Generation and preliminary characterization of an antibody library with preferential reactivity to human colorectal cancer cells as compared to normal human blood cells

Williams, Brent R.; Daley, Kylle M.; Sharon, Jacqueline

doi:10.1016/j.imlet.2003.12.002

Cited by 11 publications

(12 citation statements)

References 28 publications

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“…The results showed that Lib-Col2.1 bound to SW480 cells at higher density than MAb CO17-1A and that its binding to normal human blood cells was reduced 2.4-24 fold relative to an unabsorbed anti-SW480 serum. Furthermore, Lib-Col2.1 was more effective than MAb CO17-1A at inhibiting the growth of SW480 cells in culture [Liebman et al, 2004].…”

Section: Prototypic Pcals To Human Colorectal Cancermentioning

confidence: 90%

“…In addition to production of MAbs, our laboratory has adapted the Fab phage display system to generate polyclonal antibody libraries (PCALs) [Sharon et al, 2002;Williams and Sharon, 2002a;Williams et al, 2002b;Chen et al, 2003a,b;Liebman et al, 2004]. Repertoires of expressed light and heavy chain variable (VL and VH) region genes are obtained by RT-PCR from B cell and plasma cell RNA derived from immune humans or experimental animals, and cloned as VL-VH region gene pairs into a phage display vector to generate a combinatorial Fab phage display library.…”

Section: Polyclonal Antibody Libraries (Pcals)mentioning

confidence: 99%

“…The absorbed phage as well as a sample of the unabsorbed phage were then separately subjected to positive selection on the SW480 colon cancer cell line, in native form, by the density gradient centrifugation method [Williams and Sharon, 2002a]. This yielded a negatively selected (absorbed) library of 9.2 Â 10 4 members and a paired, non-negatively selected (unabsorbed) library of 3.8 Â 10 5 members [Liebman et al, 2004], demonstrating a fourfold reduction in library size due to negative selection.…”

Section: Prototypic Pcals To Human Colorectal Cancermentioning

confidence: 99%

“…The H and L chain V region gene pairs of the absorbed library were then transferred in mass from the Fab phage display vector to a mammalian vector and expressed as murine IgG2b following transfection, by spheroplast fusion, into Sp2/0 myeloma cells [Liebman et al, 2004]. Transfected cells were plated in 96-well plates, and ELISA analysis of cell supernatants showed that 79% of transfectants expressed IgG and of those 74% were positive for binding to the SW480 human colorectal cancer cell line.…”

Section: Prototypic Pcals To Human Colorectal Cancermentioning

confidence: 99%

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Recombinant polyclonal antibodies for cancer therapy

Sharon¹,

Liebman²,

Williams³

2005

J of Cellular Biochemistry

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Although monoclonal antibodies are increasingly used for cancer therapy, remissions are only temporary due to emergence of tumor cell escape variants that are no longer affected by the antibody. The emergence of escape variants could be minimized by multi-targeting of tumor cells with polyclonal antibodies, which would also be more efficient than monoclonal antibodies at mediating effector functions for target destruction. A technology for generating recombinant polyclonal antibodies for cancer therapy has been developed based on the construction and selection of tumor-reactive Fab phage display libraries. The selected Fabs are mass-converted to full-length polyclonal antibody libraries (PCALs) of any isotype and any species. Prototypic PCALs generated against human colorectal cancer cell lines showed that libraries of diverse recombinant antibodies, enriched for reactivity to the cancer cells compared to normal human cells, can be obtained. The success of recombinant polyclonal antibodies as cancer therapeutics will depend on the ability to generate, characterize, and mass-produce PCALs with high ratios of cancer-to-normal reactivities that crossreact with many cancers of the same type.

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Section: Prototypic Pcals To Human Colorectal Cancermentioning

