concanavalin-a and Colorectal-Neoplasms

The effect of essential fatty acids (EFA), given orally as dietary supplements, on the responsiveness in vitro of peripheral blood lymphocytes (PBL), to the mitogen concanavalin A have been studied in 10 patients with localized and 14 patients with advanced colorectal cancer. The degree of lymphocyte activation was assessed by measuring the amount of tritiated [3H]thymidine incorporated into newly synthesised lymphocyte DNA. The results were expressed as stimulation indices. T cell responses to concanavalin A stimulation showed a significant reduction of stimulation indices following EFA supplementation, in both the localized (P = 0.026) and advanced (P = 0.016) tumour groups, when compared with pretreatment activity in vitro. Mixing experiments, using EFA-supplemented and non-EFA-supplemented lymphocytes with concanavalin A, suggest no enhancement of T suppressor cell activity. Cell surface marker analysis (fluorescence-activated cell sorting for CD phenotyping) revealed a reduction of absolute numbers of CD4+ and CD8+ lymphocytes following EFA supplementation. The stimulation indices returned to pre-supplementation values 3 months following cessation of EFA intake. There was no significant change of these indices in the control (no EFA supplementation) advanced tumour group tested. This study suggests that EFA supplementation in patients with colorectal cancer selectively reduces circulating PBL, and T cell subset (including suppressor cells) numbers and/or activity. Such effects may have an important outcome in patients with malignant disease.

Topics: Administration, Oral; Aged; Aged, 80 and over; Antigens, CD; Colorectal Neoplasms; Concanavalin A; Fatty Acids, Essential; Female; Flow Cytometry; Humans; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Stimulation, Chemical; T-Lymphocyte Subsets

Combinations of cellular immune-based therapies with chemotherapy and other antitumour agents may be of significant clinical benefit in the treatment of many forms of cancer. Gamma delta (gammadelta) T cells are of particular interest for use in such combined therapies due to their potent antitumour cytotoxicity and relative ease of generation in vitro. Here, we demonstrate high levels of cytotoxicity against solid tumour-derived cell lines with combination treatment utilizing Vgamma9Vdelta2 T cells, chemotherapeutic agents and the bisphosphonate, zoledronate. Pre-treatment with low concentrations of chemotherapeutic agents or zoledronate sensitized tumour cells to rapid killing by Vgamma9Vdelta2 T cells with levels of cytotoxicity approaching 90%. In addition, zoledronate enhanced the chemotherapy-induced sensitization of tumour cells to Vgamma9Vdelta2 T cell cytotoxicity resulting in almost 100% lysis of tumour targets in some cases. Vgamma9Vdelta2 T cell cytotoxicity was mediated by perforin following TCR-dependent and isoprenoid-mediated recognition of tumour cells. Production of IFN-gamma by Vgamma9Vdelta2 T cells was also induced after exposure to sensitized targets. We conclude that administration of Vgamma9Vdelta2 T cells at suitable intervals after chemotherapy and zoledronate may substantially increase antitumour activities in a range of malignancies.

Topics: Adenocarcinoma; Antineoplastic Agents; Apoptosis; Breast Neoplasms; Burkitt Lymphoma; Carcinoma; Cell Line, Tumor; Cisplatin; Colorectal Neoplasms; Concanavalin A; Cytotoxicity, Immunologic; Diphosphonates; Doxorubicin; Drug Screening Assays, Antitumor; Drug Synergism; Etoposide; Female; Genes, T-Cell Receptor delta; Genes, T-Cell Receptor gamma; Humans; Imidazoles; Interferon-gamma; Lovastatin; Lung Neoplasms; Male; Membrane Glycoproteins; Neoplasms; Perforin; Pore Forming Cytotoxic Proteins; Prostatic Neoplasms; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocyte Subsets; Urinary Bladder Neoplasms; Vincristine; Zoledronic Acid

Clusterin is an enigmatic protein altered in tumors of colorectal cancer patients. Because there is no information available about serum clusterin regarding this pathology, we applied proteomic techniques to analyze its isoforms in donors and patients. First we separated serum proteins through concanavalin A, obtaining a fraction with non- and O-glycosylated proteins (FI) and a second fraction enriched in N-glycoproteins (FII) wherein clusterin was supposed to elute on the basis of its glycosylation. Surprisingly analysis of the FI fraction revealed the existence of an unexpected and aberrantly N-glycosylated clusterin that was overexpressed in patients and comprised at least five isoforms with different isoelectric points. On the other hand, two-dimensional electrophoretic analysis of the clusterin eluted in FII detected one isoform that was increased and 15 isoforms that were decreased or absent in serum of patients. Finally immunoquantification by slot blot showed that in total serum and in FI the clusterin levels were significantly increased in patients, whereas in FII there was no significant variation. Therefore, serum clusterin and some of its isoforms could have a potential value as colorectal tumor markers and are interesting subjects for biomarker studies.

