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Updated: May 20, 2026
Immunoglobulin Gene Sequence Analysis In Chronic Lymphocytic Leukemia: From Patient Material To Sequence Interpretation
Published on: November 26, 2018
The identification of irreversible rituximab-resistant lymphoma caused by CD20 gene mutations
Abstract:
C-terminal mutations of CD20 constitute part of the mechanisms that resist rituximab therapy. Most CD20 having a C-terminal mutation was not recognized by L26 antibody. As the exact epitope of L26 has not been determined, expression and localization of mutated CD20 have not been completely elucidated. In this study, we revealed that the binding site of L26 monoclonal antibody is located in the C-terminal cytoplasmic region of CD20 molecule, which was often lost in mutated CD20 molecules. This indicates that it is difficult to distinguish the mutation of CD20 from under expression of the CD20 protein. To detect comprehensive CD20 molecules including the resistant mutants, we developed a novel monoclonal antibody that recognizes the N-terminal cytoplasm region of CD20 molecule. We screened L26-negative cases with our antibody and found several mutations. A rituximab-binding analysis using the cryopreserved specimen that mutation was identified in CD20 molecules indicated that the C-terminal region of CD20 undertakes a critical role in presentation of the large loop in which the rituximab-binding site locates. Thus, combination of antibodies of two kinds of epitope permits the identification of C-terminal CD20 mutations associated with irreversible resistance to rituximab and may help the decision of the treatment strategy.
Insights
C-terminal mutations in CD20 can cause resistance to rituximab therapy by altering antibody binding sites. A new antibody targeting the N-terminal region helps identify these resistant CD20 mutations.
Area of Science:
- Immunology
- Molecular Biology
- Oncology
Background:
- Rituximab resistance in B-cell malignancies is often linked to mutations in the CD20 protein.
- The L26 antibody epitope on CD20 has not been fully characterized, complicating the study of C-terminal mutations.
- Distinguishing CD20 mutations from underexpression is challenging with existing antibodies.
Purpose of the Study:
- To determine the L26 antibody binding site on CD20.
- To develop a novel antibody for detecting CD20 mutants resistant to rituximab.
- To investigate the role of the CD20 C-terminal region in rituximab binding.
Main Methods:
- Epitope mapping of the L26 monoclonal antibody.
- Development and application of a novel N-terminal targeting CD20 antibody.
- Rituximab-binding assays on patient-derived cryopreserved specimens.
Main Results:
- The L26 epitope is located in the C-terminal cytoplasmic region of CD20.
- A novel antibody targeting the N-terminal region successfully identified L26-negative CD20 mutants.
- CD20 C-terminal mutations critically affect the presentation of the rituximab-binding site.
Conclusions:
- The C-terminal region of CD20 is crucial for rituximab binding.
- Combined use of N-terminal and C-terminal targeting antibodies can identify CD20 mutations associated with rituximab resistance.
- This approach may aid in guiding treatment strategies for patients with resistant B-cell malignancies.
