In utero infection by RV of the fetus in the first trimester of pregnancy can cause fetal teratogenesis. In addition most common complications associated with Rubella virus (RV) vaccine occur in adult women and are manifested in polyarthraliga and arthritis, associated with RV persistence and localized viral replication. In a recent joint meeting (1996) organized by the Institute of Medicine and US National Academy of Sciences on Vaccine Associated Adverse Reactions, it was recommended that new strategies must be developed for live viral vaccines so as to reduce the incidence of adverse reactions associated with such vaccines. Based on these guidelines, in case of RV, there is a need for a """"""""replication-crippled"""""""" RV vaccine which is free of adverse reactions. Recently, we have shown that the cause of adverse reactions with live virus vaccine could probably be due to interaction of host proteins with RV. The host proteins (calreticulin and La) are known autoantigens and patients with natural RV infection develop significant anti-La response following prolonged infection. In addition, we have observed that rubella virus non-structural proteins have domains which interact with cell cycle regulatory protein i.e. retinoblastoma protein (RB) both in vivo and in vitro. This interaction could result in the alteration in cell growth upon RV infection and may be the probable cause of RV induced teratogenesis. Recently we have also shown that cellular calreticulin chaperones ruubella virus E1 glycoprotein and mutations in the E1 glycosilation sites affect viral assembly and release due to perturbations in E1 chaperoning to proper sites for viral assembly. Using this knowledge, we are developing following strategies to make a assembly-defective and or """"""""replication-crippled"""""""" rubella viruses to be useful as safer vaccine candidates. Construction of a RV infectious plasmid that has: 1. Altered E1 glycosylation sites so that E1 chaperoning by calreticulin is affected and resultant mutants survive only a few cycles of replication 2. Mutations in the Rb-binding domain in such a way that RV RNA can only replicate at sub optimal level and be eliminated by dilution effect after a few cycles. Our working hypothesis is that by combining the above mentioned approaches, we could create recombinant """"""""suicide"""""""" RV vaccines that contain mutations in the viral proteins that interact with celluar proteins and are therefore """"""""crippled"""""""" and would be free of adverse reactions. PROGRESS Developed several poorly replicating RV clones using recombinant DNA methods to target mutations in the Rb-binding motif of Rb NSP90 and also in the E1 glycosylation sites. To date we have created 12 different mutant rubella viruses. These recombinants will be tested for their attenuation characteristics in vero 76 cell line. Recently we have identified the Complement 1 protein's """"""""globular head"""""""" receptor as another rubella virus interacting host protein. This interaction could play a mojor role in elucidating the transient/chronic arthritis associated with rubella virus vaccine and natural infections.
