Krzysztof Grabowski

Acute-phase response proteins are related to cachexia and accelerated angiogenesis in gastroesophageal cancers

Abstract Background: Accurate outcome prediction in gastroesophageal malignancies is challenging. Acute-phase response proteins (APRPs) have been claimed to be independent prognosticators, although the basis for their association with prognosis remains unexplained. We hypothesized that, similarly to pancreatic and lung cancers, changes in APRPs in gastroesophageal malignancies are associated with cachexia and accelerated angiogenesis. Methods: C-reactive protein (CRP), albumin and transferrin serum levels were evaluated and the Glasgow Prognostic Score (GPS) calculated. These data were compared with concentrations of circulating interleukin (IL)-1, IL-6 and IL-8, tumor necrosis factor-α (TNF-α), vascular endothelial growth factor (VEGF)-A, VEGF-C and midkine in 96 gastroesophageal cancer patients (49 with cachexia) and 42 healthy subjects. Results: Albumin and CRP levels were altered in the cancer patients, with further CRP elevation in those with cachexia. Transferrin was decreased only in the cachectic patients. The interrelationships between the APRPs were strengthened in cachexia and only then were APRPs correlated with the cytokines elevated in gastroesophageal cancer-related cachexia: IL-6, IL-8, VEGF-A and midkine. GPS corresponded well to transferrin, IL-1, IL-6, IL-8, TNF-α, VEGF-A and midkine concentrations. Conclusions: Cachexia in gastroesophageal cancers is associated with changes in APRP concentrations. This, together with a direct relationship of APRPs with accelerated angiogenesis, may constitute a foundation for the association of APRPs and GPS with outcome in these malignancies. Clin Chem Lab Med 2008;46:359–64.

Elevation of circulating interleukin-8 is related to lymph node and distant metastases in esophageal squamous cell carcinomas—Implication for clinical evaluation of cancer patient

The presence of lymph node metastasis (LNM) is an important factor in clinical evaluation of esophageal cancer patients. Biological markers able to support detection of metastatic lymph nodes are sought after. Interleukin-8 (IL-8) is overexpressed by many cancers and involved in cancer dissemination. We investigated the relationship between circulating IL-8 and clinicopathological features of esophageal squamous cell carcinoma (ESCC), and evaluated the diagnostic potential of IL-8, with reference to the key angiogenic and lymphangiogenic factors: vascular endothelial growth factors A and C (VEGF-A and VEGF-C). We found elevated IL-8 levels in ESCC patients, correlated with tumor size and cancer dissemination, especially LNM. Circulating IL-8 correlated with lymphangiogenic VEGF-C rather then angiogenic VEGF-A. The association weakened in metastatic cancers, suggesting divergent mechanism of IL-8 involvement in the dissemination process. The cytokine levels correlated with platelets and neutrophils, pointing at these cells as possible sources of circulating IL-8. We demonstrated IL-8 that positively correlated with inflammation status of ESCC patients. Circulating IL-8 was a better indicator of ESCC dissemination than VEGF-A or VEGF-C. Yet, the detection rates were not satisfactory enough to allow for the recommendation of IL-8 determination as an adjunct to the clinical evaluation of lymph node involvement in ESCC patients.

Serum midkine depends on lymph node involvement and correlates with circulating VEGF-C in oesophageal squamous cell carcinoma

Abstract Lymph node metastasis (LNM) is a key factor for selection of treatment method and patients’ prognosis in oesophageal squamous cell carcinoma (ESCC). However, no biomarkers able to support the clinical detection of LNM have been reported. Recently, vascular endothelial growth factor C (VEGF-C) was found to be a more accurate marker of LNM in lung cancer than computed tomography. Midkine is a multifunctional cytokine involved in cancer development. We investigated circulating midkine levels in ESCC patients (n=73) compared with those in healthy subjects (n=42) with double-antibody-sandwich indirect enzyme-linked immunosorbent assay (DASI-ELISA). We found that midkine was elevated in ESCC and involved in metastatic disease. Serum midkine (sMK) was a good marker of LNM, evaluated both clinically and pathologically, as revealed by ROC analysis. It also correlated with serum levels of VEGF-C. The increase of sMK was related to cancer cells, although a weak correlation was observed between sMK and platelet and leucocyte counts.

