New and cost effective cell-based assay for Dialyzed Leukocyte Extract (DLE)-induced Jurkat cells proliferation under azathioprine treatment

Validation of a Cell Proliferation Assay to Assess the Potency of a Dialyzable Leukocyte Extract Intended for Batch Release

“…In our study, we confirmed that Transferon ® was capable of inducing the proliferation of Jurkat cells exposed to azathioprine in a specific concentration range. This was unlike the previous study, which evaluated the batch to batch consistency in biological activity using a single concentration of the product [16]. Usually, the assays for the evaluation of biologics exhibit a non-linear behavior, mainly sigmoidal.…”

“…Recently, the capacity of DLEs to induce Jurkat cell proliferation exposed to azathioprine has been reported without describing the mechanism of action involved for this biological effect, additionally the assay lacks potency estimation with respect to a reference standard and system suitability for a linear response [16], which are required as a routine batch release assay in a quality control environment. In this work, we report the validation of a Jurkat-cells-based assay according to the recommendations for the development, validation, and analysis of bioassays of the United States Pharmacopeia (USP), the European Pharmacopoeia (Ph.…”

Obstacles and limitations of transfer factor biological activity assay design

“…Despite strong clinical evidence of the potential use of DLE in the treatment of various diseases, the molecular mechanisms responsible for the DLE biological activity are still unclear. part of the general disinterest in the DLE basic research is the absence of an adequate and universally accepted DLE in vitro and/or in vivo model (Cardoso et al, 2017). There are several models used for DLE biological activity quantification, including: E-rosette test (Valdimarsson and mcGuire, 1977), induction of delayed type hypersensitivity in mice (Kirkpatrick et al, 1995), leukocyte migration (pizza et al, 1996a) and iFN-γ secretion (medina-Rivero et al, 2014).…”

“…The amount of generated formazan is proportional to the number of cells and their metabolic activity (Berridge and Tan, 1993;Bernas and Dobrucki, 2002). Cardoso et al (2017) tested a new way to determine the biological activity of the commercial preparation TF (immoDiN ® ) cell proliferation, by using two methods -mTT and/or BrdU (bromodeoxyuridine) incorporation assays. When the cells, A20 or jurkat were treated with +Aza/+DLE (azathioprine) they observed a significantly higher proliferation, when compared with +Aza/-DLE.…”

Sequencing Analysis and Identification of the Primary Peptide Component of the Dialyzable Leukocyte Extract “Transferon Oral”: The Starting Point to Understand Its Mechanism of Action

“…Among the immunotherapeutics, stand out the human dialyzable leukocyte extracts (hDLE) which are composed of a low-molecular mass peptide mixture with immunomodulatory properties obtained from the lysis of mammalian cells. Since their first description, made in 1949 by Henry Sherwood Lawrence who also coined the term "Transfer factor" (Lawrence, 1949), dialyzable extracts have been developed in some countries such as China, Czech Republic, Cuba, the USA, and Mexico, as auxiliary drugs to treat immune system-related diseases (Ojeda et al, 2005;Zhou et al, 2013;Cardoso et al, 2017). However, the biological effects of these cellular extracts have been controversial due to the variability of the peptide content among products, which is highly dependent on their cellular origin and the manufacturing method.…”