Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: DrugBank:EXPT01586 (G418)
 2,237 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Murine papilloma cell lines 308 and SP-1 have been used as recipients for transfected oncogenes to investigate malignant conversion. These cell lines express an activated c-rasHa gene with a codon 61 mutation and produce squamous papillomas when transplanted as skin grafts onto nude mice. They are not tumorigenic by subcutaneous injection. Both papilloma cell lines were stably transfected with plasmid DNA containing either a rearranged murine plasmacytoma-derived c-myc (minus exon 1), adenovirus 5 E1A, FBJ v-fos or a human c-fos/FBJ v-fos chimera, using cotransfection with the neomycin resistance gene contained in pSV2neo to select for transformants. Southern and northern blotting analysis confirmed the uptake and expression of exogenous DNA in both G418-selected cell lines and in the derived tumors. Unlike the E1A- and myc-containing plasmids, both fos constructs caused malignant conversion in either cell line, as defined by the squamous cell carcinoma histology of tumors from grafted cells and the development of carcinomas after subcutaneous injection into athymic nude mice. Immunofluorescence analysis for specific keratin gene expression indicated that tumors derived by introduction of either of the fos oncogenes were devoid of staining for K1, a 67 kDa epidermal keratin that is expressed in papillomas but not in squamous carcinomas. Tumors from E1A, myc, or pSV2neo transfectants expressed K1, although in a focal distribution. The malignant phenotype induced by the fos oncogene constructs was not associated with the ability to form agar colonies in vitro or to express gamma-glutamyl transpeptidase in the tumors. Since both 308 and SP-1 were sensitive to the fos oncogene for malignant conversion and insensitive to E1A or myc, it is possible that fos may cooperate with the endogenous-activated c-rasHa gene to convert these cells to malignancy. However, since gamma-glutamyl transpeptidase activity is found in the majority of chemically induced mouse skin carcinomas that possess an activated c-rasHa gene, fos activation may not be a common pathway for spontaneous malignant conversion.
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PMID:Malignant conversion of murine squamous papilloma cell lines by transfection with the fos oncogene. 247 37

Human papillomaviruses (HPVs) are known as etiologic agents of various diseases in regions of the human epithelium. Specific type HPV16 is most frequently found in association with human squamous cell carcinoma. To examine biological activity of HPV type 16 in human cells, primary foreskin epidermal cells and dermal fibroblasts were transfected by recombinant viral DNA containing neo gene with Ca2+-phosphate precipitation. Epidermal cells were maintained in 0.5% Chelex-treated fetal calf serum, low calcium medium supplemented with bovine pituitary extracts and hormone mix. Fibroblasts were cultured in DMEM plus 10% fetal calf serum. The transfected cells were then selected with G418-resistant phenotype. These cells were propagated to maintain in culture and subsequently became stable lines carrying HPV16 genomes, while mock transfected control cells died off at approximately 40-50 population doublings (PD) in a parallel experiment. We have established two independently immortalized human epidermal cell lines (PHK16-I and II) which harbor different copies of HPV16 genome and express HPV16 specific mRNA. Although younger populations of PHK16 lines were fairly sensitive to high Ca2+-level to be differentiating keratinocytes, progressive changes of the cellular phenotype were demonstrated in terms of changes in Ca2+-response and anchorage independent growth during over 300 PD. Altered Ca2+-regulation of growth and differentiation appeared to be common reliable phenotype associated with stable transformation of skin epidermal cells. In contrast, none of the HPV16-transfected fibroblast line immortalized but simply showed extended life span up to 100 PD in average. The result suggested that this biological activity of HPV16 could be reflected in HPV-tropism related to epithelial transformation. We then studied correlations between HPV16 gene expression and the regulation of growth and differentiation of PHK lines during the progressive transformation. Northern blot analysis of RNA from cells in earlier passages demonstrated that down-regulation of HPV16 E6/E7 transcription was associated with keratinocyte differentiation induced by added 1.0mM calcium. The p97 promoter for HPV 16 early genes covering E6/E7 was specifically responsible for this Ca2+-regulation. The eventual loss of Ca2+-regulation could be implicated in a process of progressive transformation of HPV16-epidermal cell system.
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PMID:[HPV16 participates in progressive transformation of normal epidermal cells]. 253 84

