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Table of Contents
Foreword by the Supervisor; Acknowledgments; Abbreviations; Contents; Summary; Zusammenfassung; 1 Introduction; 1.1 Drug Screening with New Perspectives; 1.2 Real-Time Monitoring of Living Cells Using a Novel Biosensor Chip; 1.2.1 Oxygen Consumption; 1.2.2 Extracellular Acidification; 1.2.3 Cellular Impedance; 1.3 Anticancer Agents
Effects Measured in Real Time; 1.3.1 Cisplatin
A Prominent Anticancer Drug; 1.3.2 Organometallic Compounds; 1.4 Aim of this Study; 2 Materials; 2.1 Instruments; 2.2 Laboratory Materials; 2.3 Chemicals; 2.4 Kit Systems; 2.5 Primers and Oligonucleotides
2.6 Biological Software2.7 Mammalian Cell Lines; 2.8 Vectors; 2.9 Media/Buffers/Solutions; 2.10 Antibodies for Microarrays; 3 Methods; 3.1 Cell Culture; 3.2 Real-Time Monitoring of Cellular Metabolism in Living Cells; 3.2.1 Preparation of Running Medium; 3.2.2 Bionas System Setup for the Experiment; 3.2.3 Preparation and Cultivation of Cells on the Chip; 3.2.4 Real-Time Measurement of Cellular Metabolism; 3.2.5 SIRT3 Overexpression
Experimental Setup and Real-Time Measurement; 3.3 Mitochondrial Oxygen Consumption; 3.4 Antibodies and Recombinant Protein Standards for Protein Microarrays
3.5 RNA Isolation, DNA Microarray, Real-Time PCR3.6 Intracellular ROS Determination; 4 Results; 4.1 Respiration and Glycolysis
Striking Differences Between Cancer Cell Lines; 4.2 Glucose Uptake Regulation
Online Measurements; 4.3 SIRT3 Overexpression
Respiration Profiles Measured Online; 4.4 Cisplatin
Metabolic Changes Before Onset of Cell Death; 4.4.1 Mitochondrial Oxygen Consumption; 4.4.2 Cisplatin
Cell Signaling Profiles; 4.4.3 Cisplatin
Gene Expression Profiles; 4.5 Organometallics
Real-Time Analysis of Metabolic Responses in Cancer Cells; 4.5.1 Ruthenium(II) Polypyridyls
4.5.2 Rhodium(III) Polypyridyls4.5.3 Phenylenediamine Iron; 4.5.4 Benzimidazol-2-ylidene-Gold(I); 5 Discussion; 5.1 Metabolism of Cancer Cells
Differences in the Basic Metabolic Rate; 5.2 Energy Metabolism and Glucose Availability; 5.3 Regulation of Energy Metabolism by Sirtuin Deacetylase SIRT3; 5.4 Cisplatin
Its Effects on Metabolism; 5.4.1 Time Points of Metabolic Changes in Cisplatin Treatment; 5.4.2 Signaling Molecules Akt1, GSK-3€, and ERK1 Are Affected After Long-Time Exposure to Cisplatin
5.4.3 Apoptosis Initiation by Cisplatin
Time Point of Onset as Determined by Gene Expression Profiling5.5 Organometallics
New Anticancer Candidates; 5.5.1 Ruthenium(II) Polypyridyls: Impact on Cellular Impedance in HT-29; 5.5.2 Rhodium(III) Polypyridyls: MCF-7 and HT-29 Respond Differently; 5.5.3 Phenylenediamine Iron: Response of MCF-7 Is Immediate and Irreversible; 5.5.4 Benzimidazol-2-ylidene-Gold(I): Respiration Is Strongly Affected in MCF-7; 6 Conclusion; 7 List of Publications; 8 References; Appendix
Effects Measured in Real Time; 1.3.1 Cisplatin
A Prominent Anticancer Drug; 1.3.2 Organometallic Compounds; 1.4 Aim of this Study; 2 Materials; 2.1 Instruments; 2.2 Laboratory Materials; 2.3 Chemicals; 2.4 Kit Systems; 2.5 Primers and Oligonucleotides
2.6 Biological Software2.7 Mammalian Cell Lines; 2.8 Vectors; 2.9 Media/Buffers/Solutions; 2.10 Antibodies for Microarrays; 3 Methods; 3.1 Cell Culture; 3.2 Real-Time Monitoring of Cellular Metabolism in Living Cells; 3.2.1 Preparation of Running Medium; 3.2.2 Bionas System Setup for the Experiment; 3.2.3 Preparation and Cultivation of Cells on the Chip; 3.2.4 Real-Time Measurement of Cellular Metabolism; 3.2.5 SIRT3 Overexpression
Experimental Setup and Real-Time Measurement; 3.3 Mitochondrial Oxygen Consumption; 3.4 Antibodies and Recombinant Protein Standards for Protein Microarrays
3.5 RNA Isolation, DNA Microarray, Real-Time PCR3.6 Intracellular ROS Determination; 4 Results; 4.1 Respiration and Glycolysis
Striking Differences Between Cancer Cell Lines; 4.2 Glucose Uptake Regulation
Online Measurements; 4.3 SIRT3 Overexpression
Respiration Profiles Measured Online; 4.4 Cisplatin
Metabolic Changes Before Onset of Cell Death; 4.4.1 Mitochondrial Oxygen Consumption; 4.4.2 Cisplatin
Cell Signaling Profiles; 4.4.3 Cisplatin
Gene Expression Profiles; 4.5 Organometallics
Real-Time Analysis of Metabolic Responses in Cancer Cells; 4.5.1 Ruthenium(II) Polypyridyls
4.5.2 Rhodium(III) Polypyridyls4.5.3 Phenylenediamine Iron; 4.5.4 Benzimidazol-2-ylidene-Gold(I); 5 Discussion; 5.1 Metabolism of Cancer Cells
Differences in the Basic Metabolic Rate; 5.2 Energy Metabolism and Glucose Availability; 5.3 Regulation of Energy Metabolism by Sirtuin Deacetylase SIRT3; 5.4 Cisplatin
Its Effects on Metabolism; 5.4.1 Time Points of Metabolic Changes in Cisplatin Treatment; 5.4.2 Signaling Molecules Akt1, GSK-3€, and ERK1 Are Affected After Long-Time Exposure to Cisplatin
5.4.3 Apoptosis Initiation by Cisplatin
Time Point of Onset as Determined by Gene Expression Profiling5.5 Organometallics
New Anticancer Candidates; 5.5.1 Ruthenium(II) Polypyridyls: Impact on Cellular Impedance in HT-29; 5.5.2 Rhodium(III) Polypyridyls: MCF-7 and HT-29 Respond Differently; 5.5.3 Phenylenediamine Iron: Response of MCF-7 Is Immediate and Irreversible; 5.5.4 Benzimidazol-2-ylidene-Gold(I): Respiration Is Strongly Affected in MCF-7; 6 Conclusion; 7 List of Publications; 8 References; Appendix