miR-200c in extracellular vesicles as a circulating biomarker of EGFR-TKI response and a mediator of EGFR-TKI sensitivity in heterogeneous NSCLC.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. e20547-e20547
Author(s):  
Chienchung Lin ◽  
Wu-Chou Su
Keyword(s):  
Extracellular Vesicles ◽  
Mutant Cell ◽  
Poor Response ◽  
Wild Type Cell ◽  
Egfr Mutations ◽  
Small Rna Sequencing ◽  
Wild Type ◽  
Egfr Mutant ◽  
Egfr Tki

e20547 Background: Through intercellular transfer of EV(extracellular vesicles) miRNA, tumor cells can confer drug resistance to each other and contribute to tumor heterogeneity. However, the mechanisms why EGFR-TKI can be effective in heterogeneous NSCLC with low abundances of EGFR mutations remain unknown. The aim of the study is to investigate the significance of EV miRNA in mediating the efficacy of EGFR-TKI in heterogeneous NSCLC and serving as the biomarker of response to EGFR-TKI. Methods: We first evaluate if EVs from EGFR mutant cell (PC9) can affect EGFR-TKI sensitivity of EGFR wild type cell (CL1-5, H1299) in vitro co-culture system and i n vivo. We then identified the differential miRNA panel by comparing EVs from PC9 to those from CL1-5. Finally, we verified if the expressions level of these miRNA are different in blood EVs from patient with good response compared to those with poor response to EGFR-TKI. Results: We first verified that CL1-5 can take up labelled EVs from PC9 under time-lapse microscope and EGFR mutant DNA can be detected in recipient EGFR wild-type cell using digital PCR. We found EVs from PC9 enhanced gefitinib sensitivity of CL1-5. And co-culturing PC9 with CL1-5 enhanced CL1-5 gefitinib sensitivity which was reversed by adding GW4789, the inhibitor of exosome secretion. In subcutaneous CL1-5 animal model, in comparison to treating with gefitinib or PC9 EVs alone, only the combination treatment with gefitinib and PC9 EVs delayed cancer growth. Using small RNA sequencing, we identified a unique miRNA profile allowing discriminating EV from PC9 cells to those from CL1-5; MiRNA 200 family including 200a, 200b, 200c and mir429 were up-regulated significantly in PC9 EV. From Aug 2015 to Sep 2017, sixteen patients with good response (PFS > 12M) or poor response (PFS < 6M) to EGFR-TKI were enrolled and blood were collected for EV miRNA isolation. Ten of these blood samples were qualified for miRNA analysis and mir200a and 200c were found up-regulated in good responder to EGFR-TKI. The transfection of mir200c in CL1-5 cells not only inhibited the oncogenic pathway contributing to EGFR-TKI resistance including Stat3, Akt, EMT and BIM pathway but also enhanced gefitinib sensitivity of CL1-5 cells. Conclusions: Our data suggested mir200c from blood EV serve as a biomarker of response to EGFR-TKI and mir200c may mediate EGFR-TKI sensitivity in heterogenous EGFR mutant NSCLC.

Investigation of immune microenvironment in EGFR mutant NSCLC.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. e20517-e20517
Author(s):  
Yi Hu ◽  
Longgang Cui ◽  
Xiaochen Zhao ◽  
Yuezong Bai ◽  
Fan Zhang
Keyword(s):  
T Cells ◽  
Egfr Mutation ◽  
Tumor Stroma ◽  
Median Number ◽  
Egfr Mutations ◽  
Wild Type ◽  
Immune Microenvironment ◽  
Regulatory Pathways ◽  
Significant Difference ◽  
Egfr Mutant

