Resistance to gefitinib and cross‐resistance to irreversible EGFR‐TKIs mediated by disruption of the Keap1‐Nrf2 pathway in human lung cancer cells

Abstract

The development of resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) occurs by various mechanisms and appears to be almost inevitable, even in patients with lung cancer who initially respond well to EGFR-TKIs. Consequently, considerable efforts have been made to develop more effective EGFR-TKIs. Therefore, an understanding of the mechanisms behind TKI resistance is essential for improving EGFR-TKI therapeutic efficacy in non-small cell lung cancer (NSCLC) patients. In this study, we discovered that overexpression of antioxidant-responsive element (ARE)-containing Nrf2 target genes by increased transactivation of Nrf2 occurred because of an acquired Keap1 mutation in the gefitinib-resistant (GR) NSCLC cell line we established. These GR cells also acquired cross-resistance to the irreversible EGFR-TKIs, afatinib and osimertinib, and showed increased viability, invasiveness, proliferation, and tumorigenicity both in vitro and in vivo. These results were confirmed by the fact that inhibition of Nrf2 activity, either by treatment with brusatol or by inducing expression of exogenously introduced wild-type Keap1, suppressed tumor cell proliferation and tumorigenicity in vitro and in vivo. Our data suggest that disruption of the Keap1-Nrf2 pathway is one of the mechanisms by which EGFR-TKI resistance occurs, a fact that must be considered when treating patients with EGFR-TKI.-Park, S.-H., Kim, J. H., Ko, E., Kim, J.-Y., Park, M.-J., Kim, M. J., Seo, H., Li, S., Lee, J.-Y. Resistance to gefitinib and cross-resistance to irreversible EGFR-TKIs mediated by disruption of the Keap1-Nrf2 pathway in human lung cancer cells.

The transcription factor NRF2 enhances melanoma malignancy by blocking differentiation and inducing COX2 expression

“…NRF2 is considered an oncogene, and activation of NRF2 by deleterious mutations of KEAP1 are found in several tumor types, such as non-small cell lung cancer (NSCLC) with 15-20% mutation frequency [13] (www.cbioportal.org). Mutational activation of the KEAP1/NRF2 pathway is connected to therapy resistance in NSCLC [14,15]. In melanoma, the role of NRF2 is much less understood.…”

Emerging role of nuclear factor erythroid 2-related factor 2 in the mechanism of action and resistance to anticancer therapies

“…In anticancer chemotherapy, NRF2 and NRF-dependent genes have been implicated in the cellular resistance to a wide range of anticancer agents (e.g., tamoxifen, Cisplatin, Oxaliplatin, Cisplatin, Doxorubicin, and Etoposide) and cancer types [18][19][20][21][22][23][24][25] . Likewise, the NRF2-centred system and signalling pathway is shown to modulate the action and effectiveness of certain receptor targeted therapies [26][27][28]224,231,232] and potentially promoting cancer resistance to such interventions as Trastuzumab, Pertuzumab, Erlotinib, Lapatinib, imatinib, Gefitinib, Afatinib and Osimertinib. In both anticancer chemotherapy and receptor target therapy, the inhibition of NRF2 and its function seemingly and contextually enhanced drug sensitisation of cancers and/or helped to overcome drug resistance.…”

Epigenetic Scanning of KEAP1 CpG Sites Uncovers New Molecular-Driven Patterns in Lung Adeno and Squamous Cell Carcinomas

“…The presence of genetic and epigenetic abnormalities in this pathway, such as point mutations in functional domains of KEAP1 and NFE2L2 and methylation at the KEAP1 promoter region, was firstly described in NSCLC and then widely reported in many solid tumors, with the hypothesis that the lack of KEAP1 transcription induced high NRF2 activity and aberrant overexpression of ARE-driven target genes [ 6 , 10 , 16 , 18 ]. Actually, increasing attention to the KEAP1 gene in lung cancer is mainly due to point mutations that show great translational impact in terms of increased risk of cancer progression, shorter overall survival and response of NSCLC patients’ to chemo and biological treatments [ 3 , 4 , 5 , 31 ]. This notion is also supported by a clear co-occurrence with STK11 and KRAS gene alterations with a suggested synergic role of these genes in enhancement of tumorigenesis and lung cancer progression.…”

“…Our results support the documented link between KEAP1 , EGFR, and KRAS mutations and open the debate on the role of KEAP1 methylation in the context of anti-EGFR treatments. For NSCLC patients whose tumors harbor mutations in EGFR , disruption of the KEAP1/NRF2 pathway is of the most recently reported mechanisms by which EGFR-tyrosine kinase inhibitors (EGFR-TKI) resistance occurs [ 5 ]. Coexisting mutations in the KEAP1/NFE2L2/CUL3 genes were in fact reported to be associated with significantly decreased time to TKI treatment failure [ 4 ].…”

“…Hundreds of studies have been published on discovering new prognostic molecular factors beyond the tumor-node-metastasis (TNM) non-small-cell lung cancer (NSCLC) staging system, aiming to provide some new biological and clinical insights [ 2 ]. Interestingly, a significant translational impact in terms of increased risk of cancer progression and shorter overall survival was documented for alterations in the Kelch-like ECH-associated protein 1 ( KEAP1 ) gene [ 3 , 4 , 5 ]. KEAP1 is a cytoplasmic anchor protein of Nuclear factor erythoid-2-related factor 2 (NRF2), encoded by the NFE2L2 gene, which predominantly acts as a key regulator of antioxidant stress responses also in the context of tumor resistance and progression [ 6 , 7 , 8 ].…”