metallothionein and Adenocarcinoma

The study aimed at determining levels of metallothionein (MT) and Ki-67 antigen expression in adenocarcinomas of large intestine and examining relation of the expression levels with various clinical and pathological variables. The studies were performed on 81 cases of large intestine adenocarcinoma. Using immunocytochemistry, expressions of MT (positive reaction in 73 cases) and of Ki-67 (positive reaction in 79 cases) antigen were examined and the obtained results were compared with, i.a., grade (G) of the tumour and depth to which intestinal wall was infiltrated by individual tumours. Patient survival analysis was also performed, as correlated to expression levels of the two antigens. The obtained results permitted to disclose that the lower was grade of histological differentiation (G2, G3), the more pronounced was expression of MT and Ki-67. Also, the deeper was neoplastic infiltration of intestinal wall, the more pronounced was MT and Ki-67 expression. Despite the relatively strong correlation between MT expression and Ki-67 expression (r=0.536; p<0.05), only Ki-67 antigen expression in large intestine adenocarcinomas was inversely correlated to survival of the patients. Ki-67 proved to be a better prognostic marker, as compared to MT, in large intestine adenocarcinomas.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Colonic Neoplasms; Female; Humans; Immunohistochemistry; Ki-67 Antigen; Male; Metallothionein; Middle Aged; Prognosis; Survival Analysis

Colorectal malignancies with high microsatellite instability (MSI-H), either hereditary (Lynch syndrome) or sporadic, demonstrate better prognosis and altered response to 5FU chemotherapy. It is now recommended to perform MSI testing for all new cases of colorectal cancer regardless of being categorized as hereditary or sporadic. For MSI detection, immunohistochemistry or PCR-based protocols using a cohort of various sets of STR markers are recommended. Here we aimed to evaluate a simplified protocol using just a single STR marker, MT1XT20 mononucleotide repeat, for detection of MSI in Lynch syndrome patients. A Promega five-marker MSI testing panel and immunohistochemistry (IHC) were used as the gold standard in conjunction with MT1XT20.. Colorectal patients with a positive history of familial cancers were selected by evaluating medical records. Based on Amsterdam II criteria for Lynch syndrome 20 families were short listed. DNA was extracted from formalin fixed paraffin embedded tumour and adjacent normal tissues resected from the index case in each family. Extracted DNA was subjected to MT1XT20 mononucleotide marker analysis and assessment with a commercially available five marker MSI testing kit (Promega, USA). IHC also was performed on tissue sections and the results were compared with PCR based data.. Eight (40%), seven (35%) and five (25%) cases were MSI positive using with the Promega kit, IHC and MT1XT20, respectively. Among the markers included in Promega kit, BAT26 marker showed instability in all 8 samples. NR24 and NR21 markers showed instability in 7 (87.5%), and BAT25 and MONO 27 in 6 (75%) and 5 (62.5%).. Although MT1XT20 was earlier reported as a valid standalone marker for MSI testing in CRC patients, we could not verify this in our Iranian patients. Instead BAT26 among the markers included in Promega MSI testing kit showed instability in all 8 MSI-H CRC samples. Therefore, it seems BAT26 could act well as a single marker for MSI testing in Iranian CRC patients.

Topics: Adenocarcinoma; Adult; Biomarkers, Tumor; Colorectal Neoplasms, Hereditary Nonpolyposis; Female; Follow-Up Studies; Humans; Male; Metallothionein; Microsatellite Instability; Microsatellite Repeats; Middle Aged; Neoplasm Staging; Persia; Prognosis; Young Adult

Inorganic arsenic is a human lung carcinogen. We studied the ability of chronic inorganic arsenic (2 μM; as sodium arsenite) exposure to induce a cancer phenotype in the immortalized, non-tumorigenic human lung peripheral epithelial cell line, HPL-1D. After 38 weeks of continuous arsenic exposure, secreted matrix metalloproteinase-2 (MMP2) activity increased to over 200% of control, levels linked to arsenic-induced cancer phenotypes in other cell lines. The invasive capacity of these chronic arsenic-treated lung epithelial (CATLE) cells increased to 320% of control and colony formation increased to 280% of control. CATLE cells showed enhanced proliferation in serum-free media indicative of autonomous growth. Compared to control cells, CATLE cells showed reduced protein expression of the tumor suppressor gene PTEN (decreased to 26% of control) and the putative tumor suppressor gene SLC38A3 (14% of control). Morphological evidence of epithelial-to-mesenchymal transition (EMT) occurred in CATLE cells together with appropriate changes in expression of the EMT markers vimentin (VIM; increased to 300% of control) and e-cadherin (CDH1; decreased to 16% of control). EMT is common in carcinogenic transformation of epithelial cells. CATLE cells showed increased KRAS (291%), ERK1/2 (274%), phosphorylated ERK (p-ERK; 152%), and phosphorylated AKT1 (p-AKT1; 170%) protein expression. Increased transcript expression of metallothioneins, MT1A and MT2A and the stress response genes HMOX1 (690%) and HIF1A (247%) occurred in CATLE cells possibly in adaptation to chronic arsenic exposure. Thus, arsenic induced multiple cancer cell characteristics in human peripheral lung epithelial cells. This model may be useful to assess mechanisms of arsenic-induced lung cancer.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Arsenic; Carcinogens; Cell Line, Tumor; Cells, Cultured; Epithelial Cells; Epithelial-Mesenchymal Transition; Humans; Lung Neoplasms; Metallothionein; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Phenotype; Proto-Oncogene Proteins; Proto-Oncogene Proteins p21(ras); ras Proteins

Although aberrant DNA methylation has been implicated in the pathophysiology of lung cancer, the role of methylation in multidrug resistance (MDR) of lung cancer has remained unclear. To investigate whether certain distinct DNA methylation pattern is associated with acquired MDR of lung adenocarcinoma, methylated-DNA immunoprecipitation-chromatin immunoprecipitation (MeDIP-ChIP) was utilised to compare the genome-wide promoter methylation of the human lung adenocarcinoma MDR A549/cisplatin (A549/DDP) cells with its progenitor A549 cells. The comparison identified 3617 genes with differentially methylated promoter, of which 1581 were hypermethylated and 2036 were hypomethylated. Then, bisulphite sequencing polymerase chain reaction (PCR) (BSP) and quantitative reverse transcription (RT)-PCR (Q-PCR) were used to validate the promoter methylation of five candidate genes and to determine whether the expression of genes was associated with the promoter methylation. BSP confirmed that the promoter methylation incidence of the hypermethylated genes, G protein-coupled receptor 56 isoform 3 (GPR56), metallothionein 1G (MT1G), and RAS association domain family gene 1 (RASSF1), was significantly higher in A549/DDP cells compared with A549 cells (p<0.001, p=0.0099, and p=0.0165), whereas no significant difference was found in that of the other two genes, CCNL2 and BAD (p=0.0594 and p=0.5546). Additionally, Q-PCR showed that the mRNA expression of the three hypermethylated genes was significantly lower in A549/DDP cells compared with A549 cells (all p<0.001). In conclusion, this study reported for the first time that a distinct promoter methylation pattern is associated with MDR of lung adenocarcinoma A549/DDP cells and suggested that GPR56, MT1G, and RASSF1 might be the potential methylation markers associated with acquired MDR of lung adenocarcinoma.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Cell Line, Tumor; DNA Methylation; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Genome-Wide Association Study; Humans; Lung Neoplasms; Metallothionein; Promoter Regions, Genetic; Receptors, G-Protein-Coupled; Tumor Suppressor Proteins

Metallothioneins (MTs) are a group of metal binding proteins thought to play a role in the detoxification of heavy metals. Here we showed by microarray and validation analyses that MT1h, a member of MT, is down-regulated in many human malignancies. Low expression of MT1h was associated with poor clinical outcomes in both prostate and liver cancer. We found that the promoter region of MT1h was hypermethylated in cancer and that demethylation of the MT1h promoter reversed the suppression of MT1h expression. Forced expression of MT1h induced cell growth arrest, suppressed colony formation, retarded migration, and reduced invasion. SCID mice with tumour xenografts with inducible MT1h expression had lower tumour volumes as well as fewer metastases and deaths than uninduced controls. MT1h was found to interact with euchromatin histone methyltransferase 1 (EHMT1) and enhanced its methyltransferase activity on histone 3. Knocking down of EHMT1 or a mutation in MT1h that abrogates its interaction with EHMT1 abrogated MT1h tumour suppressor activity. This demonstrates tumour suppressor activity in a heavy metal binding protein that is dependent on activation of histone methylation.

Topics: Adenocarcinoma; Animals; Cell Line, Transformed; Cell Line, Tumor; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Histone-Lysine N-Methyltransferase; Humans; Liver Neoplasms; Male; Metallothionein; Mice; Mice, SCID; Microarray Analysis; Pennsylvania; Prostatic Neoplasms; Survival Rate; Tumor Suppressor Proteins

Metallothioneins are a family of small, cysteine-rich proteins with many functions. Immunohistochemical evaluation of all metallothionein 1 + 2 isoforms in colorectal tumors has demonstrated an important down-regulation compared with normal tissue, although its prognostic significance is unclear. Moreover, the contribution of individual isoforms to overall metallothionein down-regulation is not known. To address these important issues, we analyzed the messenger RNA expression levels of all functional metallothionein 1 + 2 isoforms by quantitative reverse transcription polymerase chain reaction in 22 pairs of normal and tumor-microdissected epithelia and correlated these to the overall immunohistochemical protein expression. Our results showed that 5 isoforms (MT1G, 1E, 1F, 1H, and 1M) were lost during the transition from normal mucosa to tumor, whereas MT1X and MT2A were less down-regulated, and their expression was correlated with overall protein positivity. Second, we showed that MT1G hypermethylation occurred in cell lines and in 29% of tumor samples, whereas histone deacetylase inhibitors are able to induce most isoforms. Furthermore, we analyzed by immunohistochemistry 107 normal mucosae, 25 adenomas, 81 carcinomas, and 19 lymph node metastases to evaluate metallothionein expression during different stages of cancer development and to assess its relationship to patient survival. A lower immunohistochemical expression was associated with poorer survival, although it was not an independent predictor. Overall, this study identifies for the first time the relevant metallothionein isoforms for colorectal cancer progression, supports the concept that their loss is associated with worse prognosis, and suggests 2 mechanisms for epigenetic repression of metallothionein expression in colorectal tumors.

Topics: Adenocarcinoma; Adenoma; Aged; Aged, 80 and over; Argentina; Biomarkers, Tumor; Colorectal Neoplasms; Disease Progression; DNA Methylation; Female; Gene Expression Regulation, Neoplastic; Gene Silencing; Humans; Male; Metallothionein; Middle Aged; Protein Isoforms; RNA, Messenger; Survival Rate

Current diagnostic techniques are inefficient in distinguishing latent and low-risk forms of prostate cancer from high-risk forms. The present study is focused on determination of putative tumor markers of aggressive high-grade forms of prostate cancer. Potential markers were determined in blood sera of 133 patients (82 cases and 51 controls) and in cell lines (Gleason score 9-derived 22Rv1 and normal tissue derived PNT1A) on mRNA and protein levels. Alpha-methylacyl-CoA racemase (AMACR), metallothionein classes 1A and 2A (MT1A and MT2A) were determined and compared to prostate specific antigen (PSA) levels. On mRNA level, significantly increased expression of MT2A (2.4-fold), PSA (2.6-fold) and AMACR (8.4-fold) and insignificantly (1.9-fold) elevated MT1A in 22Rv1 compared to non-tumor PNT1A were determined. On protein level, significant enhancement of free PSA and total PSA in tumor cell line was evident. AMACR protein was 1.5-fold elevated in tumor line (below the level of significance). Contrary to mRNA, significantly (p = 0.01) reduced level of MT protein in tumor lines was determined. In the case of serum level, significantly enhanced MT level (4.5-fold) in patients' sera was found. No significant changes were observed in the case of AMACR. These findings indicate possible alternative role of MT to PSA prostate cancer marker. In addition, level of AMACR is distinctly higher in the Gleason score 9 in serum of patients and MT shows a descending trend in relation to Gleason score.

