期刊
TRANSGENIC RESEARCH
卷 32, 期 1-2, 页码 121-133出版社
SPRINGER
DOI: 10.1007/s11248-023-00341-1
关键词
Multiple sclerosis; Nicotiana benthamiana; Asparaginyl endopeptidases; Peptide; Recombinant; Plant molecular farming; Cyclotide
Multiple sclerosis (MS) is a debilitating disease that requires prolonged treatment. The experimental therapeutic [T20K]kB1, a mutant of a plant peptide, shows promise for oral dosing and stability due to its cyclic structure. This study demonstrates the production of [T20K]kB1 in the Nicotiana benthamiana plant, providing a sustainable and cost-effective production method for cyclotide-based therapeutics.
Multiple sclerosis (MS) is a debilitating disease that requires prolonged treatment with often severe side effects. One experimental MS therapeutic currently under development is a single amino acid mutant of a plant peptide termed kalata B1, of the cyclotide family. Like all cyclotides, the therapeutic candidate [T20K]kB1 is highly stable as it contains a cyclic backbone that is cross-linked by three disulfide bonds in a knot-like structure. This stability is much sought after for peptide drugs, which despite exquisite selectivity for their targets, are prone to rapid degradation in human serum. In preliminary investigations, it was found that [T20K]kB1 retains oral activity in experimental autoimmune encephalomyelitis, a model of MS in mice, thus opening up opportunities for oral dosing of the peptide. Although [T20K]kB1 can be synthetically produced, a recombinant production system provides advantages, specifically for reduced scale-up costs and reductions in chemical waste. In this study, we demonstrate the capacity of the Australian native Nicotiana benthamiana plant to produce a structurally identical [T20K]kB1 to that of the synthetic peptide. By optimizing the co-expressed cyclizing enzyme, precursor peptide arrangements, and transgene regulatory regions, we demonstrate a [T20K]kB1 yield in crude peptide extracts of similar to 0.3 mg/g dry mass) in whole plants and close to 1.0 mg/g dry mass in isolated infiltrated leaves. With large-scale plant production facilities coming on-line across the world, the sustainable and cost-effective production of cyclotide-based therapeutics is now within reach.
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