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1. Intratumor heterogeneity and cell secretome promote chemotherapy resistance and progression of colorectal cancer

Author

Julia Källberg, Alexandra Harrison, Valerie March, Santa Bērziņa, Ivan Nemazanyy, Oliver Kepp, Guido Kroemer, Sophie Mouillet-Richard, Pierre Laurent-Puig, Valérie Taly, and Wenjin Xiao

Subjects
Cytology, QH573-671
Abstract

Abstract The major underlying cause for the high mortality rate in colorectal cancer (CRC) relies on its drug resistance, to which intratumor heterogeneity (ITH) contributes substantially. CRC tumors have been reported to comprise heterogeneous populations of cancer cells that can be grouped into 4 consensus molecular subtypes (CMS). However, the impact of inter-cellular interaction between these cellular states on the emergence of drug resistance and CRC progression remains elusive. Here, we explored the interaction between cell lines belonging to the CMS1 (HCT116 and LoVo) and the CMS4 (SW620 and MDST8) in a 3D coculture model, mimicking the ITH of CRC. The spatial distribution of each cell population showed that CMS1 cells had a preference to grow in the center of cocultured spheroids, while CMS4 cells localized at the periphery, in line with observations in tumors from CRC patients. Cocultures of CMS1 and CMS4 cells did not alter cell growth, but significantly sustained the survival of both CMS1 and CMS4 cells in response to the front-line chemotherapeutic agent 5-fluorouracil (5-FU). Mechanistically, the secretome of CMS1 cells exhibited a remarkable protective effect for CMS4 cells against 5-FU treatment, while promoting cellular invasion. Secreted metabolites may be responsible for these effects, as demonstrated by the existence of 5-FU induced metabolomic shifts, as well as by the experimental transfer of the metabolome between CMS1 and CMS4 cells. Overall, our results suggest that the interplay between CMS1 and CMS4 cells stimulates CRC progression and reduces the efficacy of chemotherapy.

Published
2023
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2. Everolimus and plicamycin specifically target chemoresistant colorectal cancer cells of the CMS4 subtype

Author

Jiayin Deng, Ai-Ling Tian, Hui Pan, Allan Sauvat, Marion Leduc, Peng Liu, Liwei Zhao, Shuai Zhang, Hui Chen, Valérie Taly, Pierre Laurent-Puig, Laura Senovilla, Yingqiu Li, Guido Kroemer, and Oliver Kepp

Subjects
Cytology, QH573-671
Abstract

Abstract Colorectal cancers (CRC) can be classified into four consensus molecular subtypes (CMS), among which CMS1 has the best prognosis, contrasting with CMS4 that has the worst outcome. CMS4 CRC is notoriously resistant against therapeutic interventions, as demonstrated by preclinical studies and retrospective clinical observations. Here, we report the finding that two clinically employed agents, everolimus (EVE) and plicamycin (PLI), efficiently target the prototypic CMS4 cell line MDST8. As compared to the prototypic CMS1 cell line LoVo, MDST8 cells treated with EVE or PLI demonstrated stronger cytostatic and cytotoxic effects, increased signs of apoptosis and autophagy, as well as a more pronounced inhibition of DNA-to-RNA transcription and RNA-to-protein translation. Moreover, nontoxic doses of EVE and PLI induced the shrinkage of MDST8 tumors in mice, yet had only minor tumor growth-reducing effects on LoVo tumors. Altogether, these results suggest that EVE and PLI should be evaluated for their clinical activity against CMS4 CRC.

Published
2021
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3. Association of high SARS-CoV-2 RNAemia with diabetes and mortality in critically ill COVID-19 patients

Author

Mehran Monchi, Thomas Bruneau, Sebastien Jochmans, David Veyer, Aurelia Pitsch, Olivier Ellrodt, Marie Picque, Valérie Taly, Oumar Sy, Sandie Mazerand, Sylvain Diamantis, and Hélène Péré

Subjects
Clinical medicine, Virology, Science
Abstract

Summary: It has been suggested that during the period of respiratory worsening of severe COVID-19 patients, viral replication plays a less important role than inflammation. Using the droplet-based digital PCR (ddPCR) for precise quantification of plasma SARS-CoV-2 viral load (SARS-CoV-2 RNAemia), we investigated the relationship between plasma viral load, comorbidities, and mortality of 122 critically ill COVID-19 patients. SARS-CoV-2 RNAemia was detected by ddPCR in 90 (74%) patients, ranging from 70 to 213,152 copies per mL. A high (>1 000 copies/ml) or very high (>10,000 copies/ml) SARS-Cov-2 RNAemia was observed in 46 patients (38%), of which 26 were diabetic. Diabetes was independently associated with a higher SARS-CoV-2 RNAemia. In multivariable logistic regression models, SARS-CoV-2 RNAemia was strongly and independently associated with day-60 mortality. Early initiation of antiviral therapies might be considered in COVID-19 critically ill patients with high RNAemia.

Published
2022
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4. High throughput single cell counting in droplet-based microfluidics

Author

Heng Lu, Ouriel Caen, Jeremy Vrignon, Eleonora Zonta, Zakaria El Harrak, Philippe Nizard, Jean-Christophe Baret, and Valérie Taly

Subjects
Medicine, Science
Abstract

Abstract Droplet-based microfluidics is extensively and increasingly used for high-throughput single-cell studies. However, the accuracy of the cell counting method directly impacts the robustness of such studies. We describe here a simple and precise method to accurately count a large number of adherent and non-adherent human cells as well as bacteria. Our microfluidic hemocytometer provides statistically relevant data on large populations of cells at a high-throughput, used to characterize cell encapsulation and cell viability during incubation in droplets.

Published
2017
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5. Highly sensitive methods are required to detect mutations in histiocytoses

Author

Sarah Melloul, Zofia Hélias-Rodzewicz, Fleur Cohen-Aubart, Frédéric Charlotte, Sylvie Fraitag, Nathalie Terrones, Quentin Riller, Thibaud Chazal, Sébastien Héritier, Anne Moreau, Marianne Kambouchner, Marie Christine Copin, Jean Donadieu, Valérie Taly, Zahir Amoura, Julien Haroche, and Jean François Emile

Subjects
Diseases of the blood and blood-forming organs, RC633-647.5
Published
2019
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7. Base-Position Error Rate Analysis of Next-Generation Sequencing Applied to Circulating Tumor DNA in Non-Small Cell Lung Cancer: A Prospective Study.

Author

Nicolas Pécuchet, Eleonora Zonta, Audrey Didelot, Pierre Combe, Constance Thibault, Laure Gibault, Camille Lours, Yves Rozenholc, Valérie Taly, Pierre Laurent-Puig, Hélène Blons, and Elizabeth Fabre

