Neutralizing Antibody Sample Testing and Report Harmonization.

Immunogenicity testing and characterization is an important part of understanding the immune response to administration of a protein therapeutic. Neutralizing antibody (NAb) assays are used to characterize a positive anti-drug antibody (ADA) response. Harmonization of reporting of NAb assay performance and results enables efficient communication and expedient review by industry and health authorities. Herein, a cross-industry group of NAb assay experts have harmonized NAb assay reporting recommendations and provided a bioanalytical report (BAR) submission editable template developed to facilitate agency filings. This document addresses key bioanalytical reporting gaps and provides a report structure for documenting clinical NAb assay performance and results. This publication focuses on the content and presentation of the NAb sample analysis report including essential elements such as the method, critical reagents and equipment, data analysis, study samples, and results. The interpretation of immunogenicity data, including the evaluation of the impact of NAb on safety, exposure, and efficacy, is out of scope of this publication.

Implementation of flow cytometry testing on rare matrix samples: Special considerations and best practices when the sample is unique or difficult to obtain.

The publication of Clinical and Laboratory Standards Institute's guideline H62 has provided the flow cytometry community with much-needed guidance on development and validation of flow cytometric assays (CLSI, 2021). It has also paved the way for additional exploration of certain topics requiring additional guidance. Flow cytometric analysis of rare matrices, or unique and/or less frequently encountered specimen types, is one such topic and is the focus of this manuscript. This document is the result of a collaboration subject matter experts from a diverse range of backgrounds and seeks to provide best practice consensus guidance regarding these types of specimens. Herein, we define rare matrix samples in the setting of flow cytometric analysis, address validation implications and challenges with these samples, and describe important considerations of using these samples in both clinical and research settings.

16th GCC Closed Forum: ICH M10 implementation; NGS, qPCR/dPCR, flow cytometry validation; tissue biomarkers; IS response; immunogenicity harmonization; bioanalytical industry status.

The 16th GCC Closed Forum was held in Orlando, FL, USA, on 23 June 2023. Representatives from international bioanalytical Contract Research Organizations were in attendance in order to discuss scientific and regulatory issues specific to bioanalysis. The issues discussed at the meeting included: IS response, flow cytometry, changes to the bioanalytical industry, NGS assays, biomarker assay for tissues, dPCR validation, immunogenicity harmonization and ICH M10 implementation. Conclusions and consensus from discussions of these topics are included in this article.

Study to assess the impact of analytics software on operating room controlled substance management and drug diversion.

PURPOSE: This study investigated the impact of an advanced analytics software solution in the operating room (OR) on tracking and evaluating controlled substance discrepancies. The authors hypothesized that the software would increase identification of these discrepancies and improve the efficiency of the preexisting manual process. METHODS: In this evaluation comparing data from before to after implementation of the software, data were collected using the preexisting manual process for 50 days before implementation, followed by a 25-day period for acclimation to the new software, and ending with a 49-day postimplementation review period. Data collected included the total number of medication discrepancies, time required for discrepancy review and reconciliation by an OR analyst, types of discrepancies, and number of discrepancies leading to provider audits. RESULTS: Before implementation of the analytics software, there were 7,635 OR cases with a total of 674 charting discrepancies (8.83 discrepancies per 100 total OR cases) discovered across 439 OR cases. After implementation, there were 7,454 OR cases with a total of 930 charting discrepancies (12.48 discrepancies per 100 total OR cases; P < 0.0001) discovered across 680 OR cases. While discrepancies increased by 38%, the median review time for the OR analyst per case decreased (P < 0.0001) and the percentage of incidents resolved by the OR analyst increased by 14% while the number of cases requiring additional documentation by the provider decreased by 10%. CONCLUSION: Implementation of advanced analytics software in the OR significantly increased the number of controlled substance charting discrepancies identified compared to the preimplementation review process while increasing the efficiency of the OR analyst.

Recommendations for Method Development and Validation of qPCR and dPCR Assays in Support of Cell and Gene Therapy Drug Development.

