Specialized Session T-1 - 8am to 5:30pm, Tuesday April 10, 2018

"LBA & Flow Cytometry for Large Molecules Advanced Method Development Challenges, Solutions and New Industry Standards with NEW Case Studies"

Advanced and interactive session in LBA & Flow Cytometry where original case studies and state-of the art technologies & approaches are presented from experts in the field

Outline of Specialized Session T-1

(You can click on each blue topic title below to see details, or simply scroll down to see details)


Session Co-Chair: Dr. Chris Beaver, Senior Director Ligand Binding & Exploratory Bioanalysis, Syneos Health
Session Co-Chair: Dr. John Kamerud, AR Fellow / Director - Bioanalytical, Pfizer

Part 1: LBA in Regulated Bioanalysis
Part 2: Flow Cytometry in Regulated Bioanalysis
Part 3: What’s new in LBA PK Assays
Part 4: Novel case studies on Emerging/Established Technologies


DETAILS of Specialized Session T-1

Session Co-Chair: Dr. Chris Beaver, Senior Director Ligand Binding & Exploratory Bioanalysis, Syneos Health
Session Co-Chair: Dr. John Kamerud, AR Fellow / Director - Bioanalytical, Pfizer

Part 1: LBA in Regulated Bioanalysis
  • Lecture 1
    What’s New on LBA Validation Issues, Failures & Troubleshooting
    - Dr. Renuka Pillutla, Executive Director, Bristol-Myers Squibb
    • Dealing with unknown/unexpected biological variation & interferences
      • During Validation
      • During Sample Analysis
      • Study submitted and feedback from regulators
    • Major Target interference
      • Use of surrogate matrix and assay development challenges
      • How to prepare QCs and how to dilute study samples
      • Dealing with the presence of endogenous analyte
    • Severe ADA interference on PK
      • How to validate an PK assay with impact from ADA
      • Data interpretation
    • Case Studies: What to do if your validation fails?

  • Lecture 2
    Calibration Curves issue in LBA used in Regulated Bioanalysis: Current industry standards & lesson learnt based on case studies
    - Dr. John Kamerud, AR Fellow / Director - Bioanalytical, Pfizer
    • Focus on Quality of the LBA calibration curve
      • Current requirements
      • Lesson learnt from the selection of a regression model
        • The best fitted model
        • 4PL and 5PL on different platforms
        • Weighted and non-weighted
      • Need to have QC LLOQ and QC ULOQ
    • Issues in using surrogate matrix in LBA
    • Regulatory expectations on anchor points
    • Challenges with changes in critical reagent binding/reactivity properties
    • Case Studies: Major issues in LBA calibration curves and impact on data accuracy

  • Lecture 3
    New Development in Active Drug Assays vs. Current Guidance/Guidelines
    - Dr. Roland Staack, Senior Scientist Bioanalytics, Roche
    • Why you have to understand the importance of Active Drug Exposure to develop the correct method
      • Need for a more in-depth training for the Industry
      • Active Drug Assays and Current Guidelines
      • 2017 White Paper Part 3 Recommendations
        • Moving in the right direction
        • Active drug PK assays - continuing the discussion
        • Limitations of current PK assay Guidance/Guidelines for the BMV of Active Drug Assays
    • Understanding Active drug exposure
      • Multidomain compounds
      • Cell-based PK assays
      • Active drug: What exactly are we analyzing?
    • Understanding the biology of your analyte
      • Holistic approach
      • Biology strategy – Clinical, ADA, exposure, efficacy
      • Do we understand the analyte/assay good enough?
    • Dilutions in LBA and free drug concentration
      • Does LBA dilutions alter free drug concentration?
    • Case Studies: New data to better understand Active Drug Assays


