Crizotinib induces apoptosis of lung cancer cells through JAK-STAT pathway

Abstract

Effect of crizotinib on apoptosis of lung cancer cells was investigated. Human non-small cell lung adenocarcinoma H2228 cells were cultured in the presence of 0, 20, 40, 80, 160 and 320 nmol/l of crizotinib for 3 days, respectively. The inhibition rate of cell proliferation was measured by MTT assay, and half maximal inhibitory concentration (IC₅₀) was calculated. Cell apoptosis was detected by flow cytometry. Transwell assay was performed to detect cell migration. Expression of Janus protein tyrosine kinase (JAK) and signal transducer and activator of transcription (STAT) proteins was detected by western blot analysis. Crizotinib significantly inhibited the proliferation of human lung cancer H2228 cells, and the inhibitory effect was enhanced with the increase of the concentration of crizotinib (p<0.01). The IC₅₀ value was 311.26 nnol/l. According to IC₅₀ value, concentration of crizotinib at 300 nmol/l was selected for the study. It was found that crizotinib at 300 nmol/l significantly promoted cell apoptosis (p<0.01) and inhibited cell migration (p<0.01). Compared with pretreatment levels, crizotinib downregulated the expression of JAK and STAT (p<0.01) on the 1st day of treatment, but with the prolongation of time, no further significant difference was observed on the 1st, 2nd or 3rd day in the level of JAK protein (p=0.47); there were no statistically significant differences in the level of STAT protein (p=0.91). Crizotinib can inhibit the migration and promote cell apoptosis of human lung cancer cell line H2228 by regulating the expression of JAK and STAT proteins in JAK-STAT signaling pathway.

Keywords: JAK-STAT signal; cell apoptosis; cell migration; cell proliferation; crizotinib; lung cancer.

Figure 1.

Inhibitory effect of crizotinib on cell proliferation of H2228 at different concentrations. Results of MTT showed that crizotinib significantly inhibited cell proliferation, and the inhibitory rate increased with the increase of crizotinib concentration (*p<0.01).

Figure 2.

Effects of crizotinib on H2228 cell apoptosis. Flow cytometry results showed that crizotinib significantly promoted cell apoptosis, and with the passage of time, apoptotic rate was significantly increased (*p<0.01).

Figure 3.

Effect of crizotinib on H2228 cell migration. Results of Transwell assay showed that crizotinib inhibited cell migration, and the inhibitory effect was enhanced with the prolongation of treatment (*p<0.01).

Figure 4.

Effects of crizotinib on expression of JAK protein. Western blot analysis results showed that, compared with the level at 0 h, crizotinib significantly inhibited JAK protein expression (*p<0.01).

Figure 5.

Effects of crizotinib on expression of STAT protein. Western blot analysis results showed that, compared with the level at 0 h, crizotinib significantly inhibited STAT protein expression (*p<0.01).