Topics: Biomarkers, Tumor; Blood Proteins; Chromatography, Affinity; Clusterin; Colorectal Neoplasms; Concanavalin A; Electrophoresis, Gel, Two-Dimensional; Female; Glycosylation; Humans; Male; Middle Aged; Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; Protein Isoforms; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Expression of decay-accelerating factor (DAF, CD55), a complement-regulatory glycoprotein, is enhanced in colorectal-cancer (CC) cells and colonic epithelium in ulcerative colitis (UC), and stools from these patients contain increased amounts of DAF. Carbohydrate chains of glycoproteins are often altered during malignant transformation or inflammation. In this study, we investigated whether DAF molecules in patients with CC and those with UC differ with respect to oligosaccharide side chains. We analyzed DAF in stools and homogenates of colonic-tissue specimens obtained from patients with CC or UC using solid-phase enzyme-linked assay and Western blotting for reactivity with the lectins Ulex europaeus agglutinin I (UEA-I), wheat-germ agglutinin, peanut agglutinin, and concanavalin A. UEA-I bound to DAF in stools from patients with UC but not in that from the stools of CC patients, as demonstrated on the solid-phase enzyme-linked assay (P <.05, Mann-Whitney U test) and Western blotting. Binding of UEA-I was specifically inhibited by the addition of fucose. The difference in UEA-I reactivity with DAF was observed also in colonic-tissue homogenates from patients with UC and those with CC. DAF expressed in the mucosa and excreted into the stools of UC patients is different from that expressed in CC with regard to UEA-I reactivity. Future studies should be directed toward determining whether a qualitatively unique isoform of DAF is present, of which sugar chains are specific to CC in UC patients.

Topics: Adenocarcinoma; Adult; Aged; CD55 Antigens; Colitis, Ulcerative; Colon; Colonic Neoplasms; Colorectal Neoplasms; Concanavalin A; Feces; Female; Humans; Intestinal Mucosa; Kinetics; Lectins; Male; Middle Aged; Neoplasm Staging; Plant Lectins; Rectal Neoplasms

Tumor infiltrating lymphocytes (TIL) in colorectal cancer are a manifestation of local, cell mediated immune response to the malignant tumor. Tumor progression is due to impairment of the host ability to control tumor growth. Several studies suggested possible causes for such impairment, however, the precise factor(s) underlying such malfunction is uncertain.. To compare the possible effects of colorectal cancer (CRC) and normal colonic mucosa extracts on lectin induced blastogenesis of the same patients' peripheral blood lymphocytes (PBL) proliferation.. CRC and normal mucosa extracts were obtained from 3 patients undergoing curative surgery for colon adenocarcinoma. Proliferation assays used PBL from the CRC patients, incubated with Concanavalin A (ConA) and Phytohemoglobin (PHA) in the presence of CRC or normal mucosa extract and in medium alone. Proliferation was measured by H3 Thymidine incorporation following 48 hours on incubation.. Exposure of ConA induced PBL proliferation assay to CRC extract yielded a 98.7% inhibition measured by counts/minute (cpm) of incorporated H3 Thymidine compared to normal colonic mucosa extract (1,214 +/- 594 cpm vs. 95,335 +/- 6,997 cpm respectively, p = 0.0018). PHA stimulated proliferation exposed to CRC extract showed a 99.7% decrease in blastogenic activity compared to normal mucosa extract (362 +/- 175 cpm vs. 62,375 +/- 16,591 cpm respectively, p = 0.0234).. These preliminary results suggest that CRC extract contain factor(s) capable of profoundly inhibiting lectin induced proliferation of PBL. Shedding of suppressor substances may be one of the possible mechanisms by which the tumor evades the effector arm of the cell-mediated immune response. Characterization of such factors may aid in intratumor, local and systemic cellular immune response reconstitution.

Topics: Aged; Colorectal Neoplasms; Concanavalin A; Humans; Lectins; Lymphocyte Activation; Male; Middle Aged; Neoplasm Staging; Phytohemagglutinins

A method to improve the reactivity to specific lectins of N-glycoproteins isolated from human colorectal mucosa and from adenocarcinoma biopsies was developed using a combination of techniques. Total protein extracts were subjected to affinity chromatography, using the immobilised lectin Concanavalin A coupled to Sepharose, by fast performance liquid chromatography (FPLC). N-glycoprotein enriched fractions were resolved by SDS-PAGE, transferred to PVDF membranes and incubated with various lectins. Digoxigenin-conjugated SNA I and MAA lectins were used to detect sialic acid residues. Biotin-conjugated UEA I lectin was used to detect L-fucose residues. By this method, lectin-binding N-glycoproteins were found in a broad relative molecular mass (Mr) range (from 47 to 205 kDa). No tissue-specific N-glycoproteins were observed when human colorectal mucosa and adenocarcinoma samples were compared.