The emerging use of qPCR and dPCR in regulated bioanalysis and absence of regulatory guidance on assay validations for these platforms has resulted in discussions on lack of harmonization on assay design and appropriate acceptance criteria for these assays. Both qPCR and dPCR are extensively used to answer bioanalytical questions for novel modalities such as cell and gene therapies. Following cross-industry conversations on the lack of information and guidelines for these assays, an American Association of Pharmaceutical Scientists working group was formed to address these gaps by bringing together 37 industry experts from 24 organizations to discuss best practices to gain a better understanding in the industry and facilitate filings to health authorities. Herein, this team provides considerations on assay design, development, and validation testing for PCR assays that are used in cell and gene therapies including (1) biodistribution; (2) transgene expression; (3) viral shedding; (4) and persistence or cellular kinetics of cell therapies.

Revisiting Ebola Virus Disease infection control protocols at an academic level I trauma center: Successes, challenges, and ways ahead.

In the decade since the largest Ebola Virus Disease (EVD) outbreak in history, hospitals within the United States have discovered deficiencies in EVD infection control protocols. A large academic level I trauma medical center and frontline EVD receiving hospital in northeast Florida conducted a large-scale review and revision of the facility's EVD infection control protocols to increase preparedness. The revision process revealed opportunities for improvement and highlighted the need for excellent resource management and interdepartmental communication.

Identification and infection control response to Candida auris at an academic level I trauma center.

Candida auris, an emerging fungal pathogen with significant morbidity and mortality, can be difficult for health care facilities to identify, isolate, and control. We present our identification and infection control response to Candida auris at a 695-bed academic level I trauma center in Florida.

Validation of Anti-Adeno Associated Virus Serotype rh10 (AAVrh.10) Total and Neutralizing Antibody Immunogenicity Assays.

Immunogenicity assessment of Adeno-Associated Virus (AAV) vectors is a critical part of gene therapy drug development. Whether the assays are used for inclusion/exclusion criteria or to monitor the safety and efficacy of the gene therapy, they are critical bioanalytical assessments. While total anti-AAV assays are perceived as easier to develop and implement than neutralizing anti-AAV assays, the gene therapy field is still nascent, and it is not yet clear which of the assays should be implemented at what stage of drug development. Recently AAVrh.10 has gained interest for use in gene therapies targeting cardiac, neurological, and other diseases due to its enhanced transduction efficiency. There is limited information on anti-AAVrh.10 antibodies and their clinical impact; thus, the information presented herein documents the validation of both a total antibody assay (TAb) and a neutralizing antibody (NAb) assay for anti-AAVrh.10 antibodies. In this manuscript, the validation was performed in accordance with the 2019 FDA immunogenicity guidance with additional evaluations to comply with CLIA where applicable. The AAVrh.10 TAb and NAb assays were compared in terms of sensitivity, drug tolerance, and precision, along with a concordance analysis using the same individual serum samples. This comparison gave insight into the utility of each format as a screening assay for inclusion into clinical studies.

Neutralizing Antibody Validation Testing and Reporting Harmonization.

Evolving immunogenicity assay performance expectations and a lack of harmonized neutralizing antibody validation testing and reporting tools have resulted in significant time spent by health authorities and sponsors on resolving filing queries. A team of experts within the American Association of Pharmaceutical Scientists' Therapeutic Product Immunogenicity Community across industry and the Food and Drug Administration addressed challenges unique to cell-based and non-cell-based neutralizing antibody assays. Harmonization of validation expectations and data reporting will facilitate filings to health authorities and are described in this manuscript. This team provides validation testing and reporting strategies and tools for the following assessments: (1) format selection; (2) cut point; (3) assay acceptance criteria; (4) control precision; (5) sensitivity including positive control selection and performance tracking; (6) negative control selection; (7) selectivity/specificity including matrix interference, hemolysis, lipemia, bilirubin, concomitant medications, and structurally similar analytes; (8) drug tolerance; (9) target tolerance; (10) sample stability; and (11) assay robustness.

Utilization of a technology-assisted workflow to prepare controlled substance oral syringes.