Paraoxonase 1 (PON1) status in gastroesophageal malignancies and associated paraneoplastic syndromes — Connection with inflammation

OBJECTIVE To evaluate the status and diagnostic utility of antioxidant paraoxonase 1 (PON1) in gastroesophageal cancers. DESIGN AND METHODS PON1s arylesterase/paraoxonase activities and phenotype were determined in 82 cancers and 57 controls, and related to clinicopathological features, anemia and cachexia coexistence, cholesterol levels, liver function panel, and inflammatory and angiogenic indices. RESULTS PON1s activities were decreased in cancer. Arylesterase activity correlated with cancer T and N stages, being an N1-independent predictor. Both activities correlated with transferrin, albumin, CRP and inflammation-based Glasgow Prognostic Score, and arylesterase activity also with interleukin-6 and midkine. Phenotype A (Q192 homozygotes) was associated with lower transferrin and higher TNF-alpha concentrations. PON1s arylesterase activity reflected anemia severity, being correlated with hemoglobin, hematocrit, and iron. PON1 arylesterase activity negatively correlated with alkaline phosphatase and gamma-glutamyl transferase, but not with bilirubin, aminotransferases, HDL or LDL cholesterol. PON1 moderately indicated cancer presence and regional metastasis. CONCLUSIONS PON1 activity decreases in gastroesophageal cancers and corresponds to inflammation severity and cancer-related anemia. PON1 decrease indicates lymph node metastasis, but its moderate predictive power does not recommend PON1 determination alone for clinical application.

Circulating leptin and inflammatory response in esophageal cancer, esophageal cancer-related cachexia-anorexia syndrome (CAS) and non-malignant CAS of the alimentary tract.

We investigated the association between esophageal cancer and cachexia-anorexia syndrome (CAS) of the alimentary tract and leptin, an adipocytokine crucial for body weight regulation, a modulator of inflammatory/immune response, implication of which in cancer and CAS development remains debatable. Circulating leptin was measured in 135 esophageal cancer patients (51 non-cachectic and 84 cachectic) and 83 controls (63 non-cachectic and 20 cachectic) and referred to cancer stage, CAS, and inflammatory and nutritional indices. Leptin was down-regulated in cancer patients and cachectic controls as compared to non-cachectic controls, with more pronounced hypoleptinemia in advanced cancers. Leptin correlated directly with BMI, TNF-alpha, albumin, and hemoglobin and indirectly with IL-6, IL-8, and hsCRP. The correlations, except for hsCRP, were more pronounced in females. BMI alone (females) and BMI and hsCRP (males) were independent predictors of leptin explaining over 60% of its variability. Following adjustment for BMI and gender, cancer-related CAS but not cancer itself negatively affected leptin. Leptin and BMI were independently associated with cancer-related and non-malignant CAS with diagnostic accuracy of 93% in identifying subjects with CAS. Pro-inflammatory, angiogenic and mitogenic properties of leptin do not seem to be important for esophageal cancer development but hypoleptinemia, independently from co-occurring reduction of adiposity, appears to be strongly associated with esophageal cancer-related CAS and non-malignant CAS of the alimentary tract.

