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Suppression of MAPK Signaling and Reversal of mTOR-Dependent MDR1-Associated Multidrug Resistance by 21α-Methylmelianodiol in Lung Cancer Cells.

Aldonza MB, Hong JY, Bae SY, Song J, Kim WK, Oh J, Shin Y, Lee SH, Lee SK - PLoS ONE (2015)

Bottom Line: Interplay between PI3K/AMPK/AKT and MAPK pathways is a crucial effector in lung cancer growth and progression.Here, we described whether 21α-Methylmelianodiol (21α-MMD), an active triterpenoid derivative of Poncirus trifoliate, can display anticancer properties by regulating these signals and modulate the occurrence of multidrug resistance in NSCLC cells.Employing the established paclitaxel-resistant A549 cells (A549-PacR), we further found that 21α-MMD induced a MDR reversal activity through the inhibition of P-gp/MDR1 expressions, function, and transcription with regained paclitaxel sensitivity which might dependently correlate to the regulation of PI3K/mTOR signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: College of Pharmacy, Seoul National University, Seoul, Korea.

ABSTRACT
Lung cancer is the leading cause of cancer-related deaths worldwide and remains the most prevalent. Interplay between PI3K/AMPK/AKT and MAPK pathways is a crucial effector in lung cancer growth and progression. These signals transduction protein kinases serve as good therapeutic targets for non-small cell lung cancer (NSCLC) which comprises up to 90% of lung cancers. Here, we described whether 21α-Methylmelianodiol (21α-MMD), an active triterpenoid derivative of Poncirus trifoliate, can display anticancer properties by regulating these signals and modulate the occurrence of multidrug resistance in NSCLC cells. We found that 21α-MMD inhibited the growth and colony formation of lung cancer cells without affecting the normal lung cell phenotype. 21α-MMD also abrogated the metastatic activity of lung cancer cells through the inhibition of cell migration and invasion, and induced G0/G1 cell cycle arrest with increased intracellular ROS generation and loss of mitochondrial membrane integrity. 21α-MMD regulated the expressions of PI3K/AKT/AMPK and MAPK signaling which drove us to further evaluate its activity on multidrug resistance (MDR) in lung cancer cells by specifying on P-glycoprotein (P-gp)/MDR1-association. Employing the established paclitaxel-resistant A549 cells (A549-PacR), we further found that 21α-MMD induced a MDR reversal activity through the inhibition of P-gp/MDR1 expressions, function, and transcription with regained paclitaxel sensitivity which might dependently correlate to the regulation of PI3K/mTOR signaling pathway. Taken together, these findings demonstrate, for the first time, the mechanistic evaluation in vitro of 21α-MMD displaying growth-inhibiting potential with influence on MDR reversal in human lung cancer cells.

No MeSH data available.


Related in: MedlinePlus

21α-MMD and its mechanistic potential against lung cancer cell growth, migration, and invasion in vitro.(A) The chemical structure of 21α-Methylmelianodiol (21α-MMD), a natural triterpenoid isolated from the fruit of Poncirus trifoliata. (B) MRC-5 and L132 human normal lung cell lines and A549, H460, H358, H1299, and H292 human lung cancer cell lines were plated on 96-well plate and were treated with varying concentrations of 21α-MMD for 24 h and cell growth was analyzed by MTT assay and plotted as percentage of viable cells. Values are compared to the corresponding control value. (C) Clonal formation growth of indicated lung cancer cells was conducted after a 7-day growth period after a single administration of various concentrations of 21α-MMD. Images on the left displayed are crystal violet stained colonies while on the right are graphs representing count measurements of the colonies. (D and E) Phase-contrast microscopy was conducted on cells after exposure to 25 µM 21α-MMD to identify changes in the morphology and the DNA was observed by DAPI (lower left) and by PI (right) staining observed by confocal microscope. (F) H1299 and A549 cells were incubated with 5 μM 21α-MMD for 24 h followed by cell invasion analysis. Matrigel was diluted with serum-free culture medium and applied on the insert in the upper chambers of the multiwell and the cells were incubated to invade. Invaded cells were stained with crystal violet for apparent detection with a phase-contrast microscope. (G) Migration of cells was analyzed with the same method as in F but except without matrigel inclusion. H1299 and A549 cells were incubated with 6 μM 21α-MMD for 24 h. Columns indicate mean ± SD. (*p<0.05; **p<0.01)
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pone.0127841.g001: 21α-MMD and its mechanistic potential against lung cancer cell growth, migration, and invasion in vitro.(A) The chemical structure of 21α-Methylmelianodiol (21α-MMD), a natural triterpenoid isolated from the fruit of Poncirus trifoliata. (B) MRC-5 and L132 human normal lung cell lines and A549, H460, H358, H1299, and H292 human lung cancer cell lines were plated on 96-well plate and were treated with varying concentrations of 21α-MMD for 24 h and cell growth was analyzed by MTT assay and plotted as percentage of viable cells. Values are compared to the corresponding control value. (C) Clonal formation growth of indicated lung cancer cells was conducted after a 7-day growth period after a single administration of various concentrations of 21α-MMD. Images on the left displayed are crystal violet stained colonies while on the right are graphs representing count measurements of the colonies. (D and E) Phase-contrast microscopy was conducted on cells after exposure to 25 µM 21α-MMD to identify changes in the morphology and the DNA was observed by DAPI (lower left) and by PI (right) staining observed by confocal microscope. (F) H1299 and A549 cells were incubated with 5 μM 21α-MMD for 24 h followed by cell invasion analysis. Matrigel was diluted with serum-free culture medium and applied on the insert in the upper chambers of the multiwell and the cells were incubated to invade. Invaded cells were stained with crystal violet for apparent detection with a phase-contrast microscope. (G) Migration of cells was analyzed with the same method as in F but except without matrigel inclusion. H1299 and A549 cells were incubated with 6 μM 21α-MMD for 24 h. Columns indicate mean ± SD. (*p<0.05; **p<0.01)

