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

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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.