Rearrangement of only one human IGHV gene is sufficient to generate a wide repertoire of antigen specific antibody responses in transgenic mice

In recent years, mice carrying human IG transgenes are being generated for the production of human monoclonal antibodies as an alternative approach to the conventional use of mouse or chimeric-humanized antibodies. Theoretically, the size of the repertoire of human antibodies that these mice could produce would be critically dependent on the number of human V genes introduced in the transgene. This could be the case for BABκ and BABκ,λ transgenic mice, which carry several genes from the human IGK (BABκ), and IGK and IGL (BABκ,λ) loci, but only five human IGHV genes and the entire IGHD-IGHJ cluster linked to two human IGHC (IGHM-IGHD) genes. We analyzed the expressed human IG genes in 30 IgM-secreting hybridomas generated from transgenic mice immunized either with soluble proteins (human IgM coupled to KLH) or with cells (human PBMC, tumour cell lines or rat cells transfected with human CD69). The results show that all hybridoma cells analyzed rearranged exclusively the IGHV1-2 gene, in contrast with naive spleen B cells that used three out of the five IGHV genes present in the transgene. The configuration of the rearranged CDR3 region revealed a much higher heterogeneity in the heavy chains. A variety of IGHJ and IGHD genes were used in hybridomas, and somatic mutations were also seen in some hybrids. Regarding the rearranged light chains genes, it was a much higher variety in the use of V and J genes in both, κ and λ chains, than in the heavy chain, and also in the level of mutation. The results indicate that only one IGHV gene is sufficient to generate a wide repertoire of antigen specific antibody responses. Thus, efforts aimed at the generation of new transgenic mice should focus more on the integrity of the D/J region and on the DNA regions regulating somatic hypermutation, rather than on the number of V genes present in the transgene.