Ito2019 - Gefitinib resistance of lung adenocarcinoma caused by MET amplification
March 2021, Model of the Month by Krishna Kumar Tiwari
Original model - BIOMD0000000827
In 2020, Cancer cases has risen to 19.3 million and 10 million cancer deaths all over world 1. A plethora of chemotherapy option are available for different type of cancers at present, but development of drug resistance poses a major challenge in cancer treatment. Drug resistance can be inherent, or acquired as a result of drug exposure 2. Thus, understanding the mechanism of drug resistance cell acquire or inherit is an important aspect to analyse and can help in identification of possible drug combination to overcome drug resistance. Mathematical modelling is known to provide an excellent insight into mechanistic aspect of biological systems and also well used to study the mechanisms of drug resistance 3. Ito et al 4 developed a mechanistic ordinary differential equation-based model providing the mechanism responsible for Gefitinib (Epidermal growth factor receptor-Tyrosine Kinase inhibitor (EGFR-TKI)) resistance in MET (MET proto-oncogene, receptor tyrosine kinase) amplified lung adenocarcinoma. MET, EGFR and ErbB3 (erb-b2 receptor tyrosine kinase 3) are found to be either mutated or overexpressed in lung adenocarcinomas. Hence the authors analyzed the levels and signaling pathways of MET, EGFR and ErbB3 in Gefitinib sensitive and resistance cell lines (HCC827, HCC827-GR5, and HCC827-GR6), and predicted the mechanism of resistance.
This ODE (ordinary differential equation) based biochemical reaction model 4 examines the molecular mechanism of gefitinib resistance in lung adenocarcinoma by MET amplification. The model includes MET, EGFR, ErbB3 receptor activation mechanisms, their interactions, Gefitinib mechanism of action and interaction of MET pathway with ErbB3 signalling (Figure1). Model explains the alternate mechanism of EGFR pathway activation through amplification of MET protein level and signalling in Gefitinib sensitive and resistance cell lines.
Figure 1: EGFR, ErbB3 and MET induced signalling and their interaction along with Gefitinib/AZD8931 mechanism of action. Figure recreated from Ito et al, 2019 4.
Expression level of EGFR and ErbB3 on the cell surface was almost similar in all three cell-lines as they are derived from the same origin (Figure 2 A, B). But the expression of MET was 19 times higher (due to gene amplification) in HCC827-GR5/ HCC827-GR6 cells as compared to HCC827 cell-line (Figure 2C).
A simulation was performed to inspect gefitinib resistance by MET amplification in HCC827 (Gefitinib sensitive) and HCC827-GR5 (Gefitinib resistant) cells. Model was simulated for t = 0-200s and levels of different active and inactive complexes where measured in two cell types. Dimerization of EGFR, ErbB3, and MET attained equilibrium approximately at 1s (Figure 2 D, E, F, G), while phosphorylation of each dimer attains steady state after more than 100s (Figure 2 H, I, J), which reflects the delay in signal transduction. Also, the increased expression of MET (gene amplification) lead to enhanced phosphorylation of EGFR/ErbB3 and ErbB3/ErbB3 heterodimers in gefitinib resistant HCC827-GR5 cells.
Simulation showed that the total number of p-MET/MET (2.4 x 105 cell-1) is approximately 150 times greater than that of the sum of pEGFR/ErbB3 (1.5 x 103 cell-1) and p-ErbB3/ErbB3 (38 cell-1) in HCC827-GR5 cells (Figure 2 H, I, J). High MET signaling was forecasted to cause gefitinib resistance by activating downstream pathways. This observation poses a question whether MET can activate pAKT/pERK directly or via ErbB3 pathway activation (Figure 1). To conform this, an additional experiment was done using AZD8931, a multi-kinase inhibitor for EGFR and ErbB3 and the levels of EGFR, ErbB3 and their downstream signaling pathway i.e. pAKT and pERK were measured in HCC827 and HCC827-GR5 cells through western blot analysis. AZD8931 reduced the activity of EGFR and ErbB3 receptors in both Gefitinib sensitive (HCC827) and resistant cell lines (HCC827-GR5/HCC827-GR6) but fails to reduce levels of pAKT and pERK significantly in Gefitinib resistance cell line (HCC827-GR5/HCC827-GR6). This additional experiment demonstrates that ErbB3 inhibition in Gefitinib resistant cell line doesn’t reduce the downstream signaling (i.e. pAKT/pERK) significantly. Based on the this results, it was concluded that MET causes gefitinib resistance through activation of pAKT/pERK and ErbB3 contribution in MET induced gefitinib resistant is limited.
Figure 2: A-J: Simulation output for EGFR, ErbB3 and MET homodimers and heterodimers in Gefitinib sensitive(HCC8927) and resistant(HCC8927-GR5) cell lines.
In this study, a simulation-based analysis of gefitinib resistance by MET amplification is carried out and the model demonstrated that the role of ErbB3 is limited in the gefitinib resistance induced by MET amplification. MET can activate AKT/ERK pathway directly or through activation of ErbB3 signaling. Inhibition of ErbB3 signaling by AZD8931 didn’t impact the levels of pAKT and pERK in Gefitinib resistance cell-lines (HCC827-GR5) despite a significant change in ErbB3 activity. This observation substantiate that MET signaling activate downstream signaling pathway independent of ErbB3 activation induced by MET in MET amplified gefitinib resistance cell lines. The model developed by Ito et al, 2019  provide a quantifiable details of the EGFR, ErbB3 and MET activation levels, crosstalk and mechanism underlying gefitinib resistance induced by MET amplification in lung adenocarcinoma, which is difficult to achieve by an ordinary molecular biological approach.
1. ‘GLOBOCAN 2020: New Global Cancer Data | UICC’. https://www.uicc.org/news/globocan-2020-new-global-cancer-data.
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3. X. Sun and B. Hu, ‘Mathematical modeling and computational prediction of cancer drug resistance’, Brief. Bioinform., vol. 19, no. 6, pp. 1382–1399, Nov. 2018, doi: 10.1093/bib/bbx065.
4. T. Ito et al., ‘Mathematical analysis of gefitinib resistance of lung adenocarcinoma caused by MET amplification’, Biochem. Biophys. Res. Commun., vol. 511, no. 3, pp. 544–550, Apr. 2019, doi: 10.1016/j.bbrc.2019.02.086.