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Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities

Hypoxia-inducible factor (HIF) is one of the most important transcription factors involves in cells survival under low oxygen concentrations, termed hypoxia. Hypoxia is associated with various human diseases from ischemic injuries to cancer. The central role of HIF in these conditions has made it a...

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Main Author: Liew, Sien Yei
Format: Thesis
Language:English
Published: 2013
Online Access:http://psasir.upm.edu.my/id/eprint/39929/1/FBSB%202013%2020R.pdf
http://psasir.upm.edu.my/id/eprint/39929/
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spelling my.upm.eprints.399292016-01-21T07:28:13Z http://psasir.upm.edu.my/id/eprint/39929/ Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities Liew, Sien Yei Hypoxia-inducible factor (HIF) is one of the most important transcription factors involves in cells survival under low oxygen concentrations, termed hypoxia. Hypoxia is associated with various human diseases from ischemic injuries to cancer. The central role of HIF in these conditions has made it as one of the main target molecules for drug discovery. Demand for HIF assay system is high. Cell-based HIF assay systems are preferred since they can provide direct data for intracellular biochemical changes. HIF assay systems which are currently available in the market offer limited sensitivity with restricted reproducibility at a high cost. Therefore, in the present study, a HIF assay was succesfully developed. The system used a Saos-2 human osteosarcoma cell line which is one of the most studied cell lines to express a luciferase protein in response to HIF activation. The luciferase gene construct which was previously cloned downstream of four copies of a hypoxia response element (HRE) of the erythropoietin (EPO) gene. EPO is one of the specific target genes of HIF. Therefore this assay can be used to measure HIF activation upon induction by candidate drugs of interest, by measuring the luciferase expression. This highly sensitive assay system offers a 40-times increased sensitivity compared to the currently available systems. This aspect would be highly beneficial to researchers since the increase in sensitivity can contribute towards result consistency and reproducibility leading to reduced wastage of precious drug candidates. 2013-11 Thesis NonPeerReviewed application/pdf en http://psasir.upm.edu.my/id/eprint/39929/1/FBSB%202013%2020R.pdf Liew, Sien Yei (2013) Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities. Masters thesis, Universiti Putra Malaysia.
institution Universiti Putra Malaysia
building UPM Library
collection Institutional Repository
continent Asia
country Malaysia
content_provider Universiti Putra Malaysia
content_source UPM Institutional Repository
url_provider http://psasir.upm.edu.my/
language English
description Hypoxia-inducible factor (HIF) is one of the most important transcription factors involves in cells survival under low oxygen concentrations, termed hypoxia. Hypoxia is associated with various human diseases from ischemic injuries to cancer. The central role of HIF in these conditions has made it as one of the main target molecules for drug discovery. Demand for HIF assay system is high. Cell-based HIF assay systems are preferred since they can provide direct data for intracellular biochemical changes. HIF assay systems which are currently available in the market offer limited sensitivity with restricted reproducibility at a high cost. Therefore, in the present study, a HIF assay was succesfully developed. The system used a Saos-2 human osteosarcoma cell line which is one of the most studied cell lines to express a luciferase protein in response to HIF activation. The luciferase gene construct which was previously cloned downstream of four copies of a hypoxia response element (HRE) of the erythropoietin (EPO) gene. EPO is one of the specific target genes of HIF. Therefore this assay can be used to measure HIF activation upon induction by candidate drugs of interest, by measuring the luciferase expression. This highly sensitive assay system offers a 40-times increased sensitivity compared to the currently available systems. This aspect would be highly beneficial to researchers since the increase in sensitivity can contribute towards result consistency and reproducibility leading to reduced wastage of precious drug candidates.
format Thesis
author Liew, Sien Yei
spellingShingle Liew, Sien Yei
Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
author_facet Liew, Sien Yei
author_sort Liew, Sien Yei
title Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
title_short Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
title_full Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
title_fullStr Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
title_full_unstemmed Establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
title_sort establishment of a cell-based reporter assay for screening of hypoxia-inducible factor activities
publishDate 2013
url http://psasir.upm.edu.my/id/eprint/39929/1/FBSB%202013%2020R.pdf
http://psasir.upm.edu.my/id/eprint/39929/
_version_ 1643832566002745344
score 13.211869

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