Engineering a new generation of blood substitutes

失血的患者会出现急性低血压和氧含量下降;然后输送到组织的氧气福尔斯下降，导致灾难性的临床后果。因此，输血是治疗因创伤或手术失血的患者的主要因素。在全球范围内，每年约有8500万单位的红细胞输注，价值约160亿美元。然而，输注浓缩红细胞存在固有的问题。血型分型的需要和较短的保存期限限制了储存重大灾害所需物资的能力。它还排除了在救护车和远离大医院的人口中心（包括军队使用）进行常规输血支持。同种输血并不总是良性的：它们可能会因输错血事故或免疫功能低下的个体而导致急性输血反应。血液是一种生物制剂，不能消毒，因此总是有可能被细菌和血液传播的传染性病原体（如艾滋病毒、肝炎、vCJD和西尼罗河病毒）污染。并非所有国家都对所有病毒进行筛查，在某些情况下（vCJD），目前还没有经过验证的检测方法。新的血液传播病毒是定期发现的;对于那些具有传染性和诱导病理学的病毒，在引入新的筛查试验之前总是会有一个病理学窗口。增加筛查增加了血液制品的成本以及减少供应;人口年龄的增加了需要输血的人数，同时减少了能够献血的人口比例，从而加剧了这一问题。这些因素的结合限制了“安全”输血的使用，这些国家的医疗系统既能够管理输血服务，又能够管理相对没有传染性血液传播病毒的人口。在许多国家，特别是在非洲，这些因素排除了输血在创伤中的使用。有一个明确的全球医疗需求的寿命长，无病原体的“血液替代品”，能够增加红细胞的使用。大量的努力已经花费在生产基于血红蛋白（红细胞携氧蛋白）的化学或遗传修饰的产品上。尽管几种血红蛋白氧载体（HBOC）已被证明可以输送氧气，但由于安全问题，它们未能获得在主要医疗保健市场中使用的批准。HBOC毒性的三种主要机制已被提出。一是产品缺乏稳定性。其他两个与分子机制有关：（i）在红细胞的保护环境之外，HBOC可以释放控制血压和血流的一氧化氮（NO）（ii）HBOC可以产生有毒的自由基。所有这些问题都可以通过基因工程技术来解决。在埃塞克斯，我们得到了BBSRC和欧盟的资助，调查血红蛋白产生有毒自由基的基础科学。我们已经能够开发出新的修饰血红蛋白分子，它们更容易被人体的抗氧化剂解毒，因此引起更少的自由基损伤。我们现在希望将这些知识应用于直接解决临床问题。以前的研究人员已经开发出了血红蛋白突变体，不增加一氧化氮。我们打算将这些突变联合收割机与我们自己的优化突变结合起来，减少自由基的产生。我们的基础蛋白质将是人类胎儿血红蛋白，这是一种比成人形式稳定得多的蛋白质。解决这三个问题（产品稳定性、NO清除和自由基产生）将为下一代血液替代产品奠定基础，这些产品可供所有国家使用，并可在所有环境中使用。

Patients with blood loss suffer acute hypotension and a drop in oxygen content; oxygen delivery to tissue then falls with a catastrophic clinical outcome. Blood transfusions are therefore a major element of treating patients suffering blood loss through trauma or surgery. Globally around 85 million units of red blood cells are transfused annually in an industry worth ~$16Bn. However, there are inherent problems with transfusions of packed red blood cells. 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). Homologous blood transfusions are not always benign: they can result in acute transfusion reactions, either by mistyping accidents or in immune compromised individuals. Hence immune compromised individuals, as well as certain religious groups, are unable to benefit from blood transfusions.Blood is a biological agent; it cannot be sterilised and therefore always has the potential for contamination with bacteria and blood-borne infectious agents such as HIV, hepatitis, vCJD and West Nile virus. Not all viruses are screened for in all countries, and in some cases (vCJD) no validated tests are currently available. New blood borne viruses are discovered on a regular basis; for those that are infectious and induce pathology, there will always be a pathological window prior to the introduction of a new screening test. Increased screening raises the cost of blood products as well as decreasing supply; the increasing age of populations adds to this problem by increasing the number of people requiring transfusions, whilst decreasing the fraction of the population able to donate blood. The combination of these factors limits the use of "safe" blood transfusions to those countries with a funded and developed medical system able both to manage a blood transfusion service and having a population that is relatively free of infectious blood borne viruses. In many counties, notably in Africa, these factors preclude the use of blood transfusions in trauma.There is a clear global medical need for a long life, pathogen free "blood substitute" able to augment the use of red blood cells. A great deal of effort has been expended into producing products based on chemical or genetic modification to haemoglobin, the red cell oxygen carrying protein. Although several haemoglobin-based oxygen carriers (HBOC) have been shown to deliver oxygen they have failed to get approval for use in the major healthcare markets due to safety concerns. Three major mechanisms have been proposed for HBOC toxicity. One is a lack of product stability. The other two relate to molecular mechanisms: (i) outside the protective environment of the red cell HBOC can scavenge nitric oxide (NO) that controls blood pressure and blood flow (ii) HBOC can create toxic free radicals. All these problems can be addressed by genetic engineering techniques. At Essex we have been funded by BBSRC and the EU to investigate the basic science of toxic free radical production by haemoglobin. We have been able to develop novel modified haemoglobin molecules that are more readily detoxified by the body's antioxidants, and so induce less free radical damage.We now wish to apply this knowledge to directly address clinical issues. Previous researchers have developed haemoglobin mutants that do not scavenge nitric oxide. We intend to combine these mutations with our own optimised mutations that decrease free radical production. Our base protein will be human foetal haemoglobin, which is a far more stable protein than the adult form. Solving these three issues (product stability, NO scavenging and free radical production) will form a basis for the next generation of blood substitute products, accessible to all countries and useable in all environments.