Cultivation of circulating tumor cells in esophageal cancer

The presence of circulating tumor cells (CTCs) in patients with metastatic carcinoma is generally associated with poor clinical outcome. There have been many investigations showing a possible use of CTCs as minimally invasive predictive and prognostic biomarker in cancer medicine. In this report a size-based method (MetaCell®) for quick and easy enrichment and cultivation of CTCs is presented to enable possible CTCs use in esophageal cancer (EC) management. In total, 43 patients with diagnosed EC, 20 with adenocarcinoma (AdenoCa) and 23 with squamous cell carcinoma (SCC), were enrolled into the adaptive prospective-like study .All the patients were candidates for surgery. The CTCs were detected in 27 patients (62.8%), with a higher rate in adenocarcinoma (75%) than SCC (52%). Finally, there were 26 patients with resectable tumors exhibiting CTCs-positivity in 69.2% and 17 patients with non-resectable tumors with 41.7% CTCs-positivity. Interestingly, in the patients undergoing neoadjuvant therapy, the CTCs were detected at time of surgery in 55.5% (10/18). The overall size-based filtration approach enabled to isolate viable CTCs and evaluate to their cytomorphological features by means of vital fluorescent staining. The CTCs were cultured in vitro for further downstream applications including immunohistochemical analysis. This is the first report of the successful culturing of esophageal cancer CTCs. The detection of CTCs presence could help in the future to guide timing of surgical treatment in EC patients.

TNF-α promotes breast cancer cell migration and enhances the concentration of membrane-associated proteases in lipid rafts

PurposeTumor progression is associated with cell migration, invasion and metastasis. These processes are accompanied by the activation of specific proteases that are either linked to cellular membranes or are secreted into extracellular spaces. TNF-α is known to play an important role in various aspects of tumor progression. The aim of this work was to assess the effect of TNF-α on the migration of breast cancer cells and, in addition, to assess its association with the location of membrane-associated proteases in lipid rafts.MethodsWound scratch healing and Transwell migration assays were used to study the effect of TNF-α on the migration of both hormone-dependent and hormone-independent breast cancer-derived cells, i.e., MCF7 and MDA-MB-231, respectively. The expression and secretion of three matrix metalloproteases, MMP9, MMP2 and MT1-MMP, and two dipeptidyl peptidases, CD26 and FAP-α, was investigated using RT-PCR, Western blotting and gelatin zymography. In addition, activation of the MAPK/ERK signaling pathway was investigated by Western blotting.ResultsWe found that a TNF-α-induced enhancement of breast cancer cell migration was accompanied by an increased secretion of MMP9, but not MMP2, into the culture media. We also found that TNF-α upregulated the expression of the dipeptidyl peptidases CD26 and FAP-α in a dose-dependent manner and, in addition, enhanced the concentration of all five proteases in lipid rafts in the breast cancer-derived cells tested, regardless of cell type. Furthermore, we found that TNF-α activated the MAPK/ERK signaling pathway by increasing the ERK1/2 phosphorylation level. Application of the MEK/ERK1/2 inhibitor U-0126 resulted in down-regulation of TNF-α-induced MMP9 secretion and abrogation of the enhanced concentration of proteases in the lipid rafts.ConclusionsFrom our results we conclude that TNF-α-induced activation of the MAPK/ERK signaling pathway may promote breast cancer cell migration via both upregulation of MMP9, CD26 and FAP-α and concentration of these proteases, as also MT1-MMP and MMP2, in the lipid rafts. TNF-α may serve as a potential therapeutic target in breast cancers susceptible to TNF-α stimulation.

Serum Levels of Resistin, Adiponectin, and Apelin in Gastroesophageal Cancer Patients

The aim of the study was the investigation of relationship between cachexia syndrome and serum resistin, adiponectin, and apelin in patients with gastroesophageal cancer (GEC). Material and Methods. Adipocytokines concentrations were measured in sera of 85 GEC patients and 60 healthy controls. They were also evaluated in tumor tissue and appropriate normal mucosa of 38 operated cancer patients. Results. Resistin and apelin concentrations were significantly higher in GEC patients than in the controls. The highest resistin levels were found in cachectic patients and in patients with distant metastasis. Serum adiponectin significantly decreased in GEC patients with regional and distant metastasis. Serum apelin was significantly higher in cachectic patients than in the controls. Apelin was positively correlated with hsCRP level. Resistin and apelin levels increased significantly in tumor tissues. Weak positive correlations between adipocytokines levels in serum and in tumor tissue were observed. Conclusions. Resistin is associated with cachexia and metastasis processes of GEC. Reduction of serum adiponectin reflects adipose tissue wasting in relation to GEC progression. Correlation of apelin with hsCRP can reflect a presumable role of apelin in systemic inflammatory response in esophageal and gastric cancer.

