Recombinant IalphaIp: Treatment for Sepsis

DESCRIPTION (provided by applicant): Sepsis is the systematic and overwhelming response to infection. Once triggered, an uncontrolled cascade of coagulation, impaired fibrinolysis, and inflammation drive disease progression. This results in damage to the lungs, liver, kidney and cardiovascular system, leading to multiple organ failure and an associated high mortality rate in severe sepsis disease states. In the United States, sepsis is the leading cause of death in the non-cardiac intensive care unit and the 10th leading cause of death overall. Inter-alpha-inhibitor proteins (IaIp) are serine protease inhibitors that have recently been implicated as a treatment for sepsis. When serum IaIp values fall below 40% in septic patients, there is a high incidence of mortality. Administration of human IaIp improved the survival rate from 30% to 89% in septic animals at 10 days after cecal puncture. Presently, the only source of IaIp is to purify it from human plasma. The biochemical complexity and critical post-translation modifications of IaIp preclude expressing the recombinant protein by conventional systems. MultiCell has recently developed highly functional, immortalized human hepatocytes that continue to express plasma proteins including IaIp. The goal of this phase I SBIR project is to genetically engineer an immortalized human hepatocyte cell line designated Fa2N-4 to express high levels of recombinant IaIp. This will be accomplished by genetically introducing plasmid constructs that allow the co-expression of the 4 subunits that comprise IaIp. Conditioned medium from transiently transfected cells will be analyzed by ELISA and biologically measured via a trypsin inhibition assay for active IaIp. If successful, the goal of the Phase II project will be to create stable cell lines that express IaIp and to scale up the production and purification of recombinant IaIp for preclinical evaluation.