Mentions: Coumarins and triterpenoids from the fruits of P. trifoliata have been previously described to exhibit potent anticancer effects against a variety of cancer cell models [27–29]. Based on this information, the in vitro cytotoxic activities of 13 compounds isolated from the fruits of P. trifoliata against MDA-MB-231, T47D, SNU-638, SK-HEP-1, and A549 human cancer cell lines were evaluated using SRB assay. As shown in Table 1, triterpenoids showed the most promise by inhibiting a panel of cancer cells with IC50 values in less than 50 micro-molar ranges. Among the test compounds, triterpenoids 25-methoxyhispidol A, 21α-methylmelianodiol (21α-MMD, Fig 1A), and 21α, 25-dimethylmelianodiol exhibited the most active anti-proliferative activity. The data suggested that the anti-proliferative activity of 21α-MMD is potent when tested on A549 human lung cancer cells, which directed us to further study its mechanism of action in a larger panel of NSCLC cell models.


Suppression of MAPK Signaling and Reversal of mTOR-Dependent MDR1-Associated Multidrug Resistance by 21α-Methylmelianodiol in Lung Cancer Cells.

Aldonza MB, Hong JY, Bae SY, Song J, Kim WK, Oh J, Shin Y, Lee SH, Lee SK - PLoS ONE (2015)

21α-MMD and its mechanistic potential against lung cancer cell growth, migration, and invasion in vitro.(A) The chemical structure of 21α-Methylmelianodiol (21α-MMD), a natural triterpenoid isolated from the fruit of Poncirus trifoliata. (B) MRC-5 and L132 human normal lung cell lines and A549, H460, H358, H1299, and H292 human lung cancer cell lines were plated on 96-well plate and were treated with varying concentrations of 21α-MMD for 24 h and cell growth was analyzed by MTT assay and plotted as percentage of viable cells. Values are compared to the corresponding control value. (C) Clonal formation growth of indicated lung cancer cells was conducted after a 7-day growth period after a single administration of various concentrations of 21α-MMD. Images on the left displayed are crystal violet stained colonies while on the right are graphs representing count measurements of the colonies. (D and E) Phase-contrast microscopy was conducted on cells after exposure to 25 µM 21α-MMD to identify changes in the morphology and the DNA was observed by DAPI (lower left) and by PI (right) staining observed by confocal microscope. (F) H1299 and A549 cells were incubated with 5 μM 21α-MMD for 24 h followed by cell invasion analysis. Matrigel was diluted with serum-free culture medium and applied on the insert in the upper chambers of the multiwell and the cells were incubated to invade. Invaded cells were stained with crystal violet for apparent detection with a phase-contrast microscope. (G) Migration of cells was analyzed with the same method as in F but except without matrigel inclusion. H1299 and A549 cells were incubated with 6 μM 21α-MMD for 24 h. Columns indicate mean ± SD. (*p<0.05; **p<0.01)
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4476707&req=5

