Training Session M1 - Training Topic List
June 15, 2020, 11am to 4pm ET: Part 1
June 16, 2020, 11am to 4:30pm ET: Part 2

"Advanced Biomarkers Development & Troubleshooting by LBA & Flow Cytometry: Challenges from Exploratory to Clinical/Confirmatory Biomarkers, Fit-for-Purpose, Context of Use and New Platforms"

From Discovery/Exploratory Biomarkers to Clinical/Confirmatory Biomarkers, actual challenges in developing and implementing biomarkers assays in different phases of drug discovery/development by embedding the Fit-for-Purpose (FFP), Context of Use (COU) and new platforms in real-life situations

View M1 Agenda

  • Dr. Susan Spitz, Sr. Director Clinical Research Translational Sciences, Incyte
  • Dr. Sally Fischer, Assoc. Director BioAnalytical Sciences, Genentech

Moving Forward an Internationally Harmonized BAV Guidance - An Update

Biomarkers Assays Development & Troubleshooting (Part A)

Target Engagement (TE) & Receptor Occupancy (RO)

Biomarkers Assays Development & Troubleshooting (Part B)


Discussion Topic DETAILS of M1 – Advanced Biomarkers Development & Troubleshooting

  • Dr. Susan Spitz, Sr. Director Clinical Research Translational Sciences, Incyte
  • Dr. Sally Fischer, Assoc. Director BioAnalytical Sciences, Genentech
Moving Forward an Internationally Harmonized BAV Guidance - An Update
  • Lesson 1
    Current WRIB White Paper Recommendations on COU & FFP Biomarker Assay Development: Summary of 2019 discussions and Industry/regulators’ agreement on the need for an Internationally Harmonized BAV Guidance & Terminology
    - Dr. Lakshmi Amaravadi, VP and Head Translational Medicine, Boston Pharmaceuticals
    • Context of Use (COU) and Fit for Purpose (FFP) for Biomarker Assay Development evolved through a series of recommendations (2014-2019) until reaching its maturity in the in the 2019 White Paper in Bioanalysis
      • 2019 Link:
      • "FFP BAV analytically validates and characterizes the assay for the intended COU: The analytical error and biological variability in the measurements should be determined and related to the desired COU to set appropriate analytical validation acceptance based on the endogenous analyte: (relative) accuracy, precision, analytical measurement range, parallelism, specificity, selectivity, and stability"
    • Need for an Internationally harmonized BAV Guideline
      • Industry/Regulators' consensus and way forward
      • 2014-2019 White Paper in Bioanalysis recommendations in support of Internationally harmonized BAV Guideline
        • "fit-for-purpose" best practices in the development and validation of biomarker assays for exploratory or advanced biomarker applications
        • Strategies for different applications at various phases of biomarker development
        • Key elements in the method of development and validation
        • Decision points in Method Development and Validation
          • Context of Use & Fit for Purpose
          • Clinical significance of results Validation
          • Analytical performance
          • Acceptance criteria
    • Harmonization of BAV Terminology Clarification
      • FFP Validation vs Qualification
      • Confusion generated by the term Qualification
      • FFP Validation as current industry standard
      • Not to confuse with FDA Biomarker Qualification Program (FDA BQP)
    • Big "grey zone" in the "qualified" methods
      • What is it included?
      • Performance characteristics adequate to support the assays
      • Full validation as an official statement that assay is compliant with minimum Regulatory requirements

  • Lesson 2
    Is the newly released Industry/FDA C-Path White Paper helping perform Biomarker Assay Development & Validation and moving towards a BAV Guidance?: Case studies on application of C-Path White Paper, Industry Perspective on the new draft, and on- going discussions to turn this white paper in a BAV Guidance
    - Dr. Susan Spitz, Sr. Director Clinical Research Translational Sciences, Incyte
    • "A concise regulatory guidance/guideline for BAV is desired by many, rather than the industry relying on best practices"
      • Page 1994: 2018 White Paper in Bioanalysis
      • How to address the lack of one single Guideline to cover all types of biomarker assays and their intended uses
        • What should be taken into consideration during BAV?
        • Biomarker data used in support of the regulatory submissions
        • Industry best practice expectations on the rigor of assay qualification/validation
    • Updates on the FDA/C-Path initiative: "Points to Consider Document: Scientific and Regulatory Considerations for the Analytical Validation of Assays Used in the Qualification of Biomarkers in Biological Matrices"
    • Current Industry/Regulators' discussion about generating a Biomarker Assay Validation (BAV) guidance document from the FDA/C-Path document