Subjects
Medicine
Abstract

BackgroundCirculating tumor DNA (ctDNA) is an approved noninvasive biomarker to test for the presence of EGFR mutations at diagnosis or recurrence of lung cancer. However, studies evaluating ctDNA as a noninvasive "real-time" biomarker to provide prognostic and predictive information in treatment monitoring have given inconsistent results, mainly due to methodological differences. We have recently validated a next-generation sequencing (NGS) approach to detect ctDNA. Using this new approach, we evaluated the clinical usefulness of ctDNA monitoring in a prospective observational series of patients with non-small cell lung cancer (NSCLC).Methods and findingsWe recruited 124 patients with newly diagnosed advanced NSCLC for ctDNA monitoring. The primary objective was to analyze the prognostic value of baseline ctDNA on overall survival. ctDNA was assessed by ultra-deep targeted NGS using our dedicated variant caller algorithm. Common mutations were validated by digital PCR. Out of the 109 patients with at least one follow-up marker mutation, plasma samples were contributive at baseline (n = 105), at first evaluation (n = 85), and at tumor progression (n = 66). We found that the presence of ctDNA at baseline was an independent marker of poor prognosis, with a median overall survival of 13.6 versus 21.5 mo (adjusted hazard ratio [HR] 1.82, 95% CI 1.01-3.55, p = 0.045) and a median progression-free survival of 4.9 versus 10.4 mo (adjusted HR 2.14, 95% CI 1.30-3.67, p = 0.002). It was also related to the presence of bone and liver metastasis. At first evaluation (E1) after treatment initiation, residual ctDNA was an early predictor of treatment benefit as judged by best radiological response and progression-free survival. Finally, negative ctDNA at E1 was associated with overall survival independently of Response Evaluation Criteria in Solid Tumors (RECIST) (HR 3.27, 95% CI 1.66-6.40, p < 0.001). Study population heterogeneity, over-representation of EGFR-mutated patients, and heterogeneous treatment types might limit the conclusions of this study, which require future validation in independent populations.ConclusionsIn this study of patients with newly diagnosed NSCLC, we found that ctDNA detection using targeted NGS was associated with poor prognosis. The heterogeneity of lung cancer molecular alterations, particularly at time of progression, impairs the ability of individual gene testing to accurately detect ctDNA in unselected patients. Further investigations are needed to evaluate the clinical impact of earlier evaluation times at 1 or 2 wk. Supporting clinical decisions, such as early treatment switching based on ctDNA positivity at first evaluation, will require dedicated interventional studies.

Published
2016
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8. Multiplex Detection of Rare Mutations by Picoliter Droplet Based Digital PCR: Sensitivity and Specificity Considerations.

Author

Eleonora Zonta, Fanny Garlan, Nicolas Pécuchet, Karla Perez-Toralla, Ouriel Caen, Coren Milbury, Audrey Didelot, Elizabeth Fabre, Hélène Blons, Pierre Laurent-Puig, and Valérie Taly

Subjects
Medicine, Science
Abstract

In cancer research, the accuracy of the technology used for biomarkers detection is remarkably important. In this context, digital PCR represents a highly sensitive and reproducible method that could serve as an appropriate tool for tumor mutational status analysis. In particular, droplet-based digital PCR approaches have been developed for detection of tumor-specific mutated alleles within plasmatic circulating DNA. Such an approach calls for the development and validation of a very significant quantity of assays, which can be extremely costly and time consuming. Herein, we evaluated assays for the detection and quantification of various mutations occurring in three genes often misregulated in cancers: the epidermal growth factor receptor (EGFR), the v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) and the Tumoral Protein p53 (TP53) genes. In particular, commercial competitive allele-specific TaqMan® PCR (castPCR™) technology, as well as TaqMan® and ZEN™ assays, have been evaluated for EGFR p.L858R, p.T790M, p.L861Q point mutations and in-frame deletions Del19. Specificity and sensitivity have been determined on cell lines DNA, plasmatic circulating DNA of lung cancer patients or Horizon Diagnostics Reference Standards. To show the multiplexing capabilities of this technology, several multiplex panels for EGFR (several three- and four-plexes) have been developed, offering new "ready-to-use" tests for lung cancer patients.

Published
2016
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9. From Toxins Targeting Ligand Gated Ion Channels to Therapeutic Molecules

Author

Antoine Taly, Adak Nasiripourdori, Valérie Taly, and Thomas Grutter

Subjects
nAChR, P2X, GABA, Glycine, Serotonin, NMDA, AMPA, Kainate, Medicine
Abstract

Ligand-gated ion channels (LGIC) play a central role in inter-cellular communication. This key function has two consequences: (i) these receptor channels are major targets for drug discovery because of their potential involvement in numerous human brain diseases; (ii) they are often found to be the target of plant and animal toxins. Together this makes toxin/receptor interactions important to drug discovery projects. Therefore, toxins acting on LGIC are presented and their current/potential therapeutic uses highlighted.

Published
2011
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10. Data from Plasma Circulating Tumor DNA in Pancreatic Cancer Patients Is a Prognostic Marker

Author

Jean-Baptiste Bachet, Pierre Laurent-Puig, Valérie Taly, Jean-Christophe Vaillant, Mehdi Karoui, Francoise Imbert-Bismut, Solène Doat, Olivier Dubreuil, Audrey Didelot, Fanny Garlan, Nicolas Pécuchet, and Daniel Pietrasz

Abstract

Purpose: Despite recent therapeutic advances, prognosis of patients with pancreatic adenocarcinoma remains poor. Analyses from tumor tissues present limitations; identification of informative marker from blood might be a promising alternative. The aim of this study was to assess the feasibility and the prognostic value of circulating tumor DNA (ctDNA) in pancreatic adenocarcinoma.Experimental Design: From 2011 to 2015, blood samples were prospectively collected from all consecutive patients with pancreatic adenocarcinoma treated in our center. Identification of ctDNA was done with next-generation sequencing targeted on referenced mutations in pancreatic adenocarcinoma and with picoliter droplet digital PCR.Results: A total of 135 patients with resectable (n = 31; 23%), locally advanced (n = 36; 27%), or metastatic (n = 68; 50%) pancreatic adenocarcinoma were included. In patients with advanced pancreatic adenocarcinoma (n = 104), 48% (n = 50) had ctDNA detectable with a median mutation allelic frequency (MAF) of 6.1%. The presence of ctDNA was strongly correlated with poor overall survival (OS; 6.5 vs. 19.0 months; P < 0.001) in univariate and multivariate analyses (HR = 1.96; P = 0.007). To evaluate the impact of ctDNA level, patients were grouped according to MAF tertiles: OS were 18.9, 7.8, and 4.9 months (P < 0.001). Among patients who had curative intent resection (n = 31), 6 had ctDNA detectable after surgery, with an MAF of 4.4%. The presence of ctDNA was associated with a shorter disease-free survival (4.6 vs.17.6 months; P = 0.03) and shorter OS (19.3 vs. 32.2 months; P = 0.027).Conclusions: ctDNA is an independent prognostic marker in advanced pancreatic adenocarcinoma. Furthermore, it arises as an indicator of shorter disease-free survival in resected patients when detected after surgery. Clin Cancer Res; 23(1); 116–23. ©2016 AACR.

Published
2023

12. Supplementary Data File 2 from Plasma Circulating Tumor DNA in Pancreatic Cancer Patients Is a Prognostic Marker

Author

Jean-Baptiste Bachet, Pierre Laurent-Puig, Valérie Taly, Jean-Christophe Vaillant, Mehdi Karoui, Francoise Imbert-Bismut, Solène Doat, Olivier Dubreuil, Audrey Didelot, Fanny Garlan, Nicolas Pécuchet, and Daniel Pietrasz

Abstract

Correlation between Mutation Allelic Frequency (MAF) according to Next Generation Sequencing and picoliter droplet based digital PCR.

Published
2023

13. Data from Intratumor CMS Heterogeneity Impacts Patient Prognosis in Localized Colon Cancer

Author

Pierre Laurent-Puig, Aurélien de Reyniès, Jean-François Emile, Valérie Boige, Sophie Mouillet-Richard, Janick Selves, Audrey Rapinat, David Gentien, Valérie Taly, Hélène Blons, Alex Duval, Daniela Aust, Ramon Salazar, Côme Lepage, Karine Le Malicot, Camilla Pilati, Mira Ayadi, Julien Taieb, Yuna Blum, and Laetitia Marisa

Abstract

Purpose:The consensus molecular subtypes (CMS) represent a significant advance in the understanding of intertumor heterogeneity in colon cancer. Intratumor heterogeneity (ITH) is the new frontier for refining prognostication and understanding treatment resistance. This study aims at deciphering the transcriptomic ITH of colon cancer and understanding its potential prognostic implications.Experimental Design:We deconvoluted the transcriptomic profiles of 1,779 tumors from the PETACC8 trial and 155 colon cancer cell lines as weighted sums of the four CMSs, using the Weighted In Silico Pathology (WISP) algorithm. We assigned to each tumor and cell line a combination of up to three CMS subtypes with a threshold above 20%.Results:Over 55% of tumors corresponded to mixtures of at least two CMSs, demonstrating pervasive ITH in colon cancer. Of note, ITH was associated with shorter disease-free survival (DFS) and overall survival, [HR, 1.34; 95% confidence interval (CI; 1.12–1.59), 1.40, 95% CI (1.14–1.71), respectively]. Moreover, we uncovered specific combinations of CMS associated with dismal prognosis. In multivariate analysis, ITH represents the third parameter explaining DFS variance, after T and N stages. At a cellular level, combined WISP and single-cell transcriptomic analysis revealed that most colon cancer cell lines are a mixture of cells falling into different CMSs, indicating that ITH may correspond to distinct functional statuses of colon cancer cells.Conclusions:This study shows that CMS-based transcriptomic ITH is frequent in colon cancer and impacts its prognosis. CMS-based transcriptomic ITH may correspond to distinct functional statuses of colon cancer cells, suggesting plasticity between CMS-related cell populations. Transcriptomic ITH deserves further assessment in the context of personalized medicine.

