Isotype Switching Increases Efficacy of Antibody Protection against Cryptococcus neoformans Infection in Mice

The isotype and epitope specificities of antibodies both contribute to the efficacy of antibodies that mediate immunity to Cryptococcus neoformans, but the relationship between these properties is only partially understood. In this study, we analyzed the efficacy of protection of two sets of immunoglobulin G (IgG) isotype switch variants from two IgG3 monoclonal antibodies (MAbs) which are either not protective or disease enhancing, depending on the mouse model used. The two IgG3 MAbs 3E5 and 4H3 have different epitope specificities. Protection experiments were done with A/JCr mice infected intravenously with C. neoformans and administered with 3E5 IgG3 and its IgG1, IgG2a, and IgG2b switch variants. These experiments revealed that IgG1, IgG2b, and IgG2a were each more effective than IgG3. For 4H3 IgG3 and its IgG1 and IgG2b switch variants, the relative efficacy was IgG2b > IgG1 >> IgG3. The combination of 3E5 IgG3 and 4H3 IgG3 was more deleterious than either IgG3 alone. All IgG isotypes were opsonic for mouse bronchoalveolar cells, with the relative efficacy being IgG2b > IgG2a > IgG1 > IgG3. These results (i) confirm that a nonprotective IgG3 MAb can be converted to a protective MAb by isotype switching, (ii) indicate that the efficacy of protection of an IgG1 MAb can be increased by isotype switching to another subclass, (iii) show that protective and nonprotective IgG MAbs are opsonic, and (iv) provide additional evidence for the concept that the efficacy of the antibody response to C. neoformans is dependent on the type of MAb elicited.Cryptococcus neoformans is a fungus which is a frequent cause of life-threatening meningoencephalitis in patients with impaired immunity (22, 25). Cryptococcosis has been reported to occur in 6 to 8% of patients with AIDS (7). In immunocompromised individuals, C. neoformans infections are often incurable with conventional antifungal agents, and these patients frequently require lifelong therapy (45). The difficulties involved in the management of cryptococcosis in immunocompromised individuals have led to a reexamination of the potential of antibody-mediated immunity for prevention and therapy of cryptococcal infections. A polysaccharide-tetanus toxoid (TT) conjugate vaccine which is highly immunogenic and can elicit protective antibodies in mice has been made (3, 8, 9). In addition, several monoclonal antibodies (MAbs) have been shown to modify the course of infection in mice, and these may be useful in therapy of human infection (12, 14, 28, 42, 43).Cell-mediated immunity is generally acknowledged to provide important host defense against C. neoformans infection (4, 20, 26, 31, 42). In contrast, the role of antibody-mediated immunity in host resistance is less certain (2), but there is considerable evidence that administration of some MAbs can modify the course of infection in mice (8, 12, 14, 16, 28, 33). C. neoformans is unusual among fungal pathogens in that it has a polysaccharide capsule composed primarily of glucuronoxylomannan (GXM) (6), which is important for virulence (5). The capsular polysaccharide has been shown to produce a variety of deleterious effects including inhibition of phagocytosis (21), interference with antigen presentation (39), shedding of adhesion molecules (11), inhibition of leukocyte migration (10), and alterations in cytokine production by host effector cells (24, 40, 41). Antibodies to the C. neoformans capsular polysaccharide may contribute to host defense through multiple effects including enhanced opsonization (13, 18, 23, 30, 44), clearance of polysaccharide antigen (15), promotion of granuloma formation (14), and release of oxygen- and nitrogen-derived oxidants (27, 38).In previous studies, we demonstrated that immunoglobulin G3 (IgG3) MAbs are not protective in various mouse models of cryptococcal infection (32, 42). When one of these nonprotective IgG3 MAbs was switched to IgG1, the IgG1 significantly prolonged animal survival (32, 42). In the present study, we analyzed two families of IgG switch variants generated in vitro from two nonprotective IgG3 MAbs with different epitope specificities. We found that MAbs with different isotypes have different protective efficacies and that switching of nonprotective IgG3 MAbs to IgG1, IgG2b, and IgG2a significantly increased antibody protective efficacy. These studies demonstrate a complex relationship among efficacy of antibody protection, epitope specificity, and isotype.