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. 2021 Jan;21(1):9.
doi: 10.3892/etm.2020.9441. Epub 2020 Nov 4.

PI3K inhibition sensitizes EGFR wild-type NSCLC cell lines to erlotinib chemotherapy

Xin Zhou  1 , Xiaowen Wang  2 , Hongge Zhu  1 , Guomin Gu  1 , Yiyi Zhan  1 , Chunling Liu  1 , Gang Sun  3
Affiliations

PI3K inhibition sensitizes EGFR wild-type NSCLC cell lines to erlotinib chemotherapy

Xin Zhou et al. Exp Ther Med. 2021 Jan.

Abstract

Tyrosine kinase inhibitors (TKIs) bring significant benefits for patients with cancers harboring epidermal growth factor receptor (EGFR) mutations. However, after treatment for a certain period, most patients ultimately acquire resistance. Numerous studies indicated that PI3K has an important role in tumor cell growth and drug sensitivity. Furthermore, inhibition of PI3K may lead to sensitization of non-small cell lung cancer (NSCLC) cells to EGFR-TKIs. The aim of the present study was to explore whether LY294002, an inhibitor of PI3K, is able to improve the sensitivity of NSCLC cell lines with wild-type EGFR to the EGFR-TKI erlotinib. An MTT assay was used to examine the effect of combined treatment with LY294002 and erlotinib on cell survival of two EGFR wild-type NSCLC cell lines, NCI-H661 and NCI-H460. Furthermore, flow cytometry was used to assess apoptosis in NCI-H661 and NCI-H460 cells after treatment with erlotinib and LY294002. In addition, the expression of downstream proteins was detected by western blot analysis. The results indicated that the number of viable NCI-H661 and NCI-H460 cells was dose-dependently reduced by erlotinib or LY294002. Compared to treatment with erlotinib alone, the cell apoptosis was enhanced if combined treatment of erlotinib and LY294002 was performed in NCI-H661 cells. Furthermore, combination treatment of erlotinib and LY294002 resulted in a significant reduction of phosphorylated p70S6K levels in NCI-H661 [PI3K catalytic subunit alpha (PI3KCA) wild-type] cells. However, this phenomenon was not observed in the NCI-H460 cell line (PIK3CA mutant-type). In conclusion, the present study indicated that inhibition of PI3K may have the potential to improve the sensitivity of NSCLC cells to an EGFR-TKI. However, the therapeutic effect may depend on the mutation status of PIK3CA.

Keywords: PI3K inhibitor; epidermal growth factor receptor; inhibitor; non-small cell lung cancer; p70S6K.

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Figures

Figure 1
Figure 1
Effect of LY294002 and erlotinib on NCI-H460 and NCI-H661 cell growth. (A) Erlotinib and (B) LY294002 inhibited the growth of NCI-H460 and NCI-H661 cells in a dose-dependent manner. Values are presented as the mean ± standard error of the mean from three independent experiments.
Figure 2
Figure 2
Combined effect of erlotinib and LY294002 to inhibit the growth of NCI-H460 and NCI-H661 cells. (A) NCI-H460 and (B) NCI-H661 cells were treated with erlotinib or LY294002 alone, or a combination of erlotinib and LY294002 at a fixed concentration. Compared with the erlotinib only group and the LY294002 only group, the growth inhibition of the combined treatment of erlotinib and LY294002 on the NCI-H460 and NCI-H661 cells was enhanced.
Figure 3
Figure 3
Apoptosis of (A) NCI-H661 and (B) NCI-H460 cells was measured by flow cytometry after treatment with a combination of erlotinib and LY294002. The percentage of apoptotic (C) NCI-H661 and (D) NCI-H460 cells was compared between the monotherapy and combined treatment groups. In NCI-H661 cells, a difference in cell apoptosis was detected with the combined drug treatment. Compared with the control, erlotinib (3 µM) or LY294002 (10 µM) treatment alone, the cell apoptosis was significantly enhanced after co-treatment. However, no significantly enhanced apoptosis was observed in the NCI-H460 cell line after co-treatment compared to LY294002 treatment alone. Apoptosis of the erlotinib+LY294002 group was significantly increased compared with the Erlotinib or LY294002 groups. *P<0.05 vs. Erlotinib; #P<0.05 vs. LY294002.
Figure 4
Figure 4
Effects of combined erlotinib and LY294002 treatment on the phosphorylation of p70S6K. NCI-H661 and NCI-H460 cell lines were treated with erlotinib, LY294002 or a combination of the two drugs. The cell extracts were examined by western blot analysis for determination of p-p70S6K. Erlotinib, LY294002 or the combination treatment inhibited p70S6K phosphorylation in the NCI-H661 cell line. While the phosphorylation level of p70S6K was not reduced after monotherapy in the NCI-H460 cell line, combination treatment caused a slight decrease in the level of p-p70S6K. p-p70S6K, phospho-p70S6K.
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