pone.0127841.g001: 21α-MMD and its mechanistic potential against lung cancer cell growth, migration, and invasion in vitro.(A) The chemical structure of 21α-Methylmelianodiol (21α-MMD), a natural triterpenoid isolated from the fruit of Poncirus trifoliata. (B) MRC-5 and L132 human normal lung cell lines and A549, H460, H358, H1299, and H292 human lung cancer cell lines were plated on 96-well plate and were treated with varying concentrations of 21α-MMD for 24 h and cell growth was analyzed by MTT assay and plotted as percentage of viable cells. Values are compared to the corresponding control value. (C) Clonal formation growth of indicated lung cancer cells was conducted after a 7-day growth period after a single administration of various concentrations of 21α-MMD. Images on the left displayed are crystal violet stained colonies while on the right are graphs representing count measurements of the colonies. (D and E) Phase-contrast microscopy was conducted on cells after exposure to 25 µM 21α-MMD to identify changes in the morphology and the DNA was observed by DAPI (lower left) and by PI (right) staining observed by confocal microscope. (F) H1299 and A549 cells were incubated with 5 μM 21α-MMD for 24 h followed by cell invasion analysis. Matrigel was diluted with serum-free culture medium and applied on the insert in the upper chambers of the multiwell and the cells were incubated to invade. Invaded cells were stained with crystal violet for apparent detection with a phase-contrast microscope. (G) Migration of cells was analyzed with the same method as in F but except without matrigel inclusion. H1299 and A549 cells were incubated with 6 μM 21α-MMD for 24 h. Columns indicate mean ± SD. (*p<0.05; **p<0.01)
Mentions: Coumarins and triterpenoids from the fruits of P. trifoliata have been previously described to exhibit potent anticancer effects against a variety of cancer cell models [27–29]. Based on this information, the in vitro cytotoxic activities of 13 compounds isolated from the fruits of P. trifoliata against MDA-MB-231, T47D, SNU-638, SK-HEP-1, and A549 human cancer cell lines were evaluated using SRB assay. As shown in Table 1, triterpenoids showed the most promise by inhibiting a panel of cancer cells with IC50 values in less than 50 micro-molar ranges. Among the test compounds, triterpenoids 25-methoxyhispidol A, 21α-methylmelianodiol (21α-MMD, Fig 1A), and 21α, 25-dimethylmelianodiol exhibited the most active anti-proliferative activity. The data suggested that the anti-proliferative activity of 21α-MMD is potent when tested on A549 human lung cancer cells, which directed us to further study its mechanism of action in a larger panel of NSCLC cell models.

Bottom Line: Interplay between PI3K/AMPK/AKT and MAPK pathways is a crucial effector in lung cancer growth and progression.Here, we described whether 21α-Methylmelianodiol (21α-MMD), an active triterpenoid derivative of Poncirus trifoliate, can display anticancer properties by regulating these signals and modulate the occurrence of multidrug resistance in NSCLC cells.Employing the established paclitaxel-resistant A549 cells (A549-PacR), we further found that 21α-MMD induced a MDR reversal activity through the inhibition of P-gp/MDR1 expressions, function, and transcription with regained paclitaxel sensitivity which might dependently correlate to the regulation of PI3K/mTOR signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: College of Pharmacy, Seoul National University, Seoul, Korea.

ABSTRACT
Lung cancer is the leading cause of cancer-related deaths worldwide and remains the most prevalent. Interplay between PI3K/AMPK/AKT and MAPK pathways is a crucial effector in lung cancer growth and progression. These signals transduction protein kinases serve as good therapeutic targets for non-small cell lung cancer (NSCLC) which comprises up to 90% of lung cancers. Here, we described whether 21α-Methylmelianodiol (21α-MMD), an active triterpenoid derivative of Poncirus trifoliate, can display anticancer properties by regulating these signals and modulate the occurrence of multidrug resistance in NSCLC cells. We found that 21α-MMD inhibited the growth and colony formation of lung cancer cells without affecting the normal lung cell phenotype. 21α-MMD also abrogated the metastatic activity of lung cancer cells through the inhibition of cell migration and invasion, and induced G0/G1 cell cycle arrest with increased intracellular ROS generation and loss of mitochondrial membrane integrity. 21α-MMD regulated the expressions of PI3K/AKT/AMPK and MAPK signaling which drove us to further evaluate its activity on multidrug resistance (MDR) in lung cancer cells by specifying on P-glycoprotein (P-gp)/MDR1-association. Employing the established paclitaxel-resistant A549 cells (A549-PacR), we further found that 21α-MMD induced a MDR reversal activity through the inhibition of P-gp/MDR1 expressions, function, and transcription with regained paclitaxel sensitivity which might dependently correlate to the regulation of PI3K/mTOR signaling pathway. Taken together, these findings demonstrate, for the first time, the mechanistic evaluation in vitro of 21α-MMD displaying growth-inhibiting potential with influence on MDR reversal in human lung cancer cells.

No MeSH data available.


Related in: MedlinePlus