High-molecular-weight DNA isolated from eight fresh human skin cancers occurring on sun-exposed body sites were assayed for their ability to transform NIH 3T3 cells. A cotransfection protocol using pSV2-neo DNA, which confers resistance to the antibiotic G418, was used to select cells that had taken up the transfected DNA. About 2 weeks after transfection, G418-resistant colonies were pooled and injected s.c. into athymic nude mice. The NIH 3T3 cells transfected with DNA from six of the human skin cancers induced tumors in nude mice. DNAs from all six tumor cell lines contained human alu sequences. Southern blot hybridization with ras-specific probes revealed that DNAs from the four alu-rich tumors contained the human Ha-ras oncogene, in addition to that of the NIH 3T3 controls. In contrast, DNAs from the other two tumors did not contain any of the known oncogenes tested, except those endogenous to NIH 3T3 cells. DNAs from three of four first cycle tumorigenic transformants gave rise to morphologically transformed foci when assayed in a second cycle of transfection. DNAs from all three secondary transformants contained discrete human alu sequences, and in addition, contained Ha-ras sequences similar to those present in their respective primary transformants. Interestingly, DNA from both primary and secondary transformants of one particular human squamous cell carcinoma contained highly amplified copies of the Ha-ras oncogene. These results suggest that activation of the Ha-ras oncogene may be common in human skin cancers originating on sun-exposed body sites. Further characterization of the Ha-ras oncogenes present in these human skin cancers may provide information on the molecular mechanisms by which UV radiation of the sun induces human neoplasms on exposed body sites.
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PMID:Detection and identification of activated oncogenes in human skin cancers occurring on sun-exposed body sites. 337 Jun 35

We investigated the effects of transfection of wild-type TP53 on the growth properties of a human gingival carcinoma cell line, KOSC-3, in which the TP53 gene is mutated at codon 248 and overexpressed. The wild-type TP53 expression plasmid, pCDM8-p53/neo and the control plasmid, pCDM8/neo, were each stably transfected into KOSC-3 cells by using the calcium phosphate method. The number of G418-resistant colonies from wild-type TP53-transfected cells was approximately half that from plasmid controls. Exogenous wild-type TP53 transcripts were identified in four of the 20 G418-resistant clones analysed by reverse transcription PCR. Although the growth rates of the wild-type TP53+ clones did not drastically change during log phase, their saturation density was significantly reduced. The wild-type TP53+ cells were morphologically flat and enlarged when cultured in vitro, and were less able to form colonies in soft agar. In nude mice, the wild-type TP53+ clones formed subcutaneous tumours with conspicuous keratinisation and notable cell death that was not manifested in the parental and plasmid control cells. These findings indicate that the wild-type TP53 gene, even when it coexists with a mutated form, may function as a growth suppressor and differentiation inducer under restricted conditions in gingival squamous cell carcinoma.
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PMID:Transfection of wild-type TP53 induces differentiation in human gingival carcinoma cells. 881 3