e20517 Background: Immune checkpoint inhibitor therapy has made great achievements in NSCLC, but patients with EGFR mutations have poor efficacy with immunotherapy. Previous studies have explored the expression of PD-L1, neo-antigen, co-mutation and regulatory pathways in EGFR mutate NSCLC. This work compared the immune microenvironment of EGFR mutant and wild-type NSCLC. Methods: Patients: NSCLC. Using multi-color immunohistochemistry (multi-IHC) to evaluate the expression of 2 indicators in tumors and tumor stroma, namely CD8+ T cell and macrophage. Shapiro-Wilk was used for normality test, and t-test or Mann-Whitney U test was used according to the results. Two-sided P < 0.05 was considered a significant difference. Results: The study included 119 NSCLC patients, including 59 women (49.6%) and 60 men (50.4%), with a median age of 57. There were 68 patients (57.1%) with EGFR mutations, 19 patients (16%) with KRAS mutations, and 58 patients (48.7%) with TP53 mutations. multi-IHC results showed that, EGFR mutation Vs EGFR wild type samples, (1) the number and proportion of CD8+ T cells in tumor were not statistically different. The median number of CD8+ T cells in tumor stroma was 231.5 vs 359, p = 0.05 and the proportion of CD8+ T cells was 3.92% vs 5.64%, p = 0.02; (2) The median number of macrophage in tumors was 1522 vs 110, p < 0.01, and the proportion of macrophage cells was 24.93% Vs 1.38%, p < 0.01. The median value of macrophages in tumor stroma was 617.5 Vs 208, p < 0.01, and the proportion of macrophages cells was 10.88% vs 3.03%, p < 0.01. Conclusions: Compared with EGFR wild-type patients, EGFR mutation patients have a lower proportion of CD8+ T cells and a higher proportion of macrophages in the immune microenvironment.

Factor VIIa induces extracellular vesicles from the endothelium: A potential mechanism for its hemostatic effect

Blood ◽  
2021 ◽  
Author(s):  
Kaushik Das ◽  
Shiva Keshava ◽  
Shabbir A Ansari ◽  
Vijay Kumar Reddy Kondreddy ◽  
Charles Esmon ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Extracellular Vesicles ◽  
Factor Viia ◽  
Mutant Mice ◽  
In Vivo Studies ◽  
Cell Protein ◽  
Wild Type ◽  
Hemostatic Effect

Recombinant FVIIa (rFVIIa) is used as a hemostatic agent to treat bleeding disorders in hemophilia patients with inhibitors and other groups of patients. Our recent studies showed that FVIIa binds endothelial cell protein C receptor (EPCR) and induces protease-activated receptor 1 (PAR1)-mediated biased signaling. The importance of FVIIa-EPCR-PAR1-mediated signaling in hemostasis is unknown. In the present study, we show that FVIIa induces the release of extracellular vesicles (EVs) from endothelial cells both in vitro and in vivo. Silencing of EPCR or PAR1 in endothelial cells blocked the FVIIa-induced generation of EVs. Consistent with these data, FVIIa treatment enhanced the release of EVs from murine brain endothelial cells isolated from wild-type, EPCR overexpressors, and PAR1-R46Q mutant mice, but not EPCR-deficient or PAR1-R41Q mutant mice. In vivo studies revealed that administration of FVIIa to wild-type, EPCR overexpressors, and PAR1-R46Q mutant mice, but not EPCR-deficient or PAR1-R41Q mutant mice, increase the number of circulating EVs. EVs released in response to FVIIa treatment exhibit enhanced procoagulant activity. Infusion of FVIIa-generated EVs and not control EVs to platelet-depleted mice increased thrombin generation at the site of injury and reduced blood loss. Administration of FVIIa-generated EVs or generation of EVs endogenously by administering FVIIa augmented the hemostatic effect of FVIIa. Overall, our data reveal that FVIIa treatment, through FVIIa-EPCR-PAR1 signaling, releases EVs from the endothelium into the circulation, and these EVs contribute to the hemostatic effect of FVIIa.

scholarly journals Concurrent Genetic Alterations and Other Biomarkers Predict Treatment Efficacy of EGFR-TKIs in EGFR-Mutant Non-Small Cell Lung Cancer: A Review

2020 ◽  
Vol 10 ◽  
Author(s):  
Yijia Guo ◽  
Jun Song ◽  
Yanru Wang ◽  
Letian Huang ◽  
Li Sun ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Treatment Efficacy ◽  
Genetic Alterations ◽  
Small Cell ◽  
Egfr Mutations ◽  
Small Cell Lung ◽  
Egfr Mutant ◽  
Egfr Tki ◽  
Egfr Tkis ◽  
Gene Alterations

Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) greatly improve the survival and quality of life of non-small cell lung cancer (NSCLC) patients with EGFR mutations. However, many patients exhibit de novo or primary/early resistance. In addition, patients who initially respond to EGFR-TKIs exhibit marked diversity in clinical outcomes. With the development of comprehensive genomic profiling, various mutations and concurrent (i.e., coexisting) genetic alterations have been discovered. Many studies have revealed that concurrent genetic alterations play an important role in the response and resistance of EGFR-mutant NSCLC to EGFR-TKIs. To optimize clinical outcomes, a better understanding of specific concurrent gene alterations and their impact on EGFR-TKI treatment efficacy is necessary. Further exploration of other biomarkers that can predict EGFR-TKI efficacy will help clinicians identify patients who may not respond to TKIs and allow them to choose appropriate treatment strategies. Here, we review the literature on specific gene alterations that coexist with EGFR mutations, including common alterations (intra-EGFR [on target] co-mutation, TP53, PIK3CA, and PTEN) and driver gene alterations (ALK, KRAS, ROS1, and MET). We also summarize data for other biomarkers (e.g., PD-L1 expression and BIM polymorphisms) associated with EGFR-TKI efficacy.

EGFR-TKI benefit for NSCLC patients with EML4-ALK rearrangement.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. e18035-e18035
Author(s):  
Zhijie Wang ◽  
Jie Wang ◽  
Yi Long Wu ◽  
Hua Bai ◽  
Xu-Chao Zhang ◽  
...  
Keyword(s):  
Molecular Subtype ◽  
Objective Response ◽  
Stage Iv ◽  
Progression Free Survival ◽  
Egfr Mutations ◽  
Alk Rearrangement ◽  
Mutant Type ◽  
Egfr Mutant ◽  
Nsclc Patients ◽  
Egfr Tki

e18035 Background: EML4-ALK rearrangement defines a new molecular subtype of non-small-cell lung cancer (NSCLC). To identify the biological profiles of these patients, we examined the clinico-pathologic characteristics and treatment outcomes of NSCLC patients based on EML4-ALK and EGFR mutations. Methods: Patients with stage IV NSCLC were screened for EML4-ALK rearrangement and EGFR mutations at Peking University Cancer Hospital. EML4-ALK was identified using fluorescent in situ hybridization (FISH) confirmed by immunohistochemistry (IHC), and EGFR mutations were determined using denaturing high-performance liquid chromatography (DHPLC). Results: Of the 151 patients screened, 113 had complete follow-up data as an analysis set. The incidence of EML4-ALK was 9.7% (11/113) using FISH, in which 10 cases had sufficient specimens for IHC confirmation and all were positive. Overall, EML4-ALK and EGFR mutations were largely mutually exclusive (p = 0.033), although two patients harbored concurrent mutations. EML4-ALK rearrangement was associated with resistance to EGFR-TKIs compared with the EGFR mutant type and WT/Nonrearrangement type (p = 0.001 for objective response rate; p = 0.004 for disease control rate; p = 0.021 for progression-free survival [PFS]). In terms of patients who received platinum-based doublet chemotherapy, no significant differences were observed in PFS between the EML4-ALK type, EGFR mutant type, and WT/Nonrearrangement type. Moreover, two patients with concurrent EML4-ALK and EGFR mutations had superior PFS after EGFR-TKI compared with single EML4-ALK-rearranged patients. Conclusions: This study presents several biological features of EML4-ALK NSCLC. It is largely mutually exclusive to EGFR mutations, resistant to EGFR-TKI. Coexistence of ALK rearrangement and EGFR mutation in patients with advanced NSCLC might represent a separate genotype with unique biological characteristics.

Optimisation of EGFR TKI efficiency in the therapeutic scheme of EGFR wild-type lung cancer.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e18532-e18532
Author(s):  
Mathilde Cabart ◽  
Judith Raimbourg ◽  
Lisenn Lalier ◽  
Jaafar Bennouna ◽  
Francois Vallette
Keyword(s):  
Lung Cancer ◽  
Tyrosine Kinases ◽  
Receptor Tyrosine Kinases ◽  
Wild Type ◽  
Short Term ◽  
Egfr Ligands ◽  
Egfr Mutated ◽  
Egfr Tki

