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The sensitivity of togaviruses to the effects of interferon (IF) appears to permit successful exploitation of IFF inducers, whereas the relative insensitivity of arenaviruses to IF in vitro appears to correlate with the complete lack of success of lysine-stabilized poly(I)-poly(C) - (poly(ICLC)) therapy in the monkey model. Noteworthy is the successful use of poly(ICLC) against fatal encephalitis caused by Japanese encephalitis virus in monkeys. Additional studies appear to be warranted to characterize further the role of IF and/or IF inducers in other model infections prior to studies in man.

We have studied model arboviruses and arenaviruses to develop effective combination therapies using antiviral drugs and immunomodulators. The compounds were examined using Pichinde' (PIC) an arenavirus; Banzi (BZ) and West Nile (WN), two flaviviruses; Semliki Forest (SF), an alphavirus; and Bunyamwera (BW) and LaCrosse (LAC), two Bunyaviruses. Ribavirin was very effective against PIC, allowing for the development of lasting immunity to reinfection in surviving animals. Interferon (IFN) and IFN inducers had a consistent prophylactic antiviral action against all arboviruses tested, while not affecting PIC infection. The combination of ribavirin and IFN-Alpha/Beta provided a synergistic antiviral effect against BZ, SF, and LAC; tumor necrosis factor (TNF) and IFN-Alpha/Beta provided greater than additive antiviral effect against BZ, while the combination of interleukin-2 with IFN-Alpha/Beta erratic results against the same viruses. Finally, successful combined therapy of SF or BZ infected mice was achieved with poly I:CLC and antibodies, when the intervention was started at the time of virus spread to the target organs. This treatment provided a strong, long lasting immunity in treated animals. Keywords: Immunomodulators; Antivirals; Arboviruses; Arenaviruses; Combination therapy; RAI.

The first major finding was the appearance of Dengue antigens on the surface of infected cells. This was of major importance since it has been hypothesized that dengue hemorraghic fever (DHF) is the result of DEN antigen interaction with anti Dengue antibody resulting complement mediated vascular endothelial injury. Findings then led us to postulate that Dengue antigens on the cell surface were composed of ordered arrays of Dengue polypeptides. As a first approach we proposed to examine the surface of the virus itself and then proceed to a study of the cell surface. To do this we studied the growth of Dengue in BHK cells. Our findings established new growth conditions and assay procedures for the virus. Since our approach was to use crosslinking reagents to study the organization of the Dengue polypeptides we examined the charge properties of viral glycoproteins on two other togaviruses. Results of these studies showed that the glycoproteins of two closely related viruses were quite different but that these differences probably would not significantly alter the organization of polypeptides on the surface of the virion. Protein-protein interactions on Japanese encephalitis virus (JEV), a virus closely related to Dengue, were analyzed. These results demonstrated that the major viral glycoprotein on the surface of the virion had as its nearest neighbor either one of its kind or another newly defined viral glycoprotein. Thus the clustering of viral antigens on flaviviruses had been demonstrated., Supersedes AD-A124 306.

Several mosquito cell proteins have been identified as being associated with (or incorporated in) Banzi virus grown in the polyclonal cell line of A. albopictus cells. This fact has been demonstrated by several observations. First, Banzi virions purified by polyethylene glycol precipitation and equilibrium density sucrose gradient centrifugation contain four mosquito cell protein. Second, anti-mosquito cell serum aggregates mosquito cell-grown Banzi virus, but not Banzi virus propagated in BHK cells. Third, anti-mosquito cell serum blocks the agglutination of erythrocytes by purified Banzi virus. Uninfected mosquito cells secrete a complex of proteins which is able to agglutinate goose erythrocytes. The density of the complex and the pH optimum of its hemagglutinating activity are different than that of Banzi virus grown in mosquito cells for 24 hours. Upon prolonged infection of mosquito cells with Banzi virus, the progeny virions differ in several respects. An apparent increase in the amount of one cellular protein is accompanied by a change in the pH optimum of agglutination of erythrocytes by Banzi virus. As a result of the incorporation of mosquito cell proteins into Banzi virions, mice immunized with uninfected mosquito cells or with purified A. albopictus hemagglutinin are protected against death due to Banzi viral infection.

Monoclonal MC antibodies specific for the E1 and E2 envelope glycoproteins of AR 339 strain Sindbis virus (SV) have been tested individually for their ability to passively protect adult C57BL/6J mice from fatal paralytic disease produced by a neuroadapted variant (na-SV) of the non-lethal parent (p) strain. To date, protection has been obtained with 3 anti-E1 MC antibodies and is associated with their capacity to neutralize infectivity of na-SV, in addition to that of p-SV; three others which neutralize only p-SV are non-protective. In the presence of complement (C), anti-E2 IgG2a MC antibodies exhibit greater virus-neutralizing activity and they also mediate C-dependent lysis of cultured SV-infected cells. Results suggested at least two specific humoral mechanisms of alphavirus elimination in vivo: (1) neutralization of infectivity by antibodies binding to E2 determinants on free virions, a process enhanced by, but not requiring C, and (2) C-dependent lysis of virus-infected cells mediated by antibodies which bind to either E1 or E2 determinants expressed on cell surfaces during viral morphogenesis. On the other hand, the possibility that a waning humoral immunity may increase host-susceptibility to reinfection is suggested by the ability of low concentrations of E2-specific antibodies to enhance SV infectivity in vitro.