Topics: Adenocarcinoma; Adult; Aged; Biomarkers, Tumor; Blotting, Western; Case-Control Studies; Humans; Male; Metallothionein; Middle Aged; Neoplasm Grading; Prognosis; Prostate-Specific Antigen; Prostatic Neoplasms; Racemases and Epimerases; Real-Time Polymerase Chain Reaction; RNA, Messenger

The study aimed at the evaluation of the effects of radiotherapy on expression of metallothionein (MT) isoforms, both in the form of quantitative alterations in mRNA, and differences in expression of MTI/II proteins in rectal tumours.. Material for the study originated from 21 patients with rectal cancer at stage II or III. Material for immunohistochemical studies [MTI/II, Minichromosome Maintenance Protein 3 (MCM3), Ki-67] and real-time polymerase chain reaction (PCR) (mRNA of MT1F, MT1X and MT2A) was sampled twice: during rectoscopic examination before the start of the preoperative radiotherapy (samples A) and from the post operative specimen, following radiotherapy (samples B).. The level of mRNA expression for each of the studied MT isoforms was higher in cancer cells subjected to irradiation. The most extensive differences were observed for the MT2A isoforms (p=0.09). No differences were disclosed between samples A and B in expression of MT I/II protein. The material sampled after radiotherapy manifested a tendency for reduced proliferative activity of the tumour cells: the decrease of MCM3 expression was significant (p=0.022), while in the case of Ki-67, the difference approached statistical significance (p=0.096).. Application of radiotherapy to rectal adenocarcinoma cells is followed by an increase in MT mRNA expression level, affecting first of all the MT2A isoform. However, we failed to note an increased expression of MTI/II protein coded by the gene. Moreover, application of radiotherapy was followed by a decrease in expression of MCM3 protein. Our results cannot clearly confirm induction of MT after irradiation of human adenocarcinoma cells. The role of MT in radioprotection remains ambiguous.

Topics: Adenocarcinoma; Aged; Aged, 80 and over; Cell Cycle Proteins; DNA-Binding Proteins; Female; Gene Expression; Gene Expression Profiling; Humans; Immunohistochemistry; Ki-67 Antigen; Male; Metallothionein; Middle Aged; Minichromosome Maintenance Complex Component 3; Nuclear Proteins; Protein Isoforms; Real-Time Polymerase Chain Reaction; Rectal Neoplasms; RNA, Messenger

Gastrointestinal stromal tumors (GISTs) are mesenchymal tumors that account for about 2% of gastric tumors. Metallothioneins (MTs) are multifunctional proteins associated with carcinogenesis and known to be coded by 10 functional MT genes. This study evaluated MT mRNA and protein expression in GISTs and compared the expression levels with gastric carcinomas. An immunohistochemical study of MT protein expression was performed in 15 GISTs (specifically located in the stomach) and 38 early stage gastric carcinomas. The percentage of cells stained and intensity of staining were determined. MT-2A mRNA expression was investigated in 6 GISTs and 6 early stage gastric carcinoma patients. All GISTs displayed positive nuclear immunostaining, with most GISTs having predominantly mildly stained nuclei (93.3%). On the other hand, 37 out of 38 gastric carcinoma cases were positively stained for nuclear MT with 24 cases (63.2%) exhibiting predominantly mild nuclear staining, 7 cases (18.4%) moderate nuclear staining, and 6 cases (15.8%) strong nuclear staining. Nuclear MT expression was found to be significantly lower in GIST samples when compared with gastric carcinoma tissues based on the percentage stained and immunoreactive score. We then established that the MT-2A gene transcript was the most abundant MT isoform in MKN28 gastric cancer cells and analyzed its expression in GIST and gastric carcinoma tissues. We found that GISTs had significantly lower MT-2A mRNA levels than gastric carcinoma tissues. Lower MT-2A gene expression and nuclear MT protein expression in GISTs when compared with gastric carcinomas may reflect their different underlying biology and divergent histogenesis.

Topics: Adenocarcinoma; Biomarkers, Tumor; Cell Line, Tumor; Cell Nucleus; Gastrointestinal Stromal Tumors; Humans; Metallothionein; Neoplasm Staging; RNA, Messenger; Stomach Neoplasms

The disturbance of zinc homeostasis featured with a significant decrease of cellular zinc level was well documented to associate with the development and progression of human prostate malignancy. We have previously reported that zinc treatment induces prostate malignant cell apoptosis through mitochondrial pathway. Metallothionein (MT) is a major receptor/donor of zinc in the cells. However, the studies on the expression of MT in association with the prostate pathological and malignant status are very limited, and the zinc regulation of MT isoform expression in prostate cells remains elusive. The goals of this study were to define the expression of endogenous MTs, the isoforms of MT 1, 2, 3 at both messenger ribonucleic acid (mRNA) and protein levels; and to investigate the zinc effect on MT expression in normal prostate, benign prostatic hyperplasia (BPH) and malignant PC-3 cells, and in relevant human tissues. Cellular MT proteins were detected by immunohistochemistry, fluorescence staining and Western blot analysis; reverse transcription polymerase chain reaction (RT-PCR) was used to determine the MT isoform-specific mRNAs.. Our results demonstrated a significant suppression of endogenous levels of MT1/2 in malignant PC-3 cells (95% reduction compared to the normal prostate cells) and in human adenocarcinoma tissues (73% MT1/2 negative). A moderate reduction of MT1/2 expression was observed in BPH. Zinc treatment remarkably induced MT1/2 expression in PC-3 and BPH cells, which was accordant with the restored cellular zinc level. MT 3, as a growth inhibitory factor, was detected and up-regulated by zinc mainly in BPH cells.. This study provided evidence of the association of attenuated MT1/2 with prostate tumor progression, and the zinc induction of MT1/2 expression resulting in cellular zinc restoration. The results suggest the potential of MT1/2 as a candidate biomarker for prostate cancer and the utilization of zinc in prostate cancer prevention and treatment.

Topics: Adenocarcinoma; Blotting, Western; Cell Line, Tumor; Fluorescent Antibody Technique; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Male; Metallothionein; Organ Specificity; Prostate; Prostatic Hyperplasia; Prostatic Neoplasms; RNA, Messenger; Zinc

Perineural invasion (PNI) is the dominant pathway for local invasion in prostate cancer. To date, only few studies have investigated the molecular differences between prostate tumors with PNI and those without it.. To evaluate the involvement of both microRNAs and protein-coding genes in PNI, we determined their genome-wide expression with a custom microRNA microarray and Affymetrix GeneChips in 50 prostate adenocarcinomas with PNI and 7 without it. In situ hybridization (ISH) and immunohistochemistry was used to validate candidate genes.. Unsupervised classification of the 57 adenocarcinomas revealed two clusters of tumors with distinct global microRNA expression. One cluster contained all non-PNI tumors and a subgroup of PNI tumors. Significance analysis of microarray data yielded a list of microRNAs associated with PNI. At a false discovery rate (FDR)<10%, 19 microRNAs were higher expressed in PNI tumors than in non-PNI tumors. The most differently expressed microRNA was miR-224. ISH showed that this microRNA is expressed by perineural cancer cells. The analysis of protein-coding genes identified 34 transcripts that were differently expressed by PNI status (FDR<10%). These transcripts were down-regulated in PNI tumors. Many of those encoded metallothioneins and proteins with mitochondrial localization and involvement in cell metabolism. Consistent with the microarray data, perineural cancer cells tended to have lower metallothionein expression by immunohistochemistry than nonperineural cancer cells.. Although preliminary, our findings suggest that alterations in microRNA expression, mitochondrial function, and cell metabolism occur at the transition from a noninvasive prostate tumor to a tumor with PNI.

Topics: Adenocarcinoma; Cell Differentiation; Cell Lineage; Coxsackie and Adenovirus Receptor-Like Membrane Protein; Gene Expression Regulation, Neoplastic; Humans; Male; Metallothionein; MicroRNAs; Mitochondria; Neoplasm Invasiveness; Neoplasm Proteins; Oligonucleotide Array Sequence Analysis; Peripheral Nerves; Prostatic Neoplasms; Receptors, Virus

Elevated metallothionein (MT) expression in ovarian cancers treated with cisplatin-based schemes represents an unfavorable prognostic index. MT expression is significantly higher in tumor samples obtained after chemotherapy. The present study aimed at examining MT expression in ovarian carcinoma cells sensitive (A2780) or resistant (A2780RCIS) against platinum drug treatment as well as examining effects of exposure to cisplatin on MT expression. Subcellular expression of MT was evaluated also in samples originating from 73 ovarian tumors. Cisplatin-resistant A2780RCIS cells were exposed to increasing cisplatin concentrations, and the subcellular expression of MT was determined by immunocytochemistry. The studies demonstrated that cisplatin-resistant A2780RCIS cells exposed to cisplatin typically manifested a nuclear MT expression. The study demonstrated also that exposure to cisplatin was paralleled by growing MT expression in cell nuclei. The nuclear expression of MT was also found to be specific for ovarian cancers of poor clinical outcome. No relationship could be demonstrated between cytoplasmic expression of MT and clinical variables. Nuclear MT expression is induced by cisplatin and seems to protect DNA in the cells from toxic effects of the drug.

Topics: Adenocarcinoma; Antineoplastic Agents; Cell Line, Tumor; Cell Nucleus; Cell Proliferation; Cisplatin; Drug Resistance, Neoplasm; Drug Screening Assays, Antitumor; Female; Humans; Metallothionein; Ovarian Neoplasms; Prognosis; Survival Rate

alpha-fetoprotein-producing adenocarcinoma of the digestive organs (APAD) is known to show a poor prognosis. To clarify the characteristics of chemoresistance in APAD, three proteins of fluoropyrimidine chemotherapy association [dihydropyrimidine dehydrogenase (DPD), thymidine phosphorylase (TP) and thymidylate synthase (TS)] and one protein of cisplatin association [metallothionein (MT)] were immunohistochemically evaluated. Tissue samples were taken from 12 AFP-positive gastric cancers and 94 AFP-negative gastric cancers. Four AFP-positive cancer xenografts (one colonic, two pancreatic, and one biliary tract) and 17 AFP-negative cancer xenografts were also examined. In gastric cancers, high expression of TP was observed in 30% of AFP-negative tumors but in none of AFP-positive tumors (p=0.03). High expression of MT was found in 30% of AFP-negative tumors but in only one of the AFP-positive tumors. The TP-low and MT-low phenotype was noted in 92% of AFP-positive tumors and in 46% of AFP-negative tumors (p=0.004). None of the AFP-positive cancer xenografts revealed high TP expression and only one showed high MT expression. In the cellular level, TP and MT were scarcely co-expressed with AFP in either gastric cancer or xenograft series, using double immunostaining and serial sectioning techniques. There were no significant differences in the expression of DPD and TS between AFP-positive group and -negative group. However, DPD was frequently co-expressed with AFP in poorly differentiated medullary areas of the AFP-positive gastric cancers. The data presented herein suggest that APAD should be sensitive to cisplatin, but resistant to capecitabine and 5'-deoxyfluorouridine, fluoropyrimidines which are converted to 5-fluorouracil by TP. S-1, a fluoropyrimidine containing a strong DPD inhibitor, may be effective for AFP-positive gastric cancers with poorly differentiated medullary growth pattern.