Two human squamous cell carcinoma of the head and neck (SCCHN) cell lines, PCI-13 and PCI-52, were transduced with the retroviral construct containing human interleukin-2 (IL-2) cDNA and selected for neomycin resistance in G418 medium. Stably transduced SCCHN cells produced and secreted IL-2, which was shown to have biologic activity in a bioassay, using an IL-2-dependent CTLL-2 cell line. By immunohistochemistry, IL-2 gene-transduced PCI-13 cells were strongly positive for IL-2, and by flow cytometry showed both cell surface and intracytoplasmic expression of IL-2 protein. Expression of IL-2 mRNA was measured by quantitative RT-PCR and found to be considerably increased in transduced SCCHN relative to that in parental cells. There was no difference in expression of IL-2R between the parental and IL-2 gene-transduced cells. In vitro proliferation of IL-2 gene-transduced tumor cells was consistently more rapid than that of parental cells. Sensitivity of the parental and IL-2 gene-transduced targets to lysis or apoptosis mediated by purified human natural killer (NK) cells or IL-2-activated NK (A-NK) cells was comparable as measured in 4-hour 51Cr-release and 1-hour [3H]thymidine-release assays, respectively. However, transduced cells were significantly more sensitive than parental cells to these effectors in 24-hour MTT assays, most likely due to IL-2 production by the transduced targets. PCI-52 cells selected for in vivo experiments formed large subcutaneous tumors in immunosuppressed nude mice. Tumors established by subcutaneous injections of 1 x 10(7) IL-2 gene-transduced cells regressed completely by day 25, while those formed by parental or LacZ gene-transduced tumor cells grew progressively. Tumor regression was mediated by numerous mononuclear cells, identified as murine NK cells and macrophages by immunohistochemistry, which accumulated around the IL-2-secreting, but not parental, tumors within 5-6 days after tumor cell injections. Thus, IL-2 gene-transduced SCCHN cells produce functional IL-2 in vivo in amounts sufficient to support the recruitment to the tumor site and antitumor activity of cytotoxic effector cells. IL-2-secreting SCCHN cells may be a useful component of vaccines designed to induce and sustain effector cell activation at the tumor site.
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PMID:In vitro and in vivo characteristics of human squamous cell carcinoma of the head and neck cells engineered to secrete interleukin-2. 940 7

Recently, it has been suggested that chemokine/receptor interactions determine the destination of the invasive tumor cells in several types of cancer. It has also been proposed that the stromal cell-derived factor-1 (SDF-1; CXCL12)/CXCR4 system might be involved lymph node metastasis in oral squamous cell carcinoma (SCC). In order to further clarify the role of the SDF-1/CXCR4 system in oral SCC, we generated CXCR4 stable transfectants (IH-CXCR4) using oral SCC cells, and compared them to IH, which did not express CXCR4 and which did not have lymph node metastatic potentials in vivo. We introduced enhanced green fluorescent protein (GFP) fused-CXCR4 into IH cells, and detected the GFP fluorescence in the cytoplasm and cell membrane in approximately 60% of the G418-resistant cells. This bulk-transfectant expressed a high level of CXCR4 mRNA and protein, and exhibited the characteristic calcium fluxes and chemotactic activity observed in treatment with SDF-1. SDF-1 biphasically activated extracellular signal-regulated kinase (ERK)1/2, but continuously activated Akt/protein kinase B (PKB) in IH-CXCR4 cells. Most importantly, IH-CXCR4 cells frequently metastasized to the cervical lymph node, but not to the distant organs in the orthotopic inoculation of nude mice. Furthermore, these lymph node metastases were inhibited by the treatment of a mitogen-activated protein kinase/ERK kinase inhibitor, U0126, or a phosphatidylinositol 3 kinase inhibitor, wortmannin. These results indicate that SDF-1/CXCR4 signaling mediates the establishment of lymph node metastasis in oral SCC via ERK1/2 or Akt/PKB pathway.
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PMID:Acquisition of lymph node, but not distant metastatic potentials, by the overexpression of CXCR4 in human oral squamous cell carcinoma. 1549 52

Gelsolin (GSN) is one of the most abundant actin-binding proteins, and is involved in several pathological processes, including Alzheimer's disease, cardiac injury and cancer. The aim of the present study was to assess the effect of GSN on the growth and motility of oral squamous cell carcinoma Tca8113 cells. The overexpression vector pcDNA3.1-GSN was transfected into Tca8113 cells and the stable GSN overexpression cell line was identified based on G418 antibiotic selection. The effect of GSN overexpression on the proliferation, apoptosis, migration and invasion of Tca8113 cells was examined using a cell counting kit-8 assay, flow cytometry and Transwell assays. The results revealed that GSN overexpression significantly promoted the cell proliferation and apoptosis of Tca8113 cells. In addition, Transwell assays demonstrated that the migration and invasion abilities of Tca8113 cells were enhanced by GSN overexpression. Therefore, the upregulation of GSN promotes cell growth and motility, indicating that it may perform a vital function in the progression of human oral cancers.
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PMID:Gelsolin regulates proliferation, apoptosis, migration and invasion in human oral carcinoma cells. 2613 26