Topics: Adenocarcinoma; Biomarkers, Tumor; Biopsy; Chromatography, Affinity; Chromatography, High Pressure Liquid; Colorectal Neoplasms; Concanavalin A; Electrophoresis, Polyacrylamide Gel; Fucose; Glycoproteins; Humans; Intestinal Mucosa; Lectins; Membranes, Artificial; Molecular Weight; N-Acetylneuraminic Acid; Organ Specificity

Tumor development or growth is accompanied by impaired immune responses, such as a poor proliferative response or down-regulated cytolytic T lymphocyte activity. Although recent reports have suggested that modification of the signal-transducing molecule is responsible for impaired immune responses in tumor-bearing hosts, the causes of defective immune function are not yet completely understood. Furthermore, the clinical significance of the findings is not yet clear. In this study, we investigated the alteration of several signal-transducing molecules in peripheral blood T lymphocytes (T-PBL) as well as in tumor-infiltrating lymphocytes (TIL) from human colorectal carcinoma patients and their relationship with the impaired host immune responses. A greater reduction in CD3zeta chain level was observed in TIL than in T-PBL from tumor-bearing hosts. CD3zeta chain reduction in T-PBL correlated with the clinicopathological stage of a tumor, especially with the status of lymph node metastasis. The levels of p56lck and p59fyn protein tyrosine kinase in T-PBL were also compared between tumor-bearing hosts and normal healthy volunteers. In T-PBL from tumor-bearing hosts, expression of protein tyrosine kinase p59fyn was significantly lower than that of p56lck. However, the level of CD3zeta chain expression did not correlate with T lymphocyte functions such as T lymphocyte proliferative response or allogeneic target cell lysis.

Topics: Carcinoma; CD3 Complex; Colorectal Neoplasms; Concanavalin A; Humans; Lymphocyte Activation; Lymphocyte Specific Protein Tyrosine Kinase p56(lck); Lymphocytes, Tumor-Infiltrating; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-fyn; Signal Transduction; T-Lymphocytes

Compared with normal colonic mucosa, lectin receptor expression is increased in hyperplastic and neoplastic tissues; some lectins have been shown to influence human colonic epithelial cell proliferation. The aim was to assess further the influence of five lectins (Phaseolus vulgaris (PNA), Griffonia simplicifolia (GSA), concanavalin A (Con A), wheat germ (WGA), and peanut (PHA-L) agglutinins) on cellular growth in three human colorectal cancer cell lines (LoVo, HCT-15 and SW837).. Cells were cultured in four lectin concentrations (0.1, 1.0, 10, and 100 micrograms/ml) and growth assessed at days 2, 3, 5, and 7. The experiments were performed in media supplemented with either 1% or 10% fetal calf serum (FCS). Growth was assessed using the MTT (3-(4,5)-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) colorimetric assay.. Growth in each cell line was greatly affected by at least two of the lectins tested. There was some variation in the effect of a given lectin on different cell lines. Lectin effects showed a dose-response and the greatest effects generally resulted from the highest concentrations at the longest culture time. WGA and Con A induced large effects in all cell lines; the effects of Con A were partly blocked by the higher concentration of FCS. PNA had modest and uniform stimulatory effects overall. The effects of GSA and PHA-L varied between cell lines.. The lectins studied all have the potential to affect colonic cancer growth in vitro. Many dietary lectins are resistant to digestion and may have important effects in vitro but the definition of their role in human colonic cancer biology must take into account the variability in lectin response.

Topics: Cell Division; Cell Line; Colorectal Neoplasms; Concanavalin A; Dose-Response Relationship, Drug; Humans; Lectins; Peanut Agglutinin; Phytohemagglutinins; Plant Lectins; Time Factors; Wheat Germ Agglutinins

We studied several blood coagulation parameters and tumor tissue procoagulant activity (PCA) in nude mice bearing human colorectal carcinomas (HCC). In a control group of 51 tumor-free nude mice, platelet number was 1.2 +/- 0.03 x 10(6)/microliters, thrombotest activity 90% +/- 2.6 and fibrinogen 172 +/- 11 mg/dl. The same parameters were studied in nude mice (n = 71) bearing 7 different HCC lines subcutaneously (s.c.). The results did not significantly differ from those in control mice but there was broad variability among groups of mice injected with different HCC lines, ranging from 0.36 to 2.55 x 10(6)/microliters for platelets, from 100 to 28% for thrombotest activity and from 42 to 460 mg/dl for fibrinogen. The results were significantly (p less than 0.05) different from those in the tumor-free group when each group of HCC-bearing animals was analyzed individually. A malignant HCC line that grew in the liver of nude mice (n = 24) significantly (p less than 0.001) reduced thrombotest activity (58% +/- 5.9). The PCA of tissue extracts from tumors grown s.c. in nude mice was assayed. All the HCC xenografts expressed PCA which differed significantly for the various tumor lines (from 25.5 +/- 1.9 to 2.8 +/- 0.6 unit/mg in tumor tissue). Cancer procoagulant (CP), a cysteine proteinase with a direct factor-X-activating effect, was present in different amounts (84.7 +/- 4.3 to 59.5 +/- 9.0%) in the tumors. Our results indicates that the nude mouse is a suitable model for evaluating the hemostatic changes induced by human tumors and may represent a tool for investigating the underlying biochemical mechanisms.

Topics: Animals; Blood Coagulation; Carcinoma; Colorectal Neoplasms; Concanavalin A; Factor VII; Fibrinogen; Humans; Iodoacetamide; Mercuric Chloride; Mice; Mice, Nude; Neoplasm Transplantation; Platelet Count; Tumor Cells, Cultured