Over the past five years, we have cloned a number of genes encoding protein antigens that appear to play a role in the host-parasite interactions in lymphatic filariasis. We have also conducted some preliminary protection studies using some of the recombinant fusion proteins encoded by these genes and obtained some promising data. In the current proposal we intend to continue and refine these studies. Specifically we would like to: 1. Continue the molecular biological studies of the collagen gene encoding the most promising candidate antigen we have identified thus far. We would like to complete the DNA sequence analysis of the genomic clones that encode the 3' and 5' ends of this gene; use recombinant DNA techniques to identify the stage and tissue of its expression; use PCR (in collaboration with Dr. John Donelson) to isolate cDNA clones that can encode this antigen; and to characterize these cDNA clones to confirm the structure of the gene we have inferred thus far from analysis of the genomic clones. 2. Conduct immunochemical studies on the various protein antigens. While we will focus mostly on the filarial collagen, we have other candidate antigens that can justifiably be studied in greater detail for their effect on the host-parasite relationship. We will optimize the expression of these antigens by using various expression vector-host combinations; purify proteins to homogeneity or near-homogeneity; generate monospecific polyclonal antisera to these antigens; use these antisera with or without immuno-affinity purification to determine the topographical localization of the protein antigens. 3. Investigate the potential of the antigens we have identified thus far in protection studies. As mentioned above, we have obtained some exciting preliminary data using the jird model suggesting that a filarial collagen we have cloned may have some protective value. We would like to confirm these studies, again in the jird model, and at the same time test the effect of preimmunization with other candidate antigens. We would like to determine the optimal method to deliver the immunogen, including purified protein; use of adjuvants other than Complete Freund's (CFA) which will not be permitted in human use; the effect of preimmunization on 'trickle"""""""" infections; and the longevity of the protection provided by preimmunization.