Topics: Adenocarcinoma; alpha-Fetoproteins; Animals; Cell Line, Tumor; Dihydrouracil Dehydrogenase (NADP); Drug Resistance, Neoplasm; Gastrointestinal Neoplasms; Gene Expression Regulation, Neoplastic; Humans; Metallothionein; Mice; Mice, Nude; Neoplasm Transplantation; Thymidine Phosphorylase; Thymidylate Synthase

Zinc is involved in several physiologic processes, including cell growth and proliferation. Although in normal prostate tissue zinc levels are high, there is a marked decrease in prostate cancer. Metallothioneins control the bioavailability of zinc and one isoform, MT1G, was reported down-regulated in prostate cancer. Here, we investigated whether promoter methylation might cause MT1G silencing in prostate cancer.. The MT1G promoter was assessed by quantitative methylation-specific PCR on prospectively collected tissue samples from 121 patients with prostate cancer, 39 paired high-grade prostatic intraepithelial neoplasias (HGPIN), 29 patients with benign prostatic hyperplasia, 13 normal prostate tissue samples from cystoprostatectomy specimens, and prostate cancer cell lines. The methylation levels were calculated and were correlated with clinical and pathologic variables. Reverse transcription-PCR was done in cell lines to assess MT1G mRNA expression before and after demethylating treatment.. MT1G promoter hypermethylation was found in 29 of 121 prostate cancer, 5 of 39 HGPIN, 3 of 29 benign prostatic hyperplasia, and 0 of 13 normal prostate tissue samples. No significant differences in methylation frequencies or levels were found (P = 0.057, for both). Methylation levels were found to correlate with tumor stage but not with Gleason grade. MT1G hypermethylation was more frequent in prostate cancer that spread beyond the prostate capsule. All prostate cancer cell lines tested showed MT1G promoter methylation, but no differences in expression were apparent after demethylation.. Our findings suggest that MT1G promoter methylation is associated with tumor aggressiveness in prostate cancer and it might be a marker of locally advanced disease.

Topics: Adenocarcinoma; Aged; Analysis of Variance; Biomarkers, Tumor; Cell Line, Tumor; Cell Proliferation; Disease Progression; DNA Methylation; Gene Expression Regulation, Neoplastic; Gene Silencing; Humans; Male; Metallothionein; Middle Aged; Neoplasm Staging; Polymerase Chain Reaction; Promoter Regions, Genetic; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; RNA; Zinc

Endometrium is a specialized organ in which phenomena controlling the level of cell proliferation and apoptosis are marked. The aim of our study was to determine the presence of proteins involved in apoptosis and proliferation: RCAS1, MT and the number of CD56-positive cells and their activity to elucidate their possible role in the development of adenocarcinoma and endometriosis.. MT, RCAS1, CD56-positivity and CD69 expression were assessed in 55 tissue samples by Western blot and immunohistochemistry methods.. We found that endometrium during secretory menstrual cycle phase is characterized by significantly higher RCAS1 and higher MT expression than in proliferative phase. The number of CD56-positive cells and the CD69 antigen expression was significantly increased. Endometrial adenocarcinoma was characterized by significantly increased RCAS1 expression, while MT expression was comparable to the level found in the secretory phase. The number of CD56-positive cells was significantly decreased and their activity was comparable to the level found in the secretory phase. Endometriosis was accompanied by significantly lower RCAS1 and MT expressions, with lower number of CD-56 positive cells and lower expression of CD69 antigen in comparison to the secretory phase.. The ability of endometrium to determine cytotoxic activity (RCAS1 expression changes) and high protection against DNA damage (MT expression) with concomitant changes in the number of immune cells and their activity, observed in normal endometrium during the menstrual cycle phases seems to be fundamental for pathological features of endometrial adenocarcinoma and endometriosis.

Topics: Adenocarcinoma; Adult; Antigens, CD; Antigens, Differentiation, T-Lymphocyte; Antigens, Neoplasm; CD56 Antigen; DNA Damage; Endometrial Neoplasms; Endometriosis; Endometrium; Female; Humans; Immunochemistry; Lectins, C-Type; Menstrual Cycle; Metallothionein; Middle Aged

An accumulation of genetic alterations forming the field of cancerization is an important event for the transformation from normal to cancer cell in multistep carcinogenesis. Histopathologically healthy tumor adjacent tissue might be considered as a cancerization field which is typified by genetic changes required for the development of cancer. Metallothionein (MT) is considered to be a protective and anti-apoptotic protein. The aim of our study was to evaluate the MT expression in head and neck squamous cells carcinoma and breast adenocarcinoma and their histologically healthy adjacent tissue.. We have sampled 29 tissue samples in total derived from head and neck cancers and 29 samples of their clear surgical margins, 33 breast adenocarcinomas and 33 clear surgical margins. Antibody recognizing MT-1 was used for immunohistochemical analysis.. MT expression was revealed in 85,7% of head and neck cancers and 94% of breast adenocarcinomas. It was found in all tumor adjacent tissue. MT expression was statistically significantly higher in tumor adjacent tissue than in cancer tissue in cases with the presence of lymph node metastases in both, breast adenocarcinoma and head and neck squamous cell carcinoma. Generally stroma seems to respond to the presence of cancer by the expression of MT, even in tissues which normally do not express MT.. MT might be a normal or protective reaction of healthy adjacent tissue to the presence of tumor.

Topics: Adenocarcinoma; Adult; Aged; Breast Neoplasms; Carcinoma, Squamous Cell; Female; Head and Neck Neoplasms; Humans; Immunohistochemistry; Lymphatic Metastasis; Male; Metallothionein; Middle Aged; Stromal Cells

Zinc is essential for the structural and functional integrity of cells and plays a pivotal role in the control of gene expression. To identify genes with altered mRNA expression level after zinc depletion, we employed oligonucleotide arrays with approximately 10,000 targets and used the human colon adenocarcinoma epithelial cell line HT-29 as a model. A low intracellular zinc concentration caused alterations in the steady-state mRNA levels of 309 genes at a threshold factor of 2.0. Northern blot analysis and/or real-time RT-PCR confirmed the array results for 12 of 14 selected targets. Genes identified as regulated based on microarray data encode mainly proteins involved in central pathways of intermediary metabolism (79 genes) including protein metabolism (21). We also identified five groups of genes important for basic cellular functions such as signaling (30), cell cycle control and growth (15), vesicular trafficking (15), cell-cell interaction (13), cytoskeleton (10) and transcription control (19). The latter group comprises several zinc finger-containing transcription factors of which the Kruppel-like factor 4 showed the most pronounced changes. Western blot analysis confirmed the increased expression level of this protein in cells grown under low zinc conditions. Our findings in a homogeneous cell population demonstrate that the molecular mechanisms by which cellular functions are altered at a low zinc status, occur via pleiotropic effects on gene expression. In conclusion, the pattern of zinc-affected genes may represent a reference for further studies to define the zinc regulon in mammalian cells.

Topics: Adenocarcinoma; Apoptosis; Blotting, Northern; Cell Division; Chelating Agents; Colonic Neoplasms; DNA-Binding Proteins; Epithelial Cells; Ethylenediamines; Gene Expression; Gene Expression Regulation; Humans; Intestines; Kruppel-Like Factor 4; Kruppel-Like Transcription Factors; Metallothionein; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription Factors; Tumor Cells, Cultured; Zinc

Tumour growth and expansion are the result of proliferative activity and the capacity to eliminate cells by apoptosis and/or necrosis. The present study was aimed at comparing the apoptosis and proliferation intensity in cells of adenocarcinomas of the large intestine with the expression of metallothionein (MT), the grade of the tumour and the depth to which the tumour infiltrated the intestinal wall. The TUNEL technique and immunocytochemical reactions (expression of caspase-3, Ki-67, MT) were used to detect apoptosis. The results demonstrated augmented levels of all the variables examined, positively correlated with grade of malignancy, G, and with the depth of intestinal wall infiltration by the tumour cells. The testing of apoptosis, proliferation and MT expression may prove useful in the appraisal of the growth and progression of primary adenocarcinomas in the large intestine.

Topics: Adenocarcinoma; Apoptosis; Caspase 3; Caspases; Cell Division; Humans; Immunohistochemistry; In Situ Nick-End Labeling; Intestinal Neoplasms; Ki-67 Antigen; Metallothionein; Neoplasm Invasiveness

The present study aimed at analysing the intensity of apoptosis, as related to the expression of pro- and anti-apoptotic cellular markers (cas-3, MT, Ki-67 antigen), and at evaluating their expression in relation to the survival time of patients with colorectal adenocarcinoma. Material for the studies was obtained from 40 patients with primary colorectal adenocarcinomas (G2, T3N0M0), treated at the Lower Silesia Centre of Oncology in Wrocław. Tumour samples were fixed in 4% buffered formalin and embedded in paraffin blocks. In obtained paraffin sections, TUNEL reaction (detection of apoptosis) and immunocytochemical reactions were performed (detection of cas-3, MT and Ki-67 antigen expression). The results disclosed a weak correlation between the intensity of apoptosis and the expression of MT, cas-3 and Ki-67 antigen (r = 0.18; r = 0.33; r = 0.15, respectively) in cells of colorectal adenocarcinomas. The survival time of the patients was shorter, when the apoptosis and expression of Ki-67 antigen were highly pronounced. The time periods of the patients' survival showed no correlation with the expression of cas-3 or MT. The obtained results point to the key role of apoptosis and proliferation processes in the clinical course of colorectal adenocarcinomas.

Topics: Adenocarcinoma; Apoptosis; Caspase 3; Caspases; Colorectal Neoplasms; Humans; Ki-67 Antigen; Metallothionein; Survival Analysis

Metallothionein (MT) is a low molecular weight protein, which participates in differentiation and proliferation of normal and tumour cells. In some malignant tumours (mammary, renal, ovarian cancers), its increased expression is thought to represent an unfavourable prognostic factor. Non-small-cellular lung cancers (mainly squamocellular cancer and adenocarcinoma) are characterised by ill-defined prognosis, which poses problems in the selection of effective post-surgical therapy. The present study aimed at demonstration of the prognostic significance of MT expression in cells of non-small cell lung cancers, attempting to correlate the intensity of MT expression with G grade and with the intensity of proliferation-associated antigen, Ki-67 expression. The studies were performed on archival paraffin blocks with samples of 25 cases of non-small cell lung cancers (5 squamous cell cancers, 20 adenocarcinomas). In paraffin sections of the studied tumours, immunocytochemical reactions were performed, using mouse monoclonal anti-MT and anti-Ki-67 antibodies. The expressions of MT and Ki-67 were demonstrated in all the studied tumours. An analysis of correlation between the expression of MT, Ki-67 antigen and G grade demonstrated a strong positive relation between the latter two parameters (r=0.70; p<0.05). Less pronounced positive correlations were disclosed between MT expression and G grade (r=0.44; p<0.05) and between MT expression and the expression of Ki-67 antigen (r=0.41; p<0.05). In addition, in 15 cases of examined tumours, survival analysis was performed, which disclosed a shorter survival in patients with high MT expression. The obtained results confirmed the relationship between MT expression and Ki-67 antigen expression, indicating an involvement of the proteins in processes of tumour cell proliferation. In turn, the shorter survival of patients with high expression of MT pointed to prognostic significance of the protein in non-small cell lung cancers.

