The objective of this proposal is to study the molecular basis for the decreased ability of the immune system in many elderly individuals, to produce high affinity antibodies and thus mount an effective immune response to vaccines or infections. To pursue this objective, the principal investigator will study the age-associated changes in the relationship between structure and function of immunoglobulin VH and VL genes which produce the natural anti-Gal antibody. Anti-Gal constitutes 1% of circulating IgG in humans and interacts specifically with the carbohydrate structure Gala1-3GalB1-4GlcNAc-R (termed alpha-galactosyl epitope). As many as 1% of circulating B lymphocytes can produce anti- Gal. The applicant has found that most of the sequenced VH genes in anti-Gal producing lymphoid clones, cluster within a distinct region of the VH3 immunoglobulin gene family, and that these genes undergo somatic mutations. Analysis of age-associated changes in activity of anti-Gal have demonstrated loss of affinity of this antibody in many elderly individuals. The principal investigator proposes to study two possible mechanisms which may cause the age-associated impairment in anti-Gal affinity in humans: 1) different utilization of VH and VL genes repertoire for anti-Gal synthesis; and 2) impaired affinity maturation in anti-Gal genes because of age-associated changes in the somatic mutations process. For this purpose he will construct combinatorial phage display libraries in young and in elderly individuals and isolate the anti-Gal displaying phage. Comparative analysis of the anti-Gal VH and VL gene repertoire in the young and elderly, and determination of the resulting antibody affinities, will enable the applicant to establish whether the age-associated decrease in antibody affinity correlates with the gene repertoire used for anti-Gal synthesis. Comparison of the V gene sequences with the corresponding germline genes will allow the investigators to establish whether the age-associated changes reflect the excess or paucity of somatic mutations in anti-Gal producing genes. The applicant will further determine whether elderly individuals with low affinity serum anti-Gal retain some high affinity clones, by selecting high affinity anti-Gal phage from the libraries of these individuals. Lastly, by correlating anti-Gal affinity with the immune response to an influenza virus vaccine, the principal investigator will determine whether anti-Gal can predict the competence of the immune system in elderly individuals to produce high affinity antibodies. Thus, by exploiting the anti-Gal system, the applicant aims to understand the molecular basis of the well-documented age-associated impairment of humoral immune responses in humans.
|Wang, L; Radic, M Z; Siegel, D et al. (1997) Cloning of anti-Gal Fabs from combinatorial phage display libraries: structural analysis and comparison of Fab expression in pComb3H and pComb8 phage. Mol Immunol 34:609-18|