Abstract
Simple SummaryLung cancer is the leading cause of cancer-related deaths, with a low (<21%) 5-year survival rate. Lung cancer is often driven by the misfunction of molecules on the surface of cells of the epithelium, which orchestrate mechanisms by which these cells grow and proliferate. Beyond common non-specific treatments, such as chemotherapy or radiotherapy, among molecular-specific treatments, a number of small-molecule drugs that block cancer-driven molecular activity have been developed. These drugs initially have significant success in a subset of patients, but these patients systematically develop resistance within approximately one year of therapy. Substantial efforts towards understanding the mechanisms of resistance have focused on the genomics of cancer progression, the response of cells to the drugs, and the cellular changes that allow resistance to develop. Fluorescence microscopy of many flavours has significantly contributed to the last two areas, and is the subject of this review. Non-small cell lung cancer (NSCLC) is a complex disease often driven by activating mutations or amplification of the epidermal growth factor receptor (EGFR) gene, which expresses a transmembrane receptor tyrosine kinase. Targeted anti-EGFR treatments include small-molecule tyrosine kinase inhibitors (TKIs), among which gefitinib and erlotinib are the best studied, and their function more often imaged. TKIs block EGFR activation, inducing apoptosis in cancer cells addicted to EGFR signals. It is not understood why TKIs do not work in tumours driven by EGFR overexpression but do so in tumours bearing classical activating EGFR mutations, although the latter develop resistance in about one year. Fluorescence imaging played a crucial part in research efforts to understand pro-survival mechanisms, including the dysregulation of autophagy and endocytosis, by which cells overcome the intendedly lethal TKI-induced EGFR signalling block. At their core, pro-survival mechanisms are facilitated by TKI-induced changes in the function and conformation of EGFR and its interactors. This review brings together some of the main advances from fluorescence imaging in investigating TKI function and places them in the broader context of the TKI resistance field, highlighting some paradoxes and suggesting some areas where super-resolution and other emerging methods could make a further contribution.
Highlights
Beyond common non-specific treatments, such as chemotherapy or radiotherapy, among molecularspecific treatments, a number of small-molecule drugs that block cancer-driven molecular activity have been developed
By imaging in the physiological cell context, and in the presence and absence of tyrosine kinase inhibitors (TKIs), the combination fluorescently tagged proteins (e.g., epidermal growth factor receptor (EGFR) and its signalling and regulatory effector machinery) and fluorescence microscopy methods have facilitated investigation of the crosstalk between canonical functions affected by catalytically competent EGFR with those that may be exerted by TKIbound EGFR independently of its kinase activity
Fluorescence imaging has helped to decipher some of the unintended consequences that the removal of kinase activity has on EGFR endocytic trafficking, and/or the regulation of pro-survival functions, revealing the significance of these changes in helping the cell to survive therapeutic insults
Summary
Lung cancers are classified in two main histological groups: small-cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) [1]. Gene sequencing technologies have allowed the identification of driver oncogenic gene alterations in the EGFR gene itself (reviewed in [5]), and/or of genes expressing oncogenic proteins within EGFR’s downstream signalling pathways, especially those that regulate cell survival and proliferation, on which tumour initiation and growth critically depend [6] (examples in Box 1). A striking response was found in a subset of ~10–40% of patients who harboured NSCLC tumours driven by somatic activating mutations in the first 4 exons of the tyrosine kinase domain of the EGFR gene [32,33] (Figure 1B). The intrinsic resistance of wtEGFR-expressing tumours to TKIs is recapitulated by many other solid tumour types (reviewed in [39]) This is so even in the absence of mutations in effectors downstream of EGFR that decouple growth and survival pathways from EGFR signalling [15] (Box 1). Simulations, it can report on the dimer and oligomer structure [85,86]
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