Topics: Adenocarcinoma; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Humans; Ki-67 Antigen; Lung Neoplasms; Metallothionein; Survival Analysis

This study is conducted to detect metallothionein (MT) distribution in the epithelial cells of prostate gland from patients with benign prostatic hypertrophy and adenocarcinoma.. Prostatic tissues from patients with benign prostatic hypertrophy and adenocarcinoma were processed for immunocytochemistry using indirect peroxidase antiperoxidase procedure and primary antibody against MT. The samples were collected over a period of 2-3 years and were processed at Jordan University of Science and Technology, Irbid, Jordan in the year 2002.. All prostatic tissues showed a positive reaction for MT. In benign prostatic hypertrophy, MT was mainly localized in the nuclei of epithelial cells while in the adenocarcinoma; MT was mainly localized in the cytoplasm of the epithelial cells.. Metallothionein expression may be affected by the pathological status of the prostate. In addition, these findings could be used in diagnosing and evaluating the prognosis of different pathological conditions of the prostate.

Topics: Adenocarcinoma; Cell Nucleus; Cytosol; Epithelial Cells; Humans; Immunohistochemistry; Male; Metallothionein; Prostate; Prostatic Hyperplasia; Prostatic Neoplasms; Zinc

Metallothioneins (MT) represent low molecular weight proteins that are supposed to fulfil several functions. They participate in the cell cycle, protect cells from oxidative stress, control levels of heavy metals and participate in multidrug resistance processes, particularly in cases of alkylating drugs. The present study aimed at evaluation of proliferation intensity (Ki67, PCNA) in ovarian cancers treated using cisplatin and paclitaxel, as related to expression of MT. The experiments were performed on samples originating from 10 patients operated on due to ovarian cancer. The material originated from the first operations or second-look operations. All the patients were treated with cisplatin and paclitaxel. Immunocytochemical reactions using antibodies to MT, Ki67 and PCNA were performed in paraffin sections originating from the cases studied. Statistical analysis was performed using Statistica software. The studies demonstrated no relation between expression of MT on the one hand and intensity of proliferation before or after chemotherapy on the other hand (gamma correlation, p > 0.05). The results indicate that expression of MT is not related to resistance to treatment using cisplatin and paclitaxel.

Topics: Adenocarcinoma; Antineoplastic Combined Chemotherapy Protocols; Biomarkers, Tumor; Cell Count; Cell Division; Cisplatin; Combined Modality Therapy; Female; Humans; Immunohistochemistry; Ki-67 Antigen; Metallothionein; Ovarian Neoplasms; Paclitaxel; Proliferating Cell Nuclear Antigen

The potential of the metal-binding protein, metallothionein, in assessing the progression of normal oesophagus through Barrett's to adenocarcinoma was investigated. Metallothionein was quantitatively determined in resected tissues from patients undergoing oesophagectomy for high grade dysplasia/adenocarcinoma and in biopsies from patients with Barrett's syndrome. In 10 cancer patients, metallothionein concentrations in adenocarcinoma were not significantly different from normal oesophagus, although six had elevated metallothionein concentrations in the metaplastic tissue bordering the adenocarcinoma. In 17 out of 20 non-cancer patients with Barrett's epithelium, metallothionein was significantly increased by 108% (P<0.004). There was no association between the metallothionein levels in Barrett's epithelium and the presence of inflammatory cells, metaplasia or dysplasia. Metallothionein is a marker of progression from normal to Barrett's epithelium but is not increased in oesophageal adenocarcinoma.

Topics: Adenocarcinoma; Adult; Aged; Barrett Esophagus; Biomarkers; Biomarkers, Tumor; Biopsy; Disease Progression; Esophageal Neoplasms; Esophagitis; Esophagus; Female; Gastroesophageal Reflux; Humans; Hyperplasia; Male; Metallothionein; Metaplasia; Middle Aged; Neoplasm Metastasis; Neoplasm Proteins; Precancerous Conditions

Some recent evidence has suggested a protective role of zinc against cancer. The mechanism by which zinc exerts this action has not been defined and, in particular, it has not been clarified whether zinc may directly act on cancer cells and the molecular mechanisms involved in this effect. In this study, we examined the in vitro effect of zinc on the apoptosis of mouse TS/A mammary adenocarcinoma cells, studying the zinc-dependent modulation of the intracellular levels of reactive oxygen species (ROS) and of p53 and Fas/Fas ligand pathways. We showed that zinc concentrations ranging from 33.7 to 75 muM Zn(2+) induced apoptosis in mammary cancer cells. The apoptosis was associated with an increased production of intracellular ROS, and of p53 and Fas/Fas ligand mRNA and protein. Zn(2+) induced a faint metallothionein response in TS/A cells in comparison with mouse lymphocytes. The treatment of tumor cells with the antioxidant N-acetylcysteine was able to prevent Zn(2+)-induced apoptosis, as well as the increase of p53 and Fas ligand protein induced by zinc. The data demonstrate that zinc exerts a direct action on mammary cancer cells inducing ROS-mediated apoptosis and that the effect may be mediated by the ROS-dependent induction of p53 and Fas/Fas ligand.

Topics: Acetylcysteine; Adenocarcinoma; Animals; Apoptosis; Bacterial Proteins; Blotting, Western; Cell Division; Cell Survival; DNA Primers; Fas Ligand Protein; Free Radical Scavengers; Gene Expression; Humans; In Situ Nick-End Labeling; In Vitro Techniques; Mammary Neoplasms, Experimental; Membrane Glycoproteins; Metallothionein; Mice; Mice, Inbred BALB C; Reactive Oxygen Species; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Superoxides; Transcription Factors; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Zinc

Over-expression of cellular metallothionein occurs frequently in human tumours but the underlying mechanism remains unknown. The aim of this study was to assess metallothionein expression in cases of lung carcinoma and to correlate it with histopathological parameters.. Tumour tissue samples from 89 patients with lung carcinoma were immunostained by the streptavidin-biotin-peroxidase technique, using a monoclonal antibody against both metallothionein-1 and -2 isoforms. Positive matallothionein immunostaining was prominent in 44 out of 89 (49%) and negative in 45 out of 89 (51%) cases of lung carcinoma examined. Metallothionein positivity was prominent in 32 out of 43 (74%) cases of squamous cell lung carcinoma, and in 12 out of 35 (34%) cases of adenocarcinoma, while it was negative in all 11 cases of small-cell lung carcinoma examined, presenting a statistically significant difference between the different histological types. The intensity of metallothionein staining revealed a statistically significant difference between the squamous cell and adenocarcinoma cases examined. The pattern and extent of metallothionein staining in tumour cells and the expression of metallothionein in stromal cells were not correlated with histopathological parameters (type and grade) in metallothionein-positive cases of lung carcinoma examined. No association was found between metallothionein expression and lymph node status in the examined cases of lung carcinoma.. Our findings indicate that expression of metallothionein was evident in squamous cell lung carcinoma and adenocarcinoma, but absent in small-cell lung carcinoma, supporting evidence for participation of this protein in the biological mechanisms underlying the carcinogenic evolution in the lung.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Carcinoma, Small Cell; Carcinoma, Squamous Cell; Female; Humans; Immunoenzyme Techniques; Lung Neoplasms; Male; Metallothionein; Middle Aged

Metallothioneins (MTs) are zinc-binding proteins whose overexpression may lead to sequestration of zinc ions and consequently to functional inactivation of the p53 tumor suppressor gene. The aim of the study was to investigate the potential role of MTs in the carcinogenesis of ulcerative colitis (UC) as well as possible effects on p53 function. The monoclonal antibodies E9 (anti-MT), DO-7, and 1801 (anti-p53) and the polyclonal antibody CM-1 (anti-p53) were used to stain formalin-fixed, paraffin-embedded colon specimens obtained from 14 patients with UC-associated colorectal carcinoma (CAC), 13 with high-grade dysplasia (HGD), 10 with low-grade dysplasia (LGD), and 30 with UC without dysplasia or carcinoma. Statistical significance (p <0.05) was assessed using Fisher's exact test. Positive MT staining (> 20% of tumor, dysplastic, or epithelial cells) was found in most UC and LGD but in only a small percentage of HGD and CAC (p <0.01 for CAC vs. UC and LGD vs. HGD). Positive p53 immunoreactivity was observed predominantly in HGD and CAC but not in LGD and UC (p <0.01 for CAC vs. UC and HGD vs. LGD). In histologically normal tissue neighboring CAC, significant MT expression was found in six of seven specimens with simultaneous lack of p53 expression. MT overexpression may represent an important early step in the development of CAC independent of p53 expression and should be investigated in the long term as an independent cancer risk factor in UC.

Topics: Adenocarcinoma; Adult; Aged; Biomarkers; Cell Transformation, Neoplastic; Colitis, Ulcerative; Colorectal Neoplasms; Humans; Immunohistochemistry; Metallothionein; Middle Aged; Precancerous Conditions; Risk Factors; Tumor Suppressor Protein p53

Metallothionein (MT) is a small protein with a high affinity for divalent heavy metal ions. This metalloproteinis involved in many (patho)physiological processes, like metal homeostasis and detoxification, cell proliferation, apoptosis, therapy resistance, and protection against oxidative damage. Alterations in the immunohistochemical expression of MT have been reported for various human tumors, and a high expression has been found to be associated with a poor clinical outcome. We showed previously that gastrointestinal cancer is accompanied by a decrease in MT expression, but the most malignant phenotypes had the highest MT levels. The purpose of the present study was to assess the clinical relevance of MT in gastrointestinal cancer.. In this study, we determined the MT levels, by radioimmunoassay, in intestinal tissue of 251 patients with colorectal cancer and 81 patients with gastric cancer and assessed the relation with the overall survival of these patients.. More than 74% of the carcinomas were found to have a lower MT level than their corresponding normal mucosa. In colorectal cancer patients, but not in gastric cancer patients, a high MT level in both the carcinomas and normal mucosa was, however, significantly associated with a poor overall survival, independently from clinicopathological features.. Overexpression of MT in intestinal tissue of colorectal cancer patients is a prognostic marker for a poor overall survival. In gastric cancer, however, MT expression in the gastric mucosa is not of prognostic significance. This observation emphasizes the clinical relevance of this multifunctional metalloprotein in colorectal carcinogenesis and therapy.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Case-Control Studies; Colorectal Neoplasms; Female; Gastrointestinal Neoplasms; Humans; Male; Metallothionein; Middle Aged; Mucous Membrane; Prognosis; Prospective Studies; Radioimmunoassay; Survival Rate

The purpose of this study was to define the prognostic value of a group of molecular tumor markers in a well-staged population of patients treated with trimodality therapy for esophageal cancer. The original pretreatment paraffin-embedded endoscopic esophageal tumor biopsy material was obtained from 118 patients treated with concurrent cisplatin + 5-fluorouracil (5-FU) + 45 Gy radiation followed by resection from 1986 until 1997 at the Duke University Comprehensive Cancer Center. Three markers of possible platinum chemotherapy association [metallothionein (MT), glutathione S-transferase-pi (GST-pi), P-glycoprotein (P-gp or multidrug resistance)] and one marker of possible 5-FU association [thymidylate synthase (TS)] were measured using immunohistochemistry. The median cancer-free survival was 25.0 months, with a significantly improved survival for the 38 patients who had a complete response (P < 0.001). High-level expression of GST-pi, P-gp, and TS were associated with a decreased survival. MT was not significant in this population. Multivariate analysis identified high-level expression in two of the platinum markers (GST-pi and P-gp) and the 5-FU marker TS as independent predictors of early recurrence and death. In conclusion, this investigation measured three possible markers associated with platinum and one possible marker associated with 5-FU in a cohort of esophageal cancer patients. Independent prognostic significance was observed, which suggests that it may be possible to predict which patients may benefit most from trimodality therapy. These data need to be reproduced in a prospective investigation.

