CD20 Overexpression Lysate

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

CD20 is a non-glycosylated phosphoprotein that is expressed on the surface of normal and malignant B cells and functions in mediating calcium transport and B cell differentiation (1,2). CD20 is encoded by the membrane-spanning 4-domain family A member 1 (MS4A1) gene and, in humans, is located on chromosome 11q12 (1). The CD20 protein is 297 amino acids (aa) in length with a theoretical molecular weight (MW) of 33 kDa (1,2). Structurally, the CD20 protein has four membrane-spanning domains, two extracellular loop domains, and intracellular N- and C-terminal domains (1,2). CD20 is expressed at specific stages of B cell maturation including pre-B cells, mature naive and activated B cells, and memory B cells, but is absent from plasmablasts and plasma cells (1,3,4). Expression of CD20 is often increased on malignant B cells associated with various B cell disorders such as chronic lymphocytic leukemia (CLL), multiple sclerosis (MS), and rheumatoid arthritis (2-4). Anti-CD20 monoclonal antibody (mAb)-based therapies have become an appealing target for treating these immune-related disorders and cancers (1-5). Rituximab, a chimeric mAb, was the first FDA approved CD20 monoclonal antibody for the treatment of non-Hodgkin's lymphoma that is now commonly used to treat MS (2,3). Since its initial approval in 1997, several other chimeric and humanized anti-CD20 mAbs have been developed including Ofatumumab, Ublituximab, and Obinutuzumab (1-5). B cell depletion via CD20 mAbs can occur under different mechanisms such as antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), complement-dependent cytotoxicity (CDC), and direct induction of apoptosis (1-4). Many ongoing studies are focused on combination therapies with CD20 mAbs as an addition to chemotherapy, B cell receptor (BCR) signaling inhibitors, or BH3 mimetics (1,2,4). Additionally, the effects of bispecific antibodies and CD20 chimeric antigen receptor (CAR) T cell therapies are under investigation for the treatment of B cell malignancies (4,5).

References

1. Pavlasova G, Mraz M. The regulation and function of CD20: an "enigma" of B-cell biology and targeted therapy. Haematologica. 2020; 105(6):1494-1506. https://doi.org/10.3324/haematol.2019.243543

2. Payandeh Z, Bahrami AA, Hoseinpoor R, et al. The applications of anti-CD20 antibodies to treat various B cells disorders. Biomed Pharmacother. 2019; 109:2415-2426. https://doi.org/10.1016/j.biopha.2018.11.121

3. Margoni M, Preziosa P, Filippi M, Rocca MA. Anti-CD20 therapies for multiple sclerosis: current status and future perspectives. J Neurol. 2022; 269(3):1316-1334. https://doi.org/10.1007/s00415-021-10744-x

4. Klein C, Jamois C, Nielsen T. Anti-CD20 treatment for B-cell malignancies: current status and future directions. Expert Opin Biol Ther. 2021; 21(2):161-181. https://doi.org/10.1080/14712598.2020.1822318

5. Sharman JP. Targeting CD20: teaching an old dog new tricks. Hematology Am Soc Hematol Educ Program. 2019; 2019(1):273-278. https://doi.org/10.1182/hematology.2019000031

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Array NBL1-13311

• CD20 Antibodies
• CD20 Antibody Pair
• CD20 ELISA Kits
• CD20 Lysates
• CD20 Proteins