Purification and NH2-terminal amino acid sequence of a T-cell-derived lymphokine with growth factor activity for B-cell hybridomas.

Snick, Jacques Van; Cayphas, Sylvie; Vink, Anne; Uyttenhove, Catherine; Coulie, Pierre G.; Rubira, Michael R.; Simpson, Robert J.

doi:10.1073/pnas.83.24.9679

Cited by 603 publications

(263 citation statements)

References 24 publications

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“…One thousand cells of the fibroblastic cell line Cl1D were added. Fourty-eight hours later, supernatants were collected and concentrations of induced IL-6 were determined with the 7TD1 bioassay as previously described [35]. Inhibitory titers correspond to the serum dilution that reduces the induced IL-6 secretion by 50%.…”

Section: Assesment Of Anti-il-17 Inhibitory Ab In Vitromentioning

confidence: 99%

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Development of an anti‐IL‐17A auto‐vaccine that prevents experimental auto‐immune encephalomyelitis

2006

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IL-17 has been associated with multiple inflammatory disorders such as rheumatoid arthritis, asthma and multiple sclerosis. As these diseases require long-term treatment we turned to an auto-vaccine strategy for IL-17 neutralization in vivo. Mouse IL-17A was covalently linked to ovalbumin and used to immunize C57BL/6 mice. This vaccine induced the production of antibodies that blocked IL-17A bioactivity in vitro but did not react with the other IL-17 isoforms, including IL-17F. As the half-life of the Ab titers after the last immunogen administration was approximately 4 months, the vaccine provides for long lasting and selective inhibition of IL-17A activity in vivo. A monoclonal Ab (mAb) derived from these mice showed the same specificity for IL-17A. To test the ability of the vaccine to confer protection against an IL-17-dependent disorder, SJL mice were vaccinated with IL-17-OVA and encephalomyelitis (EAE) was induced by proteolipid protein (PLP) peptide 139-151. Vaccinated mice were completely protected against the disease. The above-mentioned anti-IL-17A mAb also prevented EAE development. The absence of clinical symptoms contrasted with unaltered PLP-induced cytokine production in vitro and unmodified anti-PLP IgG titers and isotypes. These results suggest that an anti-IL-17A auto-vaccine offers new perspectives for therapy of autoimmune diseases.

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Section: Assesment Of Anti-il-17 Inhibitory Ab In Vitromentioning

confidence: 99%

“…IFN-c, IL-13 and IL-17 concentrations were measured by ELISA (R&D). IL-6 and IL-9 activities were measured by bioassays [35,39].…”

Section: Assesment Of Anti-il-17 Inhibitory Ab In Vitromentioning

confidence: 99%

Development of an anti‐IL‐17A auto‐vaccine that prevents experimental auto‐immune encephalomyelitis

2006

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“…A murine IL-6-dependent hybridoma cell line, 7TD1 (Van Snick et al 1986), was cultured in RPMI 1640 medium (Nissui Pharma, Tokyo, Japan) supplemented with 10% FBS (Iwaki, Tokyo, Japan) and 2 ng/ml of murine IL-6 (Genzyme/Techne, Cambridge, MA). A retroviral packaging cell line, Plat-E (Morita et al 2000), was cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% FBS, 1 lg/ml puromycin (Sigma, St. Louis, MO) and 10 lg/ml blasticidin (Kaken Pharmaceutical, Tokyo, Japan).…”

Section: Cell Culturementioning

confidence: 99%

Growth control of hybridoma cells with an artificially induced EpoR-gp130 heterodimer

