Integrated platform for liquid biopsy-based …Integrated platform for liquid biopsy-based...
Transcript of Integrated platform for liquid biopsy-based …Integrated platform for liquid biopsy-based...
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Integrated platform for liquid biopsy-basedpersonalized cancer medicine
Dr. Bernhard Polzer
Fraunhofer ITEM-RegensburgPersonalized Tumor Therapy
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Personalized cancer therapy
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Primary tumor Micrometastasissingle tumor cells Metastasis
Molecular Analysis &Therapy Selection
Molecular Analysis &Therapy Selection
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Polzer and Klein, Nat Med 2013
Cellular heterogeneity is the basis for therapy resistance
Selection for resistant, aggressive subclones by applied therapies
Cancer is an evolving disease
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Personalized cancer therapy
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Surgery
Primary tumor Micrometastasissingle tumor cells Metastasis
Information on systemic cancer?
metastatic disease
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Personalized cancer therapy
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Surgery
Primary tumor Micrometastasissingle tumor cells Metastasis
Information on systemic cancer? Liquid Biopsy
metastatic disease
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Diaz and Bardelli, JCO 2014
Information on systemic cancerin a simple blood test
- Circulating tumor cells (CTCs)
- Circulating tumor DNA (ctDNA)
- miRNA (and other RNAs)
- Extracellular vesicles(e.g. exosomes)
Concept of liquid biopsy
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2. Detection of CTC
1. Enrichment for CTC
4. Whole Genome amplification (WGA)
5. Unbiased molecular analysis
Ampli1TM
WGA
e.g.CellSearch®
3. Isolation of single CTC e.g.DepArrayTM
Analysis of circulating tumor cells
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CK HER2 CD45 DAPI PIK3CA mutHER2 qPCR
amplified
amplified
amplified
amplified
amplified
amplified
Patient MU09 Primary Her2 negative, CTC count 42 (Veridex)
Molecular heterogeneity in breast cancer
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T02
T04
T05
T06
T10
T07
CK HER2 CD45 DAPI PIK3CA mut
wt
wt
H1047R
H1047R
H1047R
H1047R
HER2 qPCR
amplified
amplified
amplified
amplified
amplified
amplified
Patient MU09 Primary Her2 negative, CTC count 42 (Veridex)
Molecular heterogeneity in breast cancer
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cfDNA isolationfrom plasma
CTC enrichmentfrom buffy coat
Expertise inpreanalytical workflows andsample logisticsfor liquid biopsy studies
Comprehensive liquid biopsy concept
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cfDNA isolationfrom plasma
CTC enrichmentfrom buffy coat
Expertise inpreanalytical workflows andsample logisticsfor liquid biopsy studies
Protocols available for
customized strategies(e.g. epitope based/marker-free)
High blood volumes(50-100 ml, leukapheresis
Other body fluids (e.g. bonemarrow, cerebro-spinal fluid)and organs (e.g. lymph nodes)
Comprehensive liquid biopsy concept
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Whole transcriptomeamplification (WTA)
Whole genomeamplification (WGA)
Cell lysis
Quality control forpatient-derived cells
Whole GenomeSequencing (HiSeq)
RNASeq analysis(HiSeq)
CNV analysis(MiSeq)
Whole Exome/PanelSequencing (HiSeq)
Molecular analysis of single cells of patients
lncRNASeq analysis(HiSeq)
Small RNASeq analysis(MiSeq)
Error free single cell Seq(PCTEP2015052436)
Single celltranscriptomicsbioinformatics
(tailored to WTAtechnology)
Single cellgenomics
bioinformatics(tailored to WGA
technology)
Automatedworkflow
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liquid biopsy models for:
• testing novel therapies
• personalized therapy
selection
• biomarker/ drug targets
• understanding
metastastic progression
in vivo
in vitro
tumor cells
Preclinical models from liquid biopsy samples
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Cell culture 384-wellassay plate
Distribution of patient-derived cellularmodels on assay plates
Compoundadministration (nl)
incubation (37oC)Dispensing of detection reagent on cells
readout(e.g. luminescence)
Compound libraries
MTT, AlamarBlue, protease, ATP
High-throughput screening of compoundsin patient-derived cell models
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Poster Z9
Dr. Bernhard [email protected]
Dr. Kamran [email protected]
Fraunhofer Institute for Toxicology and Experimental Medicine - ITEMDivision of Personalized Tumor Therapy, RegensburgAm Biopark 993053 RegensburgGermany