Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0278883 (metastatic melanoma)
 6,224 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In this study we used differential display technology in an attempt to obtain an insight into the mechanisms underlying escape of tumour cells to the specific cytotoxic T cell response. A primary tumour cell line and autologous tumour infiltrating lymphocytes were raised from a metastatic melanoma sample (ME15). Upon co-culture of tumour infiltrating lymphocytes with irradiated tumour cells, CTL specific for neoplastic cells were generated and cloned. Using a CTL clone, a cytotoxicity resistant tumour subline (ME15R) was immunoselected. We applied a PCR-based differential display technique to amplify DNA sequences differentially expressed in ME15 sublines sensitive (S) or resistant (R) to specific CTL killing. 10 different sequences whose expression was exclusively detectable in ME15S cells were identified. Five of them matched with known expressed sequence tags encoding products of unidentified function. 2 showed high homology with a mitochondrial mRNA and with the gene encoding the S24 ribosomal protein. Most interestingly, genes coding for glutamine synthetase, TGF-beta-3 and PAX3, a well-characterised transcription factor, were only expressed in ME15S cells. The latter gene was found to be transcribed in all healthy tissues tested, but only in a subgroup of established melanoma cell lines. Taken together, our data underline the relevant potential of differential display technology in the molecular analysis of paired tumour lines endowed with different phenotypic characteristics. Cloning of entire open reading frames and transfection studies are warranted to clarify the role of individual differentially displayed genes in the escape of tumour cells from cellular immune response.
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PMID:Identification of genes differentially expressed in melanoma sublines derived from a single surgical specimen characterised by different sensitivity to cytotoxic T-lymphocyte activity. 1082 99

There are currently no effective therapies for metastatic melanoma and targeted immunotherapy results in the remission of only a very small percentage of tumors. In this study, we show that the noncanonical Wnt ligand, Wnt5A, can increase melanoma metastasis in vivo while down-regulating the expression of tumor-associated antigens important in eliciting CTL responses (e.g., MART-1, GP100, and tyrosinase). Melanosomal antigen expression is governed by MITF, PAX3, and SOX10 and is inhibited upon signal transducers and activators of transcription 3 (STAT3) activation, via decreases in PAX3 and subsequently MITF expression. Increasing Wnt5A in Wnt5A-low cells activated STAT3, and STAT3 was decreased upon Wnt5A knockdown. Downstream targets such as PAX3, MITF, and MART-1 were also affected by Wnt5A treatment or knockdown. Staining of a melanoma tissue array also highlighted the inverse relationship between MART-1 and Wnt5A expression. PKC activation by phorbol ester mimicked Wnt5A effects, and Wnt5A treatment in the presence of STAT3 or PKC inhibitors did not lower MART-1 levels. CTL activation studies showed that increases in Wnt5A correspond to decreased CTL activation and vice versa, suggesting that targeting Wnt5A before immunotherapy may lead to the enhancement of current targeted immunotherapy for patients with metastatic melanoma.
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PMID:Wnt5A regulates expression of tumor-associated antigens in melanoma via changes in signal transducers and activators of transcription 3 phosphorylation. 1907 88

PAX3 and MITF are important transcriptional activators in the melanocyte lineage and PAX3 is thought to control MITF expression during normal melanocyte differentiation. However, it is not clear whether this is still true in melanoma and whether the effects of knockdown of PAX3 on the inhibition of melanoma growth or survival are by its regulation of MITF. By western blot and quantitative real-time reverse transcription-PCR, we investigated the relationship between PAX3 and MITF expression in 27 metastatic melanoma and one immortalized melanocyte cell lines. All lines were found to express both PAX3 and MITF proteins but levels varied by 15 fold and more than 100 fold, respectively. The expression of PAX3 protein was correlated with that of MITF (r=0.75; P<0.001) but the expression of PAX3 protein and MITF mRNA was not. Immunofluorescence microscopy showed that individual cells expressed widely differing relative amounts of PAX3 and MITF protein. By MTT cell proliferation and flow cytometry assays, both MITF and PAX3 proteins seemed to be functional, as knockdown with siRNA led to reduced proliferation and induction of apoptosis. However, knockdown of PAX3 with small interfering RNA did not decrease MITF expression and vice versa. In one cell line (NZM15), silencing of PAX3 induced terminal differentiation whereas silencing of MITF induced expression of FOXD3, a repressor of melanogenesis. The results suggest that the melanoma lines used in this study show considerable phenotypic variation of expression of these two transcriptional activators and reflect a deregulation of the developmental process operating in the genesis of the melanocyte lineage, and that they probably function independently to enhance the survival of melanoma cells.
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PMID:PAX3 knockdown in metastatic melanoma cell lines does not reduce MITF expression. 2116 69

Comparing the transcriptomes of primary and metastatic tumour tissues is a useful strategy for studying tumour progression. One factor limiting the interpretation of tissue-based transcriptomic data is the lack of cell-type purity. Laser capture microdissection (LCM) has been shown to be useful for overcoming this limitation. We established an efficient protocol for gene expression profiling of LCM and matched metastatic melanomas using a transgenic mouse model. This optimized workflow combines microsurgical recovery of mouse lungs, appropriate tissue freezing, laser microdissection of homogeneous tumour cell populations from cryosections, isolation of high-quality RNA and gene expression analysis. The RNA isolated from laser-microdissected material was not contaminated by stroma cells, was of excellent quality, and the synthesis of cDNAs was homogeneous and highly reproducible. Subsequent custom-based Taqman-low-density-array (TLDA)-based gene expression profiling identified stronger expression of five genes (M-MITF, TYR, STAT3, CCND1 and PAX3) in primary than metastatic melanoma. We detected only minor transcriptomic differences between primary and metastatic melanoma tissue. This optimized workflow could be very valuable for various studies requiring cell type-specific transcriptomic analysis.
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PMID:Efficient gene expression profiling of laser-microdissected melanoma metastases. 2293 21