et al. 2006

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IL-6 has been known to modulate the growth of many hybridoma cells and also promote resultant antibody productivity. However, IL-6 is so expensive that the use of IL-6-dependent hybridomas for industrial antibody production is not practical. In this study, we aimed at designing antibody/gp130 and antibody/EpoR chimeras which could tightly control cell growth in response to more affordable cognate antigen. Retroviral vectors encoding V H or V L region of anti-hen egg lysozyme (HEL) antibody HyHEL-10 tethered to a pair of extracellular D2/transmembrane domains of erythropoietin receptor (EpoR) and cytoplasmic domains of either EpoR or gp130, were constructed, and a homodimeric or a heterodimeric pair of chimeric receptor combinations (V H -gp130 and V L -gp130 or V H -gp130 and V L -EpoR) were expressed in an IL-6-dependent hybridoma 7TD1. The chimeric receptor-derived growth signal was observed in both combinations, while some residual growth signal was observed in the absence of HEL. To reduce interchain interaction between the two receptor chains, we introduced mutations to the transmembrane domain of both chimera combinations. Consequently, the heterodimeric combination of V H -gp130 and V L -EpoR showed clear HEL-dependent cell growth, while the homodimeric combination of V H -gp130 and V L -gp130 showed reduced cell growth in the absence of HEL. This is the first report that an EpoR-gp130 cytoplasmic domain heterodimer could transduce a growth signal in hybridoma cells, indicating tight and economical growth control of hybridoma cells via our chimeric receptors.

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“…The concentrations of the proteins were determined by amino acid anaIysis using a Beckman amino acid analyzer (model 6300) equipped with a model 7000 integrator, as described (Simpson et al, 1986).…”

Section: Protein Estimationmentioning

confidence: 99%

“…Murine IL-6, pMC5, and pMC5H titers were determined in a hybridoma growth factor assay, as previously described (Van Snick et al, 1986). Briefly, IL-6-dependent murine hybridoma 7TD1 cells (2,000 cells/microwell) were incubated with serial dilutions of the test fractions in a final volume of 0.2 mL.…”

Section: Hybridoma Growth Factor Assaymentioning

confidence: 99%

Role of the C‐terminus in the activity, conformation, and stability of interleukin‐6

et al. 1993

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Two murine interleukin-6 (mIL-6) variants were constructed using the polymerase chain reaction (PCR), one lacking the last five residues (183-187) at the C-terminus (pMC5) and another with the last five residues of mIL-6 substituted by the corresponding residues of human IL-6 (pMC5H). The growth stimulatory activity of pMC5 on the mouse hybridoma cell line 7TD1 was <0.05% of mIL-6, whereas pMC5H and mIL-6 were equipotent. The loss of biological activity of pMCS correlated with its negligible receptor binding affinity on 7TD1 cells, while the binding of pMC5H was comparable to that of mIL-6. Both pMC5 and pMCSH, like mIL-6, failed to interact with recombinant soluble human IL-6 receptor when assayed by surface plasmon resonance-based biosensor analysis. These studies suggest that the C-terminal seven amino acids of human IL-6, alone, do not define species specificity for receptor binding.A variety of biophysical techniques, as well as the binding of a conformational-specific monoclonal antibody, indicated that the global fold of the mIL-6 variants was similar to that of mIL-6, although small changes in the NMR spectra, particularly for pMC5, were observed. Some of these changes involved residues widely separated in the primary structure. For instance, interactions involving Tyr-22 were influenced by the C-terminal amino acids suggesting that the N-and C-termini of mIL-6 are in close proximity. Equilibrium unfolding experiments indicated that pMC5 was 0.8 kcal/mol less stable than mIL-6, whereas pMCSH was 1.4 kcal/mol more stable. These studies emphasize the structural importance of the C-terminal amino acids of IL-6 and suggest that truncation or mutation of this region could lead to small but significant alterations in other regions of the molecule.

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Purification and NH2-terminal amino acid sequence of a T-cell-derived lymphokine with growth factor activity for B-cell hybridomas.

Cited by 603 publications

References 24 publications

Development of an anti‐IL‐17A auto‐vaccine that prevents experimental auto‐immune encephalomyelitis

Development of an anti‐IL‐17A auto‐vaccine that prevents experimental auto‐immune encephalomyelitis

Growth control of hybridoma cells with an artificially induced EpoR-gp130 heterodimer

Role of the C‐terminus in the activity, conformation, and stability of interleukin‐6

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