Targeting antigen-specific receptors on B lymphocytes to generate high yields of specific monoclonal antibodies directed against biologically active lower antigenic peptides within presenilin 1

Tomita, Masahiro; Sugi, Hiroyuki; Ozawa, Kazuharu; Tsong, Tian Yow; Yoshimura, Teizo

doi:10.1016/s0022-1759(01)00299-x

Cited by 12 publications

(17 citation statements)

References 39 publications

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“…BSANPs modification with MBS was performed as described by Masahiro Tomita and coworkers. 27 BSANPs were first activated by heterobiofunctional cross-linker, MBS (10 mg/ml in DMF) for half an hour at room temperature with gentle agitation. After activation, the suspension was incubated with cyclic RGD peptide at a molar ratio of 10:1 for 3 h at room temperature.…”

Section: Methodsmentioning

confidence: 99%

RGD-conjugated albumin nanoparticles as a novel delivery vehicle in pancreatic cancer therapy

Zhang

et al. 2012

Cancer Biology & Therapy

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These authors contributed equally to this work. Keywords: nanoparticles, albumin, RGD, FITC, delivery, pancreatic cancer, therapyIntegrin avb3 receptor is expressed on several types of cancer cells, including pancreatic cancer cells, and plays an important role in tumor growth and metastasis. The ability to target the integrin avb3 receptor on cancer cells increases the efficacy of targeted therapy and reduces the side effects. The aim of this study is to develop a novel arginine-glycineaspartic acid (RGD) peptide-conjugated albumin nanoparticle to enhance the intracellular uptake of anticancer drug into the pancreatic cancer cells through receptor-mediated endocytosis. In cellular uptake studies, the fluorescent signal of RGD-conjugated BSANPs in BxPC3 cells was higher than that of BSANPs without RGD conjugation as determined by fluorescence spectrophotometer. We also found that BSANPs bound to BxPC3 cells in a time-and concentration-dependent manner. The uptake of RGD-conjugated BSANPs by pancreatic cancer cells was inhibited by an excess amount of free RGD peptide, indicating that the binding and/or uptake were mediated by the avb3 receptor. Furthermore, the nanoparticles were found to be located close to the nuclei by using laser scanning confocal microscopy. Besides, no significant in vitro cytotoxicity was observed as measured with MTT assay. Both in vitro and in vivo antitumor efficacy was improved by targeting gemcitabine-loaded nanoparticles to BxPC-3 cells using RGD peptides. Therefore, the RGD-conjugated BSANPs hold great potential as an effective drug delivery system to deliver therapeutic agents to pancreatic cancer.

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Section: Methodsmentioning

confidence: 99%

RGD-conjugated albumin nanoparticles as a novel delivery vehicle in pancreatic cancer therapy

Zhang

et al. 2012

Cancer Biology & Therapy

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“…(5,10) In vivo immunization BALB/c mice at 4-to 10-weeks of age were primed with 40 g of human insulin emulsified in Freund's complete adjuvant (FCA) (ratio 1:1) and injected intraperitoneally. The mice were given a second injection of 30 g human insulin emulsified in Freund's incomplete adjuvant (FIA) at the ratio of 1:1, and two or three additional intraperitoneal injections of 20 g human insulin were carried out at 1-to 2-week intervals.…”

Section: Preparation Of Spleen Cellsmentioning

confidence: 99%

“…(10)(11)(12) B-cell targeting can yield monoclonal antibodies against antigens with high efficiency and specificity and provide a versatile methodology. Theoretically, any desired antigens can be targets and the advantages of this new technique are that the immunized B lymphocytes can be strictly targeted on the basis of surface immunoglobulin receptors, and that the antigen-selected B lymphocytes can be selectively fused with myeloma cells by application of an electrical pulse.…”

Section: Introductionmentioning

confidence: 99%

“…This new technology has also been verified by other groups. (13)(14)(15) Lately, we have succeeded in generating monoclonal antibodies to functional peptide sequences of presenilin 1, (10) that is believed to be related to the early onset of familial Alzheimer's disease. Until now, however, no direct evidence or proof of an entire pathway has been presented, except a preliminary result indicating pre-selection of B lymphocytes by an antigen.…”

Section: Introductionmentioning

confidence: 99%

“…Until now, however, no direct evidence or proof of an entire pathway has been presented, except a preliminary result indicating pre-selection of B lymphocytes by an antigen. (10) To address this point and further develop this promising technology, we here used immunofluorescence analysis, focusing on theoretical quantification of fusion efficiency.…”

Section: Introductionmentioning

confidence: 99%

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Antigen-Based Immunofluorescence Analysis of B-Cell Targeting: Advanced Technology for the Generation of Novel Monoclonal Antibodies with High Efficiency and Selectivity

et al. 2006

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On the basis of immunofluorescence analysis, an entire pathway of B-cell targeting was successfully identified, which can drive selective production of monoclonal antibodies with high efficiency and selectivity. The technique comprises three critical steps, antigen-based preselection of B lymphocytes, formation of antigenselected B lymphocyte and myeloma cell complexes, and selective fusion of B-cell-myeloma cell complexes with electrical pulses. Intriguingly, expression of surface immunoglobulin receptors on B lymphocytes was recognized even after immunization in vitro. The number of the antigen-selected B lymphocytes after in vitro immunization was in fact higher than that obtained after in vivo immunization, suggesting that such shortterm immunization is applicable for B-cell targeting. Immunofluorescence analysis revealed the targeting technique to demonstrate fivefold to tenfold higher efficiency for formation of hybridoma cells than a polyethylene glycol (PEG)-mediated method. This efficiency is in good agreement with production of hybridoma cells secreting desired monoclonal antibodies determined by an enzyme-linked immunosorbent assay (ELISA). The addition of a low concentration of PEG brought about enhanced fusion efficiency and reduced cell damage at even electric intensities as high as 4.0 kV/cm and 5.0 kV/cm. Here we demonstrate that immunofluorescence analysis can successfully clarify an entire pathway of B-cell targeting and provide advantages over the PEG-mediated method. This advanced technology may be applicable for rapid production of monoclonal antibodies based on in vitro immunization.

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Conformation-specific monoclonal antibodies recognizing the native structure of G protein-coupled receptor (GPCR)

Isozaki

Tsumoto

Tomita

2021

International Immunopharmacology

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Targeting antigen-specific receptors on B lymphocytes to generate high yields of specific monoclonal antibodies directed against biologically active lower antigenic peptides within presenilin 1

Cited by 12 publications

References 39 publications

RGD-conjugated albumin nanoparticles as a novel delivery vehicle in pancreatic cancer therapy

RGD-conjugated albumin nanoparticles as a novel delivery vehicle in pancreatic cancer therapy

Antigen-Based Immunofluorescence Analysis of B-Cell Targeting: Advanced Technology for the Generation of Novel Monoclonal Antibodies with High Efficiency and Selectivity

Conformation-specific monoclonal antibodies recognizing the native structure of G protein-coupled receptor (GPCR)

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