Mutation and drug-specific intracellular accumulation of EGFR predict clinical responses to tyrosine kinase inhibitors

Wit, Maurice de; Gao, Yinjie; Mercieca, Darlene; Heer, Iris de; Valkenburg, Bart; Royen, Martin E. van; Aerts, Joachim; Smitt, Peter Sillevis; French, Pim J.

doi:10.1016/j.ebiom.2020.102796

Cited by 11 publications

(10 citation statements)

References 39 publications

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“…and while its abundance was larger in the nuclear fraction, upregulation in the nuclear fraction was not observed. The accumulation of EGFR in the cytoplasm was noted consistently and was in agreement with prior findings that emerged from the treatment of various EGFR mutant cancers with Tyr kinase inhibitors where the cytoplasmic accumulation of EGFR was used as a predictor of clinical efficacy [82].…”

Section: Upregulated Biological Processessupporting

confidence: 89%

Proteomic Assessment of SKBR3/HER2+ Breast Cancer Cellular Response to Lapatinib and Investigational Ipatasertib Kinase Inhibitors

Karcini,

Mercier,

Lazar

2024

Preprint

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Modern cancer treatment approaches aim at achieving cancer remission by using targeted and personalized therapies, as well as harnessing the power of the immune system to recognize and eliminate the cancer cells. To overcome a relatively short-lived response due to the development of resistance to the administered drugs, combination therapies have been pursued, as well. To expand the outlook of combination therapies, the objective of this study was to use high-throughput data generation technologies such as mass spectrometry and proteomics to investigate the response of HER2+ breast cancer cells to a mixture of two kinase inhibitors that has not been adopted yet as a standard treatment regime. The broader landscape of biological processes that are affected by inhibiting two major pathways that sustain the growth and survival of cancer cells, i.e., EGFR and PI3K/AKT, was investigated by treating SKBR3/HER2+ breast cancer cells with Lapatinib or a mixture of Lapatinib/Ipatasertib small molecule drugs. Changes in protein expression and/or activity in response to the drug treatments were assessed by using two complementary quantitative proteomic approaches based on peak area and peptide spectrum match measurements. Over 900 proteins matched by three unique peptide sequences (FDR<0.05) were affected by the exposure of cells to the drugs. The work corroborated the anti-proliferative activity of Lapatinib and Ipatasertib, and, in addition to cell cycle and growth arrest processes enabled the identification of several multi-functional proteins with roles in cancer-supportive hallmark processes. Among these, immune response, adhesion and migration emerged as particularly relevant to the ability to effectively suppress the proliferation and dissemination of cancer cells. The supplementation of Lapatinib with Ipatasertib further affected the expression or activity of additional transcription factors and proteins involved in gene expression, trafficking, DNA repair, and development of multidrug resistance. Furthermore, over fifty of the affected proteins represented approved or investigational targets in the DrugBank database, which through their protein-protein interaction networks can inform the selection of effective therapeutic partners. Altogether, our findings exposed a broad plethora of yet untapped opportunities that can be further explored for enhancing the anti-cancer effects of each drug as well as of many other multi-drug therapies that target the EGFR/ERBB2 and PI3K/AKT pathways. The data are available via ProteomeXchange with identifier PXD051094.

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Section: Upregulated Biological Processessupporting

confidence: 89%

Proteomic Assessment of SKBR3/HER2+ Breast Cancer Cellular Response to Lapatinib and Investigational Ipatasertib Kinase Inhibitors

Karcini,

Mercier,

Lazar

2024

Preprint

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“…Another recent study reports that TKI treatment induces intracellular accumulation of mutated-EGFR. The authors found a positive correlation between EGFR accumulation and clinical benefit in patients presenting with tumors harboring these EGFR mutations, suggesting that blocked EGFR-membrane recycling contributes to TKI sensitivity and a positive therapy outcome [ 67 ]. Our results demonstrate not only that erlotinib has a different effect on tumor and normal cells but also that PDAC cells respond differently to erlotinib treatment when cultured in a 3D microenvironment.…”

Section: Discussionmentioning

confidence: 99%

Erlotinib Promotes Ligand-Induced EGFR Degradation in 3D but Not 2D Cultures of Pancreatic Ductal Adenocarcinoma Cells

