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Query: UMLS:C0027651 (tumor)
 685,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interleukin-12 (IL-12) is a heterodimeric cytokine originally defined by its ability to induce the maturation of cytolytic lymphocytes and by its capacity to effectively synergize with IL-2 in the induction of cytolytic activity. Recent studies in mice have demonstrated the ability of IL-12 to cause tumor regression and stimulate long-term antitumor immunity in treated animals. To examine the antitumor effect of direct gene transfer of IL-12 into tumors, we have developed retroviral vectors that coordinately express both subunits of IL-12. An MFG-based retroviral vector was used to generate a recombinant retrovirus in which a long terminal repeat (LTR)-driven polycistronic transcript encodes both subunits of human IL-12: hp35 and hp40 cDNAs are linked and coexpressed using the internal ribosome entry site (IRES) from the encephalomyocarditis virus (DFG-hIL-12). In addition, two IRES sequences were used to express both subunits of IL-12 and a neomycin resistance (neoR) selectable marker gene from the same polycistronic message (TFG-hIL-12). The amphotropic DFG-hIL-12 and TFG-hIL-12 viruses were used to infect both human and murine cell lines as well as primary tumor cultures. The production of human IL-12 by the nonselected, infected cells was measured in both a PHA blast proliferation bioassay and an ELISA and ranged from 15 to 40 ng/10(6) cells per 24 hr. Following G418 selection of TFG-hIL-12-infected cells, the level of expression of IL-12 was significantly higher (up to 120 ng/10(6) cells per 24 hr). The IL-12 protein secreted by the infected cells exhibited all of the biologic activities of recombinant hIL-12: proliferation of activated natural killer (NK) and T cells, stimulation of interferon-gamma (IFN-gamma) induction by NK and T cells, and enhancement of lymphokine-activated killer (LAK) activity. These retroviral vectors expressing human IL-12 should be useful in evaluating the biological properties of IL-12 as well as for use in clinical trials for gene therapy of patients with cancer.
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PMID:Construction and characterization of retroviral vectors expressing biologically active human interleukin-12. 771 Nov 42

A mouse/human chimeric antibody cACT19 derived from the murine ACT19 antibody was constructed; it recognizes an epitope different from the sialosyl-Tn on the TAG72 antigen as defined by the B72.3 antibody. This chimeric cACT19 antibody was constructed by using two expression vectors, the heavy chain expression vector mpSV2neo-EP1-VHC gamma 1 and the light chain expression vector mpSV2gpt-EP1-VKCK. These vectors contain the following: (i) the neo or gpt gene as a selection marker, (ii) the murine immunoglobulin promoter and enhancer (EP1), (iii) the genomic DNA fragments of human immunoglobulin constant region (CK and C gamma 1) and (iv) the murine cDNA fragments of VH or VK region cloned from the murine ACT19 cDNA library. These two vector DNAs were sequentially transfected into the SP2/0Ag14 cell line. Transfectants were selected in media containing both G418 and mycophenolic acid. The chimeric cACT19 antibody was purified from the transfectant supernate by Protein A Sepharose chromatography. We confirmed that the chimeric cACT19 antibody was reactive for an epitope that differed from the sialosyl-Tn on the TAG72 antigen. This was achieved by using the TAG72-binding inhibition ELISA utilizing various monosaccharides and disialyllato-N-tetraose with the latter containing the NeuAc2-6 alpha Ga1NAc structure. The immunohistochemical double-staining technique provided further evidence of the difference between the epitope defined by the chimeric cACT19 antibody and the sialosyl-Tn epitope by illustrating complementary as well as noncomplementary expression of these two epitopes in different areas of colon carcinoma tissues. We also demonstrated that the chimeric cACT19 antibody displayed much more effective ADCC and CDC for the human OVAR3 tumor cells than the murine ACT19 antibody. Therefore, the mouse/human chimeric anti-colorectal carcinoma cACT19 antibody may prove to be useful in cancer immunotherapy in its own right, or especially when used in combination with the chimeric B72.3 antibody.
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PMID:Recombinant mouse/human chimeric anti-colorectal carcinoma antibody cACT19. 775 75

