Piperine Causes G1 Phase Cell Cycle Arrest and Apoptosis in Melanoma Cells through Checkpoint Kinase-1 Activation

Abstract

In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR) and checkpoint kinase 1 (Chk1). Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb). Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length) and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis.

Nanoemulsion formulations for anti-cancer agent piplartine—Characterization, toxicological, pharmacokinetics and efficacy studies

“…Cytotoxicity assays were performed by sulforhodamine B (SRB) assay as previously described by us (Fofaria et al, 2014). A375 or B16-F0 cells were plated in a 96-well plate at a density of 5000 cells per well and were allowed to attach overnight.…”

“…Morphological changes due to chronic treatment with nanoemulsions were evaluated by hematoxylin and eosin staining according to the procedure described previous (Fofaria and Srivastava, 2014). …”

Quantitative Structure–Cytotoxicity Relationship of Newly Synthesized Piperic Acid Esters

“…Among eleven piperic acid ester derivatives, 2-(3,4-dihydroxyphenyl)ethyl ester [4] showed the highest cytotoxicity against OSCC cell lines (mean CC 50 =17 μM), followed by (4-methoxyphenyl)methyl ester (38 μM) [1], 2-(4-methoxyphenyl)ethyl ester [5] (42 μM), 2-(4-hydroxyphenyl)ethyl ester (46 μM) [3], 2-phenylethyl ester [7] (61 μM) and 2-(3,4-dimethoxyphenyl)ethyl ester [6] (102 μM). On the other hand, (4-hydroxy-3-methoxyphenyl)methyl ester [2] (>368 μM), 3-phenylpropyl ester [8], 4-phenyl-2-butyl ester [9], 4-phenylbutyl ester [10] and decyl ester [11] (>400 μM) were essentially inactive (Table I).…”

“…Culture plastic dishes and plates (96-well) were purchased from Becton Dickinson (Franklin Lakes, NJ, USA). [6], (2E,4E)-5-(3,4-methylenedioxyphenyl)-2,4-pentadienoic acid 2-phenylethyl ester [7], (2E,4E)-5-(3,4-methylenedioxyphenyl)-2,4-pentadienoic acid 3-phenylpropyl ester [8], (2E,4E)-5-(3,4-methylenedioxyphenyl)-2,4-pentadienoic acid 4-phenyl-2-butyl ester [9], (2E,4E)-5-(3,4-methylenedioxyphenyl)-2,4-pentadienoic acid 4-phenylbutyl ester [10] and (2E,4E)-5-(3,4-methylenedioxyphenyl)-2,4-pentadienoic acid decyl ester [11] (structures shown in Figure 1) were synthesized by coupling of piperic acid with the appropriate alcohols by means of a modified procedure described elsewhere (18). To a mixture of piperic acid (1.1 mmol) and the appropriate alcohol (1.0 mmol) in dry tetrahydrofuran (4 ml) were added triphenylphosphine (1.5 mmol) and diisopropyl azodicarboxylate (1.5 mmol).…”

“…Piperine induced apoptotic cell death in human colon cells (9), melanoma cells (10), and lung (11) and rectal carcinoma cell lines (12), whereas it induced autophagy in human prostate cancer cells (13). On the other hand, piperine protected normal cells from apoptosis induced by cadmium (14), cisplatin (15) and glutamate (16).…”