Published
2023

14. Data from Prognostic Value and Relation with Adjuvant Treatment Duration of ctDNA in Stage III Colon Cancer: a Post Hoc Analysis of the PRODIGE-GERCOR IDEA-France Trial

Author

Pierre Laurent-Puig, Thierry André, Dewi Vernerey, Aimery de Gramont, Thibault Mazard, Kariman Chaba, Katia Hormigos, Claire Mulot, Olivier Bouche, Joëlle Egreteau, May Mabro, Céline Lepere, Christophe Louvet, Jérôme Desrame, Jaafar Bennouna, Laurent Mineur, Camille Bourreau, Julie Henriques, Valérie Taly, and Julien Taieb

Abstract

Purpose:Circulating tumor DNA (ctDNA) has been suggested as a major prognostic factor in resected stage-III colon cancer. We analyzed ctDNA of patients randomized in the phase III IDEA-France trial.Experimental Design:ctDNA was tested for WIF1 and NPY by droplet digital PCR with method developed and validated for colorectal cancer. Disease-free survival (DFS) and overall survival (OS) were analyzed via multivariable analysis in patients with ctDNA samples and in sub-groups according to treatment duration (3/6 months) and disease stage (high/low-risk stage III).Results:Of 2,010 randomized patients, 1,345 had available ctDNA samples (1,017 collected both post-surgery and pre-chemotherapy). More Eastern Cooperative Oncology Group performance status (ECOG PS) of 0 (78% versus 69%) and T4 and/or N2 (40% versus 36%) were observed in patients studied (n = 1017) versus not analyzed (n = 993). There were 877 ctDNA-negative (86.2%) and 140 ctDNA-positive (13.8%) patients; their baseline characteristics were similar. With a median follow-up of 6.6 years, the 3-year DFS rate was 66.39% for ctDNA-positive patients and 76.71% for ctDNA-negative patients (P = 0.015). ctDNA was confirmed as an independent prognostic marker for DFS (adjusted HR = 1.55, 95% CI 1.13–2.12, P = 0.006) and OS (HR = 1.65, 95% CI 1.12–2.43, P = 0.011). ctDNA was prognostic in patients treated for 3 months and with T4 and/or N2 tumors, but not in those treated for 6 months and with T1–3/N1 tumors.Conclusions:In this first ctDNA assessment of a large series of patients with stage III colon cancer enrolled in phase III trial, post-surgery ctDNA was found in 13.8% of them and was confirmed as an independent prognostic marker.See related commentary by Bent and Kopetz, p. 5449

Published
2023

18. Supplementary Data from Prognostic Value and Relation with Adjuvant Treatment Duration of ctDNA in Stage III Colon Cancer: a Post Hoc Analysis of the PRODIGE-GERCOR IDEA-France Trial

Author

Pierre Laurent-Puig, Thierry André, Dewi Vernerey, Aimery de Gramont, Thibault Mazard, Kariman Chaba, Katia Hormigos, Claire Mulot, Olivier Bouche, Joëlle Egreteau, May Mabro, Céline Lepere, Christophe Louvet, Jérôme Desrame, Jaafar Bennouna, Laurent Mineur, Camille Bourreau, Julie Henriques, Valérie Taly, and Julien Taieb

Abstract

Supplementary Data

Published
2023

19. 2022-RA-1480-ESGO TRANSLACOL project: digital-PCR human papilloma virus (HPV) detection for recurrence prediction in early cervical cancer patients without pelvic lymph node invasion

Author

Rosa Montero Macías, Nicolas Robillard, Thomas Bruneau, Coronado Pluvio, Mélanie Boulhic, Marie-Aude Le Frére-Belda, Ivana Stankovic, Martina Aida Angeles, Eliane Mery, Pascal Rigolet, Cecile Badoual, Patrice Mathevet, Anne-Sophie Bats, Valérie Taly, David Veyer, Fabrice Lecuru, and Helene Pere

Published
2022

20. Early detection of pancreatic cancer by liquid biopsy 'PANLIPSY': a french nation-wide study project

Author

Thomas Bardol, Antoine M. Dujon, Valerie Taly, Catherine Dunyach-Remy, Jean-Philippe Lavigne, Bruno Costa-Silva, Keerthi Kurma, Zahra Eslami-S, Laure Cayrefourcq, Cindy Canivet, Fabrice Muscari, Barbara Bournet, and Catherine Alix-Panabières

Subjects
Liquid biopsy, Pancreatic cancer, Circulating tumor cells, Circulating tumor DNA, Exosomes, Multi-omic approach, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, RC254-282
Abstract

Abstract Background Pancreatic cancer, predominantly characterized by ductal adenocarcinoma (PDAC) accounts for 90% of cases and is the fourth leading cause of cancer-related deaths globally. Its incidence is notably increasing. This poor prognosis is primarily due to late-stage diagnosis (approximately 70% to 80% of patients are diagnosed at an advanced stage), aggressive tumor biology, and low sensitivity to chemotherapy. Consequently, it is crucial to identify and develop a simple, feasible and reproducible blood-based signature (i.e., combination of biomarkers) for early detection of PDAC. Methods The PANLIPSY study is a multi-center, non-interventional prospective clinical trial designed to achieve early detection of PDAC with high specificity and sensitivity, using a combinatorial approach in blood samples. These samples are collected from patients with resectable, borderline or locally advanced, and metastatic stage PDAC within the framework of the French Biological and Clinical Database for PDAC cohort (BACAP 2). All partners of the BACAP consortium are eligible to participate. The study will include 215 PDAC patients, plus 25 patients with benign pancreatic conditions from the PAncreatic Disease Cohort of TOuLouse (PACTOL) cohort, and 115 healthy controls, totaling 355 individuals. Circulating biomarkers will be collected in a total volume of 50 mL of blood, divided into one CellSave tube (10 mL), two CELL-FREE DNA BCT® preservative tubes (18 mL), and five EDTA tubes (22 mL in total). Samples preparation will adhere to the guidelines of the European Liquid Biopsy Society (ELBS). A unique feature of the study is the AI-based comparison of these complementary liquid biopsy biomarkers. Main end-points: i) to define a liquid biopsy signature that includes the most relevant circulating biomarkers, ii) to validate the multi-marker panel in an independent cohort of healthy controls and patients, with resectable PDAC, and iii) to establish a unique liquid biopsy biobank for PDAC study. Discussion The PANLIPSY study is a unique prospective non-interventional clinical trial that brings together liquid biopsy experts. The aim is to develop a biological signature for the early detection of PDAC based on AI-assisted detection of circulating biomarkers in blood samples (CTCs, ctDNA, EVs, circulating immune system, circulating cell-free nucleosomes, proteins, and microbiota). Trial registration ClinicalTrials.gov Identifier: NCT06128343 / NCT05824403. Registration dates: June 8,2023 and April 21, 2023.

