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)

CD40/CD40L interactions play a pivotal role in T cell activation, and take part in many physiologic and pathologic procedures and different levels. In this article, stable CHO transformants secreting human CD40-Ig fusion protein were established through transfection and selection with Lipofectamaine and G418, respectively. In order to obtain great valume of recombinant protein, big batch serum-free cultures of engineered CHO cells were performed in roller-bottle using CHO-II-SFM medium. After cultures, the cell-culture supernatants were harvested, concentrated through ultra-filtration, and finally purified by affinity choromatography with Protein G Sepharose Fast Flow. Human peripheral bloods were collected freshly and seperated with Ficoll, CFU-T was cultured in semi-solid culture system with peripheral blood mononuclear cells (PBMNC). Effect of human CD40-Ig fusion protein on the formation of CFU-T was observed in vitro. The results showed that the yield of human CD40-Ig fusion protein was 30 mg in total 3 liter CHO-II-SFM culture supernatant, and it supposed that the expression level of CD40-Ig in CHO cells was more than 10 micro g/ml. The purity of purified fusion protein is above 95%. Furthermore, compared with human IgG, human CD40-Ig fusion protein significantly inhibited the formation of CFU-T at dose 0.25, 1.0, 4.0, and 10 micro g/ml, it lays a good foundation to evaluate its potential functions in vivo.
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PMID:[Purification of Human CD40-Ig Fusion Protein and Its Effect on Human Peripheral Blood CFU-T Formation In Virto] 1257 91

To modulate alloreactivity after hematopoietic stem cell (HSC) transplantation, suicide gene-expressing donor T cells can be administered with an allogeneic T cell-depleted HSC graft. Immune competence of such cells is a critical issue. We have examined the impact of our ex vivo gene transfer protocol (12-day culture period including CD3/IL-2 activation, retrovirus-mediated gene transfer, and G418-based selection) on the phenotype and functional properties of gene-modified cells (GMC). GMC were compared with control cells that had been cultured in parallel with GMC, but nontransduced and nonselected, as well as with peripheral blood mononuclear cells (PBMC). Our data show that phenotypical modifications are similar in control cells and GMC, demonstrating that alterations result from the 12-day culture rather than from the transduction and/or selection process itself. Such modifications include a reversal of CD4/CD8 ratio, activated phenotype (increased expression of CD45RO, CD95, and HLA-DR), and acquisition or increased expression of co-stimulatory molecules (CD80, CD86, and CD40). This led to an enhanced allostimulating potential of GMC, as compared with resting T cells, when used as stimulating cells in mixed lymphocyte reactions. Conversely, when using them as responder cells in mixed lymphocyte reactions, GMC exhibited a rapid loss of alloreactivity that resulted both from culture-dependent and from transduction and/or selection-dependent events. In conclusion, the retrovirus-mediated gene transfer can be associated with major phenotypical and functional alterations that could have strong clinical implications (increased immunogenicity, reduced anti-leukemic effect). Thus, future T cell expansion protocols should try to improve not only cell expansion or gene transfer efficiency, but also T cell functions.
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PMID:Influence of ex vivo expansion and retrovirus-mediated gene transfer on primary T lymphocyte phenotype and functions. 1259 Jul 8