modified on the screen-printed electrode surface as the sensing platform and GPC3 aptamer as recognize molecule. In the existence of GPC3, the aptamer can specifically bind with the target GPC3 and form GPC3-aptamer conjugations on the sensing surface, which would increase the resistance of the electron transfer on the electrode and make the decrease of electrochemical signals of Hemin in RGO-Hemin nanocomposites. The electrochemical current change was recorded by differential pulse voltammetry (DPV). Scanning electron microscopy (SEM), Raman microscope (RM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the GPC3 electrochemical aptasensor. Under the optimum conditions, the current response of the electrochemical aptasensor is linearly correlated with the concentration of GPC3 (0.001-10.0 µg/mL) with the detection limit of 2.86 ng/mL (S/N = 3) and the sensitivity of 0.134 µA/µM/cm2. In addition, the aptasensor was applied to the determination of GPC3 in spiked human plasma and the recoveries fluctuated from 102.68% to 117.29%. All these results show that the aptasensor has good specificity, sensitivity, stability and reproducibility for GPC3 detection.

Soluble cluster of differentiation 26/soluble dipeptidyl peptidase-4 and glypican-3 are promising serum biomarkers for the early detection of Hepatitis C virus related hepatocellular carcinoma in Egyptians.

BACKGROUND AND STUDY AIMS: Many patients are diagnosed with hepatocellular carcinoma (HCC) in the late stage when it is already untreatable. Therefore, there is an increased need for sensitive biomarkers to detect HCC at an earlier stage in high risk patients with hepatitis C virus (HCV)-induced cirrhosis. This study aimed to evaluate the diagnostic performance of soluble cluster of differentiation 26/dipeptidyl peptidase 4 (sCD26/sDPP4) and glypican-3 (GPC3) as serum biomarkers for the early detection of HCV related HCC and compare it with that of the conventional tumor marker serum alpha fetoprotein (AFP). PATIENTS AND METHODS: The study included 80 participants, 30 patients diagnosed with HCV infection without HCC (HCV group), 30 patients diagnosed with HCV- related HCC (HCV group), and 20 healthy volunteers (control group). The serum levels of GPC3 and sCD26 were measured using specific enzyme linked immunosorbant assay (ELISA) kits, whereas AFP levels were determined using chemiluminescence. RESULTS: The serum levels of both sCD26 and GPC3 were found to be significantly higher in patients with early-stage HCC than in the HCV group, (1450 and 1.16 ng/mL, respectively). sCD26 at a cutoff value of > 1000 ng/ml, showed a high sensitivity (83.3%) and 63.3% specificity with an area under curve (AUC) of 0.811 and a 95% confidence interval (CI) of (0.682-0.94). While, the combination of GPC3 and sCD26 exhibited the best diagnostic performance for early-stage-HCC because it increased the sensitivity and specificity (85% and 93.3% respectively), with an AUC of 0.986 and a 95% CI of (0.899-1.00) compared to sCD26 alone. CONCLUSION: We conclude that serum sCD26 could be a sensitive biomarker for the early detection of HCC among HCV patients. Moreover, the combination of sCD26 and GPC3 increases both the sensitivity and specificity for the early detection of HCV related HCC compared with AFP and could help in the monitoring of HCC in high risk patients with HCV induced cirrhosis.

Glypican-3 electrochemical aptamer nanobiosensor based on hemin/graphene nanohybrids peroxidase-like catalytic silver deposition.