Published
2024
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21. TRANSLACOL project: Nodal human papillomavirus tumoral DNA detection by ddPCR for survival prediction in early cervical cancer patients without pelvic lymph node invasion

Author

Rosa Montero-Macías, David Veyer, Thomas Bruneau, Nicolas Robillard, Marie-Aude Le Frère-Belda, Pascal Rigolet, Mélanie Boulhic, Ivana Stankovic, Laurent Bélec, Martina Aida Angeles, Eliane Mery, Cécile Badoual, Pluvio J Coronado, Anne-Sophie Bats, Patrice Mathevet, Valérie Taly, Fabrice Lécuru, and Hélène Péré

Subjects
Infectious Diseases, Virology
Published
2023

22. Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma

Author

Guillaume Beinse, Bruno Borghese, Marie Métairie, Pierre-Alexandre Just, Geoffroy Poulet, Simon Garinet, Beatrice Parfait, Audrey Didelot, Camille Bourreau, Natacha Agueeff, Alexandre Lavollé, Benoit Terris, Charles Chapron, François Goldwasser, Karen Leroy, Helene Blons, Pierre Laurent-Puig, Valérie Taly, Jérôme Alexandre, Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Cancer Research and Personalized Medicine - CARPEM [Paris], Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Prévention et traitement de la perte protéique musculaire en situation de résistance à l'anabolisme (PRETRRAM (URP_4466)), Université Paris Cité (UPCité), TALY, Valerie, and Laurent-Puig, Pierre

Subjects
DNA methylation, Biochemistry (medical), Clinical Biochemistry, endometrial carcinoma, [SDV.CAN]Life Sciences [q-bio]/Cancer, personalized medicine, Polymerase Chain Reaction, Circulating Tumor DNA, Endometrial Neoplasms, molecular characterization, [SDV.CAN] Life Sciences [q-bio]/Cancer, Biomarkers, Tumor, Humans, Female, Prospective Studies, digital droplet PCR, Retrospective Studies
Abstract

Background No circulating biomarker is available for endometrial carcinoma (EC). We aimed to identify DNA positions universally hypermethylated in EC, and to develop a digital droplet PCR (ddPCR) assay for detection of hypermethylated circulating tumor DNA (meth-ctDNA) in plasma from patients with EC. Methods DNA positions hypermethylated in EC, and without unspecific hypermethylation in tissue/cell types releasing circulating cell-free DNA in plasma, were identified in silico from TCGA/Gene Expression Omnibus (GEO) data. A methylation-specific ddPCR (meth-ddPCR) assay following bisulfite conversion of DNA extracted from plasma was optimized for detection of meth-ctDNA according to dMIQE guidelines. Performances were validated on a retrospective cohort (n = 78 tumors, n = 30 tumor-adjacent tissues), a prospective pilot cohort (n = 33 stage I–IV patients), and 55 patients/donors without cancer. Results Hypermethylation of zinc finger and SCAN domain containing 12 (ZSCAN12) and/or oxytocin (OXT) classified EC samples from multiple noncancer samples with high diagnostic specificity/sensitivity [>97%; area under the curve (AUC) = 0.99; TCGA/GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC = 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank = 2) and sensitivity (absolute lower threshold of detection = 50 pgmethDNA/mLplasma). In the pilot cohort, meth-ctDNA was detected in pretreatment plasma samples from 9/11 and 5/20 patients with advanced and non-advanced EC, respectively. 2 of 9 patients had ctDNA detected after macroscopic complete surgery and experienced progression within 6 months. No healthy donors had any copy of hypermethylated DNA detected in plasma. Conclusions Meth-ddPCR of ZSCAN12/OXT allows a highly specific and sensitive detection of ctDNA in plasma from patients with EC and appears promising for personalized approaches for these patients.

Published
2022

23. Prognostic value of circulating tumour DNA in metastatic pancreatic cancer patients: post-hoc analyses of two clinical trials

Author

Aurélien de Reyniès, Claire Mulot, Laetitia Dahan, Jean-Baptiste Bachet, Shu-Fang Wang-Renault, Valérie Taly, Jean-Marc Phelip, Pierre Laurent-Puig, Daniel Pietrasz, Yves Rinaldi, Julien Taieb, Jerome Durand-Labrunie, Hélène Blons, Solene Doat, Mathilde Postel, Karine Le Malicot, Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Centre Hépato-Biliaire [Hôpital Paul Brousse] (CHB), Hôpital Paul Brousse-Assistance Publique - Hôpitaux de Paris, Cancer Research and Personalized Medicine - CARPEM [Paris], Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hôpital de la Timone [CHU - APHM] (TIMONE), Fédération Francophone de Cancérologie Digestive (FFCD), Equipe EPICAD (LNC - U1231), Lipides - Nutrition - Cancer [Dijon - U1231] (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Agro Dijon, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Agro Dijon, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Epigenetec, Centre de Ressources Biologiques (CRB), Hôpital Européen [Fondation Ambroise Paré - Marseille], CHU Saint-Etienne, CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Université Pierre et Marie Curie - Paris 6 (UPMC), (le programme) Cartes d'identité des tumeurs (CIT), Ligue Nationales Contre le Cancer (LNCC), and TALY, Valerie

Subjects
Male, Oncology, Cancer Research, medicine.medical_specialty, Multivariate analysis, Post hoc, medicine.medical_treatment, [SDV.CAN]Life Sciences [q-bio]/Cancer, Article, Circulating Tumor DNA, [SDV.CAN] Life Sciences [q-bio]/Cancer, Internal medicine, Metastatic pancreatic cancer, Biomarkers, Tumor, Humans, Medicine, Prospective Studies, Neoplasm Metastasis, Aged, Chemotherapy, Plasma samples, business.industry, Hazard ratio, DNA Methylation, Middle Aged, Metastatic Pancreatic Adenocarcinoma, Prognosis, Pancreatic Neoplasms, Survival Rate, Clinical trial, Mutation, Female, business
Abstract

International audience; Objective: The prognostication of metastatic pancreatic adenocarcinoma (mPDAC) patients remains uncertain, mainly based on carbohydrate antigen 19-9 (CA19-9), with limited utility. Circulating tumour DNA (ctDNA) has been suggested as a prognostic factor, but its added value has been poorly explored. The objective was to determine whether ctDNA is an independent factor for the prognostication of mPDAC.Design: Translational study based on two prospective collections of plasma samples of mPDAC patients naïve for chemotherapy. One used as a test series and the other as validation series coming from two randomised trials (Prodige 35 and Prodige 37). CtDNA was assessed by digital droplet PCR targeting two methylated markers (HOXD8 and POU4F1) according to a newly developed and validated method. Univariate and multivariate analyses were performed according to ctDNA status.Results: Of 372 plasma samples available, 354 patients were analyzed for survival. In the validation series, 145 of 255 patients were found ctDNA positive (56.8%), Median PFS and OS were 5.3 and 8.2 months in ctDNA-positive and 6.2 and 12.6 months in ctDNA-negative patients, respectively. ctDNA positivity was more often associated with young age, high CA19-9 level and neutrophils lymphocytes ratio. In multivariate analysis including these previous markers, ctDNA was confirmed as an independent prognostic marker for PFS (adjusted hazard ratio (HR) 1.5, CI 95% [1.03-2.18], p = 0.034) and OS (HR 1.62, CI 95% [1.05-2.5], p = 0.029).Conclusions: In this first ctDNA assessment in a large series of mPDAC derived from clinical trials, ctDNA was detectable in 56.8% of patients and confirmed as an independent prognostic marker.

