Optimization and revision of the production process of the Necator americanus glutathione S-transferase 1 (Na-GST-1), the lead hookworm vaccine recombinant protein candidate.
Optimization and revision of the production process of the Necator americanus glutathione S-transferase 1 (Na-GST-1), the lead hookworm vaccine recombinant protein candidate.
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DOI：
10.4161/hv.28872
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发表时间：
2014
影响因子：
4.8
通讯作者：
Elena Bottazzi M
中科院分区：
文献类型：
--
作者：
Curti E;Seid CA;Hudspeth E;Center L;Rezende W;Pollet J;Kwityn C;Hammond M;Matsunami RK;Engler DA;Hotez PJ;Elena Bottazzi M
在非洲、亚洲和美洲的发展中国家，人类钩虫Necator americanus的感染是贫血和残疾的主要原因。为了在资源贫乏的环境中预防儿童钩虫病，萨宾疫苗研究所和德克萨斯儿童医院疫苗开发中心正在开发一种重组疫苗，这是一个产品开发伙伴关系（PDP）。在此之前，我们报道了一种非常有希望的钩虫疫苗候选物Na-GST-1的表达和纯化。在巴斯德毕赤酵母（酵母）中表达的美洲人谷胱甘肽S-转移酶，这导致以20 L规模生产1.5 g 95%纯的重组蛋白。Na-GST-1的产率和纯度足以用于早期中试生产和初始1期临床试验。然而，基于允许大规模疫苗接种所需的剂量数和尽可能便宜地递送疫苗的潜在目标，高度期望以尽可能低的成本表达重组抗原的较高产量。在这里，我们报告了对在巴斯德毕赤酵母中表达的抗原的发酵（上游过程）和重组蛋白的纯化（下游过程）的修改，其允许Na-GST-1纯化蛋白的最终产率提高2-3倍。主要改进包括上游工艺变化，例如在甲醇诱导期间添加山梨醇脉冲和共进料，以及将诱导阶段延长至约96小时;下游工艺变化包括将UFDF改性为具有10 kDa分子量截止值（MWCO）的平板，利用与原始步骤洗脱相反的梯度洗脱调节离子交换色谱步骤的容量，并改变疏水相互作用色谱条件。描述了目标Na-GST-1及其在Alhydrogel®上的制剂的完整工艺以及纯度和稳定性特征。
Infection by the human hookworm Necator americanus is a leading cause of anemia and disability in the developing countries of Africa, Asia, and the Americas. In order to prevent childhood hookworm disease in resource poor settings, a recombinant vaccine is under development by the Sabin Vaccine Institute and Texas Children’s Hospital Center for Vaccine Development, a Product Development Partnership (PDP). Previously, we reported on the expression and purification of a highly promising hookworm vaccine candidate, Na-GST-1, an N. americanus glutathione s-transferase expressed in Pichia pastoris (yeast), which led to production of 1.5 g of 95% pure recombinant protein at a 20L scale. This yield and purity of Na-GST-1 was sufficient for early pilot manufacturing and initial phase 1 clinical testing. However, based on the number of doses which would be required to allow mass vaccination and a potential goal to deliver a vaccine as inexpensively as possible, a higher yield of expression of the recombinant antigen at the lowest possible cost is highly desirable. Here we report on modifications to the fermentation (upstream process) of the antigen expressed in P. pastoris, and to the purification (downstream process) of the recombinant protein that allowed for a 2–3-fold improvement in the final yield of Na-GST-1 purified protein. The major improvements included upstream process changes such as the addition of a sorbitol pulse and co-feed during methanol induction as well as an extension of the induction stage to approximately 96 hours; downstream process changes included modifying the UFDF to flat sheet with a 10 kDa Molecular Weight cut-off (MWCO), adjusting the capacity of an ion-exchange chromatography step utilizing a gradient elution as opposed to the original step elution, and altering the hydrophobic interaction chromatography conditions. The full process, as well as the purity and stability profiles of the target Na-GST-1, and its formulation on Alhydrogel®, is described.
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影响因子：
5.5
作者：
Hotez, Peter J.;Diemert, David;Bacon, Kristina M.;Beaumier, Coreen;Bethony, Jeffrey M.;Bottazzi, Maria Elena;Brooker, Simon;Couto, Artur Roberto;Freire, Marcos da Silva;Homma, Akira;Lee, Bruce Y.;Loukas, Alex;Loblack, Marva;Morel, Carlos Medicis;Oliveira, Rodrigo Correa;Russell, Philip K.
通讯作者：
Russell, Philip K.