Topics: Adenocarcinoma; Adult; Age Factors; Aged; Antineoplastic Combined Chemotherapy Protocols; ATP Binding Cassette Transporter, Subfamily B, Member 1; Biopsy; Carcinoma, Squamous Cell; Cisplatin; Disease-Free Survival; Esophageal Neoplasms; Fluorouracil; Glutathione S-Transferase pi; Glutathione Transferase; Humans; Immunohistochemistry; Isoenzymes; Metallothionein; Middle Aged; Multivariate Analysis; Prognosis; Proportional Hazards Models; Thymidylate Synthase; Time Factors; Treatment Outcome

The aim of this study is to clarify whether the expression of metallothionein (MT) is related with the malignant potential in primary colorectal cancer and/or synchronous liver metastasis. Immunohistochemical staining for MT was performed on the specimens of adenocarcinoma of the colon and rectum and its liver metastases in 34 patients treated with curative surgery, respectively. Expression of MT was compared with clinicopathological variables and patient survival. In patients with primary colorectal cancer, positive expression was found in 7 of 34 (20.6%) patients, but MT was not detected in any of the cases of liver metastases (0%; p = 0.0111). In the primary tumor, positive MT expression was significantly associated with a higher degree of lymph node involvement (mean +/- SD: 48.4 +/- 33.8 vs. 18.6 +/- 24.4% in MT-positive and MT-negative tumors, respectively; p = 0.0122). The survival rate in the patients with MT-negative tumors was significantly better than that in those with MT-positive tumors as primary sites (p = 0.0198). MT expression in colorectal cancer may be a potential marker affecting lymph node metastases and may be a predictor of a poor prognosis, particularly in patients with synchronous liver metastases.

Topics: Adenocarcinoma; Aged; Antineoplastic Combined Chemotherapy Protocols; Chemotherapy, Adjuvant; Cisplatin; Colorectal Neoplasms; Combined Modality Therapy; Doxorubicin; Drug Resistance, Neoplasm; Female; Fluorouracil; Humans; Life Tables; Liver Neoplasms; Male; Metallothionein; Middle Aged; Mitomycin; Multivariate Analysis; Neoplasm Proteins; Prognosis; Survival Analysis

Because the status of the inherent drug-resistance of colorectal cancers remains obscure, human colorectal cancers with no neoadjuvant therapy were retrospectively investigated regarding the expression of three drug-resistant proteins: metallothionein, glutathione S-transferase-pi, and P-glycoprotein.. Paraffin-embedded tissues of 130 colorectal cancers (Dukes A, 20; B, 49; C, 41; D, 20) obtained by surgical resections from 1982 to 1989 were used. The three proteins were immunostained by the streptavidin-biotin complex method. The immunostaining was judged to be positive if more than 5 percent of cells showed positive staining by use of cell analysis system. The data were compared with clinicopathologic features (Dukes A-D) and patients' prognosis (Dukes AC).. Metallothionein, glutathione S-transferase-pi, and P-glycoprotein were positively expressed in 91 (70 percent), 30 (23 percent), and 98 (75 percent), respectively. A total of 120 (86 percent) expressed at least one drug-resistant protein. No intergroup differences were observed between positive and negative expressions of the proteins and their clinicopathologic features except tumor location. Rectal cancers positively expressed P-glycoprotein and three proteins more frequently. Twenty-six (20 percent), 65 (50 percent), and 21 (16 percent) cancers positively expressed one, two, and three proteins, respectively. The disease-free survival rates of patients with Dukes A through C cancer with positive staining for one, two, and three proteins were 100, 94, and 83 percent (at 1 year); 100, 72, and 51 percent (at 3 years); and 94, 66, and 38 percent (at 5 years), respectively (Kaplan-Meier with log-rank test; P = 0.016). In the multivariate Cox analysis, age, Dukes stage, tumor size, and glutathione S-transferase-pi were independent prognostic factors.. The patients with concurrent expression of drug-resistant proteins in their cancers had worse prognoses. Examining drug-resistant proteins in colorectal cancers may be useful in selecting adjuvant chemotherapy and in predicting prognosis more accurately.

Topics: Adenocarcinoma; Aged; ATP Binding Cassette Transporter, Subfamily B, Member 1; Biomarkers, Tumor; Colorectal Neoplasms; Combined Modality Therapy; Disease-Free Survival; Female; Glutathione S-Transferase pi; Glutathione Transferase; Humans; Immunoenzyme Techniques; Isoenzymes; Male; Metallothionein; Neoplasm Invasiveness; Retrospective Studies

Maspin is a recently described member of the serpin family of protease inhibitors that is consistently expressed at high levels in mammary myoepithelial cells. This feature was used in the immunohistochemical evaluation of tubular carcinoma (TC) and radial sclerosing lesion (RSL) of the breast, and compared with other markers of myoepithelial cells. Ten cases of TC and 11 cases of RSL were studied for the expression of maspin, alpha-smooth muscle actin (alpha-SMA), metallothionein (MT), and S-100 protein by immunohistochemistry. Myoepithelial cells stained strongly and diffusely for maspin creating a pattern of an outer continuous ring surrounding the epithelium of tubules of all RSLs. This pattern was absent in all TCs; however, the single-layered epithelium comprising the tubules of two TCs was positive for maspin with a moderate to strong intensity. Myoepithelial cells were not positive for MT in a consistent manner. Benign nonproliferative epithelium stained focally and weakly for maspin in four of 11 cases of RSL and was negative for MT in all 11 cases. Foci of mild to moderate epithelial hyperplasia noted in five of 11 cases of RSL stained diffusely with a weak to moderate intensity for maspin and focally with a strong intensity for MT. alpha-SMA was consistently expressed in myoepithelial cells but also in stromal myofibroblasts and blood vessels, creating a pattern that was less satisfactory than maspin in distinguishing RSL from TC. Immunohistochemical staining for S-100 protein was of no differential diagnostic value. In conclusion, immunohistochemical staining for maspin is diagnostically useful and superior to MT, S-100, and alpha-SMA, in distinguishing RSL from TC. The epithelial immunoreactivity for maspin in two of 10 TCs merits further investigation from a prognostic viewpoint.

Topics: Adenocarcinoma; Biomarkers, Tumor; Breast; Breast Neoplasms; Diagnosis, Differential; Epithelium; Female; Genes, Tumor Suppressor; Humans; Immunohistochemistry; Metallothionein; Proteins; Sclerosis; Serpins

Metallothionein (MT) is a small thiol-rich metalloprotein with antioxidant properties, involved in tumour pathophysiology and therapy resistance. In order to assess the contribution of MT in gastrointestinal carcinogenesis, this study examined both the MT content by radioimmunoassay and the MT localization by immunohistochemistry in pairs of neoplastic and normal-appearing human gastrointestinal tissues. In addition, the relationship between MT expression and major clinicopathological parameters was assessed. The MT concentration of gastric carcinomas and of colorectal adenomas, carcinomas, and liver metastases was found to be significantly lower than that of corresponding normal-appearing tissue. A relatively high MT content, however, was found to be associated with the villous character of colorectal adenomas and with the Dukes' stage of colorectal carcinomas, indicating a relationship between MT level and malignant potential. Immunohistochemical evaluation showed a fairly good correlation with these quantitative data. MT was found to be expressed at a low level and in a patchy pattern in the gastrointestinal neoplastic and metastatic tissues, whereas in normal-appearing gastrointestinal mucosa MT was uniformly distributed in the cytoplasm and/or nucleus of apical cells. Although in the gastric cancer patients no association was found between the MT concentration and the clinicopathological parameters, the strong MT expression in areas with intestinal metaplasia, known to have neoplastic potential, further points to a relationship between this antioxidant metalloprotein and the malignant character of cells. Gastrointestinal neoplasms are apparently accompanied by a low level and decreased expression of MT, but those with a relatively high level seem to have an increased malignant potential. Further studies will be required to determine the clinical relevance of these observations.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Colorectal Neoplasms; Female; Fluorescent Antibody Technique, Indirect; Humans; Liver Neoplasms; Male; Metallothionein; Middle Aged; Radioimmunoassay; Stomach Neoplasms

Metallothioneins (MTs) are a group of ubiquitous low molecular weight proteins with a high affinity for heavy metal ions. The aim of the present study was to investigate MT expression in a series of endometrial carcinomas. We wished to determine whether MT expression in endometrial carcinoma was related to established prognostic factors such as tumour grade, stage and histological type. We also wanted to establish if high MT expression in curettings of endometrial carcinoma was predictive of high expression in the subsequent hysterectomy specimen.. Sixty-three cases of endometrial carcinoma were included in the study. These comprised 57 endometrioid adenocarcinomas (15 grade 1, 25 grade 2, 17 grade 3), three papillary serous adenocarcinomas, two mucinous adenocarcinomas and one clear cell adenocarcinoma. Forty-five tumours were stage I, 10 were stage II and eight were stage III. In 28 cases, diagnostic endometrial curettings, performed prior to hysterectomy, were available for study. Immunohistochemical staining was performed using the anti-MT monoclonal antibody E9. The intensity and distribution of MT staining were assessed using a semiquantitative method. This resulted in an intensity distribution (ID) score out of a maximum of 300. The mean ID score of grade 1 and 2 endometrioid adenocarcinomas was 67 and 63, respectively, while for grade 3 tumours the mean ID score was 114. This was statistically significant (P = 0.05). The three papillary serous adenocarcinomas had high ID scores with a mean of 208. The mean ID score of stage I tumours was 69. This was lower than those of stage II and III tumours which had mean ID scores of 116 and 128, respectively. However, these differences were not statistically significant (P = 0.288). A significant correlation was observed between MT ID scores in endometrial curettings and in the subsequent hysterectomy (P = 0.013).. MT isoforms can be demonstrated in most endometrial adenocarcinomas. High MT ID scores are associated with high grade and high stage endometrial adenocarcinomas and with the aggressive papillary serous adenocarcinoma. Whether this is of value as an independent prognostic factor has yet to be established.

