![]() Repeated administration of recombinant biopharmaceuticals can lead to a break in immune tolerance (1) and the production of anti-drug antibodies (ADA). Thus, cell-based assays are used in drug-discovery, potency assays, the assessment of neutralizing antibodies against protein-based biopharmaceuticals and antibody mediated effector functions including complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and antibody-dependent cell-mediated cytotoxicity (ADCC). BACKGROUND OF THE INVENTIONĬell-based assays are required for the detection and quantification the activity of pharmacologically active substances whether small chemical entities or large protein based biopharmaceuticals. Importantly, the system according to the invention may be used to determine the effectiveness of a treatment based on the use of a small synthetic drug, large therapeutic protein, recombinant or naturally occurring, AAV transgene, cellular therapy including CAR-T cells, or a therapeutic antibody or Fc fusion protein, or the immune response elicited by the therapeutic protein, therapeutic antibody, AAV vector, AAV transgene, or cells used in adoptive therapy, that may be advantageous as in the case of the induction of effector cell function or disadvantageous as in the case of a break in immune tolerance and the production of anti-drug antibodies. The system according to the invention may be used in a kit or a kit of parts that may be used in a diagnostic context. The system according to the invention can be used to screen libraries of small chemical entities, quantify the potency of small synthetic drugs, large protein-based biopharmaceuticals, adeno associated virus (AAV) transgenes, or cellular therapy including CAR-T cells, and to quantify the neutralizing antibody response to biopharmaceuticals, AAV vectors, AAV transgenes, or cellular therapy including CAR-T cells, as well as the effector cell function of therapeutic antibodies or Fc fusion proteins using immuno-assay platforms that are readily automated. The present invention relates to a novel system and its use in methods for determining the activity of pharmacologically active molecules including small chemical entities and large protein-based biopharmaceuticals. Said ASCII copy, created on May 5, 2021, is named 38526-251_ST25.txt and is 5,247 bytes in size. The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. INCORPORATION-BY-REFERENCE OF MATERIAL SUBMITTED ELECTRONICALLY 27, 2017, each of which are herein incorporated by reference. 25, 2018, which claims the benefit of European Application No. § 371 of PCT International Application No. This application is a national phase application under 35 U.S.C. The present invention also provides a means of increasing the dynamic range, sensitivity and reducing the cost of cell-based assays and can be applied to existing engineered cell lines, such as those containing a reporter- gene such as a luciferase reporter-gene obviating the necessity to extensively re-engineer cell lines containing multiple molecular constructs. a substantial improvement of cell-based assays for analysis using automated immune-detection platforms and allows simultaneous analysis of multiple analytes, multiple sampling from a single cell culture, and obviates the necessity to lyse cells and remove cell debris by centrifugation prior to analysis on an automated immuno-detection platform. The detection sequence is unique to each secreted reporter-peptide and may be labelled with a Sulfo-Tag that permits detection of the peptide on the MSD platform, or Alexa that permits detection on the Gyros platform, or digoxigenin that permits detection of on the PerkinElmer AlphaLISA platform, or left unlabeled for detection by SPR on a Biacore platform. The anchor sequence may differ from one peptide to another or may be common to multiple secreted reporter-peptides such that all the peptides can be analyzed simultaneously by ELISA using a detection antibody labelled with for example HRP, or by the use of a commonly available immuno-detection platform such MSD, Gyros, AlphaLisa, or Biacore. The present invention relates to small secreted reporter-peptides 15 to 150 amino-acids in length comprising a response element, activated by one or more transcription factors induced by the pharmacology active substance to be analyzed following its interaction with a specific intracellular or cell surface molecule, functionally linked to a response element, a TATA box, a signal peptide, anchor and detection sequences to which antibodies can be raised, and a poy-A tail. ![]()
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