The rapid spread of the Zika virus, which can have a significant impact on neurological disorders in unborn fetuses and potentially adults, the recent outbreak of the extremely virulent Ebola virus, periodic emergence of SARS, recurrent outbreaks of potentially pandemic strains of influenza such as H5N1, the continuing epidemic of MERS and the worldwide AIDS epidemic have highlighted a persistent concern in the health-care community -- the need for effective pathogen inactivation and removal techniques for human blood plasma and plasma-derived products. There is no commercially available, FDA-approved technology for the inactivation of non-enveloped viruses in pooled human plasma and biologics, and only one approved method for units of plasma, which can inactivate some, but not all known non-enveloped viruses. This dearth of FDA-approved pathogen inactivation technologies could pose a significant future threat for known and new viruses in human plasma and biologics. We propose to develop a physical pathogen inactivation technology, CFI?, for the inactivation of both non-enveloped and enveloped viruses as well as pathogenic bacteria and parasites in human plasma, plasma protein products and biologics. CFI? technology is applicable to both pooled human plasma and units of plasma, the more globally significant focus of the current application. CFI? (critical fluid inactivation) utilizes supercritical and near-critical fluids (SuperFluids? or SFS). SuperFluids? are normally gases which, when compressed, exhibit enhanced thermodynamic properties of solvation, penetration, selection and expansion. These gases are used to permeate and saturate virus and pathogen particles. The SFS-saturated particles then undergo decompression and, as a result of rapid phase conversion, virus inflation and rupture at their weakest points. We have demonstrated that the CFI? (critical fluid inactivation) process inactivates both enveloped viruses such as MuLV, VSV, Sindbis, HIV (all completely inactivated), TGE, and BDVD, and the non-enveloped viruses Polio, Adeno, EMC (complete inactivation), Reo, and Parvo viruses, while preserving biological activity of the CFI-treated product. In research collaboration with the National Institute of Biological Standards and Control (NIBSC), London, England, our CFI inactivated more than 4 logs of human Parvovirus B19 (one of the smallest and toughest viruses) in human plasma in a two-stage CFI? unit in less than 20 seconds. We have also demonstrated that SFS can disrupt and inactivate microorganisms such as E. coli, thick-walled prokaryotes such as Bacillus subtilis and tough eukaryotes such as Saccharomyces cerevisiae at viral inactivation SFS conditions. CFI can be used with viral reduction methods such as nanofiltration as an orthogonal method of pathogen clearance, and is versatile for refinement to treat cellular blood. The present data have been generated using prototypes of our pilot-scale CFI unit. Our Phase I Specific Aims are to: (1) Design and construct a portable and comparably versatile bench-top CFI unit for parallel treatment of single units of human plasma; and (2) CFI treat human plasma in the designed bench-top device using a customized blood bag, and rigorously characterize the CFI-treated units of human plasma for protein and enzyme activities. Our Phase II Specific Aims are to: (3) Complete characterization of CFI-treated units of human plasma using customized blood bag in the bench-top CFI prototype; (4) Conduct toxicological and neoantigenic studies of CFI treated human plasma in small animal models; and select best CFI design in terms of performance, operations and cost; and (5) Draft IND, establish and conduct pre-IND meeting with FDA, prepare and file a Drug Master File (DMF) for the manufacturing of CFI treated human plasma units. In Phase III, we will construct bench-top CFI units for blood banks, and through licensing agreements, provide equipment and technology transfer as well as prevention and maintenance support to blood banks.
There are a number of emerging viruses such as Zika, West Nile, Ebola, SARS, potential pandemic strains of influenza (H5N1), the Mexican swine flu, bacteria, parasites and a number of potential bioterrorism pathogens such as smallpox that are of concern to the safety of the human plasma supply. Current approaches for pathogen inactivation in biologics are not always effective against a wide spectrum of human and animal viruses, are sometimes encumbered by process-specific deficiencies, and often result in denaturation of the biologicals that they are designed to protect. CFI pathogen inactivation technology gives pathogens the ?bends,? inactivating them without damaging proteins and enzymes in medically important transfusion fluids such as human plasma. This purely physical technique does not involve the use of heat, chemicals and/or irradiation, each of which has significant drawbacks in the viral inactivation of human plasma. As such, while CFI is capable of inactivating wide classes of viruses, bacteria and parasites, it has negligible negative impact on biological integrity and potency of the treated fluids. We have developed, tested and validated this process using continuous-flow laminar flow devices, and now plan to scale this technology down for units of human plasma using a novel, portable bench-top CFI device using blood bags. The potential impact of a generally- applicable physical technology for inactivating viruses and emerging pathogens with high retention of biological activity will be very significant. CFI technology will also be impactful in developing countries and hot zones for the clearance of viruses from human plasma.
