Optimisation of the manufacture of a homogeneous synthetic haemoglobin as a novel Oxygen Therapeutic / Blood Substitute

输血是一种挽救生命的技术;然而，同种异体红细胞具有固有的缺点，例如保质期有限，需要血型分型，并且它们不能用于免疫受损的个体和那些由于宗教原因而拒绝输血的人。历史上使用艾滋病毒和肝炎污染的血液仍然显示出今天通过公众调查和幸存患者的持续健康问题的影响。如今，血液制品更加安全，全球每年约有1.08亿单位的红细胞输血，价值超过100亿美元。然而，新的血液传播疾病是定期发现的，有可能在发现之前就渗透到输血人群中。此外，血型分型的需要和较短的保质期限制了储存重大灾害所需物资的能力。它还排除了救护车和远离大医院的人口中心的常规输血支持（包括军事使用）。血红蛋白（Hb）是红细胞中的蛋白质，将氧气从肺部运送到组织。因此，合成血红蛋白基氧载体（HBOC）乍一看似乎是一个理想的起点，更安全的替代红细胞输血。然而，HBOCs的临床试验通常结果不佳。最近，HBOCs已被推广为患者的氧气“桥梁”，有希望实现即使循环失败也将氧气输送到组织的治疗的可能性，从而保护或维持重要器官。这可以针对具有经历手术的医学状况的患者，例如脑损伤、心脏手术、肾移植和创伤性损伤导致的失血后休克。此外，HBOC可用于实体瘤的氧合，以增强败血症、镰状细胞病、溶血性贫血等的治疗。尽管如此，对HBOCs的安全性和有效性的担忧仍然存在，越来越多的人认识到，HBOCs的固有毒性，与一氧化氮（NO）清除和氧化损伤有关，可能在很大程度上是导致不良临床结果的原因。虽然一些焦点已经被给予解决NO清除，但很少有研究集中在HBOCs的“流氓”酶活性的氧化损伤问题上。在埃塞克斯，我们有一个专家的机制和血红蛋白的破坏性氧化反应的后果的理解。通过之前的MRC DPFS资助，我们成功地开发了一种基于更稳定的胎儿Hb的基因工程HBOC，解决了无细胞Hb的固有毒性。这是第一个利用重组技术来重新设计Hb分子的基本特性来解决NO清除和氧化损伤问题的HBOC。我们的新型突变Hb解决了-并解决了-在前几代HBOC中引起不良副作用的无细胞Hb的固有毒性。用聚乙二醇（PEG）涂覆HBOC的表面延长了HBOC在血液中的寿命，增加了其粘度并降低了其免疫原性。我们已经开发了一种独特的聚乙二醇化的统一系统，与以前的技术不同，它不会间接影响氧结合和协同性。该奖项的目的是跟进我们独特的稳定和无反应的HBOC产品的成功开发，为该产品的临床生产提供途径。这一发展将达到临床所需的标准，并将寻求克服与蛋白质生产相关的先前问题。此外，我们还将进行旨在证明产品安全性和有效性的测试，并提供证据证明我们的产品已经克服了前几代HBOC的主要问题，从而在临床前和临床试验的完整计划中提供积极结果的信心。

Blood transfusion is a lifesaving technology; however, allogeneic red blood cells have inherent disadvantages such as limited shelf life, need for blood group typing, and they cannot be used in immune compromised individuals and those that reject transfusions due to religious reasons. The historical use of HIV- and hepatitis-contaminated blood still shows ramifications today through public inquiries and the ongoing health issues of surviving patients. Blood products are much safer today, with globally around 108 million units of red blood cells transfused annually in an industry worth >$10Bn. However, new blood borne diseases are discovered on a regular basis with the potential of permeating the transfusion population before detection. Additionally, the need for blood group typing and the short shelf life limits the ability to stockpile the supplies necessary for major disasters. It also precludes routine transfusion support in ambulances and in population centres distant from major hospitals (including use by the military).Haemoglobin (Hb) is the protein in the red blood cell that carries oxygen from the lungs to the tissues. Therefore, a synthetic Haemoglobin-based oxygen carrier (HBOC) at first glance appears to be an ideal starting point for a safer alternative to erythrocyte transfusions. However, clinical trials of HBOCs have typically seen poor outcomes. More recently HBOCs have been promoted as an oxygen 'bridge' for patients promising the possibility of a therapy that takes oxygen to the tissues even though the circulation is failing, thereby protecting or maintaining vital organs. This may be targeted to patients with medical conditions undergoing surgery, for example brain injuries, heart surgery, kidney transplantation and shock following blood loss resulting from trauma injury. Additionally, HBOCs may be used for oxygenation of solid tumours to enhance therapies, treatments for sepsis, sickle cell disease, haemolytic anaemia and many more. Nonetheless concerns over the safety and efficacy of HBOCs remain.There is a growing realisation that the inherent toxicity of HBOCs, relating to nitric oxide (NO) scavenging and oxidative damage, may largely be responsible for the poor clinical outcomes. While some focus has been given to tackling NO scavenging, very little research has focused on the issue of oxidative damage from 'rogue' enzymatic activities of HBOCs. At Essex we have an expert understanding of the mechanisms and consequences of the damaging oxidative reactions of Hb. Through previous MRC DPFS funding we have successfully developed a genetically engineered HBOC based on the more stable foetal Hb that addresses the inherent toxicities of cell-free Hb. This is the first HBOC to tackle both issues of NO scavenging and oxidative damage utilising recombinant technologies to re-engineer the fundamental properties of the Hb molecule. Our novel mutant Hb addresses -and resolves- the inherent toxicities of cell-free Hb that caused adverse side effects in previous generations of HBOC. Coating the surface of the HBOC with polyethylene glycol (PEG) extends the lifespan of the HBOC in blood, increases its viscosity and decreases its immunogenicity. We have developed a uniquely uniform system of PEGylation that, unlike previous techniques, does not detrimentally effect oxygen binding and cooperativity. The purpose of this award is to follow up on the successful development of our uniquely stable and unreactive HBOC product to provide a route to clinical manufacture of the product. This development will be to the clinically required standards and will seek to overcome the previous issues associated with protein production. In addition, we will conduct tests designed to show safety and efficacy of the product and to provide evidence that our product has overcome the major issues of previous generations of HBOC and hence provide confidence of positive outcomes in a full program of preclinical and clinical trials.