Importance of the Keap1-Nrf2 pathway in NSCLC: Is it a possible biomarker?

Abstract

Worldwide, lung cancer remains the most common cause of cancer-related mortality, with non-small cell lung cancer (NSCLC) accounting for 85% of all diagnosed lung cancer cases. Chemotherapy is considered the standard of care for patients with advanced NSCLC; however, the tumors can develop mechanisms that inactivate these drugs. Comparative genomic analyses have revealed that disruptions in the kelch-like ECH-associated protein 1 (Keap1)-nuclear factor erythroid-2-related factor-2 (Nrf2) pathway are frequent in NSCLC, although Nrf2 mutations occur less frequently than Keap1 mutations. As the Keap1-Nrf2 pathway appears to be a primary regulator of key cellular processes that aid to resist the action of chemotherapy drugs, the clinical implementation of Nrf2 inhibitors in patients with advanced NSCLC may be a useful therapeutic approach for patients harboring KEAP1-NRF2 mutations. The aim of the present review was to highlight findings of how constitutive Nrf2 activation may be a specific biomarker for predicting patients most likely to benefit from classical chemotherapy drugs, overall improving patient survival rate.

Keywords: biomarker; drug resistance; kelch-like ECH-associated protein 1-nuclear factor erythroid-2-related factor-2 pathway; lung cancer; non-small cell lung cancer.

Figure 1.

Keap1-Nrf2 pathway. (A) Under physiological/basal conditions, the Keap1-Cullin3 complex promotes ubiquitin E3-ligase activity and proteasomal degradation of Nrf2. (B) Upon exposure to chemicals or ROS, reactive cysteine residues of Keap1 are modified leading to a disruption of the Keap1-Nrf2 complex and the release of Nrf2, which also allows de novo synthesized Nrf2 to accumulate in the cytoplasm. The phosphorylation of Ser-40 in the Neh2 domain of Nrf2 by PKCδ results in Nrf2 activation and translocation to the nucleus, where Nrf2-mediated transcriptional activity of ARE-responsive genes is initiated. GSK-3β phosphorylates the tyrosine kinase Fyn and induces its nuclear accumulation. Fyn phosphorylates Nrf2 at tyrosine-568, facilitating its nuclear export and degradation. (C) Loss-of-function mutations in Keap1 or Nrf2 lead to constitutive activation of Nrf2 by disrupting the Keap1-Nrf2 interaction. In the nucleus, Nrf2 heterodimerizes with sMaf proteins and regulates the expression of >200 oxidative stress-related genes. ARE, antioxidant response element; Fyn, proto-oncogene Src-family tyrosine kinase; GSK-3β, glycogen synthase kinase 3; Keap1, kelch-like ECH-associated protein 1; sMaf, musculoaponeurotic fibrosarcoma oncogene; Nrf2, nuclear factor erythroid-2-related factor-2; PKCδ, protein kinase C-δ; ROS, reactive oxygen species.

Figure 2.

Schematic representation of domain architecture of the Keap1 and Nrf2 proteins and mutations found in NSCLC. (A) Human NRF2 is a polypeptide of 605 amino acids and contains 7 Neh domains. Neh1 contains the signature CNC motif, which is a highly conserved bZIP domain for DNA binding. The Neh1 CNC-bZIP domain is responsible for dimerization with sMaf protein and is required for binding to ARE sequences in DNA; the Neh2 domain controls the interaction with Keap1 through the DLG and ETGE motifs; the Neh6 domain contains two binding sites (DSGIS and DSAPGS motifs), and the phosphorylation of the DSGIS motif by GSK-3β increases binding ability to the β-TrCP1 adaptor protein. (B) Human Keap1 is a polypeptide of 644 amino acids. The BTB domain is required for the formation of Keap1 homodimers as well as the recruitment of Cul3-based E3-ligase. The Kelch-repeat domain controls Nrf2 interaction. The region between the BTB and Kelch repeat domains constitutes the IVR. Amino acid positions of the identified mutations of Nrf2 and Keap1 are shown for ADC, ADC/SqCC and SqCC. All the indicated mutations are listed in the Catalogue of Somatic Mutations in Cancer database (http://cancer.sanger.ac.uk/cosmic). ADC, adenocarcinoma; ADC/SqCC, adenocarcinoma/squamous cell carcinoma; ARE, antioxidant response element; bZIP, basic leucine zipper; BTB, broad complex, tram-track and bric-a-brac; CNC, cap'n'collar; Cul3, Cullin3; GSK-3β, glycogen synthase kinase 3; IVR, intervening linker; KEAP1, kelch-like ECH-associated protein 1; sMaf, musculoaponeurotic fibrosarcoma oncogene; Neh, Nrf2-ECH homologous structure; NRF2, nuclear factor erythroid-2-related factor-2; SqCC, squamous cell carcinoma; β-TrCP1, β-T-complex protein 1.