Expression of bioactive single-chain murine IL-12 in transgenic plants.
Interleukin-12 (IL-12), an important immunomodulator for cell-mediated immunity, shows significant potential as a vaccine adjuvant and anticancer therapeutic. However, its clinical application is limited in part by lack of an effective bioproduction system for this complex heterodimeric glycoprotein. Transgenic plants show promise as scalable bioproduction platforms for challenging biopharmaceutical proteins. To test the potential of plants to effectively produce bioactive IL-12, we developed transgenic tobacco plant lines and derived root cultures yielding high levels of mouse IL-12 (MuIL-12). Functional IL-12 is a heterodimer consisting of two disulfide-linked subunits, p35 and p40. To ensure the stoichiometric expression and assembly of p35 and p40, we expressed a single-chain version of MuIL-12. Plant-derived single-chain MuIL-12 was characterized and purified for in vitro bioactivity assays. Our results demonstrated precise cleavage of the endogenous mouse p40 signal peptide in plants as well as addition of N-linked glycans. Plant-derived MuIL-12 triggered induction of interferon-gamma (IFN-gamma) secretion from mouse splenocytes and stimulated splenocyte proliferation with comparable activities to those observed for commercially available animal cell-derived MuIL-12. These studies indicate that plants produce fully functional MuIL-12 at levels compatible with commercial production and may serve as an effective bioproduction platform for bioactive IL-12s from other species for human or veterinary vaccine and therapeutic applications.
Liu
Jianyun
J
Arkansas Biosciences Institute, Arkansas State University, Jonesboro, AR 72467, USA.
Dolan
Maureen C
MC
Reidy
Michael
M
Cramer
Carole L
CL
D016960:Agrobacterium tumefaciens; D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000276:immunology; D006801:Humans; D007167:Immunotherapy; D018664:Interleukin-12 / Q000235:genetics* / Q000276:immunology / Q000627:therapeutic use; D053765:Interleukin-12 Subunit p35 / Q000235:genetics / Q000276:immunology / Q000627:therapeutic use; D053762:Interleukin-12 Subunit p40 / Q000235:genetics / Q000276:immunology / Q000627:therapeutic use; D051379:Mice; D008810:Mice, Inbred C57BL; D009369:Neoplasms / Q000276:immunology / Q000628:therapy; D018517:Plant Roots / Q000276:immunology / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics* / Q000276:immunology / Q000627:therapeutic use; D014026:Nicotiana / Q000235:genetics* / Q000276:immunology
D053765:Interleukin-12 Subunit p35; D053762:Interleukin-12 Subunit p40; D011994:Recombinant Proteins; D018664:Interleukin-12