Part 2: Flow Cytometry in Regulated Bioanalysis
  • Lecture 4
    Current Best Practices in Performing Flow Cytometry in a Regulated Bioanalysis
    - Dr. Cherie Green, Senior Scientific Manager, Genentech
    • Importance of Flow Cytometry standardization in a regulated environment
      • Current industry standards to
        • Develop robust methods
        • Ensure consistent and reproducible data
        • Standardization using hard-dyed microspheres
        • Optimization of Flow Cytometer performance
    • What to consider in a regulated environment
      • Minimizing the high biological variability of cells
      • Minimizing the limited stability of samples
        • How to improve samples stability
        • PBMC vs Whole blood
      • Perfect understanding of
        • Complexity of the data output
        • Interpretation of results
    • No regulatory guidance/guidelines on Flow Cytometry
      • Working with recognized industry standards
      • Recent discussions in Bioanalysis
        • 2016 White Paper Part 3 Recommendations
    • Critical reagents
      • Lot-to-lot differences
      • Variation in conjugation and staining intensities
      • Dye degradation
    • Industry/Regulators’ interaction and feedbacks
    • Case Studies: State-of-the-art approaches in the use of Flow Cytometry in a regulated environment and Regulators’ feedback

  • Lecture 5
    Challenges with Receptor Occupancy (RO) Assays by Flow Cytometry: How to successful validate them?
    - Dr. Meina Liang, Director, MedImmune
    • Advanced understanding of Receptor Occupancy (RO) as a critical component of drug development
      • Issues in RO assays
        • Standardization
        • Validation
        • Data modelling and interpretation
      • Challenges and novel solutions in validating Flow Cytometry RO assays
        • Different approaches for biological fluids vs tissues
        • Dose-response curve
        • Improving Reproducibility
        • Stained Cell stability
        • Acceptance criteria for “Unoccupied” Receptors
    • Why is Flow Cytometry such a powerful tool for Receptor Occupancy?
      • Demonstration of target engagement in the compartment of interest
      • Focus on
        • Drug Activity
        • Optimal dosing
        • Safety assessment
    • Importance of monitoring changes in the number of receptor on the cell
      • Receptor binding by drug vs pre-drug administration receptor levels
      • Extra complexity of RO assays
        • Surface binding
        • Receptor internalization/shedding
    • Case Studies: In-depth evaluation of the issues, how to fix them for Receptor Occupancy assays used clinical trial

  • Lecture 6
    Advanced Applications of Flow Cytometry in CAR T-Cell Therapy (Chimeric Antigen Receptor modified T cells)
    - Dr. Vellalore Kakkanaiah, Associate Director, PPD
    • Recent increase in CAR T-Cell Therapy development after US FDA approval of Kymriah
      • CAR-T landscape
      • CAR-T mechanism of action (MOA)
    • Challenges with the bioanalytical requirements for CAR-T development
      • Bioanalytical development for measuring CAR T-Cells
        • Cellular kinetics
        • Post-antigen exposure
        • Persistence and immunogenicity
    • Flow Cytometry assay development and FFP validation
      • Challenges & solutions
      • Method Robustness
    • Case Studies: Fit-for-Purpose (FFT) Validation of a Flow Cytometry Assay for Anti- B cell maturation antigen (BCMA) CAR-T Cells


Part 3: What’s new in LBA PK Assays
  • Lecture 7
    Insights on a Fully Integrated Bioanalytical Strategies for Early Stage Clinical Studies: Impact of immunogenicity on PK Assays and its correlation to PK & PD data
    - Dr. Eginhard Schick, Principal Scientist, Roche
    • Based on the intended use of the biotherapeutic and the immunogenicity risk assessment, a comprehensive bioanalytical strategy comprising a dedicated assay battery for PK and ADA analysis is of utmost importance, especially at the early clinical phase
    • Understanding of the drug’s pharmacokinetic properties
      • Knowledge of the drug species present in the patient’s blood
        • Fully functional
        • Partially degraded
        • ADA bound
      • Knowledge of the drug’s integrity in the patient’s blood
        • Safety concerns
    • Determination of the ADA domain specificity and possible resulting clinical consequences
      • To know if and which functionally important drug moiety is affected
        • Targeted exposure
        • Systemic exposure
      • To evaluate possible safety concerns
        • Cross-reactivity with endogenous homologue
    • Investigation of the onset of immune responses
      • Immunostimulatory effect of drug on ADA onset
    • Immunocytokines can elicit immune responses in patients resulting in the generation of ADA with multiple specificities
      • Depending on their specificity, ADAs can have an impact on exposure, efficacy and safety in patients
    • Case Studies: Interferences issues - targeted delivery of cytokines using highly potent antibody-cytokine fusion proteins (immunocytokines) for cancer therapy

