Cell transformation assays for prediction of carcinogenic potential: state of the science and future research needs

Cell transformation assays (CTAs) have long been proposed as in vitro methods for the identification of potential chemical carcinogens. Despite showing good correlation with rodent bioassay data, concerns over the subjective nature of using morphological criteria for identifying transformed cells and a lack of understanding of the mechanistic basis of the assays has limited their acceptance for regulatory purposes. However, recent drivers to find alternative carcinogenicity assessment methodologies, such as the Seventh Amendment to the EU Cosmetics Directive, have fuelled renewed interest in CTAs. Research is currently ongoing to improve the objectivity of the assays, reveal the underlying molecular changes leading to transformation and explore the use of novel cell types. The UK NC3Rs held an international workshop in November 2010 to review the current state of the art in this field and provide directions for future research. This paper outlines the key points highlighted at this meeting.

[1]  C. Fenoglio-Preiser,et al.  Acetylcholinesterase supports anchorage independence in colon cancer , 2008, Clinical & Experimental Metastasis.

[2]  J. Carter,et al.  CANCER BIOLOGY: Isolation and biological characterization of morphological transformation-sensitive Syrian hamster embryo cells , 1996 .

[3]  L. Sachs,et al.  Cell susceptibility to transformation and cytotoxicity by the carcinogenic hydrocarbon benzo[a]pyrene. , 1966, Proceedings of the National Academy of Sciences of the United States of America.

[4]  M. Stampfer,et al.  Stepwise DNA methylation changes are linked to escape from defined proliferation barriers and mammary epithelial cell immortalization. , 2009, Cancer research.

[5]  R. Isfort,et al.  Comparison of the standard and reduced pH Syrian hamster embryo (SHE) cell in vitro transformation assays in predicting the carcinogenic potential of chemicals. , 1996, Mutation research.

[6]  R. E. Kouri,et al.  Morphological transformation of BALB/3T3 cells by various procarcinogens in the presence of a rat liver S‐9 activation system , 1990, Environmental and molecular mutagenesis.

[7]  William C Hahn,et al.  Oncogenic transformation and experimental models of human cancer. , 2008, Frontiers in bioscience : a journal and virtual library.

[8]  Lutz Müller,et al.  Evaluation of the ability of a battery of three in vitro genotoxicity tests to discriminate rodent carcinogens and non-carcinogens II. Further analysis of mammalian cell results, relative predictivity and tumour profiles. , 2006, Mutation research.

[9]  R. Marcos,et al.  No increase in micronuclei frequency in cultured blood lymphocytes from a group of filling station attendants. , 1996, Mutation research.

[10]  P. Yaswen,et al.  Human epithelial cell immortalization as a step in carcinogenesis. , 2003, Cancer letters.

[11]  D E Robinson,et al.  Background and Framework for ILSI's Collaborative Evaluation Program on Alternative Models for Carcinogenicity Assessment , 2001, Toxicologic pathology.

[12]  P. Carmichael,et al.  Non-Animal Approaches for Consumer Safety Risk Assessments: Unilever's Scientific Research Programme , 2009, Alternatives to laboratory animals : ATLA.

[13]  R. E. Kouri,et al.  A method for the amplification of chemically induced transformation in C3H/10T1/2 clone 8 cells: its use as a potential screening assay. , 1987, Journal of the National Cancer Institute.

[14]  E. Lalli,et al.  Specific Immunoassays for Placental Alkaline Phosphatase As a Tumor Marker , 2006, Journal of biomedicine & biotechnology.

[15]  J. Campisi,et al.  Cellular senescence: when bad things happen to good cells , 2007, Nature Reviews Molecular Cell Biology.

[16]  Kumiko Hayashi,et al.  A Bhas 42 cell transformation assay on 98 chemicals: the characteristics and performance for the prediction of chemical carcinogenicity. , 2010, Mutation research.

[17]  M. Hayashi,et al.  Detection of initiating as well as promoting activity of chemicals by a novel cell transformation assay using v-Ha-ras-transfected BALB/c 3T3 cells (Bhas 42 cells). , 2005, Mutation research.

[18]  Francis L Martin,et al.  Biospectroscopy to metabolically profile biomolecular structure: a multistage approach linking computational analysis with biomarkers. , 2011, Journal of proteome research.

[19]  Heidi S Feiler,et al.  Molecular distinctions between stasis and telomere attrition senescence barriers shown by long-term culture of normal human mammary epithelial cells. , 2009, Cancer research.

[20]  L. Sachs,et al.  In Vitro Cell Transformation with Chemical Carcinogens , 1963, Nature.

[21]  Nicola J Hewitt,et al.  A tiered approach to the use of alternatives to animal testing for the safety assessment of cosmetics: genotoxicity. A COLIPA analysis. , 2010, Regulatory toxicology and pharmacology : RTP.

[22]  N. Tanaka,et al.  An international validation study of a Bhas 42 cell transformation assay for the prediction of chemical carcinogenicity. , 2011, Mutation research.

[23]  J. Harvey,et al.  The results of five coded compounds: genistein, metaproterenol, rotenone, p-anisidine and resorcinol tested in the pH 6.7 Syrian hamster embryo cell morphological transformation assay. , 2005, Mutagenesis.

[24]  Michio Sato,et al.  Improvement of carcinogen identification in BALB/3T3 cell transformation by application of a 2-stage method. , 1989, Mutation research.

[25]  Lutz Müller,et al.  Evaluation of the ability of a battery of three in vitro genotoxicity tests to discriminate rodent carcinogens and non-carcinogens I. Sensitivity, specificity and relative predictivity. , 2005, Mutation research.

