Caenorhabditis elegans as an expression system for vaccine candidates of parasitic nematodes

Parasitic nematode (worm) infections of humans and livestock cause major health, welfare and economic problems worldwide. The efficiency of drugs currently available to control parasitic nematode infections is decreasing due to the emergence of drug-resistant parasites and alternative control approaches are urgently needed. Validation of new drug or vaccine targets of parasitic nematodes is hampered by the difficulty of obtaining sufficient parasite material for testing. Parasite proteins produced artificially in bacteria, yeast or insect cells are often folded incorrectly and/or fail to be modified in the same way as the normal parasite protein. This affects the proteins' ability to stimulate protective immune responses to infection, required for an effective vaccine, and their ability to interact with inhibitors, required for drug design. Development of a system which can synthesise protein in a similar form to the protein present within the worm would, therefore, be advantageous to parasite vaccine and drug research. Parasitic nematodes are closely related to the free-living nematode Caenorhabditis elegans and many of the proteins made in free-living and parasitic nematodes are similar. We have successfully used C. elegans to synthesise a parasite enzyme and shown that the enzyme is active, can interact with inhibitors and is modified correctly. Sufficient protein was expressed for vaccine studies to be carried out. We now propose to use the C. elegans system to make a previously identified, highly effective vaccine candidate of a major sheep parasitic nematode. We will compare the folding and modifications of the C. elegans synthesised protein to the normal parasite protein and test its ability to protect sheep against nematode disease. This work can be developed to generate other important parasite proteins and will be important both to basic research of nematode proteins and to commercial vaccine development. We will link with other researchers with expertise in animal parasite control and with industry (Pfizer) to achieve our aims.

At present there is no recombinant vaccine available for any parasitic nematode. This is predominantly due to the difficulty of expressing parasite proteins in a correctly folded and/or post-translationally modified form to induce protection. The Haemonchus contortus H11 gut aminopeptidase in native form is one of the most effective parasitic nematode vaccine candidates identified, but recombinant H11 is not protective. Glycan analysis of H11 identified di- and tri-fucosylated structures, not present on mammalian glycoproteins, which are thought to confer strong immunogenicity to the antigen. Similar structures have been identified in the free-living nematode Caenorhabditis elegans. We have recently used C. elegans as an alternative expression system for an H. contortus cysteine protease; the C. elegans expressed enzyme is active and glycosylated and could be purified in sufficient amounts for vaccination studies. The proposed study aims to use the C. elegans system to express previously identified vaccine candidates of H. contortus, focussing initially on H11. H. contortus H11 will be expressed under the control of the promoter of the related C. elegans aminopeptidase T07F10.1. His-tagged protein purified from transgenic worms will be examined biochemically and immunochemically, including deglycosylation and mass spectrometry comparison of C. elegans expressed and native H11. C. elegans expressed and native H11 will be tested in protection trials, the immune response analysed and antigenic epitopes characterised. This work will establish the suitability of C. elegans not only for H11 expression, but for expression of other complex, post-translationally modified parasite proteins. It has important relevance to our understanding of parasite immunity and to commercial development of parasitic nematode vaccines.