The role of Kirsten-ras in colorectal cancer
Aims: To determine the contribution of Kirsten-ras and other abnormalities in the signalling pathway from growth factor to RAS in colorectal cancer and to develop a means of targeting a ras mutation of probable prognostic importance. Methods: Sections of 105 consecutive early colorectal cancers were...
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Institute of Cancer Research (University Of London)
2009
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ndltd-bl.uk-oai-ethos.bl.uk-5412942015-10-03T03:17:55ZThe role of Kirsten-ras in colorectal cancerTilsed, J. V. T.2009Aims: To determine the contribution of Kirsten-ras and other abnormalities in the signalling pathway from growth factor to RAS in colorectal cancer and to develop a means of targeting a ras mutation of probable prognostic importance. Methods: Sections of 105 consecutive early colorectal cancers were obtained. DNA was extracted and codons 110-134 of the TGF- β RII receptor gene were amplified using PCR and screened for microsatellite instability. Codons 12 and 13 of the Kirsten-ras gene were identified by direct sequencing. Immunohistochemical staining was used to identify expression of pan RAS proteins, the growth factors EGF and TGF- β, and their common receptor, EGF-R. A ribozyme transcription unit was constructed to target the mRNA produced from Kirsten ras encoding a valine mutation at codon 12. The ribozyme was transcribed in colorectal cell lines using a novel expression/reporter plasmid. Two control ribozyme constructs were also used: one lacked ribozyme activity, the other was not transcribed in colorectal cells. Ribozyme expression and Kirsten-ras mRNA levels in transfected cells were identified by an RNase protection assay. Cell viability after transfection was determined by labelled thymidine uptake. Results: A ras pathway abnormality was identified in almost 90% of the tumours while microsatellite instability at the TGF- β RII gene was identified in only 3 (2.9%). Kirsten-Ras mutations were present in 30 (28.6%). Cleavage of target mRNA by the ribozyme in vitro and transcription in transfected cells were confirmed. However, there was no demonstrable reduction in Kirsten-ras mRNA in treated cells and no specific effect on cell viability.616.99Gastrointestinal UnitInstitute of Cancer Research (University Of London)http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.541294http://publications.icr.ac.uk/10067/Electronic Thesis or Dissertation |
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616.99 Gastrointestinal Unit |
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616.99 Gastrointestinal Unit Tilsed, J. V. T. The role of Kirsten-ras in colorectal cancer |
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Aims: To determine the contribution of Kirsten-ras and other abnormalities in the signalling pathway from growth factor to RAS in colorectal cancer and to develop a means of targeting a ras mutation of probable prognostic importance. Methods: Sections of 105 consecutive early colorectal cancers were obtained. DNA was extracted and codons 110-134 of the TGF- β RII receptor gene were amplified using PCR and screened for microsatellite instability. Codons 12 and 13 of the Kirsten-ras gene were identified by direct sequencing. Immunohistochemical staining was used to identify expression of pan RAS proteins, the growth factors EGF and TGF- β, and their common receptor, EGF-R. A ribozyme transcription unit was constructed to target the mRNA produced from Kirsten ras encoding a valine mutation at codon 12. The ribozyme was transcribed in colorectal cell lines using a novel expression/reporter plasmid. Two control ribozyme constructs were also used: one lacked ribozyme activity, the other was not transcribed in colorectal cells. Ribozyme expression and Kirsten-ras mRNA levels in transfected cells were identified by an RNase protection assay. Cell viability after transfection was determined by labelled thymidine uptake. Results: A ras pathway abnormality was identified in almost 90% of the tumours while microsatellite instability at the TGF- β RII gene was identified in only 3 (2.9%). Kirsten-Ras mutations were present in 30 (28.6%). Cleavage of target mRNA by the ribozyme in vitro and transcription in transfected cells were confirmed. However, there was no demonstrable reduction in Kirsten-ras mRNA in treated cells and no specific effect on cell viability. |
author |
Tilsed, J. V. T. |
author_facet |
Tilsed, J. V. T. |
author_sort |
Tilsed, J. V. T. |
title |
The role of Kirsten-ras in colorectal cancer |
title_short |
The role of Kirsten-ras in colorectal cancer |
title_full |
The role of Kirsten-ras in colorectal cancer |
title_fullStr |
The role of Kirsten-ras in colorectal cancer |
title_full_unstemmed |
The role of Kirsten-ras in colorectal cancer |
title_sort |
role of kirsten-ras in colorectal cancer |
publisher |
Institute of Cancer Research (University Of London) |
publishDate |
2009 |
url |
http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.541294 |
work_keys_str_mv |
AT tilsedjvt theroleofkirstenrasincolorectalcancer AT tilsedjvt roleofkirstenrasincolorectalcancer |
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