Metastatic melanoma is an aggressive and deadly disease. The chemokine receptor CXCR4 is active in melanoma metastasis, although the mechanism for the promotion and maintenance of CXCR4 expression in these cells is mostly unknown. Here, we find melanoma cells express two CXCR4 isoforms, the common version and a variant that is normally restricted to cells during development or to mature blood cells. CXCR4 expression is driven through a highly conserved intronic enhancer element by the transcription factors PAX3 and FOXD3. Inhibition of these transcription factors slows melanoma cell growth, migration, and motility, as well as reduces CXCR4 expression. Overexpression of these transcription factors drives the production of increased CXCR4 levels. Loss of PAX3 and FOXD3 transcription factor activity results in a reduction in cell motility, migration, and chemotaxis, all of which are rescued by CXCR4 overexpression. Here, we discover a molecular pathway wherein PAX3 and FOXD3 promote CXCR4 gene expression in melanoma.
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PMID:PAX3 and FOXD3 Promote CXCR4 Expression in Melanoma. 2620 21

Using MIRA-seq, we have characterized the DNA methylome of metastatic melanoma and normal melanocytes. Individual tumors contained several thousand hypermethylated regions. We discovered 179 tumor-specific methylation peaks present in all (27/27) melanomas that may be effective disease biomarkers, and 3113 methylation peaks were seen in >40% of the tumors. We found that 150 of the approximately 1200 tumor-associated methylation peaks near transcription start sites (TSSs) were marked by H3K27me3 in melanocytes. DNA methylation in melanoma was specific for distinct H3K27me3 peaks rather than for broadly covered regions. However, numerous H3K27me3 peak-associated TSS regions remained devoid of DNA methylation in tumors. There was no relationship between BRAF mutations and the number of methylation peaks. Gene expression analysis showed upregulated immune response genes in melanomas presumably as a result of lymphocyte infiltration. Down-regulated genes were enriched for melanocyte differentiation factors; e.g., KIT, PAX3 and SOX10 became methylated and downregulated in melanoma.
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PMID:The DNA methylation landscape of human melanoma. 2638 56

The role of the Pax3 gene in embryonic development of pigment cells is well characterized. By contrast, the function of Pax3 in melanoma development is controversial. Indeed, data obtained from cultured cells suggest that PAX3 may contribute to melanomagenesis. PAX3 is found to be overexpressed in melanomas and also in nevi compared with normal skin samples. Pax3 homozygous loss of function is embryonic lethal. To assess the role of Pax3 in melanoma development in vivo, we analyzed Pax3 haploinsufficiency in a mouse model of melanoma predisposition. The Pax3(GFP/+) knock-in reporter system was combined with the Tyr::NRAS(Q61K); Cdkn2a(-/-) mouse melanoma model. Melanoma development was followed over 18 months. Histopathological, immunohistochemical, and molecular analyses of lesions at different stages of melanoma progression were carried out. Fluorescence-activated cell sorting on GFP of cells from primary or metastatic melanoma was followed by ex-vivo transformation tests and in-vivo passaging. We report here that Tyr::NRAS(Q61K); Cdkn2a(-/-); Pax3(GFP/+) mice developed metastasizing melanoma as their Tyr::NRAS(Q61K); Cdkn2a(-/-); littermates. Histopathology showed no differences between the two genotypes, although Pax3 mRNA and PAX3 protein levels in Pax3(GFP/+) lesions were reduced by half. The Pax3(GFP) allele proved to be a convenient marker to identify and directly sort heterogeneous populations of melanoma cells within the tumor bulk at each stage of melanoma progression. This new mouse model represents an accurate and reproducible means for identifying melanoma cells in vivo to study the mechanisms of melanoma development.
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PMID:Haplosufficiency of PAX3 for melanoma development in Tyr: NRASQ61K; Cdkn2a-/- mice allows identification and sorting of melanoma cells using a Pax3GFP reporter allele. 2658 92

Circulating Tumour Cells (CTCs) are promising cancer biomarkers. Several methods have been developed to isolate CTCs from blood samples. However, the isolation of melanoma CTCs is very challenging as a result of their extraordinary heterogeneity, which has hindered their biological and clinical study. Thus, methods that isolate CTCs based on their physical properties, rather than surface marker expression, such as microfluidic devices, are greatly needed in melanoma. Here, we assessed the ability of the slanted spiral microfluidic device to isolate melanoma CTCs via label-free enrichment. We demonstrated that this device yields recovery rates of spiked melanoma cells of over 80% and 55%, after one or two rounds of enrichment, respectively. Concurrently, a two to three log reduction of white blood cells was achieved with one or two rounds of enrichment, respectively. We characterised the isolated CTCs using multimarker flow cytometry, immunocytochemistry and gene expression. The results demonstrated that CTCs from metastatic melanoma patients were highly heterogeneous and commonly expressed stem-like markers such as PAX3 and ABCB5. The implementation of the slanted microfluidic device for melanoma CTC isolation enables further understanding of the biology of melanoma metastasis for biomarker development and to inform future treatment approaches.
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PMID:Isolation and detection of circulating tumour cells from metastatic melanoma patients using a slanted spiral microfluidic device. 2897 38