First-generation epidermal growth factor receptor (EGFR) targeted kinase inhibitors (TKIs) are still used in selected non-small cell lung cancer (NSCLC) patients despite the resistance. Based on the correlation of programmed cell death receptor ligand 1 (PD-L1) and EGFR signaling pathway, whether continuous TKIs treatment will affect PD-L1 expression after disease progression remains unclear. To investigate the potential change of PD-L1 expression in TKI-resistant NSCLC after continuous TKIs treatment, we treated H1975 and HCC827 for more than one month and explored the possible effect on immune cells as well as underlying biological mechanisms. We found that continuous exposure to TKIs induced upregulation of PD-L1 in H1975 and HCC827. Moreover, PD-L1 upregulation significantly inhibited proliferation and slightly promoted apoptosis of T cells. We observed the activation of STAT3 and ERK1/2 along with the PD-L1 upregulation. With the pathway inhibitors, we found ERK1/2 pathway involved in inducing PD-L1 in resistant lung cancer. This study provides preclinical evidence that continuous TKIs treatment may induce PD-L1 expression in resistant NSCLC, resulting in the suppression of T cell function and immune escape. ERK1/2 pathway inhibitors, PD-L1/PD-1 inhibitors or combination strategies should be considered to reverse the resistance to TKIs in NSCLC patients.