Limits...
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.

Show MeSH

Related in: MedlinePlus

In vivo antitumor activity of hinokitiol.(A) The growth curves of subcutaneous xenografts of H1975 are shown. (B) The excised tumors were weighed and imaged. All results are given as the mean ± SD; n = 5 - 7 for each group. *indicates a significant difference at the level of p<0.05 compared with the control group. (C) Hematoxylin and eosin-stained tumor sections at days 14 or 21 from each group were analyzed. Arrow heads indicate the atypical nuclei or abnormal mitosis. Immunohistochemically stained tumor sections at days 14 or 21 from each group were analyzed to assess γ-H2AX and LC3 expression (D). The atypical nuclei, abnormal mitosis, and positive cells were quantified at 400× magnification under a standard light microscope (Olympus BX51, Japan). Each value is the mean ± SD of 5–10 fields of triplicate tumor sections. *, ** and *** indicate a significant difference compared with its' own control at the level of p<0.05, p<0.01, and p<0.001, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4126702&req=5

pone-0104203-g007: In vivo antitumor activity of hinokitiol.(A) The growth curves of subcutaneous xenografts of H1975 are shown. (B) The excised tumors were weighed and imaged. All results are given as the mean ± SD; n = 5 - 7 for each group. *indicates a significant difference at the level of p<0.05 compared with the control group. (C) Hematoxylin and eosin-stained tumor sections at days 14 or 21 from each group were analyzed. Arrow heads indicate the atypical nuclei or abnormal mitosis. Immunohistochemically stained tumor sections at days 14 or 21 from each group were analyzed to assess γ-H2AX and LC3 expression (D). The atypical nuclei, abnormal mitosis, and positive cells were quantified at 400× magnification under a standard light microscope (Olympus BX51, Japan). Each value is the mean ± SD of 5–10 fields of triplicate tumor sections. *, ** and *** indicate a significant difference compared with its' own control at the level of p<0.05, p<0.01, and p<0.001, respectively.

Mentions: The in vivo antitumor activity of hinokitiol was evaluated using H1975 cell xenografts in NOD-SCID mice. The intra-peritoneal administration of hinokitiol at low (2 mg/kg/day) and high (10 mg/kg/day) doses for 21 days significantly reduced the tumor volume (47.58% and 47.59%, respectively; Fig. 7A) compared with the control group. The size and weight of the excised tumors showed that hinokitiol effectively inhibited tumor growth in vivo (Fig. 7B). The histological examination of the tumor sections revealed that hinokitiol was able to reduce abnormal mitosis compared with the control group at days 14 and 21 (Fig. 7C). To further investigate whether the tumor growth inhibition was related to DNA damage and autophagy, we confirmed the presence of γ-H2AX and LC3 expression in the tumor tissue using immunohistochemical staining. The histological analysis revealed that hinokitiol induced γ-H2AX and LC3 at days 14 and 21 (Fig. 7D) compared with the control group levels. These in vivo data suggest that hinokitiol reduced tumor growth, potentially through the attenuation of tumorigenicity, and induced DNA damage and autophagy to suppress tumor progression.


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)

In vivo antitumor activity of hinokitiol.(A) The growth curves of subcutaneous xenografts of H1975 are shown. (B) The excised tumors were weighed and imaged. All results are given as the mean ± SD; n = 5 - 7 for each group. *indicates a significant difference at the level of p<0.05 compared with the control group. (C) Hematoxylin and eosin-stained tumor sections at days 14 or 21 from each group were analyzed. Arrow heads indicate the atypical nuclei or abnormal mitosis. Immunohistochemically stained tumor sections at days 14 or 21 from each group were analyzed to assess γ-H2AX and LC3 expression (D). The atypical nuclei, abnormal mitosis, and positive cells were quantified at 400× magnification under a standard light microscope (Olympus BX51, Japan). Each value is the mean ± SD of 5–10 fields of triplicate tumor sections. *, ** and *** indicate a significant difference compared with its' own control at the level of p<0.05, p<0.01, and p<0.001, respectively.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0104203-g007: In vivo antitumor activity of hinokitiol.(A) The growth curves of subcutaneous xenografts of H1975 are shown. (B) The excised tumors were weighed and imaged. All results are given as the mean ± SD; n = 5 - 7 for each group. *indicates a significant difference at the level of p<0.05 compared with the control group. (C) Hematoxylin and eosin-stained tumor sections at days 14 or 21 from each group were analyzed. Arrow heads indicate the atypical nuclei or abnormal mitosis. Immunohistochemically stained tumor sections at days 14 or 21 from each group were analyzed to assess γ-H2AX and LC3 expression (D). The atypical nuclei, abnormal mitosis, and positive cells were quantified at 400× magnification under a standard light microscope (Olympus BX51, Japan). Each value is the mean ± SD of 5–10 fields of triplicate tumor sections. *, ** and *** indicate a significant difference compared with its' own control at the level of p<0.05, p<0.01, and p<0.001, respectively.
Mentions: The in vivo antitumor activity of hinokitiol was evaluated using H1975 cell xenografts in NOD-SCID mice. The intra-peritoneal administration of hinokitiol at low (2 mg/kg/day) and high (10 mg/kg/day) doses for 21 days significantly reduced the tumor volume (47.58% and 47.59%, respectively; Fig. 7A) compared with the control group. The size and weight of the excised tumors showed that hinokitiol effectively inhibited tumor growth in vivo (Fig. 7B). The histological examination of the tumor sections revealed that hinokitiol was able to reduce abnormal mitosis compared with the control group at days 14 and 21 (Fig. 7C). To further investigate whether the tumor growth inhibition was related to DNA damage and autophagy, we confirmed the presence of γ-H2AX and LC3 expression in the tumor tissue using immunohistochemical staining. The histological analysis revealed that hinokitiol induced γ-H2AX and LC3 at days 14 and 21 (Fig. 7D) compared with the control group levels. These in vivo data suggest that hinokitiol reduced tumor growth, potentially through the attenuation of tumorigenicity, and induced DNA damage and autophagy to suppress tumor progression.

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