Evaluation of 8-hydroxydeoxyguanosine, thiobarbituric acid-reactive substances and total antioxidant status as possible disease markers in oesophageal malignancies.

OBJECTIVES Evaluation of oxidative stress and diagnostic utility of its markers in oesophageal squamous cell carcinoma (OSCC). DESIGN Serum 8-hydroxydeoxyguanosine, thiobarbituric acid-reactive substances (TBARS) and total antioxidant status (TAS) were measured in OSCC (n=75), non-malignant oesophageal diseases (n=30), and healthy subjects (n=79). Three months following oesophagectomy the measurements were repeated. RESULTS Exclusively in OSCC, 8-hydroxydeoxyguanosine and TBARS were elevated. TAS was reduced in non-malignancies compared to controls, and in OSCC compared to non-malignancies and controls. Only 8-hydroxydeoxyguanosine was associated with disease progression, lymph node involvement in particular. All indices were good indicators of cancer presence (ROC analysis) and normalized following oesophagectomy. A positive linear relationship between 8-hydroxydeoxyguanosine and TBARS, and negative non-linear between TAS and both 8-hydroxydeoxyguanosine and TBARS was demonstrated. CONCLUSION OSCC is associated with oxidative stress, attenuated following oesophagectomy. Consumption of serum antioxidants prevents accumulation of oxidatively modified molecules in non-malignancies. High accuracy of oxidative stress markers in indicating cancer presence warrants further investigation on their possible application as discriminatory markers and in monitoring treatment efficacy.

Soluble urokinase-type plasminogen activator receptor and ferritin concentration in patients with advanced alimentary tract carcinoma. Relationship to localization, surgical treatment and the stage of the disease--preliminary report.

BACKGROUND The urokinase plasminogen activation system is associated with metastatic potential of cancer in several tumors. Its specific membrane receptor (uPAR) is released from cancer cells and can be detected as the soluble fraction of the plasminogen activator receptor (suPAR). Ferritin (FRT) is a poor prognostic factor in various neoplasms. OBJECTIVES We analyzed the serum concentrations of suPAR and FRT in patients with gastrointestinal cancer. Tumor localization, stage of the disease, possibility of surgery and histopathological diagnosis were considered. MATERIAL AND METHODS The analysis involved 48 patients (8 females/40 males) treated in the Department of Gastrointestinal and General Surgery, Wroclaw Medical University. Thirty two patients had esophageal, 7-gastric, 9-colorectal cancer. Fifteen patients underwent resection surgery, 33 palliative therapy. The control comprised 10 healthy donors. The serum concentration of suPAR was determined by enzyme-linked immunosorbent assay (ELISA), expressed in pg/mL. Concentration of the serum FRT was detected using immunonephelometry method, expressed in µg/L. RESULTS Serum concentration of suPAR ranged from 1789-7320, x=3676.2, SD=1042 and was significantly higher (p=0.0002) than in the control group. In patients who underwent palliative therapy, the concentration of suPAR was significantly higher (p=0.05) than in those after resection, also in patients with esophageal cancer compared to those with colorectal one (p=0.02). Serum concentration of FRT in patients with gastrointestinal cancer was significantly higher than in control group. Serum FRT concentration was higher in patients with esophageal cancer compared to patients with gastric cancer (p=0.05), in persons with IV compared to patients with I-III stage of the disease, patients who underwent palliative compared to surgical therapy. CONCLUSIONS In patients with gastrointestinal cancer the level of suPAR is high, with highest values in advances disease with remote metastases. The FRT concentration is sensitive indicator of the disease process: its level is highest in pts with IV stage who underwent palliative therapy.