e18532 Background: EGFR tyrosine kinase inhibitors (EGFR TKI) have improved the therapeutic care of lung cancer patients but only a small sub-population of patients, namely those harboring EGFR-mutated tumors, benefit from this therapy. The observation that EGFR TKI enhance prognosis and quality of life in all patients when used as second line or maintenance treatment impelled us into the search of potential markers of the optimal introduction kinetics of EGFR TKI in the therapeutic scheme. Methods: We used lung cancer cell lines harboring either wild-type or mutated EGFR (NCI-H1650, NCI-H1975) to study the consequences of cisplatin treatment in vitro on the consecutive sensitivity to erlotinib. Results: Sub-lethal cisplatin pretreatment (3µM) enhances erlotinib toxicity in EGFR wild-type, but not EGFR mutated cells (A549 IC50 drops from 28 to 15µM for short-term or 12µM for long-term exposure). This correlates with EGFR activation following short-term or prolonged cisplatin treatment through the secretion of EGFR ligands. This activation of EGFR is concomitant to the decrease in other receptor tyrosine kinases phosphorylation including Met. Conclusions: The sensitivity of EGFR wild-type lung cancer cells to erlotinib in vitro is enhanced by cisplatin pretreatment. We identified potential markers of this sensitization, namely EGFR ligands, which serum level might be predicitive of the optimal efficiency of EGFR TKI. In vivo validation of these markers is under investigation. The concomitant decrease in other receptor tyrosine kinases phosphorylation suggests that the targeting of other receptor tyrosine kinases might potentiate EGFR TKI efficiency.

Impact of HER2 aberrations on EGFR-TKI treatment outcomes in lung tumors harboring EGFR mutations: A HER2-CS STUDY subset analysis.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. 9056-9056 ◽  
Author(s):  
Hiroe Kayatani ◽  
Keisuke Aoe ◽  
Kadoaki Ohashi ◽  
Hiroshige Yoshioka ◽  
Akihiro Bessho ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor Receptor ◽  
Egfr Mutations ◽  
Her2 Protein ◽  
Epidermal Growth ◽  
Tki Treatment ◽  
Egfr Mutant ◽  
Egfr Mutated ◽  
Egfr Tki ◽  
Egfr Tkis

9056 Background: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are a key treatment for EGFR-mutated non-small-cell lung carcinoma (NSCLC). To date, a biomarker to predict whether NSCLC will exhibit a short- or long-term response to first- or second-generation EGFR-TKIs has not been established for clinical use. Human epidermal growth factor receptor-2 (HER2) aberrations are mechanisms for acquired resistance to EGFR-TKIs; however, their impact on EGFR-TKI therapy outcomes in EGFR-mutant NSCLC has not yet been systematically evaluated. Methods: Patients with advanced NSCLC were prospectively registered from more than 35 institutes (HER2-CS STUDY UMIN 000017003). EGFR mutations or anaplastic lymphoma kinase gene translocations were assessed at each institution using a commercially approved test. HER2 protein expression levels were determined by immunohistochemistry (IHC) using the Ventana I-VIEW PATHWAY anti-HER-2/neu (4B5). The IHC status scoring system applied to gastric cancer was used. Results: Of 1,126 screened patients with NSCLC, 354 (31.8%) had EGFR-mutated tumors, and the HER2 protein statuses were as follows: IHC0 (n = 71, 26%), IHC1+ (n = 148, 53%), IHC2+ (n = 51, 18%), and IHC3+ (n = 7, 3%). The patients’ demographics were almost identical in those with lung tumors harboring EGFR mutations and HER2-IHC2+/3+ (group P) or EGFR mutations and HER2-IHC0/1 (group N). The EGFR-TKI response rates were not different between these groups (Table). However, group P showed significantly shorter time to EGFR-TKI treatment failure than group N (median 19.1 vs. 13.3 months; log rank p = 0.038). Conclusions: These data from a large prospective cohort show that HER2 protein expression in EGFR-mutant NSCLC may have a negative impact on the effect of EGFR-TKIs. A clinical trial of EGFR/HER2-TKIs (e.g., afatinib) is warranted for this population. [Table: see text]