Topics: Adenocarcinoma; Endometrial Neoplasms; Female; Humans; Immunohistochemistry; Metallothionein; Prognosis

Immunocytochemical demonstration of metallothionein (MT) has been reported as a useful prognostic tool in human breast cancer. The aim of this study was to determine the immunohistochemical location of MT in canine mammary tumours and its possible correlation with the morphologic characteristics of these tumours. Surgical specimens from spontaneous malignant (n = 20) and benign mammary neoplasms (n = 20) were processed for routine histological examination and immunohistochemical study. An indirect immunoperoxidase technique, using monoclonal antibody E9 against horse MT was employed. Intensity of the stain, the percentage of immunoreactive tumour cells and immunohistochemical overexpression of MT was estimated for each case. Metallothionein over-expression, defined as those cases with more than 10% immunopositive cells, was detected in both benign and malignant mammary tumours. However, strong immunostaining intensity was seen in benign tumours, whereas in malignant tumours immunopositive cells stained weakly. Positive MT immunostaining occurred in neoplastic epithelial cells, and some chondrocytes present in mixed mammary tumours. However, staining intensity was variable in immunopositive cells. Differences in staining intensity between the primary malignant mammary tumour, tumour emboli and metastatic cells within a lymph node were also noted. Myoepithelial cells and connective tissue did not stain for MT. We concluded that metallothionein immunostaining cannot be used as a diagnostic or prognostic tool in canine mammary neoplasms. However, results of this study support the hypothesis that MT has a role in tumour proliferation and tumour progression.

Topics: Adenocarcinoma; Adenoma; Animals; Biomarkers, Tumor; Carcinoma; Dogs; Female; Immunoenzyme Techniques; Mammary Neoplasms, Animal; Metallothionein; Mixed Tumor, Malignant; Neoplasms, Complex and Mixed; Neoplasms, Glandular and Epithelial

The insulin-like growth factor II receptor (IGFIIR) has been implicated as a tumor suppressor gene in human malignancy. Frequent mutation, loss of heterozygosity, and microsatellite instability (MSI) directly affecting the IGFIIR gene have been reported in several primary human tumor types. However, to our knowledge, dynamic functional evidence of a growth-suppressive role for IGFIIR has not yet been provided. We identified one MSI-positive colorectal carcinoma cell line, SW48, with monoallelic mutation in IGFIIR identical to that seen in primary colorectal carcinomas. A zinc-inducible construct containing the wild-type IGFIIR cDNA was stably transfected into SW48 cells. Growth rate and apoptosis were compared between zinc-treated, untreated, and untransfected cells. A twofold increase in IGFIIR protein expression was detected after zinc treatment in discrete clonal isolates of transfected SW48 cells. Moreover, zinc induction of exogenous wild-type IGFIIR expression reproducibly decreased growth rate and increased apoptosis. These data prove that wild-type IGFIIR functions as a growth suppressor gene in colorectal cancer cells and provide dynamic in vitro functional support for the hypothesis that IGFIIR is a human growth suppressor gene.

Topics: Adenocarcinoma; Apoptosis; Cell Division; Colorectal Neoplasms; DNA, Complementary; Gene Expression Regulation, Neoplastic; Humans; Insulin-Like Growth Factor II; Loss of Heterozygosity; Metallothionein; Microsatellite Repeats; Neoplasm Proteins; Polymerase Chain Reaction; Promoter Regions, Genetic; Receptor, IGF Type 2; Tumor Cells, Cultured; Zinc

Human kidneys and their associated tumors (nonneoplastic kidney tissues from patients with a transitional cell carcinoma or an adenocarcinoma and the adenocarcinomas themselves) were evaluated for their Zn, Cd, and Cu contents as well as for their metallothionein (MT) level. The total Cd content was correlated with the MT content, and both values were significantly decreased in the adenocarcinomas in comparison with the other tissues. After extraction and separation by anion-exchange chromatography, MT-0 was identified in the nonneoplastic tissues from both the adenocarcinomas as well as the transitional cell carcinomas. Since until now MT-0 protein was only found in human fetal liver and in Zn-stimulated human monocytes, a possible role for this isoform as an oncofetal marker is hypothesized. Separation of the isoforms of MT by reversed-phase high-performance liquid chromatography and sequence analysis showed besides MT-1e and MT-1l the isoform-MT-1g, which is not expressed in the healthy kidney, and MT-1k, an isoform which is not yet demonstrated in renal tissues. We conclude that the expression profile of the MT isoforms in the kidney changes due to the presence of a tumor.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Amino Acid Sequence; Base Sequence; Cadmium; Carcinoma, Transitional Cell; Copper; DNA; Female; Humans; Kidney; Kidney Neoplasms; Male; Metallothionein; Middle Aged; Molecular Sequence Data; Protein Isoforms; Sequence Homology, Amino Acid; Sequence Homology, Nucleic Acid; Zinc

To investigate the role of metallothionein in colorectal tumours and the possible relation with other factors associated with tumour progression: expression of cathepsin D (CD), CD44, p53, Rb, bcl-2, c-erbB-2, epidermal growth factor receptor (EGFR), proliferation indices (Ki-67, proliferating cell nuclear antigen (PCNA)), and conventional clinicopathological variables.. The immunohistochemical expression of metallothionein was investigated in 23 cases of colorectal adenoma and 94 adenocarcinomas. Metallothionein expression was examined by the avidinbiotin peroxidase immunoperoxidase (ABC) using the monoclonal mouse antibody E9, on formalin fixed, paraffin embedded tissue.. Positive metallothionein expression (> 5% of neoplastic cells) was observed in 30.4% of adenomas and 25.5% of adenocarcinomas, while 8.7% of adenomas and 14.9% carcinomas showed focal metallothionein positivity. In contrast, 60.9% of adenomas and 59.6% of carcinomas almost completely lacked metallothionein expression. In the series of adenocarcinomas, metallothionein expression was inversely correlated with CD44 in neoplastic cells (p = 0.01). There was no statistically significant difference of metallothionein expression, or the other variables examined, between adenocarcinomas and adenomas.. Metallothionein expression does not seem to indicate aggressive biological behaviour in colorectal adenocarcinomas, in comparison with the other types of carcinoma. The inverse correlation with CD44 could suggest that the decreased metallothionein expression may contribute to the metastatic spread of the lymph node involvement in colorectal cancer. Metallothionein expression does not seem to represent an independent prognostic marker in colorectal cancer.

Topics: Adenocarcinoma; Biomarkers, Tumor; Cathepsin D; Colorectal Neoplasms; Female; Humans; Hyaluronan Receptors; Male; Metallothionein; Middle Aged; Prognosis

To study the neurophysiological functions of metallothioneins (MTs), localization of MT-I and -II was examined immunohistochemically in a variety of brain lesions in dogs, including infarct, laminar cortical necrosis, hemorrhage, invasive growth of tumour, inflammatory lesions in granulomatous meningoencephalitis and distemper encephalitis. MT-I and -II were demonstrated in both nucleus and cytoplasm of hypertrophic astrocytes in most brain lesions examined regardless of the type, size, localization and duration of the lesions. In addition, MT expression was stronger in a population of hypertrophic astrocytes localizing inside of the surviving brain tissue rather than those localizing at the boundary between the surviving brain tissue and necrotic area, where severe inflammatory changes were developing. These results suggest that MT-I and -II may play roles not only in protection of neurons from metals and free radicals ubiquitous in the inflammatory lesions but also in repair of injured neural tissues.

Topics: Adenocarcinoma; Animals; Astrocytes; Brain Diseases; Brain Neoplasms; Cerebral Hemorrhage; Cerebral Infarction; Distemper; Dog Diseases; Dogs; Encephalitis, Viral; Female; Glial Fibrillary Acidic Protein; Hypertrophy; Inflammation; Male; Meningoencephalitis; Metallothionein; Necrosis; Neoplasm Invasiveness; Pituitary Neoplasms

Differences in expression of metallothionein (MT) have been reported in various human tumours. MT is mainly expressed in proliferating epithelial tumour cells but in human hepatocellular carcinoma (HCC) there is only a minimal expression of MT. Since MT is a zinc binding protein and certain inducers of MT including zinc play a role in apoptosis, studies were undertaken to compare the expression of MT and the presence of apoptotic cells (APPC) in both primary HCC and metastatic adenocarcinoma.. Histological sections of 13 cases of primary HCC and eight cases of metastatic adenocarcinoma were obtained from archival samples. They were stained for MT using a polyclonal antibody which crossreacts readily with human MT and for APPC by the TUNEL technique. Normal human liver had consistent MT staining with no detectable APPC. The primary HCC showed moderate MT staining with a small number of APPC while metastatic adenocarcinoma showed no MT staining with a large number of APPC.. These results suggest a relationship between absence of MT and appearance of APPC in human liver tumours, especially in metastatic adenocarcinomas.

Topics: Adenocarcinoma; Adult; Aged; Apoptosis; Carcinoma, Hepatocellular; Female; Humans; Immunohistochemistry; Liver Neoplasms; Male; Metallothionein; Middle Aged

Zinc exhibits inhibitory effects on apoptosis, and a deficiency in this metal generally causes this type of cell death to occur. In the present study, we found that exposure to zinc results in necrosis of prostate carcinoma cells. When zinc acetate was added to LNCaP or PC-3 cells in monolayer culture, they began to detach from the culture dishes, and viability was lost after 4-8 h. Most of the cell death was found to be due to necrosis as determined by double staining with fluorescein-isothiocyanate-labeled annexin V and ethidium bromide, and by detection of hypodiploid cells. Associated with the induction of necrosis was an increase in low molecular-mass proteins, identified by HPLC analysis to be thymosin beta10, parathymosin and GAGE in LNCaP cells, and thymosin beta4, parathymosin and metallothionein in PC-3. The time course of the increase of thymosin beta10 in LNCaP cells and thymosin beta4 in PC-3 cells was consistent with that of appearance of cell detachment and dead cells. These results indicate that zinc can induce necrosis and suggest that production of proteins including beta-thymosins is involved in induction of processes leading to cell detachment.

Topics: Adenocarcinoma; Annexin A5; Apoptosis; Cell Adhesion; Cell Division; Cell Survival; Copper; DNA, Neoplasm; Humans; Male; Metallothionein; Necrosis; Neoplasm Proteins; Prostatic Neoplasms; Thymosin; Tumor Cells, Cultured; Zinc

Topics: Adenocarcinoma; Colorectal Neoplasms; Humans; Immunohistochemistry; Metallothionein

Prostate carcinoma is the second leading cause of death from malignancy in men in the United States. Prostate cancer cells express type I insulin-like growth factor receptor (IGF-IR) and prostate cancer selectively metastazises to bone, which is an environment rich in insulin-like growth factors (IGFs), thereby supporting a paracrine action for cancer cell proliferation. We asked whether the IGF-IR is coupled to tumorigenicity and invasion of prostate cancer. When rat prostate adenocarcinoma cells (PA-III) were stably transfected with an antisense IGF-IR expression construct containing the ZnSO4-inducible metallothionein-1 transcriptional promoter, the transfectants expressed high levels of IGF-IR antisense RNA after induction with ZnSO4, which resulted in dramatically reduced levels of endogenous IGF-IR mRNA. A significant reduction in expression both of tissue-type plasminogen activator and of urokinase-type plasminogen activator occurred in PA-III cells accompanying inhibition of IGF-IR. Subcutaneous injection of either nontransfected PA-III or PA-III cells transfected with vector minus the IGF-IR insert into nude mice resulted in large tumors after 4 weeks. However, mice injected with IGF-IR antisense-transfected PA-III cells either developed tumors 90% smaller than controls or remained tumor-free after 60 days of observation. When control-transfected PA-III cells were inoculated over the abraded calvaria of nude mice, large tumors formed with invasion of tumor cells into the brain parenchyma. In contrast, IGF-IR antisense transfectants formed significantly smaller tumors with no infiltration into brain. These results indicate an important role for the IGF/IGF-IR pathway in metastasis and provide a basis for targeting IGF-IR as a potential treatment for prostate cancer.

