Panoply™ Human JAK1 Over-expressing Stable Cell Line

AZD3759 is a novel epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) based on gefitinib, which has been shown to penetrate the central nervous system. Although clinical trials have demonstrated the good antitumor effect of AZD3759 in non-small cell lung cancer (NSCLC), its modulatory effect on the antitumor efficacy of radiotherapy (RA) remains unclear. Here, researchers compared the effects of AZD3759 and osimertinib on the efficacy of radiotherapy in NSCLC and explored the potential mechanisms of action of AZD3759. They found that at a radiation dose of 8 Gy, both 500 nM AZD3759 and osimertinib significantly reduced the survival rate of irradiated NSCLC cells. The combination of AZD3759 or osimertinib with radiotherapy significantly enhanced apoptosis, cell cycle arrest, and DNA damage in irradiated NSCLC cells, as well as brain metastasis in irradiated xenograft nude mice. Furthermore, the efficacy of AZD3759 was significantly stronger than that of osimertinib. Mechanistic studies showed that receptor tyrosine kinase signaling pathway antibody array analysis revealed that AZD3759 specifically inhibited Janus kinase-1 (JAK1), while osimertinib did not. Overexpression of JAK1 significantly attenuated the enhancing effect of AZD3759 on the efficacy of radiotherapy in PC-9 cells and the brain metastasis animal model. Therefore, these findings suggest that AZD3759 enhances the antitumor effect of radiotherapy in NSCLC by synergistically blocking both EGFR and JAK1.

To investigate the mechanistic basis for AZD3759's superior antitumor efficacy compared to osimertinib, researchers performed receptor tyrosine kinase (RTK) signaling pathway antibody array analysis. Compared to the radiotherapy (RA) group, both AZD3759 and osimertinib downregulated EGFR expression (Figure 1a). However, a decrease in JAK1 expression was only observed in the AZD3759 + RA group, but not in the osimertinib + RA group. The researchers hypothesized that the inhibition of JAK1 might be the reason why AZD3759 exhibits stronger antitumor properties than osimertinib. Therefore, they constructed JAK1-overexpressing PC-9 cells and confirmed the overexpression of JAK1 by Western blotting (Figure 1b). Colony formation assays (Figure 1c) showed that, compared to the control group, the survival rate of JAK1-overexpressing PC-9 cells was significantly increased at all concentrations of AZD3759 treatment. Compared to the RA group, AZD3759 combined treatment significantly increased the number of γ-H2AX foci per cell, while JAK1 overexpression significantly inhibited this increase (Figure 1d).

Figure 1. JAK1 overexpression abolishes effects of AZD3759 on clone formation and DNA damage in RA-treated PC-9 cells. (Zhao R, et al., 2022)