Published
2022

24. BRCA1 and RAD51C promotor methylation in human resectable pancreatic adenocarcinoma

Author

Raëf Abdallah, Shulin Zhao, Simon Garinet, Katia Hormigos, Delphine Le Corre, Jérôme Cros, Karla Perez Toralla, Anne Sophie Bats, Jérémy Augustin, Jean-Baptiste Bachet, Valérie Taly, Hélène Blons, Julien Taieb, Pierre Laurent-Puig, Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Hôpital Beaujon [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), and TALY, Valerie

Subjects
BRCA1 promotor, Hepatology, BRCA1 Protein, Gastroenterology, [SDV.CAN]Life Sciences [q-bio]/Cancer, Triple Negative Breast Neoplasms, Adenocarcinoma, DNA Methylation, DNA-Binding Proteins, Pancreatic Neoplasms, [SDV.CAN] Life Sciences [q-bio]/Cancer, Humans, Homologous recombination, Promoter Regions, Genetic, PARP inhibitors, Pancreatic adenocarcinoma, RAD51C promotor
Abstract

International audience; Background: Homozygous Recombination Deficiency (HRD) is associated with sensitivity to PARP-inhibitors (PARPi) in different cancer types. In pancreatic adenocarcinoma (PA) the main cause of HRD is BRCA1/2 germline mutation and patients with mutations in BRCA1/2 may benefit from PARPi. Recently other mechanisms leading to HRD were described in different cancer types, including gene mutations and epigenetic changes such as promoter hypermethylation. In PA, BRCA1 promoter hypermethylation, a known mechanism of gene silencing, was recently described. However, results are discordant between North American studies (0.7% of PA) and Asian ones (up to 60% of PA) and the association with HRD is not clear.Methods: Here, we developed 2 quantifications methods to explore BRCA1 and RAD51C promoter methylation in a series of 121 Formalin Fixed-Paraffin-Embedded (FFPE) specimens from resected PA without neoadjuvant treatment. The methylation-specific PCR was done with 2 different methods after DNA bisulfite conversion: a digital droplet PCR, and a PCR followed by capillary electrophoresis, to score the methylated / non methylated ratios in tumor samples. Methods were validated for specificity and sensibility using 100, 20, 10, 5 and 0% methylated commercial DNA for fragment analysis with a detection cutoff of 5-10%. Limit of blank was defined as 5 dropplets/20µL for RAD51C and 1 dropplet/20µL for BRCA1 for ddPCR. Samples were reviewed by a pathologist, macrodissected before DNA extraction to obtain 50-60% of tumoral cells. DNAs were treated for bisulfite conversion and analyzed using both methods in parallel to known positive and negative controls in each run.Results and conclusion: No methylation at BRCA1 or RAD51C was found in this series of PA suggesting that HRD gene promoter methylation is a rare event in European patients.

Published
2021

25. Prognostic Value and Relation with Adjuvant Treatment Duration of ctDNA in Stage III Colon Cancer: a Post Hoc Analysis of the PRODIGE-GERCOR IDEA-France Trial

Author

Pierre Laurent-Puig, May Mabro, Valérie Taly, Jaafar Bennouna, Claire Mulot, Julie Henriques, Olivier Bouché, Jérôme Desramé, Joëlle Egreteau, Thibault Mazard, C. Lepere, Laurent Mineur, Christophe Louvet, Dewi Vernerey, Camille Bourreau, Kariman Chaba, Aimery de Gramont, Katia Hormigos, Julien Taieb, Thierry André, Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Cancer Research and Personalized Medicine - CARPEM [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hôpital Saint-Jacques [CHRU de Besançon], Interactions hôte-greffon-tumeur, ingénierie cellulaire et génique - UFC (UMR INSERM 1098) (RIGHT), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté]), Institut Sainte Catherine [Avignon], Centre hospitalier universitaire de Nantes (CHU Nantes), Hôpital privé Jean-Mermoz [Lyon] (Ramsay-GDS), Institut Mutualiste de Montsouris (IMM), Hôpital Foch [Suresnes], Groupe Hospitalier Bretagne Sud (GHBS), Hôpital Robert Debré, Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Institut hospitalier Franco-Britannique [Levallois-Perret], CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), TALY, Valerie, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])-Université de Franche-Comté (UFC), and Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)

Subjects
Cancer Research, medicine.medical_specialty, Colorectal cancer, medicine.medical_treatment, Treatment duration, [SDV]Life Sciences [q-bio], Gastroenterology, 03 medical and health sciences, 0302 clinical medicine, Internal medicine, Post-hoc analysis, medicine, In patient, Stage (cooking), 030304 developmental biology, 0303 health sciences, business.industry, medicine.disease, 3. Good health, Stage III Colon Cancer, [SDV] Life Sciences [q-bio], Oncology, 030220 oncology & carcinogenesis, Baseline characteristics, business, Adjuvant
Abstract

Purpose:Circulating tumor DNA (ctDNA) has been suggested as a major prognostic factor in resected stage-III colon cancer. We analyzed ctDNA of patients randomized in the phase III IDEA-France trial.Experimental Design:ctDNA was tested for WIF1 and NPY by droplet digital PCR with method developed and validated for colorectal cancer. Disease-free survival (DFS) and overall survival (OS) were analyzed via multivariable analysis in patients with ctDNA samples and in sub-groups according to treatment duration (3/6 months) and disease stage (high/low-risk stage III).Results:Of 2,010 randomized patients, 1,345 had available ctDNA samples (1,017 collected both post-surgery and pre-chemotherapy). More Eastern Cooperative Oncology Group performance status (ECOG PS) of 0 (78% versus 69%) and T4 and/or N2 (40% versus 36%) were observed in patients studied (n = 1017) versus not analyzed (n = 993). There were 877 ctDNA-negative (86.2%) and 140 ctDNA-positive (13.8%) patients; their baseline characteristics were similar. With a median follow-up of 6.6 years, the 3-year DFS rate was 66.39% for ctDNA-positive patients and 76.71% for ctDNA-negative patients (P = 0.015). ctDNA was confirmed as an independent prognostic marker for DFS (adjusted HR = 1.55, 95% CI 1.13–2.12, P = 0.006) and OS (HR = 1.65, 95% CI 1.12–2.43, P = 0.011). ctDNA was prognostic in patients treated for 3 months and with T4 and/or N2 tumors, but not in those treated for 6 months and with T1–3/N1 tumors.Conclusions:In this first ctDNA assessment of a large series of patients with stage III colon cancer enrolled in phase III trial, post-surgery ctDNA was found in 13.8% of them and was confirmed as an independent prognostic marker.See related commentary by Bent and Kopetz, p. 5449

Published
2021

26. 1057 Development and validation of a machine-learning-derived RNAseq prognostic signature in endometrial cancer

Author

S Garinet, François Goldwasser, Meriem Koual, Cécile Badoual, N Bekmezian, Valérie Taly, Guillaume Beinse, Bruno Borghese, Jérôme Alexandre, Nicolas Delanoy, M. A. Le Frere Belda, C Gervais, Benoit Terris, J Pierre-Alexandre, Karen Leroy, Pierre Laurent-Puig, Charles Chapron, Hélène Blons, Anne-Sophie Bats, and Catherine Durdux

Subjects
Oncology, medicine.medical_specialty, Receiver operating characteristic, business.industry, Proportional hazards model, Endometrial cancer, Hazard ratio, medicine.disease, Confidence interval, Internal medicine, Cohort, Carcinoma, Medicine, Stage (cooking), business
Abstract