A Glypican-3 (GPC3) electrochemical aptamer nanobiosensor based on hemin/graphene nanohybrids (HGNs) peroxidase-like catalytic silver deposition and GPC3 aptamer has been constructed for the determination of GPC3. The HGNs were prepared by an one-step reduction method. Fourier transform infrared spectroscopy (FT-IR), ultraviolet spectroscopy (UV-vis), and transmission electron microscopy (TEM) were used to study the structure and morphological characteristics of the HGNs. The GPC3 electrochemical aptamer nanobiosensor was constructed using HGNs-aptamer (HGNs-Apt) as the signal probe and GPC3 aptamer as the capture probe. With the help of the catalytic action of peroxidase-like properties of HGNs, H2O2 reduces the silver (Ag) ions in solution to metallic Ag, which deposit on the surface of the electrode. The amount of deposited Ag, which was derived from the amount of GPC3, was quantified by differential pulse voltammetry (DPV). Under optimal conditions, the current response of Ag had a good positive correlation with the GPC3 concentration in the range 10.0-100.0 µg mL-1 with a correlation coefficient of 0.9958. The detection limit was 3.16 µg mL-1 at a signal-to-noise ratio of 3, and the sensitivity was calculated to be 0.807 µA µM-1 cm-2. The method is validated by analyzing spiked human serum samples with good recovery ranging from 101 to 122%. In addition, the GPC3 electrochemical aptamer nanobiosensor has acceptable selectivity, stability, and reproducibility. Graphical abstract A Glypican-3 electrochemical aptamer nanobiosensor based on hemin/graphene nanohybrids (HGNs) peroxidase-like catalytic silver deposition and GPC3 aptamer has been constructed for the determination of GPC3. The electrochemical aptamer nanobiosensor exhibits high selectivity, acceptance reproducibility, and good recovery performances.

Diagnostic value of serum glypican-3 alone and in combination with AFP as an aid in the diagnosis of liver cancer.

BACKGROUND: Recently, a series of studies have been published to examine the possible diagnostic and prognostic values of glypican-3 (GPC3) in liver cancer with conflicting results observed. Thus, the present study aimed to assess the values of preoperative serum GPC3 alone and in combination with AFP for the diagnosis of liver cancer. METHODS: An enzyme-linked immunoassay was used to quantify serum GPC3 in hepatocellular carcinoma group (HCC, n = 210), intrahepatic cholangiocarcinoma group (ICC, n = 36), combined hepatocellular cholangiocarcinoma group (cHCC-CC, n = 8), metastatic liver cancer group (MLC, n = 10) and normal controls (NC, n = 134). RESULTS: The area under the curve (AUC) of GPC3 for HCC versus NC was 0.879, with a sensitivity of 79.52% at an optimal cutoff value of 0.0414 ng/mL; when GPC3 was combined with AFP, the AUC and sensitivity were increased to 0.925 and 88.10%, respectively. In addition, 43 of 68 AFP-negative patients had elevated GPC3 levels. Furthermore, the positive rate of GPC3 was significantly higher than the that of AFP for HCC in early stage. CONCLUSIONS: Serum GPC3 was superior to AFP for the diagnosis of early-stage HCC, and may be complementary to AFP for distinguishing HCC from NC.

Increased syndecan-1 and glypican-3 predict poor perinatal outcome and treatment resistance in intrahepatic cholestasis.

BACKGROUND: Intrahepatic cholestasis of pregnancy (ICP) increases the risk of adverse pregnancy outcomes. This study aimed to explore the association between serum syndecan-1 and glypican-3 levels and the adverse perinatal outcome as well as the responses to the treatment of ursodeoxycholic acid (UDCA). METHODS: This prospective, case control study included 88 pregnant women (44 women with ICP and 44 healthy controls). The primary end points were the perinatal outcome and the response to UDCA therapy. A logistic regression model was used to identify the independent risk factors of adverse pregnancy outcomes and reduced response to UDCA therapy. RESULTS: Women with ICP had significantly higher serum syndecan-1 (1.27 ± 0.36 ng/mL vs. 0.98 ± 0.50 ng/mL; P = 0.003), glypican-3 (1.78 ± 0.13 ng/mL vs.1.69 ± 0.16 ng/mL; P = 0.004), AST (128.59 ± 1.44 vs. 13.29 ± 1.32 U/L; P < 0.001), and ALT (129.84 ± 1.53 vs. 8.00 ± 3.67 U/L; P < 0.001) levels compared with the controls. The increased levels of syndecan-1 (OR = 4.715, 95% CI: 1.554-14.310; P = 0.006), glypican-3 (OR = 8.465, 95% CI: 3.372-21.248; P = 0.007), ALT (OR = 1.382, 95% CI: 1.131-1.690; P = 0.002), and postprandial bile acid (PBA) (OR = 3.392, 95% CI: 1.003-12.869; P = 0.026) were correlated to ICP. The adverse neonatal outcome was related to increased glypican-3 (OR = 4.275, 95% CI: 2.726-5.635; P = 0.039), and PBA (OR = 3.026, 95% CI: 1.069-13.569; P = 0.037). Increases of syndecan-1 (OR = 7.464, 95% CI: 2.130-26.153, P = 0.017) and glypican-3 (OR = 6.194, 95% CI: 2.951-13.002; P = 0.025) were the risk factors of decreased response to UDCA treatment. CONCLUSION: Syndecan-1 and glypican-3 might be powerful determinants in predicting adverse perinatal outcome in patients with ICP, and they can be used to predict the response to the UDCA treatment.

