Target cell line characterization reveals changes in expression of a key antigen that impacts T cell dependent cellular cytotoxicity assay performance
Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved..
Cell lines are important tools regularly used for in-vitro potency assays employed in Good Manufacturing Process (GMP) lot release and stability testing of different therapeutic modalities. Characterization of these cell lines is key to understanding their performance. Bispecific T cell engager (BiTE®) molecules are an exciting modality in Amgen's therapeutic product portfolio. BiTE® molecules are engineered as two linked, single chain antibody domains. One domain targets the cluster of differentiation 3 (CD3) domain of T cells, and the other domain targets a specific antigen. The mechanism of action of the BiTE® molecule is to bring T cells into close proximity of tumor cells to facilitate tumor cell killing. One BiTE® molecule in development, AMG 757, is directed against delta-like ligand 3 (DLL3), which is expressed in small cell lung cancer tumor cells. AMG 757 is a half-life extended bispecific T cell engager (HLE BiTE®) construct. The bioassay employed to demonstrate the mechanism of action of AMG 757 is a T cell dependent cellular cytotoxicity (TDCC) cell-based assay requiring two cell lines, the effector T cell line HuT-78, and engineered tumor target cells, SHP-77-Luc. During the course of development of this bioassay, characterization of the SHP-77-Luc line showed an increase in the luminescence assay signal and Maximum-to-Minimum (Max-to-Min) ratio of the dose response curve as the passage number of the cells increased. Our research revealed an increase in not only luciferase expression but also an increase in the cell surface and intracellular expression levels of DLL3 over time in culture, which ultimately resulted in the increased assay signal window.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2022 |
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| Erschienen: |
2022 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:509 |
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| Enthalten in: |
Journal of immunological methods - 509(2022) vom: 28. Okt., Seite 113326 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Matthies, Kelli [VerfasserIn] |
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| Links: |
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| Themen: |
Antibodies, Bispecific |
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| Anmerkungen: |
Date Completed 13.09.2022 Date Revised 17.11.2022 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1016/j.jim.2022.113326 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| 520 | |a Cell lines are important tools regularly used for in-vitro potency assays employed in Good Manufacturing Process (GMP) lot release and stability testing of different therapeutic modalities. Characterization of these cell lines is key to understanding their performance. Bispecific T cell engager (BiTE®) molecules are an exciting modality in Amgen's therapeutic product portfolio. BiTE® molecules are engineered as two linked, single chain antibody domains. One domain targets the cluster of differentiation 3 (CD3) domain of T cells, and the other domain targets a specific antigen. The mechanism of action of the BiTE® molecule is to bring T cells into close proximity of tumor cells to facilitate tumor cell killing. One BiTE® molecule in development, AMG 757, is directed against delta-like ligand 3 (DLL3), which is expressed in small cell lung cancer tumor cells. AMG 757 is a half-life extended bispecific T cell engager (HLE BiTE®) construct. The bioassay employed to demonstrate the mechanism of action of AMG 757 is a T cell dependent cellular cytotoxicity (TDCC) cell-based assay requiring two cell lines, the effector T cell line HuT-78, and engineered tumor target cells, SHP-77-Luc. During the course of development of this bioassay, characterization of the SHP-77-Luc line showed an increase in the luminescence assay signal and Maximum-to-Minimum (Max-to-Min) ratio of the dose response curve as the passage number of the cells increased. Our research revealed an increase in not only luciferase expression but also an increase in the cell surface and intracellular expression levels of DLL3 over time in culture, which ultimately resulted in the increased assay signal window | ||
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