  • Lecture 8
    PK Assays Strategies for Multi-Domain Biotherapeutic (MDB): What is best to measure?
    - Dr. Priya Sriraman, Director Biologics DMPK, Celgene
    • PK assays for MDB
      • What are the current strategies to develop these assays?
      • What are regulatory expectations on the extent of the characterization required?
      • How to design multiple experiments
    • Characterization MDB mechanisms of action (MoA)
      • MDB currently in development have complex MoA
        • More than one domain
        • Each domain with a specific role or function
    • Issues with MDB containing endogenous counterparts
    • Increase the need for Cell-based Assays (CBA) for PK of MDB
      • Getting better details of MoA
      • Better understanding of what drives efficacy
      • Limitation of LBA for PK
    • Case Studies: New strategies to characterize the pharmacokinetic behavior of MDB and interaction with Regulators

  • Lecture 9
    Focus on Issues with Biosimilar PK Assays Bioanalysis: New Assay or Original Reference Product Assay?
    - Dr. Martin Ullmann, Head of Bioanalytics Biosimilars, Fresenius Kabi
    • What if the Critical Reagents for the Reference Product Assay are not available any more?
      • When is there the need for a newly redeveloped method vs the old one?
    • Reference Products are biotherapeutics often using very old assay designs
      • Obsolete platforms
      • Target capture vs anti-Id capture
    • Are biosimilar assays required to use the same LBA format as the reference product?
      • Development and validation of Biosimilar assays
      • Biosimilar 1 or 2 assays approach comparing results
    • Recent success in Biosimilar assay development
      • Where we came from
      • What we have achieved so far
    • How regulations have changed and are changing
      • Regulatory Perspectives for approval of Biosimilars
      • Absence of clinically meaningful differences
      • Challenges in demonstrating no clinically meaningful differences
    • Case Studies: Development and validation of Biosimilar assays when critical reagents for the reference product are not available


Part 4: Novel case studies on Emerging/Established Technologies
  • Lecture 10
    Gyros Rising: An industry experience, past, present and future
    - Dr. Judy Shih, Principal Scientist, Amgen
    • Gyros (Gyrolab): a unique history
      • From Emerging Technology (ET) to a well-established leading LBA platform
        • An ELISA on a disk
      • Less hands-on time
      • Reduced sample and reagent consumption
      • Broader dynamic range
      • Error minimization due to automation
    • Singleplex/Singlicate with Gyros
      • Validation of Singleplex methods on Gyros
      • What to consider - current strategies
      • 2016 White Paper Part 3 Recommendation on LBA Singleplex/Singlicate methods
    • Successful dealing with Gyros new platforms
      • Implementing fast & efficient method transfer and revalidation on new platforms
      • Approaches to ensure method changes for regulated LBA from old to newer Gyros formats and technology
    • Case Studies: Advanced Strategies for incorporating Gyros in a regulated environment

  • Lecture 11
    Ultra-sensitive LBA platforms: Application in a Regulated Environment
    - Dr. Tong-yuan Yang, Scientific Director, Janssen
    • Ultra-sensitive LBA Technologies
      • Singulex (Erenna®)
      • Quanterix (Simoa™)
      • Meso Scale Discovery (MSD)
      • Gyrolab (Gyros)
    • Faster Method Development
      • Lesson learnt
      • Future developments
    • Advantage of these ultra-sensitive ET
      • Implementation in LBA fully regulated laboratories
    • Case Studies: Direct experience from users on evaluation of ET for sensitivity in regulated bioanalysis

  • Lecture 12
    From Ultra to Extreme-sensitive LBA platforms: Performance evaluation enabling reliable quantification at fg/ml
    - Dr. Gizette Sperinde, Scientist, Genentech
    • Performance evaluation of ultra-sensitive LBA available platforms
      • Pushing the technologies to the extreme sensitivity
        • Singulex (Erenna®)
        • Quanterix (Simoa™)
        • Immuno-PCR, Imperacer®
      • Ability of ET to reach fg/ml in serum samples
        • Specificity and impact on sensitivity
        • Head-to-head comparison
    • Successful approaches used for evaluating ultra-sensitive platform
      • Assays transfer from platform to platform
    • Method development and Validation strategies
      • Lesson learnt
      • Future applications for extreme sensitivity LBA assays
    • Case Studies: Detection of circulating levels of IL-13 and need for extreme sensitivity





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