confidence: 90%

Section: Polyclonal Antibody Libraries (Pcals)mentioning

confidence: 99%

Section: Prototypic Pcals To Human Colorectal Cancermentioning

confidence: 99%

Section: Prototypic Pcals To Human Colorectal Cancermentioning

confidence: 99%

See 2 more Smart Citations

Recombinant polyclonal antibodies for cancer therapy

Sharon¹,

Liebman²,

Williams³

2005

J of Cellular Biochemistry

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“…The CO17-1A murine hybridoma cell line, which produces a monoclonal antibody specific for the human tumor associated antigen EpCam (epithelial cell adhesion molecule) (107, 108), was obtained from Dr. Dorothee Herlyn of the Wistar Institute (Philadelphia, PA) and cultured at 37°C in a humidified environment of 5% CO 2 /95% air in IMDM (containing 3.7 g NaHCO 3 ) supplemented with 5 μg/ml gentamicin and 10% FBS. LibCol2.1, a mixture of cell lines produced by transfection of vectors encoding murine recombinant polyclonal antibodies into Sp2/0 murine myeloma cells [10] was previously described [11]. Lib-Col2.1 cells were cultured under the same conditions as the CO17-1A hybridoma cells, except that the medium was supplemented with hypoxanthine, mycophenolic acid, and xanthine at 15, 6, and 250 μg/ml, respectively.…”

Section: Mammalian Cells and Cell Culturementioning

confidence: 99%

Antibody treatment of human tumor xenografts elicits active anti-tumor immunity in nude mice

et al. 2007

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Athymic nude mice bearing subcutaneous tumor xenografts of the human anti-colorectal cancer cell line SW480 were used as a preclinical model to explore anti-tumor immunotherapies. Intratumor or systemic treatment of the mice with murine anti-SW480 serum, recombinant anti-SW480 polyclonal antibodies, or the anti-colorectal cancer monoclonal antibody CO17-1A, caused retardation or regression of SW480 tumor xenografts. Interestingly, when mice that had regressed their tumors were re-challenged with SW480 cells, these mice regressed the new tumors without further antibody treatment. Adoptive transfer of spleen cells from mice that had regressed their tumors conferred antitumor immunity to naïve nude mice. Pilot experiments suggest that the transferred anti-tumor immunity is mediated by T cells of both γδ and αβ lineages. These results demonstrate that passive anti-tumor immunotherapy can elicit active immunity and support a role for extra-thymic γδ and αβ T cells in tumor rejection. Implications for potential immunotherapies include injection of tumor nodules in cancer patients with anti-tumor antibodies to induce anti-tumor T cell immunity.

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Phage antibody library screening for the selection of novel high-affinity human single-chain variable fragment against gastrin receptor: an in silico and in vitro study

et al. 2019

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Background As a membrane G protein coupled receptors (GPCRs) family, gastrin/cholecystokinin-2 receptor (CCK2R) plays a key role in the initiation and development of gastric cancer. Objectives Targeting CCK2R by immunotherapeutics such as single-chain variable fragments (scFvs) may provide an effective treatment modality against gastric cancer. Thus, the main objective of this study was to isolate scFvs specific to CCK2R. Methods To isolate scFvs specific to the CCK2R, we capitalized on a semi-synthetic diverse phage antibody library (PAL) and a solution-phase biopanning process. The library was panned against a biotinylated peptide of the second extracellular loop (ECL2) of CCK2R. After four rounds of biopanning, the selected soluble scFv clones were screened by enzyme-linked immunosorbent assay (ELISA) and examined for specific binding to the peptide. The selected scFvs were purified using immobilized metal affinity chromatography (IMAC). The binding affinity and specificity of the scFvs were examined by the surface plasmon resonance (SPR), immunoblotting and flow cytometry assays and molecular docking using ZDOCK v3.0.2. Results Ten different scFvs were isolated, which displayed binding affinity ranging from 0.68 to 8.0 (nM). Immunoblotting and molecular docking analysis revealed that eight scFvs were able to detect the denatured form of CCK2R protein. Of the isolated scFvs, two scFvs showed high-binding affinity to the human gastric adenocarcinoma AGS cells. Conclusions Based on our findings, a couple of the selected scFvs showed markedly high-binding affinity to immobilized CCK2R peptide and CCK2R-overexpressing AGS cells. Therefore, these scFvs are proposed to serve as targeting and/or treatment agents in the diagnosis and immunotherapy of CCK2R-positive tumors.

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Generation and preliminary characterization of an antibody library with preferential reactivity to human colorectal cancer cells as compared to normal human blood cells

Cited by 11 publications

References 28 publications

Recombinant polyclonal antibodies for cancer therapy

Recombinant polyclonal antibodies for cancer therapy

Antibody treatment of human tumor xenografts elicits active anti-tumor immunity in nude mice

Phage antibody library screening for the selection of novel high-affinity human single-chain variable fragment against gastrin receptor: an in silico and in vitro study

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