[26]  P. Carmichael,et al.  Syrian Hamster Embryo (SHE) cell transformation assay with and without X-ray irradiation of feeder cells using Di(2-ethylhexyl)phthalate (DEHP) and N-nitroso-N-methylnitroguanidine (MNNG). , 2010, Mutation research.

[27]  William C Hahn,et al.  Functional genetics and experimental models of human cancer. , 2004, Trends in molecular medicine.

[28]  William C Hahn,et al.  Identification of genotype-selective antitumor agents using synthetic lethal chemical screening in engineered human tumor cells. , 2003, Cancer cell.

[29]  H. Soreq,et al.  Acetylcholinesterase and butyrylcholinesterase genes coamplify in primary ovarian carcinomas. , 1990, The Journal of clinical investigation.

[30]  L. Sachs,et al.  In vitro transformation of normal cells to tumor cells by carcinogenic hydrocarbons. , 1965, Journal of the National Cancer Institute.

[31]  D. Griffin,et al.  A telomere-independent senescence mechanism is the sole barrier to Syrian hamster cell immortalization , 1998, Oncogene.

[32]  S. Bruce,et al.  Syrian hamster embryo (SHE) cell transformation assay with conditioned media (without X-ray irradiated feeder layer) using 2,4-diaminotoluene, 2,6-diaminotoluene and chloral hydrate. , 2008, Mutation research.

[33]  R. Leboeuf,et al.  The induction of transformed-like morphology and enhanced growth in Syrian hamster embryo cells grown at acidic pH. , 1986, Carcinogenesis.

[34]  J. N. Rodríguez-López,et al.  Cancer-associated differences in acetylcholinesterase activity in bronchial aspirates from patients with lung cancer. , 2008, Clinical science.

[35]  R. T. Bunch,et al.  The Syrian Hamster Embryo (SHE) Cell Transformation Assay: Review of the Methods and Results , 2001, Toxicologic pathology.

[36]  Raffaella Corvi,et al.  ECVAM prevalidation of three cell transformation assays. , 2011, ALTEX.

[37]  W. Hahn,et al.  Immortalized cells as experimental models to study cancer , 2004, Cytotechnology.

[38]  G. Evan,et al.  Cellular senescence: hot or what? , 2009, Current opinion in genetics & development.

[39]  Birgit Schoeberl,et al.  An activated ErbB3/NRG1 autocrine loop supports in vivo proliferation in ovarian cancer cells. , 2010, Cancer cell.

[40]  W. Hahn,et al.  An Activated ErbB 3 / NRG 1 Autocrine Loop Supports In Vivo Proliferation in Ovarian Cancer Cells , 2022 .

[41]  Jane E. Visvader,et al.  Cells of origin in cancer , 2011, Nature.

[42]  P. Layer,et al.  Novel functions of cholinesterases in development, physiology and disease. , 1995, Progress in histochemistry and cytochemistry.

[43]  David Kirkland,et al.  Evaluation of the ability of a battery of three in vitro genotoxicity tests to discriminate rodent carcinogens and non-carcinogens III. Appropriate follow-up testing in vivo. , 2005, Mutation research.

[44]  Ping Yang,et al.  Development of human cell models for assessing the carcinogenic potential of chemicals. , 2008, Toxicology and applied pharmacology.

[45]  R. Isfort,et al.  Application of in vitro cell transformation assays to predict the carcinogenic potential of chemicals. , 1996, Mutation research.

[46]  P. Farmer.  Committee on Mutagenicity of Chemicals in Food, Consumer Products and the Environment ILSI/HESI Research Programme on Alternative Cancer Models: Results of Syrian Hamster Embryo Cell Transformation Assay , 2002, Toxicologic pathology.

[47]  M. Fujita,et al.  Efficient immortalization of primary human cells by p16INK4a‐specific short hairpin RNA or Bmi‐1, combined with introduction of hTERT , 2007, Cancer science.

[48]  Francis L Martin,et al.  IR microspectroscopy: potential applications in cervical cancer screening. , 2007, Cancer letters.

[49]  R. Newbold,et al.  Fibroblast immortality is a prerequisite for transformation by EJ c-Ha-ras oncogene , 1983, Nature.

[50]  Albrecht Poth,et al.  Bhas42 cell transformation assay as a predictor of carcinogenicity , 2008 .

[51]  R. Newbold,et al.  Induction of immortality is an early event in malignant transformation of mammalian cells by carcinogens , 1982, Nature.

[52]  M. Aardema,et al.  The pH 6.7 Syrian hamster embryo cell transformation assay for assessing the carcinogenic potential of chemicals. , 1996, Mutation research.

[53]  Francis L Martin,et al.  Discrimination of a transformation phenotype in Syrian golden hamster embryo (SHE) cells using ATR-FTIR spectroscopy. , 2009, Toxicology.

[54]  S. Cohen,et al.  Alternative Models for Carcinogenicity Testing: Weight of Evidence Evaluations Across Models , 2001, Toxicologic pathology.

[55]  M. Aardema,et al.  A comprehensive protocol for conducting the Syrian hamster embryo cell transformation assay at pH 6.70. , 1996, Mutation research.

[56]  Francis L Martin,et al.  Syrian hamster embryo (SHE) assay (pH 6.7) coupled with infrared spectroscopy and chemometrics towards toxicological assessment. , 2010, The Analyst.

[57]  I. Cavalli,et al.  Amplification and deletion of the ACHE and BCHE cholinesterase genes in sporadic breast cancer. , 2010, Cancer genetics and cytogenetics.

[58]  Francis L Martin.  Shining a new light into molecular workings , 2011, Nature Methods.

[59]  W. Hahn,et al.  Emerging roles for the non-canonical IKKs in cancer , 2011, Oncogene.