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The clinical diagnosis of cutaneous melanoma always calls for histological confirmation. In addition to the recognition of the classic aspects of the neoplasm, immunohistochemistry is determinant, in particular in the assessment of the size of the replicative compartment. Generally, the proliferation rate is indicative of the neoplastic progression and is related to the clinical growth rate of the neoplasm. It allows to distinguish high risk melanomas showing a high growth rate from those of lower malignancy associated with a restricted growth rate. In melanoma, the recruitment and progression of neoplastic cells in the cell cycle of proliferation have lost some of their controls that are normally processed by a series of key regulatory molecules. In addition, the apoptotic pathway counteracting any hyperproliferative activity is released of the dependency of specific regulated molecular mechanisms. This review summarizes the current knowledge on key molecular components involved in the deregulation of the growth fraction, cell proliferation and apoptosis in melanocytic neoplasms. The implication of cyclins and of the mitogen-activated protein kinase pathways are scrutinized. The involvement of neoplastic stem cells in the metastatic process is also discussed.
Int J Mol Med 2009 Sep
PMID:Molecular pathways supporting the proliferation staging of malignant melanoma (review). 1963 20

In order to test whether 18S rDNA can influence positively xylanase gene effective expression in the yeast of Candida utilis, a targeting vector pGLR9K-XA was constructed by adding an interested gene xynA from Streptomyces olivaceoviridis into the vector pGLR9K which is constructed by ourselves. pGLR9K contains the 18S rDNA, GAP promoter and CYH resistance gene sequence, all of which is from C. utilis. Then the vector pGLR9K-XA was transformed into C. utilis. To test the vector and transformed system, PCR, Southern blot and DNS methods were used. The results showed that xylanase gene can be detected in the chromosome DNA of recombinant C. utilis and the enzyme activity of xylanase is up to 60 IU ml(-1) in the study. It is suggested that this system can be used to express exogenous genes in C. utilis as a bioreactors. This is the first report that xylanase gene was expressed in C. utilis.
Mol Biol Rep 2010 Jul
PMID:The effective expression of xylanase gene in Candida utilis by 18S rDNA targeted homologous recombination in pGLR9K. 1973 Oct 75

Capsicum species are very important in Brazil because of economic, cultural and biological factors, and the country is considered to be a diversity center for this genus. Collection and maintenance of the genetic diversity in Capsicum are important to avoid genetic erosion. Besides the identification of species, the characterization and evaluation of accessions maintained in gene banks are of fundamental importance. For this purpose, multivariate methods have become an important tool in the classification of conserved genotypes. The objectives of this study were: i) to identify and characterize accessions of the Capsicum spp collection and draw conclusions about the potential use of certain accessions in different production sectors; ii) to estimate the genetic divergence among accessions using the Ward-MLM procedure, and iii) to evaluate the efficiency of the analysis of continuous and categorical data using the Ward-MLM procedure. Fifty-six Capsicum spp accessions were evaluated based on 25 descriptors, 14 of which were morphological and 11 agronomic. Based on the qualitative descriptors, it was possible to identify all species and, together with the agronomic descriptors, genotypes could be indicated with potential for use in various production sectors. Five was determined as the ideal number of groups by the criteria pseudo-F and pseudo-t2. The Ward-MLM procedure allowed the differentiation of the species C. annuum, C. frutescens, C. baccatum, and C. chinense in separate groups. The Ward-MLM procedure showed some level of efficiency in clustering Capsicum species analyzing morphological and agronomic data simultaneously.
Genet Mol Res 2010 Feb 18
PMID:Genetic variability in domesticated Capsicum spp as assessed by morphological and agronomic data in mixed statistical analysis. 2019 84