  • Lesson 3
    An Update on the Point to Consider on BAV by Japan MHLW
    - Dr. Yoshiro Saito, Director Division of Medicinal Safety Science, Japan MHLW
    • Japan MHLW ongoing progress on BAV recommendations
    • Introduction of newly discussed items
      • Partial validation
      • Cross validation
      • Study sample analysis
      • Commercial kit
      • Re-analysis
      • Future perspective
    • Application of BMV guidelines is not appropriate
      • 2018 FDA BMV Guidance
      • 2019 FDA/C-Path BAV White Paper
      • Japan AMED research Industry/Regulators’ group draft of points to consider document on BAV
        • Included: scope, standards, selectivity, specificity, calibration curve, matrix effect, accuracy, precision, parallelism, dilution linearity, stability, partial validation, cross validation, study sample analysis, ISR, commercial kit and re-analysis. Its scope (molecular types and methods)
    • Similarities between FDA/C-Path & Japan AMED BAV documents


Biomarkers Assays Development & Troubleshooting (Part A)
  • Lesson 4
    New Insights on Parallelism Assessment in Biomarker Assays: Challenges and Solutions based on COU & FFP
    - Dr. Linda Terry, Group Lead Biomarker Assay Development, GlaxoSmithKline
    • Parallelism for Biomarker Assay Development evolved through a series of recommendations (2014-2019) until reaching its maturity in the in the 2019 White Paper in Bioanalysis
      • 2019 Link:
      • "Parallelism is an essential experiment to appropriately characterize the endogenous analyte in all ligand-binding (antibody dependent) biomarker assays: Parallelism cannot be replaced by spike/recovery experiments with recombinant material; If parallelism cannot be performed pre-study, it should be performed in-study as soon as samples become available to understand if the assay is appropriate for the COU or requires additional optimization"
    • Parallelism for BAV from 2019 FDA/C-Path White Paper Recommendations
    • Practical Case studies on parallelism evaluation based on COU & FFP
      • Parallelism evaluation and its impact on the determination of BAV parameters such as MRD, and sensitivity
        • How parallelisms can help meet the constant requests for higher sensitivity for biomarker assays allowing the detection of previously undetectable biomarkers
        • Issues around sensitivity when developing and validating biomarker assays
        • Correlation between sensitivity achievable vs FFP BAV achievable
        • Understand the expectations in defining the biomarker assay MRD
        • Case studies on how the MRD is calculated based on Parallelism and sensitivity of the assay
    • What to do if parallelism fails or cannot be assessed?
      • How to create confidence in you assay
      • Method development strategies to improve assay performance

  • Lesson 5
    Practical Strategies to Commercial Kits Optimization in Biomarkers Method Development: Building on WRIB recommendations, Context of Use (COU) & Fit for Purpose (FFP), new trends and case studies
    - Mr. David Lanham, Scientific Director Bioanalytical Group, Eurofins
    • Building on 2018 White Paper Recommendations on Commercial Kits Optimization with novel case studies and advances
    • RUO (research use only) commercial immunoassay kits
      • Lack of desired robustness and reproducibility
      • Depending on the COU (Context of Use)
        • Kit feasibility assessment before testing clinical samples
        • FFP Validation
    • Commercial Kits Optimization
      • Kit-to-kit variability
      • Lot-to-lot variability of reagents for assays
      • Cross-reactivity between Calibrators and Antibody reagent
    • Current strategies to optimize Kits the meet BAV requirements according to COU
      • Implementation of recent Guidance and White Papers
      • In-depth case studies on kits optimization in specific matrix/disease states
    • What is needed to minimize lot-to-lot variability?
      • How is it determined?
      • Why are different methods giving different results?
      • Use of spiking with recombinant analyte in assay validation
    • Novel case studies on modifications to a commercial assay based on COU to ensure well-performing bioanalytical method

