The development of improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals in the discovery phase of drug development

W.M.A. Westerink

Research output: Thesisinternal PhD, WUAcademic

Abstract

In drug development, toxicity is an important factor for attrition, resulting in a failure rate of 30%-40%. Hepatotoxicity, nephrotoxicity, cardiovascular safety, reproduction toxicity, developmental toxicity (teratogenicity), genotoxicity and carcinogenicity are the main causes for attrition in safety assessment.
Screening on these aspects in the early discovery phase of drug development and using these data for compound optimization and deselection might result in drug development candidates with an improved success rate. The present thesis focused on early screening for genotoxicity and carcinogenicity.
In recent years some progress has been made with assays to assess genotoxicity and non-genotoxic carcinogenicity at the end of the discovery phase. However, the time point at which these genotoxicity and carcinogenicity assays are performed is still relatively late, only a few assays for such a strategy are available, the throughput of these assays is in general still low and most of them have not yet been validated extensively. An additional drawback is that the currently used in vitro assays for the detection of genotoxicity give a high rate of false positive results, which makes application in the early discovery phase of drug development cumbersome.
The goal of the present thesis was therefore to develop improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals, validate these assays with proper reference compounds, and to develop a strategy for application of these assays in the early discovery phase of drug development.
High throughput assays based on bacteria, yeast and human/rodent cell lines were developed. In the case of human cell lines, the focus was on the HepG2 cell line as the properties of HepG2 cells are expected to give a good prediction for in vivo genotoxicity. The results in this thesis show that an early prediction can be made for bacterial mutagenicity (gene mutations), mammalian genotoxicity (chromosome damage), and non-genotoxic carcinogenic potential by aryl hydrocarbon receptor (AhR) activation. To develop a strategy for application of the HTS genotoxicity assays in the early discovery phase several combinations of assays were evaluated. The combination VitotoxTM + HepG2 p53 reporter assay was based on the presented results in this thesis the most useful for screening compounds for their genotoxic potential in the early drug discovery phase without the risk on high numbers of false positives. CYP1A induction assays in human HepG2 and rat H4IIE cells may be performed in parallel with these assays to be able to detect non-genotoxic carcinogenic potential by AhR activators. Further application of these assays may prove useful in future drug development strategies.

LanguageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Wageningen University
Supervisors/Advisors
  • Rietjens, Ivonne, Promotor
  • Groten, John, Promotor
  • Schoonen, W.G.E.J., Co-promotor, External person
Award date4 Feb 2011
Place of Publication[S.l.
Publisher
Print ISBNs9789085858478
Publication statusPublished - 2011

Fingerprint

Drug Discovery
Aryl Hydrocarbon Receptors
Hep G2 Cells
Cell Line
Mammalian Chromosomes
Pharmaceutical Preparations
Safety
Bacterial Genes
Reproduction
Rodentia
Yeasts
Bacteria
Mutation
In Vitro Techniques

Keywords

  • genotoxicity
  • carcinogenesis
  • new drugs
  • drug development
  • in vitro
  • in vitro culture

Cite this

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title = "The development of improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals in the discovery phase of drug development",
abstract = "In drug development, toxicity is an important factor for attrition, resulting in a failure rate of 30{\%}-40{\%}. Hepatotoxicity, nephrotoxicity, cardiovascular safety, reproduction toxicity, developmental toxicity (teratogenicity), genotoxicity and carcinogenicity are the main causes for attrition in safety assessment. Screening on these aspects in the early discovery phase of drug development and using these data for compound optimization and deselection might result in drug development candidates with an improved success rate. The present thesis focused on early screening for genotoxicity and carcinogenicity. In recent years some progress has been made with assays to assess genotoxicity and non-genotoxic carcinogenicity at the end of the discovery phase. However, the time point at which these genotoxicity and carcinogenicity assays are performed is still relatively late, only a few assays for such a strategy are available, the throughput of these assays is in general still low and most of them have not yet been validated extensively. An additional drawback is that the currently used in vitro assays for the detection of genotoxicity give a high rate of false positive results, which makes application in the early discovery phase of drug development cumbersome. The goal of the present thesis was therefore to develop improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals, validate these assays with proper reference compounds, and to develop a strategy for application of these assays in the early discovery phase of drug development. High throughput assays based on bacteria, yeast and human/rodent cell lines were developed. In the case of human cell lines, the focus was on the HepG2 cell line as the properties of HepG2 cells are expected to give a good prediction for in vivo genotoxicity. The results in this thesis show that an early prediction can be made for bacterial mutagenicity (gene mutations), mammalian genotoxicity (chromosome damage), and non-genotoxic carcinogenic potential by aryl hydrocarbon receptor (AhR) activation. To develop a strategy for application of the HTS genotoxicity assays in the early discovery phase several combinations of assays were evaluated. The combination VitotoxTM + HepG2 p53 reporter assay was based on the presented results in this thesis the most useful for screening compounds for their genotoxic potential in the early drug discovery phase without the risk on high numbers of false positives. CYP1A induction assays in human HepG2 and rat H4IIE cells may be performed in parallel with these assays to be able to detect non-genotoxic carcinogenic potential by AhR activators. Further application of these assays may prove useful in future drug development strategies.",
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TY - THES

T1 - The development of improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals in the discovery phase of drug development

AU - Westerink, W.M.A.