Topics: Adenocarcinoma; Animals; Base Sequence; Brain Neoplasms; Cell Division; Cell Line; DNA Primers; Gene Expression; Male; Metallothionein; Mice; Mice, Nude; Molecular Sequence Data; Neoplasm Invasiveness; Polymerase Chain Reaction; Promoter Regions, Genetic; Prostatic Neoplasms; Rats; Receptor, IGF Type 1; Restriction Mapping; RNA, Antisense; Sulfates; Transcription, Genetic; Transfection; Transplantation, Heterologous; Zinc Compounds; Zinc Sulfate

The immunohistochemical distribution pattern of metallothionein, a low molecular weight protein with strong affinity for divalent heavy metal ions, has been investigated in normal and neoplastic conditions of the large bowel. Utilizing a monoclonal mouse antibody the following formalin-fixed paraffin-embedded surgical or biopsy samples were studied: tubulo-villous adenomas (8 cases); adenocarcinomas with various degree of differentiation (85), nine of which were mucinous-type; synchronous tubular or tubulo-villous adenomas separate from carcinomas (30); transitional mucosa (45); metastases in lymph nodes (43); and distant metastases (45). Twenty biopsies from the right and left colon of 10 patients affected by irritable bowel syndrome were also analyzed. Normal colonic mucosa as well as transitional mucosa showed metallothionein immunopositivity in enterocytes at the luminal surface and crypts. Evident nuclear and cytoplasmic staining was encountered in tubulo-villous adenomas; the same reactivity was noted in the basal glandular component of colorectal carcinomas-synchronous adenomas, while less intense staining was noted in the apical villous portions. A variable metallothionein immunostaining was observed in adenocarcinomas (62.3%), in lymph node (55.8%) and distant hepatic (17.2%) and omental (43.8%) metastases, although it was not always concordant with that reported in the corresponding primary tumour. Whether the metallothionein positivity observed in normal and neoplastic cells is the result of expression of a stable form of the protein or an accumulation in the nucleus and cytoplasm remains to be clarified.

Topics: Adenocarcinoma; Adenoma; Colon; Colorectal Neoplasms; Humans; Immunohistochemistry; Intestinal Mucosa; Metallothionein; Rectum

Zinc homeostasis was studied during the induction, growth, and methotrexate (MTX) treatment of Dark Agouti rat mammary adenocarcinomas (DAMA). A progressive fall in plasma Zn concentration (pZn), significant at a tumor burden of less than 1% body weight (bw), was sustained during tumor enlargement to give a 54% reduction in pZn at 16.3% bw (n = 6/group). The hypozincemia was attributed to the increasing Zn demand for tumor growth. Zn content of the 16.3% bw tumors equaled that of muscle (normally 60% of total body Zn). Tumor metallothionein (tMT) was sufficient to bind < 3% of total tumor Zn, and hepatic MT (hMT) remained at basal concentrations during early tumor growth, doubling only in the presence of significant necrosis in large tumors. Methotrexate (MTX, 0.5 mg/Kg im x 2 d) at respective tumor burdens of 5 and 10% bw (n = 9, 10/group) gave 2 therapeutic effects, dependent on tumor size: 1.5% bw tumors in 7 rats remained close to their original size until experiment end when pZn, hMT, and tMT were typical of 5% bw untreated tumors. 2. Tumors in 5 rats given MTX at 10% bw had marked subcapsular necrosis and regression to a size similar to those in group 1; pZn returned toward normal, whereas hMT was 6 times its 5% bw counterpart. Host weight loss was significantly reduced, as were tumor-associated changes in plasma glucose and calcium. In summary, neither tMT nor hMT appears to play a role in the hypozincemia that follows DAMA Zn sequestration and growth. Critically timed MTX can result in tumor regression and return of plasma Zn, Ca, and glucose toward normal. This is associated with an increase in hMT and reduction in host weight loss, suggesting a flow of Zn from the resorbing tumor to the host, enabling the synthesis of hMT and retention of host structural proteins.

Topics: Adenocarcinoma; Animals; Blood Glucose; Body Weight; Calcium; Female; Homeostasis; Liver; Mammary Neoplasms, Experimental; Metallothionein; Methotrexate; Neoplasm Transplantation; Organ Size; Rats; Zinc

Metallothioneins (MTs), a set of ubiquitous low-molecular-weight proteins essential for the protection of cells against heavy metal ion toxicity, were demonstrated immunohistochemically using a monoclonal antibody (E9) against a conserved epitope of I and II isoforms in a series of 109 colorectal adenocarcinomas. In a semiquantitative analysis strong MT expression in the majority of tumour cells was observed in 34 (31%) cases, 24 (22%) tumours showed a focal MT positivity, and 51 (47%) almost completely lacked MT expression. These differences in MT expression were statistically significantly (P < 0.05) associated with the tumour stage (Dukes classification) and the lymph node involvement at the time of operation (pN stages). However, in contrast to previous findings obtained on a variety of tumours, MT positivity was associated with a favourable clinical outcome in colonic carcinoma, which may indicate their different biological behaviour. Survival curves of cases with MT-positive and MT-negative status differed from each other in a univariate analysis (Mantel-Haenszel: 8.9, P < 0.05) but lost significance when a multivariate analysis was carried out by means of the Cox proportional regression model with Dukes' stages as a stratification factor. It is concluded that immunohistochemically demonstrated MT expression is significantly associated with tumour stages but does not represent an independent prognostic variable in colorectal cancer. However, it may provide important information about some of the biological mechanisms underlying progression in colorectal cancer.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Colorectal Neoplasms; Female; Humans; Immunohistochemistry; Male; Metallothionein; Middle Aged; Neoplasm Staging; Prognosis; Survival Analysis

Eleven pairs of surgically resected lung cancers and corresponding non-neoplastic lung tissue were evaluated for metallothionein (MT) and metal content (cadmium and copper) by the heme/109Cd binding assay and atomic absorption spectroscopy, respectively. Tissue samples, obtained from patients ranging in age from 51 to 79, included six adenocarcinomas, two small cell carcinomas, one mixed cell carcinoma, one squamous cell carcinoma, and one carcinoma of non-primary origin (i.e., melanoma). Paired t-tests showed that metallothionein and copper concentrations in lung tumor tissue were significantly elevated when compared to non-malignant lung tissue. Cu was the major metal associated with the 10 kDa MT fraction in lung tumors whereas Cd was the primary metal bound to MT from non-neoplastic lung tissue. Since Cu-thionein is also known to be elevated in fetal lung tissue, the possibility exists that MT might represent an oncodevelopmental product which is useful as a biomarker for the early detection of lung carcinoma.

Topics: Adenocarcinoma; Aged; Biomarkers, Tumor; Cadmium; Cadmium Radioisotopes; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Chromatography, Agarose; Copper; Humans; Lung; Lung Neoplasms; Melanoma; Metallothionein; Middle Aged; Neoplasm Proteins; Protein Binding; Spectrophotometry, Atomic

The antibody to the metal-binding low molecular weight protein metallothionein (MT) stains preferentially the proliferative edge of epithelial tumors in paraffin sections. The present report demonstrates its usefulness as an epithelial cell marker in DNA flow cytometry of archival specimens. Nine control breast (mammoplasty) specimens, 10 control colonic specimens (resection edges), 12 adenocarcinomas of breast, and 13 adenocarcinomas of colon were analyzed by DNA flow cytometry after MT and DNA staining. The average percentage of cells stained by MT ranged from 12% to 27% in these groups of specimens, which contain epithelial as well as stromal and inflammatory cells. Comparing cell turnover, measured as S-phase fraction (SPF) in unstained and MT-stained preparations, it was 10% and 20%, respectively, in control tissues and 10% and 30%, respectively, in adenocarcinomas. The SPF is lower in unstained preparations because of dilution by noncycling inflammatory and stromal cells. Immunohistochemical staining of various tissues for MT showed specific staining of epithelial cells. Evaluation of aneuploid malignant epithelial cells detected in six breast and eight colonic adenocarcinomas showed that on average, 47% of cells were stained with MT and that their SPF increased by about 50% when MT staining was compared with the unstained preparations. The results suggest that MT stains epithelial cells adequately for ploidy and cell cycle evaluation and that it may stain preferentially the proliferating cell compartment, which is considered to be an index of malignancy.

Topics: Adenocarcinoma; Biomarkers, Tumor; Breast Neoplasms; Cell Compartmentation; Cell Division; Colonic Neoplasms; DNA, Neoplasm; Epithelium; Female; Flow Cytometry; Humans; Metallothionein

The expression of intrinsic resistance to cisplatin in two lung cancer cell lines, one derived from a small cell carcinoma (SW1271) and the other from an adenocarcinoma (A549), relative to a drug-sensitive small cell line SW900, was characterized by: (i) expression of cross-resistance to mitomycin C and cadmium chloride, but increased sensitivity to adriamycin and etoposide; (ii) significantly decreased cisplatin uptake; (iii) elevated levels of glutathione which could be reduced by buthionine L-sulfoximine resulting in significant sensitization of the cells to cisplatin; (iv) a lack of consistent modification of metallothionein content and expression of levels of glutathione S-transferase, glutathione reductase and glutathione peroxidase or of activities of DT-diaphorase or catalase; (v) significantly reduced total DNA-platination levels immediately following a 1 h cisplatin treatment with 10 micrograms/ml (33.3 microM); (vi) increased removal of Pt-GG and Pt-AG adducts by the A549 cells, consistent with increased repair capacity, but a lack of removal of these major adducts by the SW1271 cells indicative of tolerance of this drug-induced DNA damage. These data therefore provide evidence of differential formation, repair and tolerance of DNA damage following exposure of three human lung carcinoma cell lines to cisplatin.

Topics: Adenocarcinoma; Carcinoma, Small Cell; Catalase; Cisplatin; DNA; DNA Adducts; DNA Damage; DNA Repair; DNA, Neoplasm; Drug Resistance; Drug Screening Assays, Antitumor; Gene Expression; Glutathione; Glutathione Peroxidase; Glutathione Reductase; Glutathione Transferase; Humans; Kinetics; Lung Neoplasms; Metallothionein; NAD(P)H Dehydrogenase (Quinone); Tumor Cells, Cultured

A hybrid anti-tumour B72.3 antibody/metallothionein protein B72.3MT-1 was produced by the construction of the expression vector mpSV2neo-EP1-B72.3MT-1. This vector contained the neo gene as a selection marker, the murine immunoglobulin promoter and enhancer, and the hybrid B72.3 heavy chain gene fragment with mouse metallothionein-1 cDNA gene ligated into its CH2 domain. The expression vector was transfected to the heavy chain loss mutant B72.3Mut(K) cell line. The hybrid protein B72.3MT-1 was purified from transfectant supernates using a Protein G column. We showed that the hybrid protein retained the binding reactivity for the TAG72 antigen as the original B72.3 antibody, and the metal-binding capacity of the native metallothionein molecule. Therefore, the bifunctional hybrid protein B72.3MT-1 may be very useful in cancer imaging when labelled with radionuclides such as 99mTc.

