News and Publications

Molecular Parasitology

A.S. Kang, Y. Su, T. Stratmann,* R. Xu,** P. Mattei***

* University of Marburgh, Marburgh, Germany
** Fudan University, Shanghai, Peoples Republic of China
*** ETH, Zurich, Switzerland

Each year, more than 2.5 million persons die of malaria and more than 300 million are infected with the parasites that cause this disease. Malaria caused by Plasmodium falciparum is the worst type and the one most likely to kill. We study ways to intervene in the life cycle of parasites such as P falciparum in humans. The life cycle of Plasmodium in the human host is complex and offers a number of potential targets for attack by the immune system. Persons exposed to irradiated sporozoites are protected against subsequent challenge with live parasites. In the past 3 decades, efforts to duplicate this immunity have focused on recombinant vaccines and peptides.

When a desired immune response cannot be generated, one approach is to use passive immunization. Studies have shown that transfer of immune -globulin is effective against malaria. We used peripheral blood leukocytes derived from a person immune to malaria to construct combinatorial phage display libraries of antibodies to P falciparum. We selected an antibody (PfNPNA-1) to the repeat epitope of circumsporozoite surface protein and transferred the variable regions into vectors to produce whole human IgG1 and IgG4 antibodies. The IgG4 antibody inhibits sporozoite invasion of human hepatocytes in vivo at high concentrations. Current efforts are directed toward isolating antibodies to regions that flank the repeat epitope on the circumsporozoite surface protein and antibodies to other targets on the parasite.

In a related project, we are designing molecules to block the transmission of malaria at the mosquito stage. With the support of the McKnight Foundation, we are exploring intervention in arthropod-transmitted viral diseases in plants. Symbiotic bacteria that reside in an insect are engineered to express antibody fragments to impair the ability of the insect to transmit a pathogen.

Engineered antibodies have usefulness beyond the obvious roles in treatment and diagnosis of disease and in affinity enrichment. We have used our technology to develop novel cell-surface receptors that allow specific pathogens to enter usually nonhost cells, a process that enables us to study the early events of pathogenesis. To enter cells, viruses use a number of different cell-surface molecules. Foot-and-mouth disease virus uses a cell-surface integrin via an arginine--glycine--aspartic acid motif in the capsid protein. A mutant of the virus in which this motif has been deleted is noninfectious.

To propogate the mutant virus, we created a new receptor by fusing a virus-binding single-chain antibody to intracellular adhesion molecule 1. Cells not normally susceptible to infection with foot-and-mouth disease virus became infected after being transfected with the plasmid encoding the fusion protein. The mutant virions that are noninfectious in animals and other cell types grew to high titers and were able to form plaques on transfected cells containing the fusion protein.

These studies were the first production of of a totally synthetic cell-surface receptor for a virus. This novel approach will be useful for the development of safer vaccines against viral pathogens of animals and humans. This concept should be useful in research on viral and parasitic pathogens that are difficult to study because no appropriate in vitro cultivation systems exist. Using engineered cells for uptake and propogation of pathogens is an approach that may enable scientists to study the early events in host-pathogen interaction in detail. Finally, we have been actively involved in constructing IgG-cytokine fusion proteins for use in cancer immunotherapy.

PUBLICATIONS

Dreier, T., Lode, N.H., Xiang, R., Dolman, C., Reisfeld, R., Kang, A. Recombinant immunocytokines targeting the mouse transferrin receptor: Construction and biological activities. Bioconjugates, in press.

Lode, H., Dreier, T., Xiang, R., Varki, N., Kang, A., Reisfeld, R. Gene therapy with a single chain interleukin-12 fusion protein induces T-cell depedent protective immunity in a syngeneic model of murine neuroblastoma. Proc. Natl. Acad. Sci. U.S.A. 95:2475, 1997.

Stratmann, T., Schmid, S., Harper, J.F., Kang, A. Bacterial expression and purification of recombinant Plasmodium yoelii circumsporozoite protein. Protein Expr. Purif. 11:72, 1997.