TY - JOUR
T1 - Validation of an ADCC assay using human primary natural killer cells to evaluate biotherapeutic products bearing an Fc region
AU - González-González, Edith
AU - Camacho-Sandoval, Rosa
AU - Jiménez-Uribe, Alexis
AU - Montes-Luna, Alejandra
AU - Cortés-Paniagua, Ilselena
AU - Sánchez-Morales, Jazmín
AU - Muñoz-García, Leslie
AU - Tenorio-Calvo, Alejandra V.
AU - López-Morales, Carlos A.
AU - Velasco-Velázquez, Marco A.
AU - Pavón, Lenin
AU - Pérez-Tapia, Sonia Mayra
AU - Medina-Rivero, Emilio
N1 - Publisher Copyright:
© 2018
PY - 2019/1
Y1 - 2019/1
N2 - The development of biotherapeutics requires continuous improvement in analytical methodologies for the assessment of their quality attributes. A subset of biotherapeutics is designed to interact with specific antigens that are exposed on the membranes of target cells or circulating in a soluble form, and effector functions are achieved via recognition of their Fc region by effector cells that induce mechanisms such as antibody-dependent cell-mediated cytotoxicity (ADCC). Thus, ADCC induction is a critical quality attribute (CQA) that must be evaluated to ensure biotherapeutic efficacy. Induction of ADCC can be evaluated by employing effector cells from different sources, such as peripheral blood mononuclear cells (PBMC) and genetically modified cell lines (e.g., transfected NKs or Jurkat cells), and different approaches can be used for detection and results interpretation depending on the type of effector cells used. In this regard, validation of the assays is relevant to ensure the reliability of the results according to the intended purpose. Herein, we show the standardization and validation of ADCC assays to test the potency of three biotherapeutic proteins using primary NK cells obtained from fresh blood as effector cells and detecting cell death by flow cytometry. The advantage of using primary NKs instead of modified cells is that the response is closer to that occurring in vivo since cytotoxicity is evaluated in a direct manner. Our results indicate that in all cases, the assays exhibited a characteristic sigmoidal dose/response curve complying with accurate, precise and specific parameters. Thereby, the validated ADCC assay is an appropriate alternative to evaluate the biological activities of these type of biotherapeutics.
AB - The development of biotherapeutics requires continuous improvement in analytical methodologies for the assessment of their quality attributes. A subset of biotherapeutics is designed to interact with specific antigens that are exposed on the membranes of target cells or circulating in a soluble form, and effector functions are achieved via recognition of their Fc region by effector cells that induce mechanisms such as antibody-dependent cell-mediated cytotoxicity (ADCC). Thus, ADCC induction is a critical quality attribute (CQA) that must be evaluated to ensure biotherapeutic efficacy. Induction of ADCC can be evaluated by employing effector cells from different sources, such as peripheral blood mononuclear cells (PBMC) and genetically modified cell lines (e.g., transfected NKs or Jurkat cells), and different approaches can be used for detection and results interpretation depending on the type of effector cells used. In this regard, validation of the assays is relevant to ensure the reliability of the results according to the intended purpose. Herein, we show the standardization and validation of ADCC assays to test the potency of three biotherapeutic proteins using primary NK cells obtained from fresh blood as effector cells and detecting cell death by flow cytometry. The advantage of using primary NKs instead of modified cells is that the response is closer to that occurring in vivo since cytotoxicity is evaluated in a direct manner. Our results indicate that in all cases, the assays exhibited a characteristic sigmoidal dose/response curve complying with accurate, precise and specific parameters. Thereby, the validated ADCC assay is an appropriate alternative to evaluate the biological activities of these type of biotherapeutics.
KW - ADCC
KW - Biotherapeutic products
KW - Primary natural killer cells
KW - Validation assay
UR - http://www.scopus.com/inward/record.url?scp=85056276493&partnerID=8YFLogxK
U2 - 10.1016/j.jim.2018.11.002
DO - 10.1016/j.jim.2018.11.002
M3 - Artículo
C2 - 30395815
AN - SCOPUS:85056276493
SN - 0022-1759
VL - 464
SP - 87
EP - 94
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
ER -