This project aims to replace the use of gerbils in the large scale manufacture of a nematode protein that has immunomodulatory properties and potential therapeutic applications.
Parasitic filarial nematodes can be tolerated in human hosts for many years with little evidence of pathology. This appears to reflect a parasite-induced suppression of the host pro-inflammatory immune responses. In countries where parasitic infections are endemic, the incidence of allergic or autoimmune inflammatory diseases is relatively rare. One reason for this may be that parasitic nematodes have an important role in priming the host immune system and protecting from aggressive pro-inflammatory or aberrant inflammatory responses. The nematode glycoprotein ES-62 is thought to have a key role in this because of its immunomodulatory activities and its ability to interact directly with B lymphocytes, dendritic cells, macrophages and mast cells. In the mouse ES-62 is protective against collagen-induced arthritis, a model of rheumatoid arthritis.
ES-62 and other worm proteins are potential therapeutics for a range of diseases affecting the immune system. In the laboratory ES-62 can be obtained from Acanthocheilonema viteae, a parasitic nematode worm that grows in gerbils. Large numbers of gerbils (and mice) are euthanased to obtain the protein for research purposes. This project aims to investigate purifying ES-62 from the free-living nematode worm, Caenorhabditis elegans, as an alternative to using A. viteae grown in the gerbil. The ability to grow C. elegans in large volumes in the laboratory provides an opportunity for large scale production of ES-62 for therapeutic and commercial purposes that would require thousands of animals if rodents were used
Research details and methods
The research will investigate whether ES-62, with the appropriate post-translation modification and biological activity, can be produced and isolated from transgenic C. elegans. Activity will be tested by measuring inhibition of pro-inflammatory cytokine production in the human macrophage cell line, U937.
Parasitic nematodes have evolved methods to evade the human immune system, including the secretion of proteins that suppress host inflammatory responses. We have made significant progress in understanding the properties of one such protein, ES-62, which is produced by the rodent filarial nematode Acanthocheilonema viteae. ES-62 has been shown to be protective in mouse models of inflammatory diseases such as rheumatoid arthritis and asthma, suggesting potential therapeutic use. This progress has been possible because we can maintain A. viteae in gerbils and ticks. However, we need to sacrifice hundreds of lab rodents each year for this work, and it is expensive and time consuming. Furthermore, many more animals would have to be sacrificed to produce enough ES-62 for a clinical trial. We propose to replace lab rodents by producing ES-62 in the free-living nematode Caenorhabditis elegans. This nematode can be grown in the laboratory in large amounts simply by culturing with a bacterial food source. Moreover, C. elegans can modify proteins with phosphorylcholine (PC)-N-glycan structures as found on ES-62. This modification is important for the immunomodulatory activity of ES-62. There are several established methods to make transgenics in C. elegans to direct the expression of recombinant protein. Others have used C. elegans to produce proteins from parasitic nematodes in a limited manner before but we have developed a novel strategy to improve production and purification of recombinant proteins. The resulting proteins will be tested for PC modification and immunomodulatory activity. Our proposal is not only designed to directly replace lab animals in the manufacture of ES-62 but also to show that quantities large enough for therapeutic and commercial purposes can be manufactured. Furthermore, it can be applied to producing immunosuppressive proteins from other nematode species including human parasites, therefore potentially offering further substantial benefits.