Investigation


Each project is unique.

Especially when working with a new chemical entity (NCE) we highly recommend to do an early “quick and dirty” feasibility study in “real plasma” to get a grasp of how your compound behaves in plasma (see Feasibilities).

Other questions have to be clarified: Issues with protein binding? Enantio selective specifics?

Tissue analysis is a topic of its own. Done “correctly” it requires a lot of knowledge and experience, especially selection and preparation of tissue samples.


  • Feasibilities
  • Tissue Analysis
  • Protein-Binding
  • Enantio selective Separation
  • Trouble Shooting

  • Background
    Getting an Idea of the Path Ahead
    Having an early, rough outline of how your compound behaves in plasma is a great asset in regard to fundamental decisions in the early phases. Hazards, such as stability in plasma, or general detectability in plasma, want to be ruled out before investing vital time and resources. An early bioanalytical feasibility study is a proven approach in regard to cost- and risk-efficiency.

    In-Vitro: Before Pre-Clinical Studies
    Blank matrix (often plasma) is spiked with your compound and investigated in regard to following parameters (ballpark data):
    - stability in plasma
    - detectability
    - limit of detection
    - linearity
    - selectivity

    In-Vivo: Pilot Pre-Clinical Study
    In a pre-clinical pilot study the API is administered to 2-3 mice or rats (at a pre-clinical lab). Collected plasma is investigated (ballpark data) in regard to:
    - bioavailability
    - impression of metabolism
    - stability in plasma
    - detectability
    - limit of detection
    - linearity
    - selectivity

    Scope
    - 2-4 working days
    - ca. 30 mg API substance

     

    Deliverables
    - Chromatograms
    - Scientific statement

    Benefits
    - Reducing Risk & Disclosing latent “bad news”
    - Estimation of effort and cost
    - Knowing scope of data to expect of pre-clinics
    - Improved position towards VCs and authorities
      In-Vivo:
    - Pre-conclusions on administrative route
    - Impression of bioavailability / metabolism
    - Orientation for dose range



     

    Optimal Time Points for Bioanalytical Feasibilities

  • “Where else would you go for tissue analysis other than to pharm-analyt?” (Head of Institute of PK, Big Pharma).

    The challenge lies in the following aspects:
    - Selection of tissue segment
    - Homogenization (various methods)
    - Extraction (various methods)
    - Quantification / Identification

    Experience with the following types of tissue:
    Organ, Lung, Brain, Eye, Cornea, Bone, Skin, Intestine, Cardiovascular Tissue, Muscle

  • In-vitro tests of human or animal plasma
    Analysis of protein binding of drugs in in-vivo samples
    Combination with chiral separation: protein binding of enantiomeric drugs

    Methods used:
    - Dialysis (various ways)
    - Ultrafiltration


    Image: Dialysis preferred method

  • Chiral Separation of enantiomers of drugs in biological material

    Example Ibuprofen:

    Different protein binding of the enantiomers can occur.

  • Examples:
    Complex Molecule or complex combination of molecules
    Difficult Matrix (esp. tissue!)
    Stability issues / Half-life issues
    Selectivity issues
    Sensitivity issues
    General plausibility doubts or issues
    Regulatory issues related to bioanalysis
    Enantioselective issues
    Protein binding issues
    Issues with controlled delivery forms (PEGylation, Liposomal, etc.)
    Expert Assessments