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Hinokitiol induces DNA damage and autophagy followed by cell cycle arrest and senescence in gefitinib-resistant lung adenocarcinoma cells.

Li LH, Wu P, Lee JY, Li PR, Hsieh WY, Ho CC, Ho CL, Chen WJ, Wang CC, Yen MY, Yang SM, Chen HW - PLoS ONE (2014)

Bottom Line: Here, we found that hinokitiol, a natural monoterpenoid from the heartwood of Calocedrus formosana, exhibited potent anticancer effects.Furthermore, hinokitiol inhibited the growth of xenograft tumors in association with DNA damage and autophagy but exhibited fewer effects on lung stromal fibroblasts.In summary, we demonstrated novel mechanisms by which hinokitiol, an essential oil extract, acted as a promising anticancer agent to overcome EGFR-TKI resistance in lung cancer cells via inducing DNA damage, autophagy, cell cycle arrest, and senescence in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Toxicology, College of Medicine, National Taiwan University, Taipei, Taiwan; Department of Laboratory, Kunming Branch, Taipei City Hospital, Taipei, Taiwan.

ABSTRACT
Despite good initial responses, drug resistance and disease recurrence remain major issues for lung adenocarcinoma patients with epidermal growth factor receptor (EGFR) mutations taking EGFR-tyrosine kinase inhibitors (TKI). To discover new strategies to overcome this issue, we investigated 40 essential oils from plants indigenous to Taiwan as alternative treatments for a wide range of illnesses. Here, we found that hinokitiol, a natural monoterpenoid from the heartwood of Calocedrus formosana, exhibited potent anticancer effects. In this study, we demonstrated that hinokitiol inhibited the proliferation and colony formation ability of lung adenocarcinoma cells as well as the EGFR-TKI-resistant lines PC9-IR and H1975. Transcriptomic analysis and pathway prediction algorithms indicated that the main implicated pathways included DNA damage, autophagy, and cell cycle. Further investigations confirmed that in lung cancer cells, hinokitiol inhibited cell proliferation by inducing the p53-independent DNA damage response, autophagy (not apoptosis), S-phase cell cycle arrest, and senescence. Furthermore, hinokitiol inhibited the growth of xenograft tumors in association with DNA damage and autophagy but exhibited fewer effects on lung stromal fibroblasts. In summary, we demonstrated novel mechanisms by which hinokitiol, an essential oil extract, acted as a promising anticancer agent to overcome EGFR-TKI resistance in lung cancer cells via inducing DNA damage, autophagy, cell cycle arrest, and senescence in vitro and in vivo.

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The effects of hinokitiol on gene expression.(A) Microarray profiling of H1975 cells and PC9-IR cells treated with 5 µM hinokitiol for 48 h. (B) Q-PCR array validation of the expression of genes related to DNA damage and autophagy in H1975 cells and lung stromal fibroblasts after 5 µM hinokitiol treatment for 24 h. The results are representative of those obtained in three different experiments and are expressed as the fold change compared with control. * and ** indicate a significant difference at the level of p<0.05 and p<0.01, respectively.
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pone-0104203-g002: The effects of hinokitiol on gene expression.(A) Microarray profiling of H1975 cells and PC9-IR cells treated with 5 µM hinokitiol for 48 h. (B) Q-PCR array validation of the expression of genes related to DNA damage and autophagy in H1975 cells and lung stromal fibroblasts after 5 µM hinokitiol treatment for 24 h. The results are representative of those obtained in three different experiments and are expressed as the fold change compared with control. * and ** indicate a significant difference at the level of p<0.05 and p<0.01, respectively.

Mentions: To study the potential mechanisms of hinokitiol on gefitinib-resistant lung adenocarcinoma cells, we compared the gene expression profiles of H1975 and PC9-IR cells with or without hinokitiol using the Affymetrix human GeneChip. Here, we found that 383 genes were up-regulated over 3 fold, and 787 genes were down-regulated over 3 fold in both cell lines after 5 µM hinokitiol treatment for 48 h (Fig. 2A). The CRSD2 web server, Gene Ontology and Pathway Enrichment analysis, predicted that hinokitiol could affect certain key regulators/factors involved in DNA damage, autophagy, and cell cycle signaling in both cell lines. Furthermore, we examined DNA damage- and autophagy-related genes in H1975 cells and human lung stromal fibroblasts upon hinokitiol treatment using Q-PCR array (SuperArray Bioscience). We confirmed that two autophagy-related genes, ATG4B and DAPK1A, were up-regulated by hinokitiol treatment in H1975 cells but were down-regulated in stromal fibroblasts (>1.5-fold change; Fig. 2B). In addition, three DNA damage-related genes, ERCC1, XPC, and CRY1, were up-regulated in hinokitiol-treated H1975 cells but down-regulated in stromal fibroblasts (>1.5-fold change). These results indicated that hinokitiol induced the expression of certain DNA damage- and autophagy-related genes in cancer cells but not in human stromal fibroblasts.


