of applicant): Human antibodies to capsular polysaccharide of Streptococcus pneumoniae (PPS) express genes from the VH3 gene subgroup. SinceVH3 gene expression is often reduced in the setting of HIV infection and PPS-derived vaccines are poorly immunogenic in HIV-infected individuals, we have hypothesized that decreased expression of the immunoglobulin genes used in antibodies to PPS contributes to PPS vaccine failure in the setting of HIV infection. The structure-function relationship of human antibodies to most pneumococcal serotypes has not been fully investigated. The studies that are proposed in this application are designed to characterize the variable gene use and epitope specificity of human antibodies to two common serotypes of S. pneumoniae, serotypes 8 and 23F, and to use this information to develop probes to determine the specificity of vaccine elicited antibodies to PPS 8 and 23F from individuals with and without HIV infection.
The specific aims are: 1) To determine the molecular structure of human Mab to PPS 8 and 23F generated in transgenic mice reconstituted with human immunoglobulin loci; 2) To characterize the Mabs to PPS 8 and 23F produced in aim one as protective, non-protective or disease enhancing against pneumococcal infection in mice and as opsonic in vitro; 3 ) To use Mabs with functional efficacy as defined in aim two to isolate peptide mimetics of PPS 8 and 23F epitopes, and to use the peptides to determine if PPS-elicited antibodies in patients sera recognize protective, non-protective or disease enhancing epitopes. Studies are proposed to examine the hypothesis that protective antibodies to PPS can be distinguished by their variable gene use and specificity and that the production of antibodies to certain epitopes ,which are derived from VH3 genes, is impaired in the setting of HIV infection. Knowledge gained from these studies will help us to understand the structure-function relationship for antibodies to S. pneumoniae in patients with HIV infection, and to identify possible correlates of PPS vaccine failure and the requirements for more effective anti-pneumococcal vaccines. Significance: Infection with pneumococcus has become a major cause of morbidity and mortality in AIDS. In the past, AIDS-associated infections were easily treated with antibiotics however, penicillin resistant isolates have emerged in the 1990's. This emergence of antibiotic resistant pneumococcus has increased the need for vaccination of high-risk groups such as HIV+ persons. However, available pneumococcal vaccines are poorly immunogenic in HIV+ persons. Thus, the goal of this proposal, which is to identify the molecular and biological basis of human antibody responses to peumococcal vaccination in normal volunteers and HIV+ persons, is very significant. The information gained from the proposed studies could lead to better vaccines for pneumococcus in immunocompromised hosts and importantly, will contribute to the understanding of how antibodies function in resistance to S. pneumoniae. Approach: This is the second revision of this application. Since the last review of this application the author has published 1 manuscript and has two others in press having to do with this application. The first publication describes the generation of a protective human Mab to PPS 8. The second, describes the generation of human antibodies to PPS 3 in XenoMouse mice. The third describes the molecular response of HIV-infected and HIV-uninfected subjects to Pneumovax. Thus, the author has made excellent progress even though she is not funded to do this work. This indicates commitment, enthusiasm and capability of the author to carry out this work. he P.I. has responded to the previous criticisms and has written a strong and significant application. One previous concern was that if HIV-infected individuals can not produce VH3 and only VH3 ab are protective that little of this information will be useful to make patients more resistant to pneumococcus. The PI points out that the goal of this application is not to characterize VH3 expression in HIV-infected individuals, but to determine whether they do or do not produce antibodies to pneumococcal capsular polysaccharides with defined epitope specificity. The PI's response and revisions to other reviewer's comments also strengthen the 1 ZRG-1 AARR- 4 (01) 3 1 R01 AI45459-01A2 March 2000 Pirofski, Liise-Anne proposal. In addition, because of the subject of this application, the initial applications were difficult to follow. This revised version is clearer and the additions the PI has made are helpful. It is much more apparent what the PI really wants to accomplish. This is important since the reviewers agreed that this was a difficult application to read, mainly because of the subject matter and the specialized nature of the work. Innovation: The overall approach utilized in solving this problem takes advantage of established methods. The author has used these methods in the study of Cryptococcus so that the overall strategy is somewhat proven. Use of the XenoMouse is considered innovative. There are relatively few laboratories interested in antibody-mediated resistance. This makes this work somewhat unique. Investigator: Dr. Pirofski is well qualified to carry out the proposed research. She has published studies utilizing most of the methods in the proposal. Dr. John McKitrick is a consultant and brings expertise in the microbiology of S. pneumoniae. Environment: The P.I. is at Albert Einstein and has several other researchers nearby that have expertise in many of the methods she will utilize. She is in an excellent environment to accomplish this proposal. In summary, the subject of this proposal is timely and significant. The emergence of antibiotic resistant S. pneumoniae infections in AIDS patients requires the development of more efficient means of immunizing these individuals. The PI has made excellent progress on this project despite not being funded in this area. Studies proposed in this application could provide valuable information for developing more immunogenic vaccines for HIV+ persons. Simply put, this work needs to be done.
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