Introduction/Background* Because of inter-tumor heterogeneity of endometrial carcinoma (EC), prognostication remains challenging. We aimed to develop a RNAseq signature to stratify EC patient prognosis beyond molecular subtyping. Methodology A prognostic signature was identified using a LASSO-penalized Cox regression model on TCGA (N=543 patients). A polyA-RNAseq-based method was developed for validation of the signature in a cohort of stage I-IV EC patients treated in two Paris Hospitals between 2010 and 2017. Model performances were evaluated using time-dependent ROC curves (prediction of disease-specific-survival (DSS)). The additional value of the RNAseq signature was evaluated using uni/multivariable Cox models (hazard ratio (HR) with [95% confidence interval]) and Kaplan-Meier analysis. Result(s)* Among 209 patients included in the validation cohort (median follow-up 55 months IQR [41-69]), 61 (30%), 10 (5%), 52 (25%), and 82 (40%), had mismatch repair-deficient, POLE-mutated, TP53-mutated tumors, and tumors with no specific molecular profile, respectively. The 38-genes signature accurately predicted DSS (AUC=80%). Using a composite classifier accounting for the RNAseq signature and the TP53-mutated group, three groups were identified: good prognosis tumors based on RNAseq signature and without TP53 mutation, characterized by excellent outcome (N=103 patients, 5-years DSS rates of 99%) (reference), poor prognosis tumors based on RNAseq signature and without TP53 mutation (N=49 patients, 5-years DSS rates of 81%; HR: 5.86 [1.16; 29.7]), and TP53-mutated tumors whatever the RNAseq signature (N=52 patients, 5-years DSS rates of 52%; HR: 11.14 [2.40; 51.7]) (HR adjusted on FIGO stage). In 81 (38%) patients with adverse features (2020 ESGO/ESTRO/ESP guidelines: non-endometrioid histology or stage III-IVA or TP53-mutated tumors), TP53-mutated molecular group was not significantly associated with poor prognosis (p=0.18). A The composite classifier identified three classes within this subgroup: RNAseq-good prognosis (N=24), non-TP53/RNAseq-poor prognosis (N=16), and TP53-mutated tumors (N=41), with 5-years DSS rates of 100%, 59%, and 71%, respectively (p=0.015). Transcriptome analyses suggested the underlying involvement of immune deprivation and wound healing processes in tumors with poor prognosis. Conclusion* We demonstrate that RNAseq characterization can refine prognostication in EC beyond molecular subgroups and main prognostic features, and warrants validation for potential RNAseq-based adjuvant therapeutic strategies in EC.

Published
2021

27. Role of Circulating Tumor DNA in Gastrointestinal Cancers: Current Knowledge and Perspectives

Author

Julien Taieb, Aziz Zaanan, Simon Garinet, Valérie Taly, Emilie Moati, P. Laurent-Puig, Audrey Didelot, Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), and Gestionnaire, HAL Sorbonne Université 5

Subjects
Oncology, Cancer Research, medicine.medical_specialty, [SDV.CAN]Life Sciences [q-bio]/Cancer, Review, Somatic evolution in cancer, 03 medical and health sciences, 0302 clinical medicine, [SDV.CAN] Life Sciences [q-bio]/Cancer, Internal medicine, medicine, Adjuvant therapy, gastrointestinal cancers, RC254-282, 030304 developmental biology, circulating tumor DNA, 0303 health sciences, business.industry, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, personalized medicine, Response to treatment, Minimal residual disease, 3. Good health, 13. Climate action, Circulating tumor DNA, 030220 oncology & carcinogenesis, Cancer management, Biomarker (medicine), biomarker, Personalized medicine, business
Abstract

Simple Summary Management of gastrointestinal (GI) cancers t is a worldwide challenge and some new tools are needed to guide it. Circulating tumor DNA (ctDNA) is a fraction of tumor DNA that can be detected by a liquid biopsy through a simple blood sample. In this work, we tried to summarize in a clinical review to what extend the analysis of ctDNA can improve therapeutic support in digestive oncology and how this circulating biomarker appears as a very promising improvement in addition to classic diagnostic, prognostic and theranostic methods. Although the level of evaluation of this tool is still different between the different GI cancers locations, it is in dynamic evolution in all of them. Abstract Gastrointestinal (GI) cancers are major health burdens worldwide and biomarkers are needed to improve the management of these diseases along their evolution. Circulating tumor DNA (ctDNA) is a promising non-invasive blood and other bodily-fluid-based biomarker in cancer management that can help clinicians in various cases for the detection, diagnosis, prognosis, monitoring and personalization of treatment in digestive oncology. In addition to the well-studied prognostic role of ctDNA, the main real-world applications appear to be the assessment of minimal residual disease to further guide adjuvant therapy and predict relapse, but also the monitoring of clonal evolution to tailor treatments in metastatic setting. Other challenges such as predicting response to treatment including immune checkpoint inhibitors could also be among the potential applications of ctDNA. Although the level of advancement of ctDNA development in the different tumor localizations is still inhomogeneous, it might be now reliable enough to be soon used in clinical routine for colorectal cancers and shows promising results in other GI cancers.

Published
2021

28. Intratumor CMS Heterogeneity Impacts Patient Prognosis in Localized Colon Cancer

Author

Valérie Taly, Hélène Blons, Karine Le Malicot, David Gentien, Alex Duval, Julien Taieb, Laetitia Marisa, Jean-François Emile, Aurélien de Reyniès, Ramon Salazar, Mira Ayadi, Daniela Aust, Valérie Boige, Camilla Pilati, Sophie Mouillet-Richard, Janick Selves, Yuna Blum, Audrey Rapinat, Côme Lepage, Pierre Laurent-Puig, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Ligue Nationale Contre le Cancer - Paris, Ligue Nationale Contre le Cancer (LNCC), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Fédération Francophone de la Cancérologie Digestive, FFCD, Université de Bourgogne (UB), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), University Hospital Carl Gustav Carus [Dresden, Germany], Technische Universität Dresden = Dresden University of Technology (TU Dresden), Centre de Recherche Saint-Antoine (CRSA), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Université Paris Descartes - Faculté de Médecine (UPD5 Médecine), Université Paris Descartes - Paris 5 (UPD5), Institut Curie [Paris], Institut Universitaire du Cancer de Toulouse - Oncopole (IUCT Oncopole - UMR 1037), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Gustave Roussy (IGR), Oncologie digestive, Département de médecine oncologique [Gustave Roussy], Institut Gustave Roussy (IGR)-Institut Gustave Roussy (IGR), Biomarqueurs et essais cliniques en Cancérologie et Onco-Hématologie (BECCOH), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Université Paris-Saclay, Service de pathologie [CHU Ambroise Paré], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Ambroise Paré [AP-HP], This study was supported by the program 'Cartes d'Identité des Tumeurs' (CIT) from the Ligue Nationale Contre le Cancer (LNCC), by the Institut National du Cancer (INCa) with the program Heterogeneity of tumors and ecosystem (HTE program) within the sub program 'deciphering the heterogeneous genome-microenvironment interplay in colon and hepatocellular carcinomas (HETCOLI)', by the ITMO CANCER through the program single cell COCAHEMISCLE, by the integrated site of cancer research 'cancer research for personalized medicine' (SIRIC CARPEM, INCa-DGOS-Inserm_12561) and the Fédération Francophone de Cancérologie Digestive (FFCD). We thank the biological resource center EPIGENETEC (BB-0033-00055) for their support., Jonchère, Laurent, Ligue Nationnale Contre le Cancer, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Centre de Recherche Saint-Antoine (CR Saint-Antoine), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-CHU Toulouse [Toulouse]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), (le programme) Cartes d'identité des tumeurs (CIT), Ligue Nationales Contre le Cancer (LNCC), Cancer Research and Personalized Medicine - CARPEM [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Necker - Enfants Malades [AP-HP], Fédération Francophone de Cancérologie Digestive (FFCD), CHU Dijon, Universitat de Barcelona (UB), Centre de Recherches en Cancérologie de Toulouse (CRCT), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), Hôpital Ambroise Paré [AP-HP], and Laurent-Puig, Pierre

Subjects
Oncology, Male, Cancer Research, medicine.medical_specialty, Multivariate analysis, Colorectal cancer, [SDV]Life Sciences [q-bio], Cell, Context (language use), [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, Transcriptome, [SDV.CAN] Life Sciences [q-bio]/Cancer, Internal medicine, Cell Line, Tumor, medicine, Tumor Microenvironment, Humans, Treatment resistance, health care economics and organizations, Aged, business.industry, Gene Expression Profiling, Hazard ratio, medicine.disease, Prognosis, Survival Rate, [SDV] Life Sciences [q-bio], medicine.anatomical_structure, Colonic Neoplasms, Female, Personalized medicine, business
Abstract