The reagent l-dimethylaminonaphthalene-5-sulfonyl chloride (dansyl chloride, DNS-C1) reacts with the free amino groups of peptides and proteins as shown in Fig. 1. Total acid hydrolysis of the substituted peptide or protein yields a mixture of free amino acids plus the dansyl derivative of the N-terminal amino acid, the bond between the dansyl group and the N-terminal amino acid being resistant to acid hydrolysis. The dansyl amino acid is fluorescent under UV light and is identified by thin-layer chromatography on polyamide sheets. This is an extremely sensitive method for identifying amino acids and in particular has found considerable use in peptide sequence determination when used in conjunction with the Edman degradation (see Chapter 24 ). The dansyl technique was originally introduced by Gray and Hartley (1), and was developed essentially for use with peptides. However, the method can also be applied to proteins (see Note No. 12). Fig. 1. Reaction sequence for the labeling of N-terminal amino acids with dansyl chloride.
Methods Mol Biol 1984
PMID:The dansyl method for identifying N-terminal amino acids. 2051 90

Cutaneous malignant melanoma is the most aggressive skin cancer. It is also the most rapidly spreading cancer in terms of worldwide incidence. Although it is detected by simple inspection and can be relatively easily removed or treated, differential diagnosis to other melanocytic lesions, lack of prognostic markers, and no efficient treatment of advanced melanoma pose problems. Detection and targeting of proteases may represent a useful tool since they play a role in tumor cell metabolism, invasion, angiogenesis and metastasis. This review gives an overview of the role of proteases in development and progression of cutaneous malignant melanoma. In addition, regulation, activation, and interaction of proteases and their inhibitors are explained for tumors in general. The potential use of proteases as differential markers for melanoma mimicking melanocytic lesions, as biomarkers in tissues, and as prognostic serum markers is discussed. Current and future possibilities to target tumor proteases in therapy are presented.
Cell Mol Life Sci 2010 Dec
PMID:Proteases in cutaneous malignant melanoma: relevance as biomarker and therapeutic target. 2068 12

Skin melanoma, a life-threatening disease, has a recently reported worldwide increase in incidence, despite primary prevention. Skin melanoma statistics emphasize the need for finding markers related to the immune response of the host. The mechanisms that are able to over-power the local immune surveillance comprise molecules that can be valuable markers for diagnosis and prognosis. This article summarizes the immune markers that can monitor the disease stage and evaluate the efficacy of therapeutic interventions. Recent data regarding immunotherapy are presented in the context of tumor escape from immune surveillance and the immune molecules that are both targets and a means of monitoring. Perspectives for developing immune interventions for skin melanoma management and the position of tissue or soluble immune markers as a diagnostic/prognostic panel are evaluated. State-of-the-art technology is emphasized for developing immune molecular signatures for a complex characterization of the patient's immunological status.
Expert Rev Mol Diagn 2010 Oct
PMID:Immune-related biomarkers for diagnosis/prognosis and therapy monitoring of cutaneous melanoma. 2096 10

The FoxP3 (forkhead box P3) gene is an X-linked gene that is submitted to inactivation. It is an essential transcription factor in CD4(+)CD25(+)FoxP3 regulatory T cells, which are therapeutic targets in disseminated cutaneous melanoma. Moreover, FoxP3 is an important tumor suppressor gene in carcinomas and has putative cancer suppressor gene function in cutaneous melanoma as well. Therefore understanding the structure and function of the FoxP3 gene is crucial to gaining insight into the biology of melanoma to better develop immunotherapeutics and future therapeutic strategies.
Mol Oncol 2011 Apr
PMID:The dual role of the X-linked FoxP3 gene in human cancers. 2148 91

