CHEMICAL INACTIVATION OF RETROVIRAL INFECTIVITY BY TARGETING NUCLEOCAPSID PROTEIN ZINC FINGERS - A CANDIDATE SIV VACCINE

Although most viral vaccines used in humans have been composed of live attenuated viruses or whole killed viral particles, the latter approa ch has received little attention in research on experimental primate i mmunodeficiency virus vaccines. Inactivation procedures involving heat or formalin appear to adversely affect the viral envelope proteins. R ecently we have inactivated human immunodeficiency virus type 1 (HIV-1 ) with the compound 2,2'-dithiodipyridine (Aldrithiol-2, Aldrich, Milw aukee, WI), which inactivates infectivity of retroviruses by covalentl y modifying the nucleocapsid zinc finger motifs, HIV-1 inactivated wit h Aldrithiol-2 retained the conformational and functional integrity of the viral and virion-associated cellular proteins on the viral membra ne. We have extended our studies of zinc finger targeted inactivation to simian immunodeficiency virus (SIV) and evaluated the feasibility o f applying the procedures to large scale (>30 1) production and purifi cation of the primate immunodeficiency viruses. There was no detectabl e residual infectivity of SIV after treatment with 1 mM Aldrithiol-2 ( >5 logs inactivation). Treatment with Aldrithiol-2 resulted in extensi ve reaction with the nucleocapsid protein of treated virus, as shown b y immunoblot and high-performance liquid chromatography (HPLC) analysi s. As expected, the virion gp120(SU) appeared to be completely unreact ive with Aldrithiol-2, Sucrose gradient purification and concentration procedures resulted in little loss of viral infectivity or virion-ass ociated gp120(SU), When tested in a gp120-CD4 dependent cell binding a ssay, the inactivated virus bound to cells comparably to the untreated virus. Analysis of gp120-CD4 mediated postbinding fusion events showe d that the inactivated virus could induce CD4-dependent fusion with ef ficiencies similar to the untreated virus. Inactivation and processing of primate immunodeficiency viruses by methods described here results in highly concentrated virus preparations that retain their envelope proteins in a native configuration. These inactivated virus preparatio ns should be useful in whole killed-particle vaccine experiments as we ll as laboratory reagents to prepare antisera, including monoclonal an tibodies, and to study noninfective virion-cell interactions.