Research Paper Volume 13, Issue 18 pp 21914—21940

Metformin-induced chemosensitization to cisplatin depends on P53 status and is inhibited by Jarid1b overexpression in non-small cell lung cancer cells

Treatment with sub-lethal dose of cisplatin leads to Jarid1b overexpression and chemoresistance in A549 cells. Sub-lethal treatment with cisplatin in A549 cells increases Jarid1b expression in a dose dependent manner and keeps overexpressed even after 30 days post-treatment (A). A549Res cells become resistant to the combined treatment between metformin and cisplatin as no improvement in DNA fragmentation (B), caspase 3 and 7 activation (C) can be seen. Only through cell viability assay showed some difference in the metformin and cisplatin combination (pD). Metformin decreased the number of colonies in A549Res cells (pE). Inhibition of Jarid1b by the pharmacological inhibitor PBIT restores the ability of metformin to chemosensitize to cisplatin as measured by DNA fragmentation assay (pF), but caspase 3 and 7 activation was not observed (G). However, inhibition of Jarid1b by siRNA (pH) restores the ability of metformin to chemosensitize to cisplatin through DNA fragmentation (pI) and caspase 3 and 7 activation assay (pJ). Jarid1b inhibition by siRNA, MTT assay, DNA fragmentation and caspase 3 and 7 activation assay data represent the mean of three independent experiments. A549 cells were pre-treated with 10μM of cisplatin for 72 h to generate A549Res cells. After pre-treatment, A549Res cells were treated with 10mM of metformin for 72 h and 25μM of cisplatin (with or without metformin) for another 72 h.

Figure 2. Treatment with sub-lethal dose of cisplatin leads to Jarid1b overexpression and chemoresistance in A549 cells. Sub-lethal treatment with cisplatin in A549 cells increases Jarid1b expression in a dose dependent manner and keeps overexpressed even after 30 days post-treatment (A). A549Res cells become resistant to the combined treatment between metformin and cisplatin as no improvement in DNA fragmentation (B), caspase 3 and 7 activation (C) can be seen. Only through cell viability assay showed some difference in the metformin and cisplatin combination (p<0.001) (D). Metformin decreased the number of colonies in A549Res cells (p<0.05), and no colonies was observed upon cisplatin treatment (E). Inhibition of Jarid1b by the pharmacological inhibitor PBIT restores the ability of metformin to chemosensitize to cisplatin as measured by DNA fragmentation assay (p<0.01) (F), but caspase 3 and 7 activation was not observed (G). However, inhibition of Jarid1b by siRNA (p<0.01) (H) restores the ability of metformin to chemosensitize to cisplatin through DNA fragmentation (p<0.001) (I) and caspase 3 and 7 activation assay (p<0.001) (J). Jarid1b inhibition by siRNA, MTT assay, DNA fragmentation and caspase 3 and 7 activation assay data represent the mean of three independent experiments. A549 cells were pre-treated with 10μM of cisplatin for 72 h to generate A549Res cells. After pre-treatment, A549Res cells were treated with 10mM of metformin for 72 h and 25μM of cisplatin (with or without metformin) for another 72 h.