DOI：
10.1016/s0140-6736(12)61729-2
发表时间：
2012-12-15
期刊：
Lancet (London, England)
影响因子：
--
作者：
Vos T;Flaxman AD;Naghavi M;Lozano R;Michaud C;Ezzati M;Shibuya K;Salomon JA;Abdalla S;Aboyans V;Abraham J;Ackerman I;Aggarwal R;Ahn SY;Ali MK;Alvarado M;Anderson HR;Anderson LM;Andrews KG;Atkinson C;Baddour LM;Bahalim AN;Barker-Collo S;Barrero LH;Bartels DH;Basáñez MG;Baxter A;Bell ML;Benjamin EJ;Bennett D;Bernabé E;Bhalla K;Bhandari B;Bikbov B;Bin Abdulhak A;Birbeck G;Black JA;Blencowe H;Blore JD;Blyth F;Bolliger I;Bonaventure A;Boufous S;Bourne R;Boussinesq M;Braithwaite T;Brayne C;Bridgett L;Brooker S;Brooks P;Brugha TS;Bryan-Hancock C;Bucello C;Buchbinder R;Buckle G;Budke CM;Burch M;Burney P;Burstein R;Calabria B;Campbell B;Canter CE;Carabin H;Carapetis J;Carmona L;Cella C;Charlson F;Chen H;Cheng AT;Chou D;Chugh SS;Coffeng LE;Colan SD;Colquhoun S;Colson KE;Condon J;Connor MD;Cooper LT;Corriere M;Cortinovis M;de Vaccaro KC;Couser W;Cowie BC;Criqui MH;Cross M;Dabhadkar KC;Dahiya M;Dahodwala N;Damsere-Derry J;Danaei G;Davis A;De Leo D;Degenhardt L;Dellavalle R;Delossantos A;Denenberg J;Derrett S;Des Jarlais DC;Dharmaratne SD;Dherani M;Diaz-Torne C;Dolk H;Dorsey ER;Driscoll T;Duber H;Ebel B;Edmond K;Elbaz A;Ali SE;Erskine H;Erwin PJ;Espindola P;Ewoigbokhan SE;Farzadfar F;Feigin V;Felson DT;Ferrari A;Ferri CP;Fèvre EM;Finucane MM;Flaxman S;Flood L;Foreman K;Forouzanfar MH;Fowkes FG;Franklin R;Fransen M;Freeman MK;Gabbe BJ;Gabriel SE;Gakidou E;Ganatra HA;Garcia B;Gaspari F;Gillum RF;Gmel G;Gosselin R;Grainger R;Groeger J;Guillemin F;Gunnell D;Gupta R;Haagsma J;Hagan H;Halasa YA;Hall W;Haring D;Haro JM;Harrison JE;Havmoeller R;Hay RJ;Higashi H;Hill C;Hoen B;Hoffman H;Hotez PJ;Hoy D;Huang JJ;Ibeanusi SE;Jacobsen KH;James SL;Jarvis D;Jasrasaria R;Jayaraman S;Johns N;Jonas JB;Karthikeyan G;Kassebaum N;Kawakami N;Keren A;Khoo JP;King CH;Knowlton LM;Kobusingye O;Koranteng A;Krishnamurthi R;Lalloo R;Laslett LL;Lathlean T;Leasher JL;Lee YY;Leigh J;Lim SS;Limb E;Lin JK;Lipnick M;Lipshultz SE;Liu W;Loane M;Ohno SL;Lyons R;Ma J;Mabweijano J;MacIntyre MF;Malekzadeh R;Mallinger L;Manivannan S;Marcenes W;March L;Margolis DJ;Marks GB;Marks R;Matsumori A;Matzopoulos R;Mayosi BM;McAnulty JH;McDermott MM;McGill N;McGrath J;Medina-Mora ME;Meltzer M;Mensah GA;Merriman TR;Meyer AC;Miglioli V;Miller M;Miller TR;Mitchell PB;Mocumbi AO;Moffitt TE;Mokdad AA;Monasta L;Montico M;Moradi-Lakeh M;Moran A;Morawska L;Mori R;Murdoch ME;Mwaniki MK;Naidoo K;Nair MN;Naldi L;Narayan KM;Nelson PK;Nelson RG;Nevitt MC;Newton CR;Nolte S;Norman P;Norman R;O'Donnell M;O'Hanlon S;Olives C;Omer SB;Ortblad K;Osborne R;Ozgediz D;Page A;Pahari B;Pandian JD;Rivero AP;Patten SB;Pearce N;Padilla RP;Perez-Ruiz F;Perico N;Pesudovs K;Phillips D;Phillips MR;Pierce K;Pion S;Polanczyk GV;Polinder S;Pope CA 3rd;Popova S;Porrini E;Pourmalek F;Prince M;Pullan RL;Ramaiah KD;Ranganathan D;Razavi H;Regan M;Rehm JT;Rein DB;Remuzzi G;Richardson K;Rivara FP;Roberts T;Robinson C;De Leòn FR;Ronfani L;Room R;Rosenfeld LC;Rushton L;Sacco RL;Saha S;Sampson U;Sanchez-Riera L;Sanman E;Schwebel DC;Scott JG;Segui-Gomez M;Shahraz S;Shepard DS;Shin H;Shivakoti R;Singh D;Singh GM;Singh JA;Singleton J;Sleet DA;Sliwa K;Smith E;Smith JL;Stapelberg NJ;Steer A;Steiner T;Stolk WA;Stovner LJ;Sudfeld C;Syed S;Tamburlini G;Tavakkoli M;Taylor HR;Taylor JA;Taylor WJ;Thomas B;Thomson WM;Thurston GD;Tleyjeh IM;Tonelli M;Towbin JA;Truelsen T;Tsilimbaris MK;Ubeda C;Undurraga EA;van der Werf MJ;van Os J;Vavilala MS;Venketasubramanian N;Wang M;Wang W;Watt K;Weatherall DJ;Weinstock MA;Weintraub R;Weisskopf MG;Weissman MM;White RA;Whiteford H;Wiersma ST;Wilkinson JD;Williams HC;Williams SR;Witt E;Wolfe F;Woolf AD;Wulf S;Yeh PH;Zaidi AK;Zheng ZJ;Zonies D;Lopez AD;Murray CJ;AlMazroa MA;Memish ZA
通讯作者：
Memish ZA
影响因子：
--
作者：
Asojo, Oluwatoyin A.;Homma, Kohei;Hotez, Peter J.
通讯作者：
Hotez, Peter J.
影响因子：
5.5
作者：
Hotez, Peter
通讯作者：
Hotez, Peter
影响因子：
3.1
作者：
Zhan, Bin;Perally, Samirah;Hotez, Peter
通讯作者：
Hotez, Peter