N1 - WU thesis 4975

PY - 2011

Y1 - 2011

N2 - In drug development, toxicity is an important factor for attrition, resulting in a failure rate of 30%-40%. Hepatotoxicity, nephrotoxicity, cardiovascular safety, reproduction toxicity, developmental toxicity (teratogenicity), genotoxicity and carcinogenicity are the main causes for attrition in safety assessment. Screening on these aspects in the early discovery phase of drug development and using these data for compound optimization and deselection might result in drug development candidates with an improved success rate. The present thesis focused on early screening for genotoxicity and carcinogenicity. In recent years some progress has been made with assays to assess genotoxicity and non-genotoxic carcinogenicity at the end of the discovery phase. However, the time point at which these genotoxicity and carcinogenicity assays are performed is still relatively late, only a few assays for such a strategy are available, the throughput of these assays is in general still low and most of them have not yet been validated extensively. An additional drawback is that the currently used in vitro assays for the detection of genotoxicity give a high rate of false positive results, which makes application in the early discovery phase of drug development cumbersome. The goal of the present thesis was therefore to develop improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals, validate these assays with proper reference compounds, and to develop a strategy for application of these assays in the early discovery phase of drug development. High throughput assays based on bacteria, yeast and human/rodent cell lines were developed. In the case of human cell lines, the focus was on the HepG2 cell line as the properties of HepG2 cells are expected to give a good prediction for in vivo genotoxicity. The results in this thesis show that an early prediction can be made for bacterial mutagenicity (gene mutations), mammalian genotoxicity (chromosome damage), and non-genotoxic carcinogenic potential by aryl hydrocarbon receptor (AhR) activation. To develop a strategy for application of the HTS genotoxicity assays in the early discovery phase several combinations of assays were evaluated. The combination VitotoxTM + HepG2 p53 reporter assay was based on the presented results in this thesis the most useful for screening compounds for their genotoxic potential in the early drug discovery phase without the risk on high numbers of false positives. CYP1A induction assays in human HepG2 and rat H4IIE cells may be performed in parallel with these assays to be able to detect non-genotoxic carcinogenic potential by AhR activators. Further application of these assays may prove useful in future drug development strategies.

AB - In drug development, toxicity is an important factor for attrition, resulting in a failure rate of 30%-40%. Hepatotoxicity, nephrotoxicity, cardiovascular safety, reproduction toxicity, developmental toxicity (teratogenicity), genotoxicity and carcinogenicity are the main causes for attrition in safety assessment. Screening on these aspects in the early discovery phase of drug development and using these data for compound optimization and deselection might result in drug development candidates with an improved success rate. The present thesis focused on early screening for genotoxicity and carcinogenicity. In recent years some progress has been made with assays to assess genotoxicity and non-genotoxic carcinogenicity at the end of the discovery phase. However, the time point at which these genotoxicity and carcinogenicity assays are performed is still relatively late, only a few assays for such a strategy are available, the throughput of these assays is in general still low and most of them have not yet been validated extensively. An additional drawback is that the currently used in vitro assays for the detection of genotoxicity give a high rate of false positive results, which makes application in the early discovery phase of drug development cumbersome. The goal of the present thesis was therefore to develop improved and new in vitro assays for detecting the genotoxic and non-genotoxic carcinogenic potential of chemicals, validate these assays with proper reference compounds, and to develop a strategy for application of these assays in the early discovery phase of drug development. High throughput assays based on bacteria, yeast and human/rodent cell lines were developed. In the case of human cell lines, the focus was on the HepG2 cell line as the properties of HepG2 cells are expected to give a good prediction for in vivo genotoxicity. The results in this thesis show that an early prediction can be made for bacterial mutagenicity (gene mutations), mammalian genotoxicity (chromosome damage), and non-genotoxic carcinogenic potential by aryl hydrocarbon receptor (AhR) activation. To develop a strategy for application of the HTS genotoxicity assays in the early discovery phase several combinations of assays were evaluated. The combination VitotoxTM + HepG2 p53 reporter assay was based on the presented results in this thesis the most useful for screening compounds for their genotoxic potential in the early drug discovery phase without the risk on high numbers of false positives. CYP1A induction assays in human HepG2 and rat H4IIE cells may be performed in parallel with these assays to be able to detect non-genotoxic carcinogenic potential by AhR activators. Further application of these assays may prove useful in future drug development strategies.

KW - genotoxiciteit

KW - carcinogenese

KW - nieuwe geneesmiddelen

KW - geneesmiddelenontwikkeling

KW - in vitro

KW - in vitro kweek

KW - genotoxicity

KW - carcinogenesis

KW - new drugs

KW - drug development

KW - in vitro

KW - in vitro culture

M3 - internal PhD, WU

SN - 9789085858478

PB - S.n.

CY - [S.l.

ER -