Concurrent somatic alterations of TP53 and RB1 in Chinese lung adenocarcinoma with or without EGFR mutations.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e21526-e21526
Author(s):  
JUN WANG ◽  
Gang Chen ◽  
Bicheng Zhang ◽  
Jianguo Sun ◽  
Jing Liang ◽  
...  
Keyword(s):  
Lung Adenocarcinoma ◽  
Kinase Inhibitor ◽  
Genetic Alterations ◽  
Egfr Mutations ◽  
Wild Type ◽  
Mutational Status ◽  
Significant Difference ◽  
Somatic Alterations ◽  
Egfr Mutant ◽  
Mutant Egfr

e21526 Background: Endothelial growth factor receptor (EGFR)-mutant lung adenocarcinoma displays diverse responses to tyrosine kinase inhibitor therapy. Concurrent somatic alterations represent different biological substantial proportion of patients with EGFR mutations and impacts their prognosis. Methods: We conducted this retrospective study to clarify the comprehensive concurrent genetic alterations of TP53 and RB1 and tumor mutational burden (TMB) in 712 EGFR-mutant or EGFR-wild type lung adenocarcinoma by next generation sequencing-based gene panel tests. Results: EGFR was the most frequently mutated gene altered in 58.0% (413/712) of lung adenocarcinoma, followed by TP53 altered in 56.7%, KRAS altered in 13.3%, and RBM10 altered in 11.2% of all patients. Concurrent genetic alteration of TP53 and RB1 is more likely to be found in EGFR-mutant patients than in EGFR-wild type patients, with a frequency of 11.4% (45/413) and 4.3% (13/299), respectively (p = 0.003). However, the frequency of TP53 and RB1 concurrent alteration was similar in lung cancer with sensitive EGFR mutation compared to those with non-sensitive mutation (10.5% versus 11.6%). TP53 mutation was most frequently found in patients with RB1 mutation (58/61), irrespective of EGFR mutational status. Furthermore, significant difference was found regarding median TMB in patients with mutant EGFR compared to those with non-mutant EGFR (3.8 versus 4.3; p < 0.001). Median TMB was higher for patients with TP53 and RB1 concurrent alteration than those without concurrent alteration in all patients (5.4 versus 4.3, p = 0.018). Conclusions: Our data showed that high prevalence of concurrent somatic alterations of TP53 and RB1 genes among lung adenocarcinoma patients with EGFR mutations, which might help understand several key biological processes and develop potential therapeutic strategies.

scholarly journals Extracellular vesicles secreted during blastulation show viability of bovine embryos

Reproduction ◽  
2019 ◽  
Vol 158 (6) ◽  
pp. 477-492 ◽  
Author(s):  
Edwin A Mellisho ◽  
Mario A Briones ◽  
Alejandra E Velásquez ◽  
Joel Cabezas ◽  
Fidel O Castro ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Culture Media ◽  
Differential Expression Analysis ◽  
Binary Logistic Regression ◽  
Small Rna Sequencing ◽  
Bovine Embryos ◽  
Embryo Viability ◽  
Group V ◽  
Mean Size

Extracellular vesicles (EVs) secreted by blastocysts may be clinically relevant, as indicator of embryo viability on in vitro fertilization. We tested if the characteristics of EVs secreted during blastulation are related to embryo viability. Morulae were individually cultured in SOF media depleted of EVs until day 7.5 post IVF. Viable embryos were determined by a system of extended in vitro culture of bovine embryos until day 11 (post-hatching development). Afterward, a retrospective classification of blastocyst and culture media was performed based on blastulation time (early blastulation (EB) or late blastulation (LB)) and post-hatching development at day 11 (viable (V) or non-viable embryo (NV)). A total of 254 blastocysts and their culture media were classified in four groups (V-EB, NV-EB, V-LB, NV-LB). Group V-EB had a larger blastocyst diameter (170.8 μm), higher proportion of good-quality blastocysts (77%) and larger mean size of population of EVs (122.9 nm), although the highest concentration of EVs (5.75 × 109 particles/mL) were in group NV-EB. Furthermore, small RNA sequencing detected two biotypes, miRNA (86–91%) and snoRNA (9–14%), with a total of 182 and 32 respectively. In differential expression analysis of miRNAs between V versus NV blastocysts, there were 12 miRNAs upregulated and 15 miRNAs downregulated. Binary logistic regression was used to construct a non-invasive novel model to select viable embryos, based on a combination of variables of blastocyst morphokinetics and EVs characteristics, the ROC-AUC was 0.853. We concluded that characteristics of EVs secreted during blastulation vary depending on embryo quality.

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