Human (U251, U87, U343) and rat glioma cell lines (C6, 9L) were examined by the reverse transcriptase-polymerase chain reaction and subsequent nucleotide sequencing analysis to see whether they express wild type (wt)-p53 or mutated form (mut)-p53 messages. Results showed that U87, U343, and C6 cells expressed wt-p53 messages whereas U251 and 9L cells expressed mut-p53 messages. All these cell lines were transfected with wt-p53 cDNA or the s-myc gene linked to the mouse mammary tumor virus (MMTV) promoter. Of several G418-resistant clones obtained from each transfection, a few expressed the s-Myc or wt-p53 proteins. Independent of mutations in the intrinsic p53 gene, the cellular growth in vitro and tumorigenicity in nude mice of these clones were drastically suppressed, the extent of suppression being correlated with the expression level of the transfected gene. Flow-cytometric analysis demonstrated that both p53 and s-Myc arrested the cell cycle at the G1/S boundary. These data suggest that these genes having negative effects on tumor cell proliferation could be used in gene therapy of gliomas, which are caused by alteration of the p53 gene or by some other genetic change.
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PMID:Negative effects of wild-type p53 and s-Myc on cellular growth and tumorigenicity of glioma cells. Implication of the tumor suppressor genes for gene therapy. 780 77

Retroviral-mediated cytokine gene transfer into tumor cells is a highly effective way of inducing tumor inhibition and immunity. We analyzed the tumorigenicity of C-26 murine colon carcinoma cells transduced with genes encoding the two subunits of murine interleukin-12 (IL-12) in a polycistronic retroviral vector and selected for resistance to G418 and for IL-12 production (30-80 pg/ml). BALB/c mice injected s.c., i.v. and intrasplenically with C-26/IL-12 cells from three different IL-12-producing clones showed delayed tumor onset as compared with mice injected with control NeoR-transduced or parental tumor cells. Although C-26/IL-12 tumor-bearing mice eventually died of lung metastasis, their survival time was twice as long as that of mice injected with control cells. In experiments with mice selectively depleted of natural killer (NK) cells before tumor cell injection, the time of tumor onset and survival of mice injected with C-26/IL-12 s.c. and i.v., respectively, was reduced. CD8+ T cell depletion had no effect on latency or survival, whereas removal of CD4+ T cells led to C-26/IL-12 tumor regression in about 40% of mice. Histological and immunocytochemical characterization of leukocytes infiltrating C-26/IL-12 tumors showed only slight infiltration with few T cells in non-depleted mice but abundant infiltration by CD8+ T cells and asialo-GM1+ NK cells in tumors of mice depleted of CD4+ T cells. The lack of CD8+ T cell infiltration is not due to a CD4-mediated suppression of their activation because irradiated C-26/IL-12 cells primed for the induction of a strong cytotoxic T lymphocyte response against C-26 parental cells and induced CD8+ effector cells that protected against C-26/IL-12 in a Winn assay. Rather, the results suggest that, although C-26/IL-12 cells injected in vivo stimulate both NK and CD8+ T cells, tumor infiltration by the latter is inhibited by CD4+ T cells.
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PMID:CD4 T cells inhibit in vivo the CD8-mediated immune response against murine colon carcinoma cells transduced with interleukin-12 genes. 784 24

Hypophosphorylated Rb, the product of the tumor suppressor gene associated with hereditary retinoblastoma, is thought to act as a suppressor of cell growth and proliferation during G1 phase. We investigated whether Rb expression was dependent on the expression level of the immediate early cell growth gene, c-fos, which is transiently expressed as cells re-enter G1 phase from quiescence. To explore the functional relationship between c-fos and Rb, a eukaryotic expression plasmid was constructed containing the c-fos gene under control of the SV40 promoter complex. This plasmid was co-transfected with plasmids encoding pRSV cat and G418 resistance, into HeLa S3 cells, and clonal populations of transfected (RSfos) cells selected. High levels of c-fos expression in transfected cells were confirmed by both western and northern blot. Rb protein content per cell was determined by flow cytometry using an Rb-specific primary antibody and a FITC-conjugated secondary antibody. Higher expression of Rb (2-6 fold/cell) in approximately 20% of RSfos transfected cells was observed in comparison to parental HeLa cells. Rb content per cell increased approximately 2-fold during the cell cycle in both parental HeLa cells and RSfos cell clones which overexpressed Rb. Rb accumulation occurred in a manner consistent with normal mass accumulation during the cell cycle. Overexpression of Rb in RSfos cells was also confirmed by western blot analysis. Because one possible function of RB may be to act as a brake on cell growth, it is possible that overexpression of RB acts as an inhibitory counter activity to overexpression of the growth promoting activity of c-fos. This possible balancing activity of Rb was further suggested when Rb protein expression levels were measured in different clonal lines of RSfos transfected cells overexpressing increasing levels of c-fos. Overexpression of Rb was proportional to the level of overexpression of c-fos in each clonal cell line. Such evidence suggests that Rb was proportional to the level of overexpression of c-fos in each clonal cell line. Such evidence suggests that Rb expression may be regulated in a manner which balances the transcription stimulatory effects of c-fos overexpression and its effects on the transcription of other genes during the cell cycle.
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PMID:Overexpression of c-fos induces expression of the retinoblastoma tumor suppressor gene Rb in transfected cells. 787 73