Topics: Adenocarcinoma; Antibodies, Neoplasm; Antibody Specificity; Base Sequence; Cell Line; Colonic Neoplasms; DNA; Electrophoresis, Polyacrylamide Gel; Genetic Vectors; Humans; Immunoenzyme Techniques; Metallothionein; Molecular Sequence Data; Protein Engineering; Protein Multimerization

In patients with primary biliary cirrhosis and Wilson's disease liver copper concentrations become elevated during the evolution of the disorder. The accumulated copper is thought to be detoxified by metallothionein, a protein which binds copper and zinc. In liver metastasis of colorectal cancer, copper and zinc concentrations are usually decreased compared to normal liver tissue, but little is known about the concomitant metallothionein levels. In the present study metallothionein concentrations were determined in archival liver samples from patients with primary biliary cirrhosis and Wilson's disease, and in both normal and malignancy-containing liver samples from patients with metastasis from a colorectal adenocarcinoma. Twenty-seven control liver samples contained 3.98 +/- 1.55 mg metallothionein/g protein. From the 21 liver samples of patients with primary biliary cirrhosis, which had a mean metallothionein concentration of 6.06 +/- 5.03 mg/g protein, 6 were above the highest control level. Liver metallothionein concentrations for the 8 patients with Wilson's disease were significantly elevated (10.98 +/- 6.93 mg/g protein, p < 0.005 vs. controls and p < 0.05 vs. primary biliary cirrhosis). In the 11 liver metastases from colorectal adenocarcinomas metallothionein concentrations (1.17 +/- 0.90 mg/g protein) were significantly (p < 0.005) lower than surrounding normal liver tissue (4.25 +/- 1.75 mg/g protein). We conclude that in primary biliary cirrhosis and Wilson's disease increased liver metallothionein concentrations may detoxify the accumulated copper. Furthermore, liver metastasis of colorectal cancer contains less metallothionein than the surrounding normal liver tissue.

Topics: Adenocarcinoma; Adult; Aged; Colorectal Neoplasms; Female; Hepatolenticular Degeneration; Humans; Liver; Liver Cirrhosis, Biliary; Liver Neoplasms; Male; Metallothionein; Middle Aged; Radioimmunoassay

To develop transgenic mice, a mouse metallothionein promoter-human growth hormone (Mt-hGH) gene fragment was injected into the pronucleus of C57BI x DBA/2J F2 one-cell embryos. Six founder animals grew larger (1.3-2.2 times) than littermate controls and had higher levels of hGH in plasma (4.6-279 mU/l). At 27-43 weeks of age, 3 of the 4 female transgenic founders developed malignant papillary adenocarcinomas of mammary origin. One of the male transgenic founders was successfully mated and 2 of 3 female transgenic offspring developed mammary tumors. Forty-two female mice with the same genetic background as the transgenic animals and older than 43 weeks served as controls. No palpable tumor was found in this group. Five of the control animals were killed and examined histologically, revealing only normal mammary tissue. Earlier studies have shown that GH is important for growth and development of the mammary gland. Our study suggests that markedly elevated endogenous levels of GH cause mammary carcinoma in a specific strain of transgenic mice. The present animal model might be useful for studying molecular mechanisms that are involved in the development of hormonally induced mammary tumors.

Topics: Adenocarcinoma; Animals; Body Weight; Female; Gene Expression; Genetic Vectors; Growth Hormone; Male; Mammary Neoplasms, Experimental; Metallothionein; Mice; Mice, Transgenic; Promoter Regions, Genetic

Cu/Zn-containing proteins have recently become of interest with regard to their relation with malignant disorders. Cu Zn-superoxide dismutase (Cu/Zn-SOD) was found increased in chemically induced tumors of the large bowel whereas metallothionein (MT), containing Zn and some Cu, was shown important for the response of tumors to chemotherapy. In the present study, we evaluated the Cu/Zn-SOD and MT content of normal human colonic mucosa and colorectal carcinomas, obtained from 20 resection specimens, and of 47 adenomatous polyps. The Cu/Zn-SOD content of polyps and carcinomas was significantly (p less than 0.01) elevated when compared to normal mucosa. In the adenomatous polyps the Cu/Zn-SOD content increased significantly with increasing grade of epithelial cell dysplasia, diameter, and presence of a villous component. In the carcinomas no relation was noticed between the Cu/Zn-SOD content and the Dukes' stage or the grade of differentiation. The MT content was significantly decreased in both adenomatous polyps and carcinomas when compared to that in normal mucosa. The MT content was not related to the grade of epithelial cell dysplasia of the polyps, and to the Dukes' stage or the differentiation of the carcinomas. In conclusion, neoplasia of the colorectum is accompanied by an increase in Cu/Zn-SOD and a decrease in MT. These findings support the association between changes in Cu/Zn proteins and malignancy.

Topics: Adenocarcinoma; Adenoma; Adult; Aged; Aged, 80 and over; Colonic Neoplasms; Copper; Female; Free Radicals; Humans; Intestinal Mucosa; Intestinal Polyps; Male; Metallothionein; Middle Aged; Superoxide Dismutase; Zinc

Topics: Adenocarcinoma; Biomarkers, Tumor; Breast; Breast Neoplasms; Female; Humans; Immunoenzyme Techniques; Metallothionein; Ovarian Neoplasms; Ovary; Receptors, Estrogen; Receptors, Progesterone; Uterine Neoplasms; Uterus

The effect of preinduction of metallonthionein (MT) by bismuth subnitrate (BSN) on the adverse effects and antitumor activity of gamma-ray irradiation was investigated in mice. Preinduction of MT by oral administration of BSN significantly reduced the lethal effects and bone marrow injury caused by total body irradiation with gamma-rays. A significant increase in the MT concentration in bone marrow was observed in mice treated with BSN. In tumor-bearing mice, pretreatment with BSN did not compromise the antitumor activity of gamma-ray irradiation although bone marrow injury was remarkably suppressed. These results suggest that BSN pretreatment is an effective method for protection against side-effects in radiotherapy.

Topics: Adenocarcinoma; Animals; Bismuth; Bone Marrow; Colonic Neoplasms; Gamma Rays; Leukemia P388; Male; Metallothionein; Mice; Mice, Inbred Strains; Radiation Injuries, Experimental; Radiation, Ionizing

The protective effects of metallothionein (MT) preinduction by bismuth subnitrate (BSN) against the renal toxicity of cis-diamminedichloroplatinum (cis-DDP), the cardiotoxicity of adriamycin (ADR), and the lethal and bone marrow toxicities of these drugs were observed in mice receiving cis-DDP and ADR simultaneously. Preinduction of MT biosynthesis by BSN, which is currently used as an antidiarrheal drug, did not affect the antitumor activities of the two drugs, suggesting that the beneficial effects of the preinduction of MT biosynthesis by BSN may be applicable for therapy involving cis-DDP and ADR, either alone or in combination.

Topics: Adenocarcinoma; Animals; Antineoplastic Combined Chemotherapy Protocols; Bismuth; Carcinoma, Ehrlich Tumor; Cisplatin; Colonic Neoplasms; Doxorubicin; Heart; Kidney; Male; Metallothionein; Mice; Mice, Inbred ICR

The effect of preadministration of bismuth, a specific potent inducer of renal metallothionein (MT), on the lethal and renal toxicity of cis-DDP, an extensively used antitumor drug containing heavy metal platinum, in mice was investigated. Pretreatment of mice with two s.c. doses of bismuth nitrate (BN; 30 mumol/kg/day) at 24-hr interval prevented the lethal toxicity, the increase in BUN value and the occurrence of diarrhea caused by cis-DDP (35 mumol/kg, s.c.). This protective effect of BN-pretreatment was significantly correlated with increased MT levels in the kidney. The pretreatment of tumor-bearing mice with BN also depressed the lethal and renal toxicity of cis-DDP without compromising its antitumor activity, and allowed the administration of relatively high dose of cis-DDP. Daily five consecutive p.o. preadministration of bismuth subnitrate (BSN), one of the bismuth compounds being in use as a protectant of the gastrointestinal lining, was also effective to depress the lethal toxicity of cis-DDP. Since the effective dose of BSN is not far from that commonly used for men, this treatment will allow the increase in cis-DDP dose, promising a clinical advantage in cancer chemotherapy.

Topics: Adenocarcinoma; Animals; Bismuth; Cisplatin; Colonic Neoplasms; Kidney; Kidney Diseases; Metallothionein; Mice; Neoplasm Transplantation

Nuclear extract from Morris hepatoma 3924A was fractionated by DEAE-Sephadex chromatography. The fraction eluting with 300 mM (NH4)2SO4 (DE-C) was used for transcribing cloned mouse metallothionein-I (MT-I) gene in a run-off assay. This fraction contained the majority of RNA polymerase II as well as the transcription factor(s). Accuracy of MT-I DNA transcription was confirmed by S1 nuclease mapping. Low concentrations (1 microgram/ml) of alpha-amanitin inhibited the reaction, indicating that RNA polymerase II directed the transcription. Unfractionated nuclear extracts from the hepatoma or a rat mammary adenocarcinoma as well as whole cell extract obtained from the mammary tumor also transcribed MT-I gene. The extent of transcriptional activity was in the following order: hepatoma nuclear fraction DE-C greater than whole cell extract derived from rat mammary adenocarcinoma cells greater than nuclear extract derived from rat hepatoma or rat mammary adenocarcinoma cells. These studies have demonstrated that a fractionated nuclear extract obtained from a tissue supports efficient and accurate RNA polymerase II-mediated transcription of MT-I DNA.

Topics: Adenocarcinoma; Amanitins; Animals; Cell Extracts; Cell Fractionation; Cell Nucleus; Cell-Free System; Chromatography, Ion Exchange; DNA; Liver Neoplasms, Experimental; Mammary Neoplasms, Experimental; Metallothionein; Mice; Rats; RNA Polymerase II; Transcription, Genetic

High levels of metallothionein (MT) are present in the developing mammalian liver; however, a remarkable decrease is observed during postnatal life after weaning. This developmental profile is similar to that of certain oncofetal gene products such as alpha-fetoprotein, which is used as a tumor marker. This study deals with the reexpression of MT genes in thyroid tumors. With an immunohistochemical method, the presence of MT was investigated in tissue sections of normal and neoplastic human thyroid glands. Tissue sections of 34 thyroid tumors and 10 normal human thyroid glands were studied by means of the peroxidase-antiperoxidase method. MT was localized in 31 of the thyroid gland tumors. MT was also present in two of the normal thyroid glands. These findings indicate that although high levels of MT are mainly found in the fetal liver, it may also be expressed actively in certain human thyroid neoplastic tissues, and occasionally in normal thyroid tissue.

Topics: Adenocarcinoma; Adenocarcinoma, Papillary; Adenoma; Adolescent; Adult; Aged; Aged, 80 and over; Cell Nucleus; Cytoplasm; Female; Humans; Immunohistochemistry; Male; Metallothionein; Middle Aged; Thyroid Neoplasms

This case report describes the natural history of chronic cadmium intoxication in a woman who was exposed to excessive cadmium in her occupation. We document the clinical, laboratory, and tissue response to the toxicant.

Topics: Adenocarcinoma; Cadmium; Cadmium Poisoning; Environmental Exposure; Female; Humans; Kidney; Liver; Lung Neoplasms; Metallothionein; Microscopy, Electron; Middle Aged; Occupational Diseases

Topics: Adenocarcinoma; Cell Line; Cycloheximide; Cystine; Dexamethasone; Dihydrotestosterone; Hormones; Humans; Insulin; Kinetics; Male; Metallothionein; Prostaglandins E; Prostatic Neoplasms; Zinc