Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent

Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent

Aberrations in the phospbatidylinositol3-kinase (PI3K)IAkt pathway have been found in a wide spectrum of human cancers. Activation of Akt and inactivation of the downstream substrates such as GSK-3P, BAD and Forkhead family (FKHR) proteins are relevant to promote cell survival, proliferation and...

Full description

Saved in:
Bibliographic Details
Main Author: Hassan, Masriana
Format: Thesis
Language:English
Published: 2007
Subjects:
Online Access:http://psasir.upm.edu.my/id/eprint/6438/1/FPSK%28M%29_2007_18.pdf
Tags: Add Tag
No Tags, Be the first to tag this record!
id my-upm-ir.6438
record_format uketd_dc
spelling my-upm-ir.64382023-10-30T04:13:33Z Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent 2007-02 Hassan, Masriana Aberrations in the phospbatidylinositol3-kinase (PI3K)IAkt pathway have been found in a wide spectrum of human cancers. Activation of Akt and inactivation of the downstream substrates such as GSK-3P, BAD and Forkhead family (FKHR) proteins are relevant to promote cell survival, proliferation and growth. Another related pathway linked with PI3K/Akt is the Ras/Raf/MEK/ERK, which is known to promote cancer as a result of ras-transformation. The discovery of new drugs targeted at specific molecules of these pathways is a 'hot' field in cancer research. Blocking the constitutively active PI3WAkt pathway provides a new strategy for cancer therapy. Thus, inhibitors of this signaling pathway would be potential anti-cancer agents. The Streptomyces strain H7372 isolated h m mangrove soils in Sabah was found to inhibit the Ras/Raf-1 protein interaction in the yeast two-hybrid screening system. The present study was undertaken to determine the cytotoxic effect of H7372 hctionated extract on a breast cancer cell line, MCF-7 and a non-tumorigenic epithelial cell line, MCF-1OA and quantitatively measure kinase inhibition, apoptosis induction and cell cycle disruption. The crude iii extracts of H7372 were fractionated into eight Eractions using reverse phase HPLC. Fraction 5 was found to be the most cytotoxic in an M'IT assay. The crude extract and fraction 5 of H7372 were found to exert growth inhibition of MCF-7 at ICS0 of 15&nl and 1.4 pglml, respectively. Western blot analyses showed that activated PI3K and Akt (Thr308) but not Akt (Ser473) by stimulation of IGF-I were inhibited by the crude extract and fraction 5 after 72 hours. Interestingly, phosphorylation of Raf-1 (Ser259) and ERKl were also inhibited by fraction 5, indicated that there is a cross-talk between PI3WAkl and MAPK pathways. By using the flow cytometry technique, we found that fraction 5 inhibited the proliferation of MCF-7 cell line by causing them to arrest in the GI phase of the cell cycle. The induction of growth arrest by fraction 5 was associated with accumulation of cells in GI and decreasing cells in S and G2/M phases. The results were supported by inihibition of cyclin Dl in MCF-7 cells. The apoptosis study showed that fraction 5 but not crude extract was increased the percentage of cells in early apoptotic stage at all concentrations. Furthermore, treatment of MCF-7 cells with fraction 5 resulted in reduction in phosphorylation of GSK-3$ (Ser9), phospho-BAD (Serll2) and phospho-FKHR (Ser256). These results could contribute the apoptosis in MCF-7 cell line. Thus, we discovered that k t i o n 5 of H7372, a naturally occurring microbe, contains an inhibitor of cell proliferation, cell cycle progression and is able to induce apoptosis via the PI3WAkl and MAPK pathways. 2007-02 Thesis http://psasir.upm.edu.my/id/eprint/6438/ http://psasir.upm.edu.my/id/eprint/6438/1/FPSK%28M%29_2007_18.pdf text en public masters Universiti Putra Malaysia Faculty Medicine and Health Sciences Seow, Heng Fong
institution Universiti Putra Malaysia
collection PSAS Institutional Repository
language English
advisor Seow, Heng Fong
topic


spellingShingle


Hassan, Masriana
Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent
description Aberrations in the phospbatidylinositol3-kinase (PI3K)IAkt pathway have been found in a wide spectrum of human cancers. Activation of Akt and inactivation of the downstream substrates such as GSK-3P, BAD and Forkhead family (FKHR) proteins are relevant to promote cell survival, proliferation and growth. Another related pathway linked with PI3K/Akt is the Ras/Raf/MEK/ERK, which is known to promote cancer as a result of ras-transformation. The discovery of new drugs targeted at specific molecules of these pathways is a 'hot' field in cancer research. Blocking the constitutively active PI3WAkt pathway provides a new strategy for cancer therapy. Thus, inhibitors of this signaling pathway would be potential anti-cancer agents. The Streptomyces strain H7372 isolated h m mangrove soils in Sabah was found to inhibit the Ras/Raf-1 protein interaction in the yeast two-hybrid screening system. The present study was undertaken to determine the cytotoxic effect of H7372 hctionated extract on a breast cancer cell line, MCF-7 and a non-tumorigenic epithelial cell line, MCF-1OA and quantitatively measure kinase inhibition, apoptosis induction and cell cycle disruption. The crude iii extracts of H7372 were fractionated into eight Eractions using reverse phase HPLC. Fraction 5 was found to be the most cytotoxic in an M'IT assay. The crude extract and fraction 5 of H7372 were found to exert growth inhibition of MCF-7 at ICS0 of 15&nl and 1.4 pglml, respectively. Western blot analyses showed that activated PI3K and Akt (Thr308) but not Akt (Ser473) by stimulation of IGF-I were inhibited by the crude extract and fraction 5 after 72 hours. Interestingly, phosphorylation of Raf-1 (Ser259) and ERKl were also inhibited by fraction 5, indicated that there is a cross-talk between PI3WAkl and MAPK pathways. By using the flow cytometry technique, we found that fraction 5 inhibited the proliferation of MCF-7 cell line by causing them to arrest in the GI phase of the cell cycle. The induction of growth arrest by fraction 5 was associated with accumulation of cells in GI and decreasing cells in S and G2/M phases. The results were supported by inihibition of cyclin Dl in MCF-7 cells. The apoptosis study showed that fraction 5 but not crude extract was increased the percentage of cells in early apoptotic stage at all concentrations. Furthermore, treatment of MCF-7 cells with fraction 5 resulted in reduction in phosphorylation of GSK-3$ (Ser9), phospho-BAD (Serll2) and phospho-FKHR (Ser256). These results could contribute the apoptosis in MCF-7 cell line. Thus, we discovered that k t i o n 5 of H7372, a naturally occurring microbe, contains an inhibitor of cell proliferation, cell cycle progression and is able to induce apoptosis via the PI3WAkl and MAPK pathways.
format Thesis
qualification_level Master's degree
author Hassan, Masriana
author_facet Hassan, Masriana
author_sort Hassan, Masriana
title Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent
title_short Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent
title_full Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent
title_fullStr Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent
title_full_unstemmed Identification of Protein Kinase Inhibitory Activities Fromstreptomyces Strain H7372 for Potential Use As Anti-Canceragent
title_sort identification of protein kinase inhibitory activities fromstreptomyces strain h7372 for potential use as anti-canceragent
granting_institution Universiti Putra Malaysia
granting_department Faculty Medicine and Health Sciences
publishDate 2007
url http://psasir.upm.edu.my/id/eprint/6438/1/FPSK%28M%29_2007_18.pdf
_version_ 1783725750438854656

Similar Items