RESUMO
ALK, ROS1, and RET kinase fusions are important predictive biomarkers of tyrosine kinase inhibitors (TKIs) in non-small-cell lung cancer (NSCLC). Analysis of cell-free DNA (cfDNA) provides a noninvasive method to identify gene changes in tumor cells. The present study sought to use cfRNA and cfDNA for identifying fusion genes. A reliable protocol was established to detect fusion genes using cfRNA and assessed the analytical validity and clinical usefulness in 30 samples from 20 cases of fusion-positive NSCLC. The results of cfRNA-based assays were compared with tissue biopsy and cfDNA-based liquid biopsy (Guardant360 plasma next-generation sequencing [NGS] assay). The overall sensitivity of the cfRNA-based assay was 26.7% (8/30) and that of cfDNA-based assay was 16.7% (3/18). When analysis was limited to the samples collected at chemo-naïve or progressive disease status and available for both assays, the sensitivity of the cfRNA-based assay was 77.8% (7/9) and that of cfDNA-based assay was 33.3% (3/9). Fusion gene identification in cfRNA was correlated with treatment response. These results suggest that the proposed cfRNA assay is a useful diagnostic test for patients with insufficient tissues to facilitate effective administration of first-line treatment and is a useful tool to monitor the progression of NSCLC for consideration of second-line treatments.
Assuntos
Quinase do Linfoma Anaplásico/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Ácidos Nucleicos Livres , Fusão Gênica , Neoplasias Pulmonares/genética , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas c-ret/genética , Proteínas Proto-Oncogênicas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Biópsia , Carbazóis/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Ácidos Nucleicos Livres/isolamento & purificação , Crizotinibe/uso terapêutico , Proteínas do Citoesqueleto/genética , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Biópsia Líquida/métodos , Pulmão/patologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/isolamento & purificação , Piperidinas/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , RNA Mensageiro/isolamento & purificação , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Osimertinib (AZD9291) is a third-generation EGFR-tyrosine kinase inhibitor (TKI) that selectively inhibits the activating EGFR mutation and T790M mutation, and is currently used globally to treat EGFR-mutant non-small cell lung cancer (NSCLC). However, acquired resistance to osimertinib is inevitable. METHODS: We established osimertinib-resistant cells (PC9/T790M/AZDR and H1975/AZDR) derived from EGFR-mutant NSCLC cells harboring T790M mutation, and investigated the mechanism of acquired resistance to osimertinib by whole-exome sequencing and multiple phospho-receptor tyrosine kinase (RTK) array. A tumor specimen from an EGFR-mutant NSCLC patient with acquired resistance to osimertinib was also subjected to immunohistochemical analysis. RESULTS: Whole-exome sequencing analysis demonstrated that genetic alterations, such as acquisition of EGFR C797S, loss of T790M mutation, MET amplification, or mutated KRAS, MEK, BRAF, PIK3CA, were not detected. Analysis of phospho-RTK array revealed that insulin-like growth factor-1 receptor (IGF1R) was activated in PC9/T790M/AZDR and H1975/AZDR cells. Knockdown of IGF1R by siRNA as well as inhibition of IGF1R activation by linstinib (IGF1R inhibitor) significantly restored the sensitivity to osimertinib. Immunohistochemical analysis revealed that the expression level of phosphorylated IGF1R was higher in the tumor specimen from the EGFR-mutant NSCLC patient with acquired resistance to osimertinib than in the specimen collected prior to the treatment. CONCLUSIONS: IGF1R activation could occur following treatment with osimertinib in EGFR-mutant NSCLC with T790M mutation, and might be one of the mechanisms underlying osimertinib resistance. Combined treatment of osimertinib and IGF1R inhibitor might be effective in overcoming the acquired resistance to osimertinib induced by IGF1R activation. KEY POINTS: Significant findings of the study: Using osimertinib-resistant cells, we found that IGF1R activation induced by osimertinib treatment in EGFR-mutant NSCLC with T790M mutation is involved in resistance. Increased phosphorylation of IGF1R was observed in the tumor specimen from an EGFR-mutant NSCLC patient with acquired osimertinib resistance. WHAT THIS STUDY ADDS: IGF1R activation might be one of the mechanisms of osimertinib resistance. A combination therapy with osimertinib and an IGF1R inhibitor might be an optimal approach for overcoming the acquired resistance to osimertinib induced by IGF1R activation.
Assuntos
Acrilamidas/farmacologia , Compostos de Anilina/farmacologia , Biomarcadores Tumorais/metabolismo , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Receptor IGF Tipo 1/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Proliferação de Células , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Mutação , Inibidores de Proteínas Quinases/farmacologia , Receptor IGF Tipo 1/genética , Células Tumorais CultivadasRESUMO
Analyses of circulating tumor cells have been shown to be effective for the detection of cancer relapse and prognosis prediction. However, research regarding its utility in sarcoma remains scarce. In this study, the microfluidic chip-type cell sorter On-chip Sort was used to construct a system for detecting circulating sarcoma cells (CSCs). A pilot study using normal fibroblast or sarcoma cell lines was designed to establish a reliable protocol to separate CSCs by On-chip Sort. A single CSC was separated and recovered from 10 ml of whole blood from a patient with locally advanced myxofibrosarcoma. The nonsynonymous mutation for KMT2B p.Ile2602Val identified in the formalin-fixed paraffin-embedded tumor sample was also confirmed in the CSC. Use of the developed protocol may allow CSCs to become an early predictor for metastasis and recurrence of sarcoma. Further, it may aid in optimizing post-operative therapies for patients without metastasis.