Construction of an expression vector for improved secretion of canine IL-18

Sugiura, Kikuya; Akazawa, Takashi; Fujimoto, Mai; Wijewardana, Viskam; Mito, Kai; Hatoya, Shingo; Taketani, Shigeru; Komori, Masaharu; Inoue, Norimitsu; Inaba, Toshio

doi:10.1016/j.vetimm.2008.07.011

Cited by 5 publications

(5 citation statements)

References 8 publications

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“…To examine the bioactivity of the cIL-18 secreted from the transfected cells, the culture supernatant was cocultured with canine PBMCs, and the amount of canine IFN-γ produced by the PBMCs was measured. The supernatants of the cells expressing entire cIL-18 induced little IFN-γ production compared with the control, whereas the supernatants of cells expressing cIL-12ss–cIL-18 slightly enhanced the induction efficiency, as in the previous report [ 37 ]. Interestingly, the supernatant of cells transfected with pFuse–hIL2ss–cIL18 induced significant IFN-γ production ( Fig.…”

Section: Resultssupporting

confidence: 80%

“…Porcine IL-18 fused to the baculovirus gp67-encoded signal sequence displayed bioactivity in a baculovirus system [ 22 ]. In the present study, we evaluated the utility of the human IL-2 signal sequence and demonstrated that it is more efficient than the previously reported canine IL12-p40 signal sequence [ 37 ] and that it can be used to express both canine and mouse IL-18.…”

Section: Discussionmentioning

confidence: 91%

“…Several reports have indicated that the addition of a signal sequence derived from various secreted proteins enhances the extracellular secretion of bioactive IL-18 [ 14 , 24 , 29 ]. It has been reported that the fusion of the canine IL12-p40 signal sequence (cIL-12p40ss) to mature canine IL-18 (cIL-18) enhanced the secretion efficiency of bioactive IL-18 [ 37 ].…”

mentioning

confidence: 99%

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Construction of an Expression System for Bioactive IL-18 and Generation of Recombinant Canine Distemper Virus Expressing IL-18

Liu

Sato

Hamana

et al. 2014

The Journal of Veterinary Medical Science

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Interleukin 18 (IL-18) plays an important role in the T-helper-cell type 1 immune response against intracellular parasites, bacteria and viral infections. It has been widely used as an adjuvant for vaccines and as an anticancer agent. However, IL-18 protein lacks a typical signal sequence and requires cleavage into its mature active form by caspase 1. In this study, we constructed mammalian expression vectors carrying cDNA encoding mature canine IL-18 (cIL-18) or mouse IL-18 (mIL-18) fused to the human IL-2 (hIL-2) signal sequence. The expressed proIL-18 proteins were processed to their mature forms in the cells. The supernatants of cells transfected with these plasmids induced high interferon-γ production in canine peripheral blood mononuclear cells or mouse splenocytes, respectively, indicating the secretion of bioactive IL-18. Using reverse genetics, we also generated a recombinant canine distemper virus that expresses cIL-18 or mIL-18 fused to the hIL-2 signal sequence. As expected, both recombinant viruses produced mature IL-18 in the infected cells, which secreted bioactive IL-18. These results indicate that the signal sequence from hIL-2 is suitable for the secretion of mature IL-18. These recombinant viruses can also potentially be used as immunoadjuvants and agents for anticancer therapies in vivo.

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Section: Resultssupporting

confidence: 80%

Section: Discussionmentioning

confidence: 91%

mentioning

confidence: 99%

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Construction of an Expression System for Bioactive IL-18 and Generation of Recombinant Canine Distemper Virus Expressing IL-18

Liu

Sato

Hamana

et al. 2014

The Journal of Veterinary Medical Science

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“…Isolation of RNA, cDNA synthesis, and PCR amplification was carried out as described previously . In the PCR amplification, primers were designed to amplify a particular nucleotide sequence of mouse IFNγ and CD40 ligand (CD40L) cDNA (NM_008337.4 and NM_011616.2).…”

Section: Methodsmentioning

confidence: 99%

Manipulation of the tumor microenvironment by cytokine gene transfection enhances dendritic cell‐based immunotherapy

et al. 2019

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The tumor microenvironment strongly influences clinical outcomes of immunotherapy.By transfecting genes of relevant cytokines into tumor cells, we sought to manipulate the microenvironment so as to elicit activation of T helper type 1 (Th1) responses and the maturation of dendritic cells (DCs). Using a synthetic vehicle, the efficiency of in vivo transfection of GFP-cDNA into tumor cells was about 7.5% by intratumoral injection and about 11.5% by intravenous injection. Survival was significantly improved by both intratumoral and intravenous injection of the plasmid containing cDNA of interferongamma, followed by intratumoral injection of DCs presenting the tumor antigens. Also, tumor growth was inhibited by these treatments. A more significant effect on survival and tumor growth inhibition was observed following injection of the plasmid containing cDNA of CD40 ligand, which is a potent inducer of DC-maturation. Furthermore, the co-injection of both IFNγ-and CD40 ligand-encoding cDNA-plasmids, followed by DC treatment, gave rise to further marked and enhancement, including 100% survival and more than 50% complete remission. This treatment regimen elicited significant increases in mature DCs and types of cells contributing to Th1 responses, and significant decreases in immune suppressor cells in the tumor. In the spleen, the treatment significantly increased activities of tumor-specific killer and natural killer cells, but no alteration was observed in mature DCs or suppressor cells. These results indicate that transfection of these cytokine genes into tumor cells significantly alter the tumor microenvironment and improve the therapeutic results of DC-based immunotherapy. K E Y W O R D Scancer immunotherapy, CD40 ligand, gene therapy, immunity, interferon-gamma, synthetic vehicle 6 |

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“…Isolation of RNA, cDNA synthesis and PCR amplification was carried out as described previously (Sugiura et al, 2008). In the PCR amplification, primers were designed to amplify a particular nucleotide sequence of canine IFN␥ cDNA (accession number NM 001003174).…”

Section: Construction and Transfection Of Canine Ifn Expression Vectormentioning

confidence: 99%

Enhancement of anti-tumor immune responses by transfection of IFNγ gene into tumor using a novel type synthetic vector

Wijesekera

Sugiura

Yuba

et al. 2014

Veterinary Immunology and Immunopathology

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Construction of an expression vector for improved secretion of canine IL-18

Cited by 5 publications

References 8 publications

Construction of an Expression System for Bioactive IL-18 and Generation of Recombinant Canine Distemper Virus Expressing IL-18

Construction of an Expression System for Bioactive IL-18 and Generation of Recombinant Canine Distemper Virus Expressing IL-18

Manipulation of the tumor microenvironment by cytokine gene transfection enhances dendritic cell‐based immunotherapy

Enhancement of anti-tumor immune responses by transfection of IFNγ gene into tumor using a novel type synthetic vector

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