Recommendations on qPCR/ddPCR Assay Validation by GCC

Abstract

Gene therapy, cell therapy and vaccine research have led to an increased use of qPCR/ddPCR in bioanalytical laboratories. CROs are progressively undertaking the development and validation of qPCR and ddPCR assays. Currently, however, there is limited regulatory guidance for the use of qPCR and a complete lack of any regulatory guidelines for the use of the newer ddPCR to support regulated bioanalysis. Hence, the Global CRO Council in Bioanalysis (GCC) has issued this White Paper to provide; 1) a consensus on the different validation parameters required to support qPCR/ddPCR assays; 2) a harmonized approach to their validation and 3) a consistent development of standard operating procedures (SOPs) for all the bioanalytical laboratories using these techniques.

Novel Cell Quantification Method Using a Single Surrogate Calibration Curve Across Various Biological Samples

“…In the present study, we set the acceptance criteria for accuracy and precision to be within 35% RE and < 35% CV, respectively, based on our previous report (14). These criteria were in line with the recommendations from the Global CRO Council in Bioanalysis (GCC) (29). Although some studies have proposed different criteria (30)(31)(32)(33)(34), there is no one criterion for the spike-in method similar to our method.…”

Drug Regulatory-Compliant Validation of a qPCR Assay for Bioanalysis Studies of a Cell Therapy Product with a Special Focus on Matrix Interferences in a Wide Range of Organ Tissues

“…Although qPCR-based assays have emerged as an important bioanalytical method for assessing the pharmacokinetics of human-cell-based medicinal products [ 4 , 5 , 16 , 21 ], the regulatory guidelines on the validation of bioanalytical methods released by the EMA, FDA, and ICH [ 24 , 25 , 26 , 27 ] focus on methods suitable for small- and large-molecule drugs such as chromatographic and ligand-binding assays. While the basic concepts and parameters of method validation described in these guidelines can be adapted to cell quantification via qPCR, in the absence of specific regulatory recommendations including definitive acceptance criteria for a validated qPCR assay, researchers must rely on published evidence from bioanalytical scientists, as well as recently issued best practice recommendations [ 15 , 20 , 42 ] and white papers from scientific networks [ 21 , 22 , 23 , 43 ].…”

“…Therefore, any qPCR method must be validated to ensure the reliability of the data that will be obtained. Remarkably, however, despite the growing number of qPCR applications in regulated bioanalysis, and even though qPCR is one of the officially recognized methods for determining the pharmacokinetics of cell therapy products [ 11 ], regulatory guidance on the use of qPCR is limited [ 13 , 20 , 21 , 22 , 23 ]. The International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) Guideline M10 on Bioanalytical Method Validation and Study Sample Analysis [ 24 , 25 ], which recently superseded the guidelines on bioanalytical method validation released by the European Medicines Agency (EMA) [ 26 ] and the US Food and Drug Administration (FDA) [ 27 ], focuses on pharmacokinetic methods that are suitable for classical small-molecule drugs and large-molecule biologics, such as chromatographic methods and ligand-binding assays, but do not address the specific requirements for the proper validation of PCR assays [ 13 , 20 , 21 ].…”

PCR-based analytics of gene therapies using adeno-associated virus vectors: Considerations for cGMP method development

“…The Global Clinical Research Organization Council (GCC) recently published a meta-analysis of qPCR and ddPCR guidance documents that provides recommendation for validating methods. 16 Many of the recommendations are important to discuss in consideration with gene therapy testing.…”

“…Many reviews propose recommendations for criteria such as assay development and validation parameters and these are being adopted for use in gene therapies. 16 , 17 However, there remain many challenges in the various aspects of gene therapy treatment and vectors that will need to be considered in the context of these recommendations. This review surveys key biological, technological, and regulatory characteristics of nucleic acid-based analytical testing to for researchers and professionals considering future investigational new drugs and biologics license application filings for gene therapies.…”