Purpose: The consensus molecular subtypes (CMS) represent a significant advance in the understanding of intertumor heterogeneity in colon cancer. Intratumor heterogeneity (ITH) is the new frontier for refining prognostication and understanding treatment resistance. This study aims at deciphering the transcriptomic ITH of colon cancer and understanding its potential prognostic implications. Experimental Design: We deconvoluted the transcriptomic profiles of 1,779 tumors from the PETACC8 trial and 155 colon cancer cell lines as weighted sums of the four CMSs, using the Weighted In Silico Pathology (WISP) algorithm. We assigned to each tumor and cell line a combination of up to three CMS subtypes with a threshold above 20%. Results: Over 55% of tumors corresponded to mixtures of at least two CMSs, demonstrating pervasive ITH in colon cancer. Of note, ITH was associated with shorter disease-free survival (DFS) and overall survival, [HR, 1.34; 95% confidence interval (CI; 1.12–1.59), 1.40, 95% CI (1.14–1.71), respectively]. Moreover, we uncovered specific combinations of CMS associated with dismal prognosis. In multivariate analysis, ITH represents the third parameter explaining DFS variance, after T and N stages. At a cellular level, combined WISP and single-cell transcriptomic analysis revealed that most colon cancer cell lines are a mixture of cells falling into different CMSs, indicating that ITH may correspond to distinct functional statuses of colon cancer cells. Conclusions: This study shows that CMS-based transcriptomic ITH is frequent in colon cancer and impacts its prognosis. CMS-based transcriptomic ITH may correspond to distinct functional statuses of colon cancer cells, suggesting plasticity between CMS-related cell populations. Transcriptomic ITH deserves further assessment in the context of personalized medicine.

Published
2021

29. Detection of Brain Somatic Mutations in Cerebrospinal Fluid from Refractory Epilepsy Patients

Author

Valérie Taly, Mathilde Chipaux, Sara Baldassari, Dong Seok Kim, Won Seok Chang, Georg Dorfmüller, Seyeon Kim, Stéphanie Baulac, Jeong Ho Lee, Nam Suk Sim, Korea Advanced Institute of Science and Technology (KAIST), Institut du Cerveau = Paris Brain Institute (ICM), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Hôpital Rothschild [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Yonsei University College of Medicine [Séoul, Corée du Sud], Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), TALY, Valerie, Institut du Cerveau et de la Moëlle Epinière = Brain and Spine Institute (ICM), Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Pitié-Salpêtrière [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP)

Subjects
0301 basic medicine, Hemimegalencephaly, Mutation, Pathology, medicine.medical_specialty, Somatic cell, business.industry, [SDV]Life Sciences [q-bio], Cortical dysplasia, medicine.disease_cause, medicine.disease, 3. Good health, Ganglioglioma, [SDV] Life Sciences [q-bio], 03 medical and health sciences, Epilepsy, 030104 developmental biology, 0302 clinical medicine, Cerebrospinal fluid, Neurology, medicine, Epilepsy surgery, Neurology (clinical), business, 030217 neurology & neurosurgery
Abstract

Brain mosaic mutations are a major cause of refractory focal epilepsies with cortical malformations such as focal cortical dysplasia, hemimegalencephaly, malformation of cortical development with oligodendroglial hyperplasia in epilepsy, and ganglioglioma. Here, we collected cerebrospinal fluid (CSF) during epilepsy surgery to search for somatic variants in cell-free DNA (cfDNA) using targeted droplet digital polymerase chain reaction. In 3 of 12 epileptic patients with known somatic mutations previously identified in brain tissue, we here provide evidence that brain mosaicism can be detected in the CSF-derived cfDNA. These findings suggest future opportunities for detecting the mutant allele driving epilepsy in CSF. ANN NEUROL 2021;89:1248-1252.

Published
2021

30. Vemurafenib for Refractory Multisystem Langerhans Cell Histiocytosis in Children: An International Observational Study

Author

Johannes Visser, Anne Lutun, Elena Sieni, Fleur Cohen, Nabil Kabbara, Olga Slater, Jean-François Emile, Mathilde Jehanne, Alina Ferster, Michael Maschan, Houda Boudiaf, Pascale Schneider, Sarah Elitzur, Nathalie Aladjidi, Milen Minkov, Ahmed Idbaih, Matthew Collin, Michal Golan, Alexandra Kolenova, François Chalard, Karel Svojgr, Jean Donadieu, Anne Pagnier, Laurence Blanc, Nicolas Simon, Jean-Claude Alvarez, Islam Amine Larabi, Fanette Bernard, James Nicholson, Mohamed Barkaoui, Anne Sonntagbauer, Anne Lambilliote, Thomas Lehrnbecher, Dmitriy Evseev, Viktoria Efremova, Geneviève Plat, Martina Ahlmann, Jean Miron, Valérie Taly, Caroline Hutter, Julien Haroche, Zofia Hélias-Rodzewicz, Paul Milne, Sébastien Héritier, Andrej Lissat, Mathilde Tardieu, Service d'hématologie-immunologie-oncologie pédiatrique [CHU Trousseau], CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Hôpital Raymond Poincaré [AP-HP], Centre Hospitalier Universitaire [Grenoble] (CHU), Cambridge University Hospitals - NHS (CUH), University of Cambridge [UK] (CAM), Medizinische Universität Wien = Medical University of Vienna, Department Pediatric Hematology Oncology, Azienda Ospedaliero-Universitaria Meyer, Florence, Departments of Hematology, Université Paris Diderot - Paris 7 (UPD7), Groupe d'Etude des Histiocytoses (GEH), Groupe d'Etude des Histiocytoses, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Service de Radiologie [CHU Trousseau], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Newcastle University [Newcastle], Centre National de Référence du Lupus Systémique, Syndrome des Anticorps Anti-phospholipides et Maladies Auto-immunes Systémiques Rares [CHU Pitié Salpêtrière], Service de Médecine Interne 2, maladies auto-immunes et systémiques [CHU Pitié-Salpêtrière], Institut E3M [CHU Pitié-Salpêtrière], CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Institut E3M [CHU Pitié-Salpêtrière], Institut du Cerveau et de la Moëlle Epinière = Brain and Spine Institute (ICM), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-CHU Pitié-Salpêtrière [AP-HP], Biomarqueurs et essais cliniques en Cancérologie et Onco-Hématologie (BECCOH), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Université Paris-Saclay, Hôpital Sainte-Marguerite [CHU - APHM] (Hôpitaux Sud ), Centre Hospitalier Universitaire Félix-Guyon [Saint-Denis, La Réunion, France], Comenius University Children's Hospital Limbova 1 [Bratislava, Slovakia], Service Immuno Hémato-Onco Pédiatrique, CHU Grenoble, Service d'Hémato-oncologie Pédiatrique, CHU Bordeaux [Bordeaux]-Hôpital Pellegrin, Service de pédiatrie médicale et médecine de l'adolescent [Rouen], CHU Rouen, Normandie Université (NU)-Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU), CHU Toulouse [Toulouse], CHU Amiens-Picardie, Universitätsklinikum Frankfurt, Hôpital Universitaire des Enfants Reine Fabiola [Bruxelles, Belgique] (HUDERF), University Hospital [Minsk, Belarus], Universitätsklinikum Münster Klinik für Kinder [Münster, Germany], Centre hospitalier universitaire de Poitiers (CHU Poitiers), Hôpital Jeanne de Flandre [Lille], Hôpital Mustapha [Mustapha, Algeria], Charité - UniversitätsMedizin = Charité - University Hospital [Berlin], University Hospital Motol [Prague, Czech Republic], Hôpitaux Universitaires de Genève (HUG), Schneider Children’s Medical Center of Israel [Petah Tikva], The Edmond and Lily Safra Children's Hospital [Tel-Hahsomer, Israel], Dmitriy Rogachev National Center for Pediatric Hematology, Oncology and Immunology [Moscow, Russia], Sorbonne Université - Département de neurologie 2 - Mazarin, Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Pitié-Salpêtrière [AP-HP], Great Ormond Street Hospital for Children [London] (GOSH), Sant'Anna Children's Hospital, Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institute of Cellular Medicine [Newcastle], Service de Pharmacologie Toxicologie [Garches], Sorbonne Université - Faculté de Médecine (SU FM), Sorbonne Université (SU), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Comenius University in Bratislava, Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), Service d'Onco-neurologie = Département de neurologie 2 [CHU Pitié Salpêtrière], TALY, Valerie, Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Pitié-Salpêtrière [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut E3M [CHU Pitié-Salpêtrière], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Centre National de la Recherche Scientifique (CNRS), and Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)