We have constructed and characterized a polyoma virus-based plasmid that is maintained as an autonomously replicating extrachromosomal element (episome) in mouse embryonic stem (ES) cells. Plasmid pMGD20neo contains the polyoma origin of replication harboring a mutated enhancer (PyF101), a modified polyoma early region that encodes the large tumor (T) antigen only, and a gene that confers resistance to G418 (neo). After transfection, the plasmid replicates in ES cells and is maintained as an extrachromosomal element in 15% of G418-resistant clones. Integration of the plasmid DNA is undetectable for at least 28 cell generations. In one clone, the transfected DNA persists unaltered as an episome at 10-30 copies per cell for at least 74 cell generations in the presence of G418. Cells that maintain the autonomously replicating plasmid can efficiently replicate and maintain a second plasmid that carries the polyoma origin of replication. Independent vector-containing ES cell lines showed no significant alteration of the karyotype, and two cell lines yielded several chimeric animals when introduced into blastocysts, suggesting that the presence of an episomal element and expression of polyoma large T do not eliminate the ES cells' ability to populate an embryo. This system offers an efficient means for manipulating and analyzing various aspects of gene expression in ES cells.
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PMID:Maintenance of an extrachromosomal plasmid vector in mouse embryonic stem cells. 787 70

Tumor-infiltrating lymphocytes (TILs), T lymphocytes associated with solid tumors that can be grown with interleukin (IL)-2 in vitro, preferentially accumulate at tumor sites after adoptive transfer. Therefore, TILs can be considered for use as cellular vehicles in gene therapy. We transduced melanoma TILs with the IL-2 gene and clarified functional characteristics of the TIL transductants. TILs transduced with 3'-end-truncated IL-2 gene (480 bp) produced high amounts of IL-2 detected in supernatants when compared to TILs transduced with the native IL-2 gene containing 3'-end adenine-thymidine (AT)-rich sequences (650 bp). The level of IL-2 in supernatants was higher with the addition of anti-Tac antibody (Ab) to block the consumption of IL-2 by the TILs. These TILs could proliferate autonomously in the absence of exogenous IL-2, and the proliferation of TILs could be completely blocked by anti-IL-2 Ab or anti-IL-2 receptor Ab. Thus TILs transduced with IL-2 gene can proliferate through the autocrine loop. However, the expression of IL-2 from TILs transduced with the IL-2 gene was downregulated after 2 to 3 weeks of G418 selection. Our study indicates the feasibility of transduction and expression of a truncated 480-bp IL-2 gene into TILs and the possibility of employing adoptive immunotherapy protocols using TILs modified with this IL-2 gene.
J Immunother Emphasis Tumor Immunol 1994 Nov
PMID:Enhanced interleukin-2 production in human tumor-infiltrating lymphocytes engineered by 3'-truncated interleukin-2 gene. 788 35

