Use of the Site-Specific Retargeting Jump-In Platform Cell Line to Support Biologic Drug Discovery

Abstract

Biologics represent a fast-growing class of therapeutics in the pharmaceutical sector. Discovery of therapeutic antibodies and characterization of peptides can necessitate high expression of the target gene requiring the generation of clonal stably transfected cell lines. Traditional challenges of stable cell line transfection include gene silencing and cell-to-cell variability. Our inability to control these can present challenges in lead isolation. Recent progress in site-specific targeting of transgene to specific genomic loci has transformed the ability to generate stably transfected mammalian cell lines. In this article, we describe how the use of the Jump-In platform (Life Technologies, Carlsbad, CA) has been applied to drug discovery projects. It can easily and rapidly generate homogeneous high-expressing cell pools with a high degree of reproducibility. Their use in cell-based screening to identify specific binders, identify binding to relevant species variants, or detect functionally relevant therapeutic antibodies is central in driving drug discovery.

A GLP-1:CCK fusion peptide harnesses the synergistic effects on metabolism of CCK-1 and GLP-1 receptor agonism in mice

“…Peptide activity at GLP-1R was determined by a cAMP accumulation assay in Chinese hamster ovary (CHO) cells stably transfected with human GLP-1 receptor (AstraZeneca), as described ( Butler et al, 2015 ; Naylor, Rossi, & Hornigold, 2016 ). Eleven-point duplicate concentration response curves were generated in 3 independent experiments and data analysed as percent activation of the maximum GLP-1R ligand response.…”

Controlling the bioactivity of a peptide hormone in vivo by reversible self-assembly

“…In vitro agonist potencies of free, released, and fibrillated Oxm were determined in cAMP accumulation assays in CHO cells that were stably transfected with human GLP1R or human GCGR receptor 52 – 54 . In brief, peptide samples were incubated with cells plated at 500 cells per well in 384-well black shallow microtiter plates (Corning, USA) for 30 min prior to lysis and detection using the HTRF cAMP dynamic 2 assay kit (Cisbio, France).…”

Development and characterisation of a novel glucagon like peptide-1 receptor antibody

“…INS-1 832/3 cells were maintained in RPMI 1640 Media with GlutaMAX supplement (Thermo Fisher Scientific), 10% FBS, 10 mmol/l HEPES, 50 µmol/l 2-mercaptoethanol, 1 mmol/l sodium pyruvate and penicillin/streptomycin at 37°C in 5% CO 2 [14]. Stably transfected cell lines overexpressing GPCRs were generated at AstraZeneca (Sweden) or MedImmune (UK) using public-domain- or in-house-determined sequences for each receptor, with parental lines purchased from ATCC, ECACC or Invitrogen [15–17]. Overexpressing cell lines for experimental use were thawed from liquid nitrogen stocks into assay buffer on the day of the experiment for cAMP HTRF assays, and responses to agonist were assessed to confirm expected cell activity.…”