Publication: GLOBAL GENE EXPRESSION PROFILING OF MDA-MB-468 HUMAN BREAST CANCER CELLS FOLLOWING TREATMENT WITH GAMMA-TOCOTRIENOL: FUNCTIONAL ENRICHMENT AND GENE NETWORK-BASED ANALYSIS
Date
2023-07
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Publisher
International Medical University
Abstract
Breast cancer is the leading cause of death among women in Malaysia and worldwide. The overall percentage of breast cancer patients in Malaysia is 31% compared to other types of cancer. In Malaysia, the number of new cases of female breast cancer in 2020 was reported to be 11.7%. Recently, treatment for breast cancer has changed to a new perspective and researchers are finding evidence to prove that natural bioactive compounds can also exhibit anti-cancer activity with increased health benefits and reduced or no side effects compared to other chemoprevention drugs. One such compound known for its anti-cancer potential is tocotrienol, a member of the vitamin E family. Among the isomers of tocotrienols, gamma-tocotrienol (γ-T3) is known to mediate the intracellular signalling of cancer cells via various mechanisms such as promoting apoptosis, anti-angiogenesis, antimetastatic and other important cellular mechanisms. In this study, the triple negative MDA-MB-468 human breast cancer cells were treated with γ-T3 at its IC50 concentration established at 72 hours and the total RNA was isolated using RNeasy Mini columns (Qiagen, Valencia, CA, USA). Samples that yielded 7.4 to 8.4 for RNA integrity number, exhibited a clear gel image and for which no DNA contamination was observed on the gel were used for microarray gene expression profiling. Transcriptome Analysis Console (TAC) with expression console were further used for the identification of differential expression genes and generation of interactive data visualisations. DAVID database analysis was used to perform enrichment analysis for the differentially expressed genes in response to γ-T3. Differentially regulated genes identified through STRING and KEGG analysis showed the involvement of γ-T3 in regulating the cell cycle pathways. The MDA-MB-468 human breast cancer cells were found to have a significant response to γ-Breast cancer is the leading cause of death among women in Malaysia and worldwide. The overall percentage of breast cancer patients in Malaysia is 31% compared to other types of cancer. In Malaysia, the number of new cases of female breast cancer in 2020 was reported to be 11.7%. Recently, treatment for breast cancer has changed to a new perspective and researchers are finding evidence to prove that natural bioactive compounds can also exhibit anti-cancer activity with increased health benefits and reduced or no side effects compared to other chemoprevention drugs. One such compound known for its anti-cancer potential is tocotrienol, a member of the vitamin E family. Among the isomers of tocotrienols, gamma-tocotrienol (γ-T3) is known to mediate the intracellular signalling of cancer cells via various mechanisms such as promoting apoptosis, anti-angiogenesis, antimetastatic and other important cellular mechanisms. In this study, the triple negative MDA-MB-468 human breast cancer cells were treated with γ-T3 at its IC50 concentration established at 72 hours and the total RNA was isolated using RNeasy Mini columns (Qiagen, Valencia, CA, USA). Samples that yielded 7.4 to 8.4 for RNA integrity number, exhibited a clear gel image and for which no DNA contamination was observed on the gel were used for microarray gene expression profiling. Transcriptome Analysis Console (TAC) with expression console were further used for the identification of differential expression genes and generation of interactive data visualisations. DAVID database analysis was used to perform enrichment analysis for the differentially expressed genes in response to γ-T3. Differentially regulated genes identified through STRING and KEGG analysis showed the involvement of γ-T3 in regulating the cell cycle pathways. The MDA-MB-468 human breast cancer cells were found to have a significant response to γ-Breast cancer is the leading cause of death among women in Malaysia and worldwide. The overall percentage of breast cancer patients in Malaysia is 31% compared to other types of cancer. In Malaysia, the number of new cases of female breast cancer in 2020 was reported to be 11.7%. Recently, treatment for breast cancer has changed to a new perspective and researchers are finding evidence to prove that natural bioactive compounds can also exhibit anti-cancer activity with increased health benefits and reduced or no side effects compared to other chemoprevention drugs. One such compound known for its anti-cancer potential is tocotrienol, a member of the vitamin E family. Among the isomers of tocotrienols, gamma-tocotrienol (γ-T3) is known to mediate the intracellular signalling of cancer cells via various mechanisms such as promoting apoptosis, anti-angiogenesis, antimetastatic and other important cellular mechanisms. In this study, the triple negative MDA-MB-468 human breast cancer cells were treated with γ-T3 at its IC50 concentration established at 72 hours and the total RNA was isolated using RNeasy Mini columns (Qiagen, Valencia, CA, USA). Samples that yielded 7.4 to 8.4 for RNA integrity number, exhibited a clear gel image and for which no DNA contamination was observed on the gel were used for microarray gene expression profiling. Transcriptome Analysis Console (TAC) with expression console were further used for the identification of differential expression genes and generation of interactive data visualisations. DAVID database analysis was used to perform enrichment analysis for the differentially expressed genes in response to γ-T3. Differentially regulated genes identified through STRING and KEGG analysis showed the involvement of γ-T3 in regulating the cell cycle pathways. The MDA-MB-468 human breast cancer cells were found to have a significant response to γ-T3 treatment as the observation under phase contrast microscopy showed inhibition of cell proliferation. A total of 1091 genes were identified in MDA-MB-468 cells, of which 84 genes were found to be associated with breast cancer. Out of these 84 genes, 50% of it were downregulated after the treatment with γ-T3. From these subsets of genes, a total of 10 genes such as PLK1, CDKN1A, CDK1, PTTG1, MAD2L1, SKP2, CDC20, TTK, MYC, and ATM were identified to be involved in cell proliferation inhibition and cell cycle arrest. Furthermore, some of the histone mediated genes such as HIST1H2BI, HIST1H2BO, and HIST1H3F were also found to be downregulated with γ-T3 treatment which further strongly indicate inhibition of breast cancer cell proliferation. Overall, the results from this study suggest γ-T3 s' ability to modulate the anti-cancer mechanisms in MDA-MB-468 human breast cancer cells and have therapeutic potential.
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Keywords
Breast Neoplasms, Women, Tocotrienols, Vitamin E