The effect of B-cell depletion with an anti-CD20 antibody on the immunoglobulin heavy-chain repertoire in a patient with rheumatoid arthritis

Purpose

B-cell depletive therapy has been described to have beneficial effects to patients suffering from rheumatoid arthritis (RA). Nevertheless, the role of B cells in pathogenesis is not clear. In particular, it is not known whether B-cell depletion causes changes in the B-cell repertoire after peripheral regeneration. Therefore we analysed the distribution of the immunoglobulin heavy-chain (Ig-VH) genes usage of a RA patient before, 7 months and 17 months after anti-CD20 therapy with rituximab.

Methods

A patient with active RA has been treated with four weekly doses of 375 mg/m² rituximab. During the treatment phase, prednisolone was elevated up to 12.5 mg daily for 4 weeks. After that, the patient remained on a stable dose of 5 mg prednisolone per day without further anti-proliferate treatment. Ig-VH genes were amplified from genomic DNA of peripheral B cells by nested PCR using family-specific oligonucleotides. PCR products were subcloned and the sequences of more than 500 clones were analysed.

Results

Following antibody treatment, the patient experienced a good clinical response lasting over 1 year. Before therapy, the overall VH gene usage of the RA patient was largely similar to the distributions expected from published data for normal individuals, except for certain genes (4-34, 1-69, 3-07), which have already been reported with a biased distribution in autoimmune disorders. In addition, several clonally related sequences were found in the VH5 family.

B-cell regeneration occurred 7 months after anti-CD20 antibody therapy. Significant changes in the distribution of different genes could be observed: 4-34 and 1-69 decreased to a proportion that would be expected in healthy individuals. Clonally related sequences were not found anymore. A greater overall variety of the VH gene segments used was observed, in particular in the larger families like VH3 and VH4. Interestingly, somatic hypermutation was significantly enhanced at the time point of B-cell regeneration.

Seventeen months after therapy, the general distribution of the VH genes was comparable with that observed before treatment, except for some genes (4-34 remained at the level found after therapy). No clonally related sequences were found. Somatic hypermutation was significantly reduced to the frequency determined before therapy.

Conclusion

B-cell depletion by anti-CD20 monoclonal antibody leads to a significant modulation of the expressed immunoglobulin-VH gene repertoire after B-cell regeneration, which is accompanied with a clinical benefit in a single patient.