Betriu

Andreeva

Semino

2021

Cancers

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The epithelial growth factor receptor (EGFR) is a tyrosine kinase receptor that participates in many biological processes such as cell proliferation. In addition, EGFR is overexpressed in many epithelial cancers and therefore is a target for cancer therapy. Moreover, EGFR responds to lots of stimuli by internalizing into endosomes from where it can be recycled to the membrane or further sorted into lysosomes where it undergoes degradation. Two-dimensional cell cultures have been classically used to study EGFR trafficking mechanisms in cancer cells. However, it has been widely demonstrated that in 2D cultures cells are exposed to a non-physiological environment as compared to 3D cultures that provide the normal cellular conformation, matrix dimensionality and stiffness, as well as molecular gradients. Therefore, the microenvironment of solid tumors is better recreated in 3D culture models, and this is why they are becoming a more physiological alternative to study cancer physiology. Here, we develop a new model of EGFR internalization and degradation upon erlotinib treatment in pancreatic ductal adenocarcinoma (PDAC) cells cultured in a 3D self-assembling peptide scaffold. In this work, we show that treatment with the tyrosine kinase inhibitor erlotinib promotes EGFR degradation in 3D cultures of PDAC cell lines but not in 2D cultures. We also show that this receptor degradation does not occur in normal fibroblast cells, regardless of culture dimensionality. In conclusion, we demonstrate not only that erlotinib has a distinct effect on tumor and normal cells but also that pancreatic ductal adenocarcinoma cells respond differently to drug treatment when cultured in a 3D microenvironment. This study highlights the importance of culture systems that can more accurately mimic the in vivo tumor physiology.

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“…Such an analysis can monitor alterations in expression and translocation patterns in response to different cell stimulations. The membrane to cytoplasmatic ratio has previously been used in image-based analysis and biochemical methods [18][19][20][21] , but the method described here is novel in that the robust and rapid procedure can be used to quantify the MCR at a single-cell level. In addition, the imagebased workflow presented here enables the protein expression and translocation to be quantified in each cellular distribution region.…”

Section: Discussionmentioning

confidence: 99%

Imaging analysis to quantitate the Interplay of membrane and cytoplasm protein dynamics

Kislev

Egozi

Benayahu

2021

Preprint

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Plasma membrane proteins are extremely important in cell signaling and cellular functions. Protein expression and localization alter in response to various signals in a way that is dependent on cell type and niche. Compartmental quantification of the expression of particular proteins is a very useful means of understanding their role in cellular processes. Immunofluorescence staining is frequently used to investigate the distribution of proteins of interest. Here, we present an imaging method for quantifying the membrane to cytoplasm ratio (MCR) of proteins analyzed at single-cell resolution. This technique provides a robust quantification of membrane proteins and contributes new insights into membrane expression dynamics. We have developed a protocol that uses immunostaining to assess protein expression according to the fluorescent cellular distribution and to compute the MCR. The method was applied to measure the MCR of glucose transporter 4 (GLUT4) in response to insulin in 3T3-L1 cells, an in-vitro model for adipocyte function and adipogenesis. The results revealed informative changes in the subcellular localization of GLUT4 following insulin induction. MCR analysis is a powerful imaging tool that can be generally applied to membrane proteins to provide a rapid and efficient quantitative analysis of protein distribution and sub-cellular processes in cells.

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Mutation and drug-specific intracellular accumulation of EGFR predict clinical responses to tyrosine kinase inhibitors

Cited by 11 publications

References 39 publications

Proteomic Assessment of SKBR3/HER2+ Breast Cancer Cellular Response to Lapatinib and Investigational Ipatasertib Kinase Inhibitors

Proteomic Assessment of SKBR3/HER2+ Breast Cancer Cellular Response to Lapatinib and Investigational Ipatasertib Kinase Inhibitors

Erlotinib Promotes Ligand-Induced EGFR Degradation in 3D but Not 2D Cultures of Pancreatic Ductal Adenocarcinoma Cells

Imaging analysis to quantitate the Interplay of membrane and cytoplasm protein dynamics

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