Plasma and tumoral glypican-3 levels are correlated in patients with hepatitis C virus-related hepatocellular carcinoma.

Glypican-3 (GPC3) is a cancer antigen expressed in approximately 80% of hepatocellular carcinomas (HCC) and is secreted into the blood. To confirm the effectiveness of GPC3 as a biomarker in HCC, we analyzed the relationship between GPC3 expression levels in cancer cells and in blood in 56 patients with HCC. Preoperative plasma GPC3 levels were determined with an immunoassay, and expression of GPC3 in resected tumors was analyzed by immunohistochemical staining. Median plasma GPC3 level in all HCC cases was 4.6 pg/mL, and tended to be higher in patients with hepatitis C virus (HCV)-related HCC (HCV group) (9.9 pg/mL) than in patients with hepatitis B virus (HBV)-related HCC (HBV group) (2.6 pg/mL) or in those without virus infection (None group) (3.0 pg/mL), suggesting that the virus type most likely influences GPC3 secretion. Median percentage of GPC3+ cells in tumors was also higher in the HCV (26.2%) and HBV (11.1%) groups than in the None group (4.2%). In the HCV group, there was a positive correlation between the two parameters (r = 0.66, P < .01). Moreover, receiver operating characteristic analysis predicted >10% GPC3+ cells in a tumor if the cut-off value was 6.8 pg/mL (sensitivity 80%, specificity 100%; area under the curve 0.875, 95% confidence interval 0.726-1) in the HCV group. Plasma concentration of GPC3 could be a predictive marker of tumoral GPC3 expression in patients with HCV-related HCC, suggesting a useful biomarker for immunotherapies targeting GPC3, although larger-scale validations are needed.

Hepatocellular carcinoma, a unique tumor with a lack of biomarkers.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide. Interestingly, the great majority of individuals affected by the tumor have underlying liver disease, therefore narrowing the population to be screened. Still, however, there is a clear lack of blood biomarkers, and surveillance in those at risk is performed by frequent imaging of the liver. A variety of multinational collaborations are currently invested in finding biomarkers for HCC based on liver-produced proteins. A new approach with assessment of peripheral proteins might be necessary for the successful early detection of this malignancy.

Clinical significance of glypican-3-positive circulating tumor cells of hepatocellular carcinoma patients: A prospective study.

The utility of glypican-3 (GPC3) expression for the detection of circulating tumor cells (CTCs) in hepatocellular carcinoma (HCC) patients has not been elucidated. The aim of this study was to identify associations between the presence of GPC3-positive CTCs and clinicopathological factors of these patients, furthermore, to evaluate whether CTC can predict microscopic portal vein invasion (mPVI). This study was done on 85 patients who underwent hepatectomy as the first-line treatment and whose preoperative imaging showed no evidence of macroscopic PVI and distant metastases. Peripheral blood was collected from all patients immediately before surgery. Cells were purified initially by density gradient centrifugation followed by immunomagnetic positive enrichment based upon the expression of GPC3. The numbers of CTCs contained in the enriched samples were enumerated via flow cytometry. Protocol validation using HepG2 cells spiked into 8.0 mL of blood from a healthy volunteer indicated that we were able to recover 12.1% of the tumor cells. A median number of 3 CTCs (range: 0-27) was detected in the 8.0 mL of peripheral blood of the 85 analyzed HCC patients. Thirty-three patients had CTCs ≥5, and these patients had a higher incidence of mPVI (p < 0.001), a lower disease-free survival (p = 0.015), and a lower overall survival (p = 0.047) than those with CTCs <5. Multivariate analysis identified CTCs ≥5 as an independent predictor of mPVI (p < 0.001). In conclusion, preoperative GPC3-positive CTCs ≥5 was a risk factor of mPVI and poor prognosis, and therefore may be a useful biomarker for HCC patient outcomes.