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Case Studies on ultra-sensitiveimmunoanalytics based on Imperacer (Immuno-PCR) in clinical settings.Jan Detmers, Ph.D. (Chimera Biotec GmbH)
Agenda1.Overview Platforms2.Immuno-PCR (Imperacer ) Technology3.Applications4.Case Studies
Chimera BiotecCompany Overview Founded 2000 in Dortmund, Germany 3 Divisions: CRO, Kit & Instrument Manufacturer CRO: Assay Development, Bioanalysis Product: Diagnostic Kits (5 CE-IVD clearedproducts), Assay Development Kits, Instruments Revenues: 50% North America, 50% EU 2012 AAPS Innovation in Biotechnology OCompanionCompanionDiagnosticsDiagnostics
MS vs. LBA Platforms“typical” LLoQ of algap
Clinical LBA PlatformsAssay xGyrosECLFSCImperacer Choose the appropriate platform for your assay
Comparison: Clinical LBA vityrequirementBead-ELISALuminex highokayMicrofl. himera (-)lowmediumlowmediumlowokayokayokaygoodexcellent
Assay vs. MethodMethod Assay Matrix Effects Study Requirements- Reagent stability- Interfering factors- Oligomerization effects- Binding Proteins- Aging/Degradation effects- Storage effects- Healty vs. diseased- Drug tolerance- Patient pretreatment- Heterophilic antibodies- Sample availability (e.g.volume, aging effects)Sensitivity LLOQ in control and study groups,validated according to Lee et al. (2006)
Technology: The Principle Antibody-DNA conjugates instead of antibody-enzyme conjugates 1000-fold increase in sensitivity based on signal amplification data read-out by quantitative PCR (qPCR)8
Assay Formats & Sample Types* Imperacer supports all relevantassay formats. Imperacer has been validated inall clinically relevant samples types. Chimeras has completed 287 preclinical and clinical projects usingImperacer*Source: Adler, M. & Spengler, M., Curr Pharmac Anal (2009)9
ELISA vs. ImperacerImperacer Main Advantages:LBA SensitivityBroad Detection Range
1: PK of a low dosed DrugMistletoe Lectin Imperacer Phase I clinical study Solid tumors; Intravenous Aviscumine 41 patientsAviscumineGLP-compliant validated Imperacer Method correlation coefficient of 0.99 quantification range 0.1 - 1000 pg/mL average recovery in citrate plasma 86% LLOQ 100 fg/mLPlease also refer to: Imperacer Presentations at the EBF open Conference, 16-18th Nov. 2011, BarcelonaClinical Pharmacokinetics: Quantifying drug concentrations at ultra low levels;;Punnamoottil et al. (Chimera Biotec, Dortmund, Germany)
PK: Free vs. Complexed DrugDetermining the in-vivo equilibrium constant of monoclonalsidiotypicorIg-chainABDrug Targetequilibrium constant equationmABdrugTargetFree drugTarget spec.ABDrug-target complex in-vivo PK: Quantification of free drug levels (ELISA) Dosing: Understanding dynamic Drug-Target relationship in-vivo12
2: PD: Supportive BiomarkerCytokine Receptor BlockingImperacer monitored theendogenous cytokine levelbetween 0.1 and approx. 50pg/ml before treatment (blue) andthe elevated cytokine level aftertreatment (red) in a panel of 920individuals. Analysis of individual cytokine levels in serum after treatment with thedrug revealed an increased level of free cytokine which was no longerable to bind to the blocked receptor.13
PD: Anti-Cytokine TherapyHow much cytokine is g-chainABCytokineCytokineDrug CytokineAnti cytokine ABfree cytokinebound cytokinetotal cytokine Drugability: Quantification of levels of free target Dosing: Understanding dynamic relation between drug & target Dynamics: compare in vitro and in vivo binding affinities14
Free - Total - Complex Biomarker
In-house Assay Developmentassaymaximizing methoddevelopment assaykitsensitivity engineeringAntibodiesGLP Bioanalysisvalidation“Spirit of GLP”Study SeraRequired material & time-approx. 1 mg capture anddetection antibody- approx. 10-15 assaydevelopment kits- approx. 3-6 weeksImperacer Workstationassay development kitproject planning
Method Development Antibodies/sera:Customer/Commercial Deliverables:Detailed Report ( Kits) Timeline:10 to 12 weeks17
Summary Imperacer is the most sensitive LBA platform available on average 1000 fold better sensitivity than ELISA all options available Out contracting In-house assay development (instrument & kits) Third party CRO (PRA International) Imperacer has been reviewed by FDA and EMA 56 peer-reviewed publications
Thank you!
Imperacer Dive deeper into the ProteomeContact:Dr. Jan Detmersdetmers@chimera-biotec.comNorth AmericaPhone: (617) 861-6018Fax: (617) 939-9608EuropePhone: 49 (0)231 9742-840Fax: 49 (0)231 9742-844www.chimera-biotec.com
1.Overview Platforms 2.Immuno-PCR (Imperacer ) Technology . RIA Western ELISA Luminex Gyros ECL FSC Imperacer basic good excellent . Microfl. ELISA Gyros low okay Bead-ELISA Luminex high okay Comparison: Clinical LBA Platforms. Method Assay Matrix Effects Study Requirements-
