Ricin, a natural product of the castor bean (Ricinus communis) and a Category B toxin, is significant as a biological weapon because of its heat stability, worldwide availability, and ease in production. It can be disseminated as an aerosol, a likely route that terrorists may use. The ricin toxin is a lectin consisting of two polypeptide chains linked by a disulfide bridge and cellular entry is required for toxicity. The ricin B-chain (RCB) facilitates entry of the toxin into the cell and the ricin A-chain (RCA) possesses RNA N-glycosidase activity that attacks a specific site on the 28S rRNA, preventing polypeptide chain elongation, thereby inactivating ribosomes (ribotoxic) and leading to cell death. Since the ricin toxin is free in the circulation for only a brief period of time (hours to days) before cellular internalization, the greatest clinical benefits will be derived from therapeutics capable of blocking RCA enzymatic activity at intracellular sites. We have recent evidence that a high dose of a novel primatized anti-ricin A-chain antibody (43RCA IgG) administered directly to the lung can offer 83% and 75% survival when administered at 44 hr and 54 hr post-ricin challenge, respectively. The primatized 43RCA IgG was generated by fusing the V domains from macaque-derived 43RCA scFv to the constant regions of the human IgG1. The 43RCA scFv sequence is very similar to that for human IgG germline genes, with 90% sequence identity for the VH and VL regions, thereby increasing the potential for this human-like antibody to be used as ricin antidote in humans. In the case of human exposure, whether it is accidental or deliberate, it is anticipated that there will be delays in post-exposure treatment. It is, therefore, imperative to develop antidotes with a therapeutic window beyond 2 days to allow sufficient time for treatment of exposed individuals. We therefore hypothesize that the therapeutic index and/or window for post- exposure treatment can be improved and extended by cytosolic delivery of neutralizing anti-RCA antibodies to block intracellular ricin activity. To achieve our goal, we will develop cell-permeable monoclonal antibodies (or TransMabs) by conjugating antibodies to transport peptides or cell-penetrating peptides (CPPs) and the therapeutic effects will be evaluated as described in the following specific aims.
Specific Aim 1 : To Ascertain the Protective Effects of Cell-Permeable antibodies against Ricin Cytotoxicity in Cell Culture Models.
Specific Aim 2 : To Ascertain the Efficacy of Cell-Permeable Antibodies for Post-Exposure Treatment of Ricin-Induced Lung Injury and Lethality using the Lung Aspiration Model.
Specific Aim 3 : To Validate the Efficacy of Cell-Permeable Antibodies for Post-Exposure Treatment in an Aerosolized Ricin Mouse Model.
Since there is currently no treatment for the ricin toxin, development of a specific antidote for the treatment of ricin after exposure will contribute significantly to the protection of our civilian and military populations. Additionally, the availability of an effective antidote may also significantly reduce the threat of the use of ricin as a biological weapon.