Uveal melanoma (UM) is one of the most common primary intraocular malignant tumors in adults. Melanoma antigen recognized by T cell-1 (Mart-1), one of the melanosome-specific proteins, has been widely studied as a marker recognized by cytotoxicity T lymphocytes. Mart-1 is considered to play a critical role in the immunotherapy for melanoma. Additionally, as a biomarker, Mart-1 is often used with other tumor-associated antigens for antidiastole in cutaneous melanoma (CM), uveal melanoma (UM) and nevus. In this study, the differential expression of Mart-1 was investigated in four human UM cells (SP6.5, VUP, OCM-1 and OM431) on three levels of analysis: messenger ribonucleic acid (mRNA), protein and, eventually, morphology. The results revealed that SP6.5 cells had high Mart-1 protein expression while VUP cells had almost none. OCM-1 and OM431 cells produced less Mart-1 than SP6.5 cells according to Western blot analysis, although OM431 cells had the highest expression of Mart-1 mRNA according to real-time PCR. The results indicate the potential use of Mart-1 in the development of therapy for UM.
Mol Med Rep
PMID:Differential expression of Mart-1 in human uveal melanoma cells. 2166 26

Resistance against anticancer drugs remains a serious obstacle in cancer treatment. Here we used novel strategies to target microsomal glutathione transferase 1 (MGST1) and glutathione transferase pi (GSTP) that are often overexpressed in tumors and confer resistance against a number of cytostatic drugs, including cisplatin and doxorubicin (DOX). By synthetically combining cisplatin with a GST inhibitor, ethacrynic acid, to form ethacraplatin, it was previously shown that cytosolic GST inhibition was improved and that cells became more sensitive to cisplatin. Here we show that ethacraplatin is easily taken up by the cells and can reverse cisplatin resistance in MGST1 overexpressing MCF7 cells. A second and novel strategy to overcome GST mediated resistance involves using GST releasable cytostatic drugs. Here we synthesized two derivatives of DOX, 2,4-dinitrobenzenesulfonyl doxorubicin (DNS-DOX) and 4-mononitrobenzenesulfonyl doxorubicin (MNS-DOX) and showed that they are substrates for MGST1 and GSTP (releasing DOX). MGST1 overexpressing cells are resistant to DOX. The resistance is partially reversed by DNS-DOX. Interestingly, the less reactive MNS-DOX was more cytotoxic to cells overexpressing MGST1 than control cells. It would appear that, by controlling the reactivity of the prodrug, and thereby the DOX release rate, selective toxicity to MGST1 overexpressing cells can be achieved. In the case of V79 cells, DOX resistance proportional to GSTP expression levels was noted. In this case, not only was drug resistance eliminated by DNS-DOX but a striking GSTP-dependent increase in toxicity was observed in the clonogenic assay. In summary, MGST1 and GSTP resistance to cytostatic drugs can be overcome and cytotoxicity can be enhanced in GST overexpressing cells.
Mol Pharm 2011 Oct 03
PMID:Characterization of new potential anticancer drugs designed to overcome glutathione transferase mediated resistance. 2185 Oct 97

When cutaneous melanoma is confined to the skin, simple excision with adequate margins will usually cure the patient (1,2). Local recurrences do occur but reexcision still results in a very high cure rate. When cutaneous melanoma spreads beyond the primary site, the metastases are predominantly by way of the lymphatics. If in-transit disease or regional lymph node involvement is present, the 5-yr survival rate drops to approx 60% (1-3). Accurate staging of the locoregional lymphatic basin is thus extremely important. Pre- operative lymphoscintigraphy followed by selective lymphadenectomy has revolutionized the staging of cutaneous melanoma by delivering to the pathologist only those nodes that are most likely to contain metastatic cells (4). A close examination of these sentinel lymph nodes (SLNs) by serial sectioning and immunohistochemical staining can detect very minute quantities of melanoma. This type of detailed examination is impossible in standard lymphadenectomy specimens that can contain 20-40 lymph nodes. The standard technique used to examine large numbers of lymph nodes is to examine only 1-5% of each node using hematoxylin and eosin staining. This can obviously miss micrometastatic disease and understage the patient.
Methods Mol Med 2001
PMID:A molecular technique useful in the detection of occult metastases in patients with melanoma : rt-PCR analysis of sentinel lymph nodes and peripheral blood. 2232 65


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