Human GM-CSF: Molecular Cloning of the Complementary DNA and Purification of the Natural and Recombinant Proteins

Wong, Gordon G.; Witek, JoAnn S.; Temple, Patricia A.; Wilkens, K.; Leary, Anne C.; Luxenberg, Deborah; Jones, Simon; Brown, Eugene L.; Kay, Robert M.; Orr, Elizabeth C.; Shoemaker, Charles B.; Golde, David W.; Kaufman, Randal J.; Hewick, Rodney M.; Wang, Elizabeth A.; Clark, Steven C.

doi:10.1126/science.3923623

Cited by 1,173 publications

(319 citation statements)

References 44 publications

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“…BCG can be formulated in nanomicroparticle aerosol form that is effectively delivered to the lung and induces significant protective efficacy in animal models, and such an approach may further improve the protective effect on BCG:GM-CSF against pulmonary M. tuberculosis infection [34]. Continued evaluation in human trials will require the engineering of human GM-CSF into recombinant BCG strains, since cross-species receptor binding or biological activity does not exist between murine and human homologues of GM-CSF [35,36]. Nevertheless, the present study provides an excellent platform for the development of a safe and effective replacement for the current BCG vaccine to protect against tuberculosis.…”

Section: Discussionmentioning

confidence: 99%

Modulation of pulmonary DC function by vaccine‐encoded GM‐CSF enhances protective immunity against Mycobacterium tuberculosis infection

et al. 2009

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The rational design of new vaccines engineered to target key components of the host immune response is crucial to aid control of important infectious diseases such as tuberculosis. In this report, we determined whether modifying the function of pulmonary APC could improve protection against infection with Mycobacterium tuberculosis. Targeted delivery to the lung of the cytokine GM‐CSF, expressed by the Mycobacterium bovis BCG vaccine strain, increased pulmonary DC numbers and secretion of the immunoregulatory cytokine IL‐12, compared with parental BCG immunization. This impact on APC number by BCG:GM‐CSF resulted in accelerated priming of antigen‐specific CD4+ T cells in the mediastinal lymph nodes and increased migration of activated CD4+ T cells into the lung. i.n. administration of BCG:GM‐CSF resulted in significantly increased protection against M. tuberculosis infection compared with mice vaccinated with BCG alone. BCG:GM‐CSF exhibited an improved safety profile, as immunodeficient RAG1−/− mice vaccinated i.n. with BCG:GM‐CSF survived significantly longer than control BCG‐vaccinated mice. These data demonstrate that manipulating immune cells in the lung by BCG‐based delivery of GM‐CSF can assist the development of protective mucosal immunity against pulmonary bacterial infection.

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Section: Discussionmentioning

confidence: 99%

Modulation of pulmonary DC function by vaccine‐encoded GM‐CSF enhances protective immunity against Mycobacterium tuberculosis infection

et al. 2009

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“…Transfection of the 2.5 kb murine Gas6 cDNA, cloned into the P91023 (B) vector (Wong et al, 1985) was performed using 5 mg of plasmid using the calcium phosphate procedure. Positive clones were selected in medium containing dialyzed FCS and lacking hypoxanthine and thymidine.…”

Section: Production Of Recombinant Murine Gas6mentioning

confidence: 99%

Signaling through the ARK tyrosine kinase receptor protects from apoptosis in the absence of growth stimulation

et al. 1997

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ARK (AXL) is the prototype of a distinctive family of receptor tyrosine kinases which contain in their extracellular domains features reminiscent of cell adhesion molecules. ARK is capable of homophilic binding, which results in a degree of receptor activation, but can also be activated by a heterophilic ligand, Gas6, a member of the family of vitamin K dependent proteins that is preferentially expressed in quiescent cells. Since a number of tissues and cell lines express both ARK and Gas6, we studied the e ect of endogenous and exogenous Gas6 on the phenotype of ARK expressing cells. Here we show that constitutive expression of Gas6 in an NIH3T3 cell line that does not spontaneously express this protein does not result in cell transformation or uncontrolled growth, but protects from apoptosis induced by serum deprivation. Recombinant exogenous Gas6 was also capable of protecting cells from apoptosis at concentrations that did not result in signi®cant induction of DNA synthesis. Activation of ARK phosphorylation and a weak but signi®cant induction of MAP kinase activity accompanied the increased survival of cells treated with Gas6. The antiapoptotic e ect of ARK signaling was con®rmed by studies using ®broblasts from ARK knock-out mice, that showed that the absence of ARK resulted in higher levels of serum deprivation-induced apoptosis, that could not be rescued by the addition of Gas6. Interestingly ARK signaling protects from apoptosis induced by serum deprivation, myc overexpression, or by TNFa but not from u.v. irradiation or Staurosporine. These results suggest that a major function of Gas6 ± ARK signaling is that of increasing cell survival under conditions which do not allow cell proliferation.

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“…The HSE element -5Ј-AGAATGTTCTAGAAG-3Ј was synthesised as a consensus sequence which confers heat shock and HSF-1 responsiveness on heterologous genes as described. 9,10 To assess promoter/enhancer strengths, plasmids were constructed using standard techniques such that different promoters and promoter/enhancer fragments were placed upstream either of the chloramphenicol acetyl transferase (CAT) gene, 7 the human GM-CSF gene 27 or the cDNA of the gibbon ape leukaemia virus fusogenic membrane glycoprotein (GALV-FMG). 2,4 Levels of gene expression were assayed using either CAT assays, 7 by measuring levels of human GM-CSF secreted from the plasmids by ELISA (Pharmingen, San Diego, CA, USA) or by semi-quantitative rtPCR as described below.…”

Section: Plasmids and Cell Linesmentioning

confidence: 99%

A transcriptional feedback loop for tissue-specific expression of highly cytotoxic genes which incorporates an immunostimulatory component

et al. 2001

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Human GM-CSF: Molecular Cloning of the Complementary DNA and Purification of the Natural and Recombinant Proteins

Cited by 1,173 publications

References 44 publications

Modulation of pulmonary DC function by vaccine‐encoded GM‐CSF enhances protective immunity against Mycobacterium tuberculosis infection

Modulation of pulmonary DC function by vaccine‐encoded GM‐CSF enhances protective immunity against Mycobacterium tuberculosis infection

Signaling through the ARK tyrosine kinase receptor protects from apoptosis in the absence of growth stimulation

A transcriptional feedback loop for tissue-specific expression of highly cytotoxic genes which incorporates an immunostimulatory component

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