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德國亥姆霍茲中心感染研究所25日發表公報說，該所發現了能令胰島素前體產量翻倍的方法，并決定不注冊而公開所有技術細節，以使更多的、尤其是發展中國家的糖尿病患者能夠得到藥物治療。
目前，臨床治療中使用的胰島素制劑是由轉化胰島素前體制成的，胰島素前體主要在大腸桿菌或釀酒酵母菌中生產。該所在此基礎上發現了一種新方法，能令胰島素前體的產量翻倍，從而降低胰島素生產的成本。這一方法生產的胰島素經證實能夠安全地在人體應用。
公報說，糖尿病已經不再是以往認為的“富貴病”。發展中國家的患者人數增長迅速，但許多貧窮國家患者因買不起藥而飽受折磨甚至失去生命，其重要原因是許多新藥和方法申請了保護，導致治療費用高昂。亥姆霍茲中心感染研究所這一研究項目的目標就是降低胰島素生產成本，讓更多糖尿病患者受益。
上海勁馬生物（）推薦原文出處：
Microbial Cell Factories doi:10.1186/1475-2859-9-31
Application of simple fed-batch technique to high-level secretory production of insulin precursor using Pichia pastoris with subsequent purification and conversion to human insulin
Chandrasekhar Gurramkonda , Sulena Polez , Natasa Skoko , Ahmad Adnan , Thomas Gabel , Dipti Chugh , Sathyamangalam Swaminathan , Navin Khanna , Sergio Tisminetzky and Ursula Rinas
Background
The prevalence of diabetes is predicted to rise significantly in the coming decades. A recent analysis projects that by the year 2030 there will be ~366 million diabetics around the world, leading to an increased demand for inexpensive insulin to make this life-saving drug also affordable for resource poor countries.
Results
A synthetic insulin precursor （IP）-encoding gene, codon-optimized for expression in P. pastoris, was cloned in frame with the Saccharomyces cerevisiae alpha-factor secretory signal and integrated into the genome of P. pastoris strain X-33. The strain was grown to high-cell density in a batch procedure using a defined medium with low salt and high glycerol concentrations. Following batch growth, production of IP was carried out at methanol concentrations of 2 g L-1, which were kept constant throughout the remaining production phase. This robust feeding strategy led to the secretion of ~3 gram IP per liter of culture broth （corresponding to almost 4 gram IP per liter of cell-free culture supernatant）. Using immobilized metal ion affinity chromatography （IMAC） as a novel approach for IP purification, 95% of the secreted product was recovered with a purity of 96% from the clarified culture supernatant. Finally, the purified IP was trypsin digested, transpeptidated, deprotected and further purified leading to ~1.5 g of 99% pure recombinant human insulin per liter of culture broth.
Conclusions
A simple two-phase c*tion process composed of a glycerol batch and a constant methanol fed-batch phase recently developed for the intracellular production of the Hepatitis B surface antigen was adapted to secretory IP production. Compared to the highest previously reported value, this approach resulted in an ~2 fold enhancement of IP production using Pichia based expression systems, thus significantly increasing the efficiency of insulin manufacture.
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