Induction of apoptosis through downregulation of anti-apoptotic genes bcl-2 and Nucleolin in MCF-7 cells by Cleome gynandra leaf fractions containing Tricin, Pinocembrin, and Acacetin

This study investigates the anti-proliferative effect of Cleome gynandra leaf fractions on MCF 7 cells. Chloroform extract of Cleome gynandra leaves (CCG) was subjected to column chromatographic separation with increasing polarity of solvents. The total fractions (36) collected were subjected to TLC analysis and pooled into three fractions (F1-F3) based on their R _f value. Further, these fractions were screened for anti-proliferative and apoptotic effects on MCF7 cells. The IC ₅₀ values for the CCG fractions (F1-F3) were 82.78, 93.69, and 200µg/mL, respectively. Of these fractions, F1 demonstrated a remarkable cytotoxic effect and was subsequently analyzed using LC-MS/MS. For the first time, the plant F1 is found to contain Tricin (a methylated flavone), Pinocembrin (a flavone), 4-hydroxy-2,3,4,6-tetramethoxychalcone (an aromatic ketone), Acacetin (a methylated flavone), and Cinnamic acid. The expression of bax, CYCS, bcl-2, and nucleolin was analyzed using RT-PCR. The MCF 7 cells treated with the F1 showed downregulation of anti-apoptotic genes bcl-2 and nucleolin and upregulation of bax and CYCS. Thus, the induction of apoptosis is mediated through the downregulation of the anti-apoptotic genes. Further, an in silico approach was employed to evaluate the binding affinity of the identified compounds with the Bax, nucleolin, and bcl-2 proteins. It was observed that Acacetin exhibited a stronger binding affinity with bax, Tricin, Nucleolin, and Pinocembrin with bcl-2. Based on this study, Tricin, Pinocembrin, Chalcones, and Cinnamic acid could target the apoptotic and anti-apoptotic genes studied.