Abstract
The B-cell receptor (BCR) is a key player of the adaptive immune system. It is a unique part of immunoglobulin (Ig) molecules expressed on the surface of B cells. In case of many B-cell lymphomas, the tumor cells express a tumor-specific and functionally active BCR, also known as idiotype. Utilizing the idiotype as target for lymphoma therapy has emerged to be demanding since the idiotype differs from patient to patient. Previous studies have shown that shark-derived antibody domains (vNARs) isolated from a semi-synthetic CDR3-randomized library allow for the rapid generation of anti-idiotype binders. In this study, we evaluated the potential of generating patient-specific binders against the idiotype of lymphomas. To this end, the BCRs of three different lymphoma cell lines SUP-B8, Daudi, and IM-9 were identified, the variable domains were reformatted and the resulting monoclonal antibodies produced. The SUP-B8 BCR served as antigen in fluorescence-activated cell sorting (FACS)-based screening of the yeast-displayed vNAR libraries which resulted after three rounds of screening in the enrichment of antigen-binding vNARs. Five vNARs were expressed as Fc fusion proteins and consequently analyzed for their binding to soluble antigen using biolayer interferometry (BLI) revealing binding constants in the lower single-digit nanomolar range. These variants showed specific binding to the parental SUP-B8 cell line confirming a similar folding of the recombinantly expressed proteins compared with the native cell surface-presented BCR. First initial experiments to utilize the generated vNAR-Fc variants for BCR-clustering to induce apoptosis or ADCC/ADCP did not result in a significant decrease of cell viability. Here, we report an alternative approach for a personalized B-cell lymphoma therapy based on the construction of vNAR-Fc antibody-drug conjugates to enable specific killing of malignant B cells, which may widen the therapeutic window for B-cell lymphoma therapy.
Highlights
In recent years, monoclonal antibodies have emerged as therapeutics with exceptional relevance for inflammatory and infectious diseases and the treatment of cancer [1]
Since we could not observe the induction of apoptosis via clustering of B-cell receptor (BCR) molecules in first initial experiments, we developed an alternative approach by generating vNAR antibody-drug conjugates that induced specific killing in lymphoma cells at low nanomolar concentrations
Afterwards, the synthesized cDNA served as template for amplification of VH and VL genes using primers reported in a study by Maleshko et al [34] that elucidated a specially designed panel of primers for the rapid amplification of variable regions of tumor immunoglobulins
Summary
Monoclonal antibodies have emerged as therapeutics with exceptional relevance for inflammatory and infectious diseases and the treatment of cancer [1]. The number of antibodies in late-stage clinical trials has substantially increased to over 50 in the year 2017. In this year the number of novel antibody therapeutics that were approved in either United States or the European Union reached double-digit numbers for the first time [2]. Rituximab is active in a diversity of human lymphomas and chronic lymphocytic leukemia [3]. Rituximab is a chimeric mAB targeting the CD20 antigen present on both healthy B cells as well as on most B-cell lymphomas
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