  • Lesson 6
    Biomarker Advanced Method Development Strategies using Endogenous Quality Controls: Practical Case Studies and Lesson Learnt for COU & FFP
    - Mr. John Allinson, Vice President Biomarker Sciences, Immunologix
    • Building on 2018 White Paper Recommendations on Endogenous QC with novel case studies and advances
    • Endogenous QC determination & application for precision and stability evaluations
      • Evaluation of the differences between spiked QC with recombinant reference standard and endogenous QC
      • Strategies to prepare QC material in bulk to ensure Biomarker assays life cycle management
    • Issues in the use of non-endogenous forms of the biomarker in validation
      • Bias in assay the performance
      • non-endogenous vs endogenous forms
    • Importance to use QC endogenous biomarkers
      • Parallelism
      • Inter-assay precision
      • Stability
      • Use of Endogenous QC and ISR evaluation for Biomarker Assays
    • Case studies on the characterization of biomarker assay performance by using Endogenous QC samples


Target Engagement (TE) & Receptor Occupancy (RO)
  • Lesson 7
    Target Engagement (TE) Flow Cytometry Assays in Patient Samples Whole Blood: Method development and COU & FFP approaches
    - Ms. Lisa Patti-Diaz, Principal Scientist Clinical Flow Cytometry, Bristol-Myers Squibb
    • Building on 2018 White Paper Recommendations on Target Engagement Biomarker Assays by Flow Cytometry with novel case studies and advances
    • Flow Cytometry as the elective technique for Target Engagement (TE) assays
      • TE assays and need for an accurate measuring technique
      • TE assays as unique type of biomarker assay to evaluate the direct interaction of the drug molecule and target
    • TE assays challenges & solution in development, validation and implementation
      • Overcoming development obstacles
      • Strategies in performing a successful TE assay validation
        • COU & FFP validation design
        • Sample stability evaluation
      • Designing the correct TE assay principle
    • Importance of TE biomarkers assays as PD markers
      • Impact on rational/optimal design of dosing regimen in clinical applications
        • Key parameter to define safe starting doses for FIH clinical trials
        • Estimation in rational dose selection
      • Proof-of-mechanism
      • In-vivo activity
      • Loss of efficacy
      • Exposure
    • Case studies on challenges in the development and validation and implementation of a Flow Cytometry Target Engagement Biomarker assay to support clinical trials
      • Development of Clinical FFP functional flow cytometry Whole Blood Assay with Cross-Linking Stimulation to Evaluate Target Engagement in Patient Samples

  • Lesson 8
    Target Engagement (TE) Biomarker LBA Development Strategies: Building on WRIB recommendations, COU & FFP and new trends
    - Mr. Gregor Jordan, Lab Head, Roche
    • Building on 2018 White Paper Recommendations on TE FFP Validation with novel case studies and advances
    • COU evaluation before implementing the biomarker assay in a clinical study
      • Ongoing increased request of TE assays by clinical teams
      • Concentration time profile of the total target concentration
      • Importance of interaction of the drug with the endogenous target for decision-making (COU)
    • Challenges with TE assay for soluble target
      • Assay to detect the free portion of the soluble target
    • Active target concentration
      • KD value of the interaction partners
      • Complex stability
      • Impact of sample manipulation
        • Effect of dilutions in the increase in free target concentration
        • Risk for erroneous underestimation of TE
    • Case studies on TE assay selection by using endogenous QC samples to verify the correct selection of the assay-platform, reagents and assay conditions

  • Lesson 9
    Receptor Occupancy (RO) by Flow Cytometry:
    • Strategies for developing robust PD biomarker assays
    • Challenges in Method Development and FFP Validation Flow Cytometry Pharmacodynamics Receptor Occupancy (RO) Assay: Lesson leant and case studies
    - Dr.Christèle Gonneau, Lead Scientist Flow Cytometry, Covance
    - Dr. Fabian Junker, Senior Scientist Pharmaceutical Sciences Division, Roche
    • Building on 2018 White Paper Recommendations on Challenges with Receptor Occupancy Assays by Flow Cytometry with novel case studies and advances
    • Flow Cytometry Receptor Occupancy (RO) assays
      • Routinely used in development of antibody-based therapeutics
      • Flow Cytometry RO assays &h PK data for FIH initial dose selection
    • RO by Flow Cytometry
      • PD biomarker assay
        • RO as robust PD assay
        • Rapid assessment of the degree to which specific cellular surface targets are occupied by drug
      • Correlation with PK
        • Dose finding
    • Different strategies available for RO assay development
      • Potential limitations
      • Important lessons learned
      • RO for decision making in the clinical setting
    • Case studies on challenges in developing and implementing Flow Cytometry-based Pharmacodynamics Receptor Occupancy (RO)


Biomarkers Assays Development & Troubleshooting (Part B)

Final Agenda Agenda at a Glance