PURPOSE: Utilization of technology-assisted workflow (TAWF) systems has gained popularity in the sterile compounding setting. This study was designed to evaluate whether safety and efficiency could be seen when preparing oral controlled substance doses gravimetrically vs volumetrically. METHODS: This 2-phase observational study combined manual data collection with automated logs generated by a single TAWF. During phase I, oral controlled substance solutions were prepared volumetrically. In phase II, the same subset of medications was to be prepared gravimetrically via the same TAWF. Findings from phases I and II were compared against each another to determine safety, efficiency, and documentation differences between the volumetric and gravimetric workflows. RESULTS: Thirteen different medications were evaluated during phase I (1,495 preparations) and phase II (1,781 preparations) of this study. Mean compounding time (min:sec) increased in phase II when compared to phase I (1:49 vs 1:28; P < 0.01), with the deviation detection rate also increasing (7.9% vs 4.7%; P < 0.01). Despite a target in phase II of utilizing gravimetric analysis for more than 80% of preparations, only 45.5% (811 preparations) were prepared with this workflow, as adoption challenges and dose size limitations prevented compliance. Doses that were prepared gravimetrically had a mean accuracy rate of 100.6% (the mean achieved dose was 0.6% higher than the mean prescribed dose) and a rejection rate of 0.99% (compared to the phase I rejection rate of 1.07%; P = 0.67). CONCLUSION: The gravimetric workflow provided accuracy and additional safety checks when compared to the volumetric alternative, all while providing users with greater access to data. Health systems should consider staffing, product sourcing, patient populations, and medication safety when determining the balance between volumetric and gravimetric workflows.

Anti-drug Antibody Sample Testing and Reporting Harmonization.

A clear scientific and operational need exists for harmonized bioanalytical immunogenicity study reporting to facilitate communication of immunogenicity findings and expedient review by industry and health authorities. To address these key bioanalytical reporting gaps and provide a report structure for documenting immunogenicity results, this cross-industry group was formed to establish harmonized recommendations and a develop a submission template to facilitate agency filings. Provided here are recommendations for reporting clinical anti-drug antibody (ADA) assay results using ligand-binding assay technologies. This publication describes the essential bioanalytical report (BAR) elements such as the method, critical reagents and equipment, study samples, results, and data analysis, and provides a template for a suggested structure for the ADA BAR. This publication focuses on the content and presentation of the bioanalytical ADA sample analysis report. The interpretation of immunogenicity data, including the evaluation of the impact of ADA on safety, exposure, and efficacy, is out of scope of this publication.

Recommendations on qPCR/ddPCR assay validation by GCC.

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.

Recommendations on ELISpot assay validation by the GCC.

Gene therapy, cell therapy and vaccine research have led to an increased need to perform cellular immunity testing in a regulated environment to ensure the safety and efficacy of these treatments. The most common method for the measurement of cellular immunity has been Enzyme-Linked Immunospot assays. However, there is a lack of regulatory guidance available discussing the recommendations for developing and validating these types of assays. Hence, the Global CRO Council has issued this white paper to provide a consensus on the different validation parameters required to support Enzyme-Linked Immunospot assays and a harmonized and consistent approach to Enzyme-Linked Immunospot validation among contract research organizations.

Bioanalytical Assay Strategies and Considerations for Measuring Cellular Kinetics.

A vast evolution of drug modalities has occurred over the last several decades. Novel modalities such as cell and gene therapies have proven to be efficacious for numerous clinical indications-primarily in rare disease and immune oncology. Because of this success, drug developers are heavily investing in these novel modalities. Given the complexity of these therapeutics, a variety of bioanalytical techniques are employed to fully characterize the pharmacokinetics of these therapies in clinical studies. Industry trends indicate that quantitative PCR (qPCR) and multiparameter flow cytometry are both valuable in determining the pharmacokinetics, i.e. cellular kinetics, of cell therapies. This manuscript will evaluate the pros and cons of both techniques and highlight regulatory guidance on assays for measuring cellular kinetics. Moreover, common considerations when developing these assays will be addressed.

Deploying a rapid point-of-care polymerase chain reaction test for SARS-CoV-2 in a clinical research unit to ensure healthy volunteer safety.

The entire world was severely affected by the outbreak of the SARS-CoV-2 virus. Early phase clinical research was no exception and clinical healthy volunteer trials were halted across the globe. To enable continuation of development of new drugs, we developed a testing strategy for nonsymptomatic trial participants in an early stage of the outbreak. A point-of-care polymerase chain reaction test combined with a gold standard polymerase chain reaction test and strict social distancing and hygiene measures limited the number of infected subjects entering our clinical research units and reduced further spread for the duration of the clinical trial. Thus, proving efficacy of this strategy to allow safe and effective continuation of early phase clinical trials during the COVID-19 pandemic.