Passive immunoprophylaxis has been an important public health tool. For example, normal immunoglobulin has been important in the prevention of hepatitis A. However, monoclonal preparations could be more potent, tailored to specific neutralization epitopes and highly consistent in potency. Chimpanzee globulins are virtually identical to human immunoglobulins, making them attractive choices for immunoprophylactic and immunotherapeutic agents. We have prepared combinatorial libraries from the bone marrow of chimpanzees that had been experimentally infected in sequence with each of the five human hepatitis viruses. We isolated monoclonal immunoglobulins that react with HAV, HBV, HDV and HEV. In other studies, we recovered human monoclonal antibodies that react with HCV. Many of the monoclonal antibodies described above were neutralizing and their production was scaled up for tests of passive immunoprophylaxis in a variety of animals and, eventually, in humans. These studies were extended through 2010. Construction of combinatorial libraries from bone marrow has also been carried out for chimpanzees that have been experimentally infected with dengue viruses 1 through 4: these have yielded important neutralizing monoclonal antibodies to dengue virus types 1, 2 and 4. More importantly, we reestablished an animal model for antibody-mediated enhancement of dengue virus infections, which can lead to the more severe forms of dengue virus infection: dengue hemorrhagic fever and dengue shock syndrome. Furthermore, we identified a deletion in the antibody molecule that abrogates enhancement, possibly making antibody therapy practical for the first time. We have extended our antibody studies to other viruses and bacteria of interest that can be experimentally administered to chimpanzees. For example, in response to new concerns about bioterrorism, we prepared neutralizing monoclonal antibodies to vaccinia virus for use as immunoprophylactic/ immunotherapeutic agents in those who require immunization with vaccinia but who are susceptible to the side-effects of such immunization. More importantly, in collaboration with the CDC, we demonstrated that monoclonal antibodies that neutralize vaccinia can also neutralize variola (the smallpox virus). Thus, these neutralizing monoclonal antibodies should be useful not only for prophylaxis and therapy of the side effects of vaccination but also for the prevention and therapy of smallpox, should it ever be released into populations. Similarly, we immunized chimpanzees with anthrax toxin in an attempt to make monoclonal antibodies that could immediately neutralize anthrax in vivo and have isolated highly potent monoclonal antibodies that can neutralize all three anthrax toxins (PA, LF and EF.) In 2010 we extended the study of monoclonal antibody to protective antigen (PA) to include a metabolic study of protection in rats administered either edema toxin (EF plus PA) or lethal toxin (LF plus PA) or both toxins. Rats were administered the antibody, followed by the toxin and monitored for hypotension, acidosis and renal and liver dysfunction. The antibody increased survival and reduced hypotension, in some cases even when administered up to 12 hours after intoxication. Similarly, rats administered Bacillus anthracis spores subcutaneously had increased survival when the antibody was administered up to 48 hours after intoxication. Thus, the monoclonal antibody had the potential to reduce morbidity due to ET or LT even after toxin release and shock had developed. Also in 2010, we generated monoclonal antibodies to the capsule of B. anthracis by immunizing chimpanzees with synthetic capsular protein. These antibodies can bind to the capsule of the bacterium and opsonize it. We demonstrated that a single 30 mcg dose of monoclonal antibody administered to mice 18 hours before challenge conferred approximately 50% protection against a lethal intra-tracheall spore challenge by the virulent B. anthracis Ames strain. More importantly, monoclonal antibody given eight hours or 20 hours after challenge provided significant protection also. These and monoclonal antibodies that neutralize vaccinia and smallpox were the subjects of a CRADA with MacroGenics. We have also prepared chimpanzee monoclonal antibodies to the three serotypes of poliovirus, to rabies virus, to West Nile virus and to the tick-borne encephalitis virus complex. We have also added the seven toxins of Clostridium botulinum. Some of these will have potential utility in efforts to counteract bioterrorism and all will have immunoprophylactic and immunotherapeutic potential in the battle against emerging and re-emerging pathogens. An impediment to understanding the immune response to hepatitis C virus (HCV) has been the inability to measure neutralizing antibodies because most HCV strains do not replicate in cell culture. We previously demonstrated neutralizing antibodies in an in vivo neutralization assay utilizing chimpanzees. This has been the only accepted neutralization assay until recently, when an in vitro assay based on the neutralization of recombinant retroviruses bearing the envelope glycoproteins of HCV was developed. We demonstrated that this assay generally correlated with the in vivo assay and that the neutralizing antibodies were more broadly reactive than previously thought. We have applied the pseudo-typed virus neutralization assay to sera from chimpanzees experimentally vaccinated with two candidate antibody-based HCV vaccines and have shown that one vaccine, which was effective in preventing infection, hepatitis and chronicity in most animals following challenge with virulent HCV stimulated high levels of broadly neutralizing antibody, whereas the other vaccine, which was completely ineffective in preventing infection, hepatitis or chronicity, completely failed to stimulate neutralizing antibodies as measured by the pseudo-typed virus assay. Broadly neutralizing monoclonal antibodies were identified for HCV through a CRADA with Innogenetics, Ghent, Belgium. These monoclonal antibodies, directed against the E1 envelope glycoprotein of HCV, were recovered from a patient who had been successfully treated for chronic hepatitis C with interferon. The monoclonal antibodies were highly neutralizing and broadly reactive in the pseudo-typed virus assay. The results obtained with them (and with other polyclonal sera) suggest that hepatitis C viruses, which consist of six genotypes, may comprise two or three serotypes. It is hoped that these monoclonal neutralizing antibodies will find clinical utility in the prevention and therapy of HCV infections. In 2011 we demonstrated in collaborative studies with the FDA that monoclonal antibodies derived from chimpanzees were highly potent for the neutralization of all three poliovirus serotypes and that at least one of these antibodies could neutralize two different serotypes. Furthermore, they could protect mice from paralysis and death whether administered before or after exposure to virulent poliovirus (the mice used in this study are genetically modified to express the human receptor for polioviruses). One of the antibodies is being prepared for clinical testing. A major impediment to the eradication of poliomyelitis worldwide is the presence of immunologically compromised individuals who chronically shed the virus into the environment;these would be subjects for antibody therapy. In 2011 we have also demonstrated pre-and post-exposure protection against virulent anthrax infection in mice by humanized monoclonal antibodies to Bacillus anthracis capsule. These antibodies have potential, in concert with antibodies against B. anthracis toxins, to prevent and treat anthrax. In 2012 we markedly extended our studies of passive protection against anthrax.