Hinokitiol induces DNA damage and autophagy followed by cell cycle arrest and senescence in gefitinib-resistant lung adenocarcinoma cells.

Li LH, Wu P, Lee JY, Li PR, Hsieh WY, Ho CC, Ho CL, Chen WJ, Wang CC, Yen MY, Yang SM, Chen HW - PLoS ONE (2014)

The effects of hinokitiol on gene expression.(A) Microarray profiling of H1975 cells and PC9-IR cells treated with 5 µM hinokitiol for 48 h. (B) Q-PCR array validation of the expression of genes related to DNA damage and autophagy in H1975 cells and lung stromal fibroblasts after 5 µM hinokitiol treatment for 24 h. The results are representative of those obtained in three different experiments and are expressed as the fold change compared with control. * and ** indicate a significant difference at the level of p<0.05 and p<0.01, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126702&req=5

pone-0104203-g002: The effects of hinokitiol on gene expression.(A) Microarray profiling of H1975 cells and PC9-IR cells treated with 5 µM hinokitiol for 48 h. (B) Q-PCR array validation of the expression of genes related to DNA damage and autophagy in H1975 cells and lung stromal fibroblasts after 5 µM hinokitiol treatment for 24 h. The results are representative of those obtained in three different experiments and are expressed as the fold change compared with control. * and ** indicate a significant difference at the level of p<0.05 and p<0.01, respectively.
Mentions: To study the potential mechanisms of hinokitiol on gefitinib-resistant lung adenocarcinoma cells, we compared the gene expression profiles of H1975 and PC9-IR cells with or without hinokitiol using the Affymetrix human GeneChip. Here, we found that 383 genes were up-regulated over 3 fold, and 787 genes were down-regulated over 3 fold in both cell lines after 5 µM hinokitiol treatment for 48 h (Fig. 2A). The CRSD2 web server, Gene Ontology and Pathway Enrichment analysis, predicted that hinokitiol could affect certain key regulators/factors involved in DNA damage, autophagy, and cell cycle signaling in both cell lines. Furthermore, we examined DNA damage- and autophagy-related genes in H1975 cells and human lung stromal fibroblasts upon hinokitiol treatment using Q-PCR array (SuperArray Bioscience). We confirmed that two autophagy-related genes, ATG4B and DAPK1A, were up-regulated by hinokitiol treatment in H1975 cells but were down-regulated in stromal fibroblasts (>1.5-fold change; Fig. 2B). In addition, three DNA damage-related genes, ERCC1, XPC, and CRY1, were up-regulated in hinokitiol-treated H1975 cells but down-regulated in stromal fibroblasts (>1.5-fold change). These results indicated that hinokitiol induced the expression of certain DNA damage- and autophagy-related genes in cancer cells but not in human stromal fibroblasts.

Bottom Line: Here, we found that hinokitiol, a natural monoterpenoid from the heartwood of Calocedrus formosana, exhibited potent anticancer effects.Furthermore, hinokitiol inhibited the growth of xenograft tumors in association with DNA damage and autophagy but exhibited fewer effects on lung stromal fibroblasts.In summary, we demonstrated novel mechanisms by which hinokitiol, an essential oil extract, acted as a promising anticancer agent to overcome EGFR-TKI resistance in lung cancer cells via inducing DNA damage, autophagy, cell cycle arrest, and senescence in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Toxicology, College of Medicine, National Taiwan University, Taipei, Taiwan; Department of Laboratory, Kunming Branch, Taipei City Hospital, Taipei, Taiwan.

ABSTRACT
Despite good initial responses, drug resistance and disease recurrence remain major issues for lung adenocarcinoma patients with epidermal growth factor receptor (EGFR) mutations taking EGFR-tyrosine kinase inhibitors (TKI). To discover new strategies to overcome this issue, we investigated 40 essential oils from plants indigenous to Taiwan as alternative treatments for a wide range of illnesses. Here, we found that hinokitiol, a natural monoterpenoid from the heartwood of Calocedrus formosana, exhibited potent anticancer effects. In this study, we demonstrated that hinokitiol inhibited the proliferation and colony formation ability of lung adenocarcinoma cells as well as the EGFR-TKI-resistant lines PC9-IR and H1975. Transcriptomic analysis and pathway prediction algorithms indicated that the main implicated pathways included DNA damage, autophagy, and cell cycle. Further investigations confirmed that in lung cancer cells, hinokitiol inhibited cell proliferation by inducing the p53-independent DNA damage response, autophagy (not apoptosis), S-phase cell cycle arrest, and senescence. Furthermore, hinokitiol inhibited the growth of xenograft tumors in association with DNA damage and autophagy but exhibited fewer effects on lung stromal fibroblasts. In summary, we demonstrated novel mechanisms by which hinokitiol, an essential oil extract, acted as a promising anticancer agent to overcome EGFR-TKI resistance in lung cancer cells via inducing DNA damage, autophagy, cell cycle arrest, and senescence in vitro and in vivo.

Show MeSH
Related in: MedlinePlus