An Empirical Approach towards the Efficient and Optimal Production of Influenza-Neutralizing Ovine Polyclonal Antibodies Demonstrates That the Novel Adjuvant CoVaccine HT™ Is Functionally Superior to Freund's Adjuvant

Abstract

Passive immunotherapies utilising polyclonal antibodies could have a valuable role in preventing and treating infectious diseases such as influenza, particularly in pandemic situations but also in immunocompromised populations such as the elderly, the chronically immunosuppressed, pregnant women, infants and those with chronic diseases. The aim of this study was to optimise current methods used to generate ovine polyclonal antibodies. Polyclonal antibodies to baculovirus-expressed recombinant influenza haemagglutinin from A/Puerto Rico/8/1934 H1N1 (PR8) were elicited in sheep using various immunisation regimens designed to investigate the priming immunisation route, adjuvant formulation, sheep age, and antigen dose, and to empirically ascertain which combination maximised antibody output. The novel adjuvant CoVaccine HT™ was compared to Freund’s adjuvant which is currently the adjuvant of choice for commercial production of ovine polyclonal Fab therapies. CoVaccine HT™ induced significantly higher titres of functional ovine anti-haemagglutinin IgG than Freund’s adjuvant but with fewer side effects, including reduced site reactions. Polyclonal hyperimmune sheep sera effectively neutralised influenza virus in vitro and, when given before or after influenza virus challenge, prevented the death of infected mice. Neither the age of the sheep nor the route of antigen administration appeared to influence antibody titre. Moreover, reducing the administrated dose of haemagglutinin antigen minimally affected antibody titre. Together, these results suggest a cost effective way of producing high and sustained yields of functional ovine polyclonal antibodies specifically for the prevention and treatment of globally significant diseases.

Development of a Cost-effective Ovine Polyclonal Antibody-Based Product, EBOTAb, to Treat Ebola Virus Infection

“…In the present study, the EBOV-GP 1,2ecto used to raise the ovine antisera was proven to be an excellent immunogen, with high levels of pAb present binding with high avidity. The procedure used to manufacture EBOTAb from pools of the antisera has been used for several years to provide an antivenom that is used widely in West Africa [ 22 ]. Ovine immunoglobulin preparations also confer additional benefits, such as stability at ambient temperatures and tolerance to tropical temperatures without loss in binding activity [ 23 ].…”

Preserved antiviral adaptive immunity following polyclonal antibody immunotherapy for severe murine influenza infection

“…Sheep were immunised with baculovirus-produced recombinant HA (rHA) derived from PR8 influenza with serum stored at −20 °C as previously described 13 . Thawed serum was filtered through glass wool and pAb precipitated by addition of saturated ammonium sulfate solution to 45% (v/v).…”

“…Passive immunotherapy using neutralising Ab may be the ideal rapid treatment strategy for influenza infection that functions independently of the individual’s immunocompetency 12 as highlighted by previous animal and human studies 13 14 15 . Furthermore, a recent multicentre double-blind randomised controlled trial has found that hyperimmune intravenous immunoglobulin (IVIG) prepared from donors exposed to pandemic H1N1 influenza significantly reduced viral load in infected individuals compared to IVIG prepared from donations received before the 2009 pandemic 16 .…”

“…We have previously shown that ovine serum pAbs elicited against HA have potent neutralising capacity in vitro and are protective in an in vivo murine influenza model 13 . However, any Ab administered to prevent or treat influenza must not prevent the formation of adaptive immunity necessary for enduring protection against reinfection with future circulating strains, and this has not been explored in the context of foreign pAb delivery.…”

Targeted localized use of therapeutic antibodies: a review of non-systemic, topical and oral applications

“…Without the extremely stringent production requirements of sterile pyrogen-free parenteral products and all that this entails, large quantities of food grade sera or antibody can be manufactured at low-cost for oral use, with the potential to generate cost-effective treatment strategies. Following immunization, animals capable of producing large amounts of potent antibody at minimal cost, such as sheep, goats, cows, camels, horses and chickens (Abbott et al, 2013;Berghman et al, 2005;Carlander et al, 2000;Kovacs-Nolan & Mine, 2012;Landon & Smith, 2003;Meddeb-Mouelhi et al, 2003;Schade et al, 2005;Stevens et al, 2013), may therefore be particularly suited for plasma, serum, colostrum, milk or egg production. Any residual unwanted taste or color, which generally results from a variable hemoglobin contamination within some batches of crude serum or plasma preparations for oral use, may be easily removed by applying skimmed milk, as described previously (Jones & Marks, 2013).…”