Methods for Construction of Yeast Display Libraries of Four-Domain T-Cell Receptors

Sádio, Flávio; Stadlmayr, Gerhard; Eibensteiner, Katja; Stadlbauer, Katharina; Rüker, Florian; Wozniak‐Knopp, Gordana

doi:10.1007/978-1-4939-9853-1_13

Cited by 3 publications

(2 citation statements)

References 45 publications

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“…For pMHC, which has a low density on the cell surface (e.g., for NY-ESO-1 157-165 /HLA-A*02:01, 10-50 copies per cell [19]), a monovalent molecule with a high affinity is more effective in enriching molecules on the tumor cell side than a bivalent molecule with a low affinity, which requires a mature screening platform to obtain a specific clone with a high affinity. Compared with the TCR platform [20][21][22], the antibody platform is relatively easy to establish. Some researchers have attempted to develop TCR mimic antibodies to target pMHC [23], but most TCR mimic antibodies exhibited a greater degree of cross-reactivity due to the different recognition modes of TCRs and antibodies towards pMHC [24], which was not conducive to overall specificity.…”

Section: Discussionmentioning

confidence: 99%

T Cell Receptor-Directed Bispecific T Cell Engager Targeting MHC-Linked NY-ESO-1 for Tumor Immunotherapy

Li,

Zhao,

Shen

et al. 2024

Biomedicines

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Antibody-based bispecific T cell engagers (TCEs) that redirect T cells to kill tumor cells have shown a promising therapeutic effect on hematologic malignancies. However, tumor-specific targeting is still a challenge for TCEs, impeding the development of TCEs for solid tumor therapy. The major histocompatibility complex (MHC) presents almost all intracellular peptides (including tumor-specific peptides) on the cell surface to be scanned by the TCR on T cells. With the premise of choosing optimal peptides, the final complex peptide–MHC could be the tumor-specific target for TCEs. Here, a novel TCR-directed format of a TCE targeting peptide–MHC was designed named IgG-T-TCE, which was modified from the IgG backbone and prepared in a mammalian cell expression system. The recombinant IgG-T-TCE-NY targeting NY-ESO-1157–165/HLA-A*02:01 could be generated in HEK293 cells with a glycosylated TCR and showed potency in T cell activation and redirecting T cells to specifically kill target tumor cells. We also found that the in vitro activity of IgG-T-TCE-NY could be leveraged by various anti-CD3 antibodies and Fc silencing. The IgG-T-TCE-NY efficiently inhibited tumor growth in a tumor–PBMC co-engrafted mouse model without any obvious toxicities.

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Section: Discussionmentioning

confidence: 99%

T Cell Receptor-Directed Bispecific T Cell Engager Targeting MHC-Linked NY-ESO-1 for Tumor Immunotherapy

Li,

Zhao,

Shen

et al. 2024

Biomedicines

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“…The percentage of yeast cells binding to the anti-Xpress antibody (Thermo Fisher Scientific, Waltham, MA, USA), conformation-sensitive anti-human CH2 antibody (Bio-Rad, Hercules, CA, USA), protein A (Sigma-Aldrich, St. Louis, MO, USA) and CD64 (R&D Systems, Minneapolis, Minnesota, USA) was determined as described before [50]. The first antigen selection round proceeded with magnetic activated cell sorting (MACS) as described previously [52], using 1 µM biotinylated CD59, refolded from bacterial inclusion bodies, and further sorting rounds were performed with yeast cells stained with 1 µM biotinylated CD59 and a fluorescein isothiocyanate (FITC)-conjugated anti-human CH2 antibody (Bio-Rad, Hercules, CA, USA), diluted 1:200 in 10% bovine serum albumin (BSA)-PBS. Bound CD59 was detected using the conjugated streptavidin-Alexa Fluor ® 647 or NeutrAvidin ® -phycoerythrin (PE) (both Thermo Fisher Scientific, Waltham, MA, USA), diluted 1:1000, interchangeably in the sorting rounds.…”

Section: Yeast Libraries Quality Control and Selectionsmentioning

confidence: 99%

Bispecific mAb2 Antibodies Targeting CD59 Enhance the Complement-Dependent Cytotoxicity Mediated by Rituximab

Stadlbauer

Andorfer

Stadlmayr

et al. 2022

IJMS

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Inhibition of complement activation via the overexpression of complement-regulatory proteins (CRPs), most notably CD46, CD55 and CD59, is an efficient mechanism of disguise of cancer cells from a host immune system. This phenomenon extends to counteract the potency of therapeutic antibodies that could lyse target cells by eliciting complement cascade. The manifold functions and ubiquitous expression of CRPs preclude their systemic specific inhibition. We selected CD59-specific Fc fragments with a novel antigen binding site (Fcabs) from yeast display libraries using recombinant antigens expressed in bacterial or mammalian cells. To produce a bispecific antibody, we endowed rituximab, a clinically applied anti-CD20 antibody, used for therapy of various lymphoid malignancies, with an anti-CD59 Fcab. This bispecific antibody was able to induce more potent complement-dependent cytotoxicity for CD20 and CD59 expressing Raji cell line measured with lactate dehydrogenase-release assay, but had no effect on the cells with lower levels of the primary CD20 antigen or CD20-negative cells. Such molecules are promising candidates for future therapeutic development as they elicit a higher specific cytotoxicity at a lower concentration and hence cause a lower exhaustion of complement components.

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Yeast Mating as a Tool for Highly Effective Discovery and Engineering of Antibodies via Display Methodologies

Baek

Park

Adams

et al. 2022

Methods in Molecular Biology

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Methods for Construction of Yeast Display Libraries of Four-Domain T-Cell Receptors

Cited by 3 publications

References 45 publications

T Cell Receptor-Directed Bispecific T Cell Engager Targeting MHC-Linked NY-ESO-1 for Tumor Immunotherapy

T Cell Receptor-Directed Bispecific T Cell Engager Targeting MHC-Linked NY-ESO-1 for Tumor Immunotherapy

Bispecific mAb2 Antibodies Targeting CD59 Enhance the Complement-Dependent Cytotoxicity Mediated by Rituximab

Yeast Mating as a Tool for Highly Effective Discovery and Engineering of Antibodies via Display Methodologies

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