Human papillomavirus (HPV) 5 and HPV8 are often detected in skin cancers developed in patients suffering from epidermodysplasia verruciformis, as well as in skin cancers developed in immunosuppressed patients. In the present study, in order to examine the transforming activity of the HPV8E7 gene, the HPV8E7 and HPV8E6/E7 genes were cloned into the expression vector (pcD2-Y), under the SV40 enhancer/promoter to construct pcD2-8E7 and pcD2-8E6/E7, respectively. The E7 and E6/E7 genes of genital high-risk HPV16 were also cloned into pcD2-Y to construct pcD2-16E7 and pcD2-16E6/E7, respectively. They were tested for their ability to collaboratively transform primary rat embryo fibroblasts (REFs) with activated H-ras gene. Transfection experiments of REFs having an activated H-ras gene revealed that pcD2-8E7, as well as pcD2-16E7 and pcD2-16E6/E7, induced transformation of cells in G418-resistant colonies at efficiencies of 11.9%, 43.0% and 53.0%, respectively. Transformed cell lines induced by activated H-ras gene and pcD2-8E7 or pcD2-16E7 were named 8RE and 16RE cell lines, respectively. Tumor induction in syngeneic newborn rats by injected the 8RE cells was higher than that of the 16RE cells. In cytological and histological examination, the 8RE cell lines and their induced tumors were different from the 16RE cell lines and their induced tumors. The 8RE cell lines showed the characteristic transformation with efficient growth ability on plastic and colony formation in 0.3% soft agar. These results support the hypothesis that the HPV8E7 gene plays an important role in the carcinogenesis of skin cancers.
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PMID:[Experimental study on carcinogenesis by human papillomavirus type 8 E7 gene]. 792 81

The aim of our study was to examine the potential usefulness of transducing the protein kinase C-gamma (PKC-gamma) cDNA gene into tumor-specific T cells as a technique for facilitating the generation of large numbers of functional Ag-specific T for tumor therapy. Murine CD8+, F-MuLV gag-specific CTL clones, and CD4+, F-MuLV env-specific Th clones, as well as bulk-cultured T cell lines with defined Ag specificity to FBL-3, a Friend murine leukemia virus (F-MuLV)-induced tumor, were transduced with a retroviral vector pZipNeoPKC-gamma and selected in G418. The results demonstrated that PKC-gamma-transduced clones remained activated in culture, as evidenced by continued expression of up-regulated levels of IL-2R, which were as high after 6 mo in culture without Ag restimulation as 24 h after Ag stimulation. In vitro functional studies demonstrated that PKC-gamma-transduced CD8+ T cell clones maintained specific cytolytic activity to FBL-3, and PKC-gamma-transduced CD4+ T cell clones maintained specific proliferative activity to FBL-3 or F-MuLV Ag presented by irradiated syngeneic APC. Short-term bulk-cultured T cells specific to FBL-3 were also transduced and could be grown long term in vitro with maintenance of functional specificity. In vivo study showed that PKC-gamma-transduced CD4+ T cells were able to proliferate in response to Ag plus IL-2 stimulation in vivo in a similar pattern as the parental T cells. Therapy with adoptively transferred PKC-gamma-transduced T cell clones and lines into syngeneic mice, with or without FBL-3 tumor, showed that the PKC-gamma-transduced T cells were not tumorigenic and were effective in curing mice with disseminated FBL-3.
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PMID:Retroviral transduction of protein kinase C-gamma into tumor-specific T cells allows antigen-independent long-term growth in IL-2 with retention of functional specificity in vitro and ability to mediate tumor therapy in vivo. 793 May 83

Hamster-complement-C1s cDNA was inserted into an expression plasmid BCMGSNeo (BCMGSNeoHACS). BALB/c mouse fibroblast A31 cells, which do not produce C1s, were transfected with BCMGSNeoHACS and the transfectants were selected with G418. Normal C1s production by the transfectants was confirmed by Northern and immunoblot analysis and by an esterase assay. To examine the tumorigenicity of the transfectants, 1 x 10(6) cells were injected s.c. into 6-week-old BALB/c nu/nu mice. Three C1s cDNA transfectants (A3CS9, A3CS12, A3CS13) formed tumors whereas both A31 and A31 transfected with the vector alone (A3BCM1 and A3BCM3) did not. The tumors derived from the transfectants showed invasive growth, and many capillaries were observed in the tumors. A tumor derived from A3CS13 was examined immunohistochemically and found to be reactive with an anti-C1s monoclonal antibody. Tumor cells were cultured in vitro again and C1s secreted into the culture medium was examined by immunoblot analysis. C1s synthesized by the tumor cells derived from A3CS13 maintained its biological functions. Tumor cells derived from A3CS9 and A3CS12 cells, however, produced C1s having abnormal disulfide bonds.
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PMID:Tumorigenicity of BALB3T3 A31 cells transfected with hamster-complement-C1s cDNA. 802 91


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