Subjects
Male, Proto-Oncogene Proteins B-raf, Cancer Research, medicine.medical_specialty, Time Factors, Adolescent, Treatment outcome, Drug Resistance, [SDV.CAN]Life Sciences [q-bio]/Cancer, Severity of Illness Index, 03 medical and health sciences, 0302 clinical medicine, [SDV.CAN] Life Sciences [q-bio]/Cancer, Langerhans cell histiocytosis, Refractory, medicine, Humans, Child, Vemurafenib, Protein Kinase Inhibitors, 030304 developmental biology, 0303 health sciences, Childhood Langerhans Cell Histiocytosis, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, business.industry, Age Factors, Infant, ORIGINAL REPORTS, medicine.disease, Dermatology, 3. Good health, Europe, Histiocytosis, Langerhans-Cell, Histiocytosis, Treatment Outcome, Oncology, Multicenter study, Pediatric Oncology, Child, Preschool, 030220 oncology & carcinogenesis, Mutation, Female, Observational study, business, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Signal Transduction, medicine.drug
Abstract

PURPOSE Off-label use of vemurafenib (VMF) to treat BRAFV600E mutation–positive, refractory, childhood Langerhans cell histiocytosis (LCH) was evaluated. PATIENTS AND METHODS Fifty-four patients from 12 countries took VMF 20 mg/kg/d. They were classified according to risk organ involvement: liver, spleen, and/or blood cytopenia. The main evaluation criteria were adverse events (Common Terminology Criteria for Adverse Events [version 4.3]) and therapeutic responses according to Disease Activity Score. RESULTS LCH extent was distributed as follows: 44 with positive and 10 with negative risk organ involvement. Median age at diagnosis was 0.9 years (range, 0.1 to 6.5 years). Median age at VMF initiation was 1.8 years (range, 0.18 to 14 years), with a median follow-up of 22 months (range, 4.3 to 57 months), whereas median treatment duration was 13.9 months (for 855 patient-months). At 8 weeks, 38 complete responses and 16 partial responses had been achieved, with the median Disease Activity Score decreasing from 7 at diagnosis to 0 ( P < .001). Skin rash, the most frequent adverse event, affected 74% of patients. No secondary skin cancer was observed. Therapeutic plasma VMF concentrations (range, 10 to 20 mg/L) seemed to be safe and effective. VMF discontinuation for 30 patients led to 24 LCH reactivations. The blood BRAFV600E allele load, assessed as circulating cell-free DNA, decreased after starting VMF but remained positive (median, 3.6% at diagnosis, and 1.6% during VMF treatment; P < .001) and was associated with a higher risk of reactivation at VMF discontinuation. None of the various empirical therapies (hematopoietic stem-cell transplantation, cladribine and cytarabine, anti-MEK agent, vinblastine, etc) used for maintenance could eradicate the BRAFV600E clone. CONCLUSION VMF seemed safe and effective in children with refractory BRAFV600E-positive LCH. Additional studies are needed to find effective maintenance therapy approaches.

Published
2019

31. Microfluidic extraction and digital quantification of circulating cell-free DNA from serum

Author

Iago Pereiro, Charlotte Proudhon, Sonia Garrigou, Karla Perez-Toralla, Jean-Louis Viovy, Valérie Taly, Stéphanie Descroix, François-Clément Bidard, Fahima Di Federico, Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire Physico-Chimie Curie [Institut Curie] (PCC), Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Sorbonne Université (SU), Laboratoire des biomarqueurs tumoraux circulants [Institut Curie, Paris] (SiRIC - Département de recherche translationnelle), Institut Curie [Paris]-Université Paris sciences et lettres (PSL), Institut Curie [Paris], Physico-Chimie-Curie (PCC), Institut Curie-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Université de recherche Paris-Sciences-et-Lettres (Université de recherche PSL)-Institut Curie [Paris], and Institut Curie

Subjects
Chromatography, Chemistry, [SDV]Life Sciences [q-bio], Extraction (chemistry), Microfluidics, Metals and Alloys, 02 engineering and technology, 010402 general chemistry, 021001 nanoscience & nanotechnology, Condensed Matter Physics, 01 natural sciences, DNA extraction, Circulating Cell-Free DNA, 0104 chemical sciences, Surfaces, Coatings and Films, Electronic, Optical and Magnetic Materials, chemistry.chemical_compound, Materials Chemistry, Nucleic acid, Digital polymerase chain reaction, Multiplex, Electrical and Electronic Engineering, 0210 nano-technology, Instrumentation, DNA
Abstract

International audience; Miniaturized devices for the extraction of DNA have been used for assessing genetic material in biological, forensic and environmental samples. However, the ability to isolate trace amounts of highly fragmented DNA from biological fluids remains a challenge. The current work reports a microfluidic approach that combines on line a dynamic magnetic extraction procedure with droplet-based digital PCR (ddPCR). This strategy maximizes the surface area for DNA binding within the chip, in order to capture short DNA fragments, with the possibility of recovering the purified samples into picoliter volumes for high sensitivity mutation detection. The application of this technology to capture circulating cell-free DNA (ccfDNA) from serum samples of cancer patients is demonstrated herein, with efficiencies comparable to standard column-based DNA extraction methods. This technology uses lesser amounts of required material and reagents, and has a higher potential for automation and multiplex DNA analysis. Furthermore, this approach can also be extended for the detection of other circulating biomarkers, such as nucleic acid sequences with aberrant methylation patterns or miRNA.

Published
2019

32. HPV-circulating tumoural DNA by droplet-based digital polymerase chain reaction, a new molecular tool for early detection of HPV metastatic anal cancer? A case report

Author

Laure Gibault, David Veyer, Marie-Laure Lucas, Hélène Péré, Laurence Weiss, Valérie Taly, Sonia Garrigou, Marion Mandavit, Juliette Pavie, Simon Pernot, Laboratoire de Virologie [Hôpital Européen Georges Pompidou - APHP], Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Service d'Immunologie Clinique [Hôpital Européen Georges Pompidou - APHP], Unité Fonctionnelle de Pharmacogénétique et Oncologie Moléculaire [AP-HP Hôpital Européen Georges Pompidou] (Service de Biochimie), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Sorbonne Université (SU), Paris-Centre de Recherche Cardiovasculaire (PARCC - UMR-S U970), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Equipe labellisée Ligue contre le Cancer, Service d'Anatomo-cytopathologie [Hôpital Européen Georges Pompidou - APHP], Université Paris Descartes - Paris 5 (UPD5), This work was supported by the Assistance Publique Hopitaux de Paris (AP-HP), Ministère de l’Enseignement Supérieur et de la Recherche, the Université Paris-Descartes, the Centre National de la Recherche Scientifique (CNRS), the Institut National de la Santé et de la Recherche Médicale (INSERM), the SIRIC CARPEM and the ligue nationale contre le cancer (LNCC, Program 'Equipe labelise´e LIGUE', no. EL2016.LNCC/VaT)., and TALY, Valerie

Subjects
Cancer Research, [SDV]Life Sciences [q-bio], HPV ctDNA, Early detection, [SDV.CAN]Life Sciences [q-bio]/Cancer, Metastasis, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, [SDV.CAN] Life Sciences [q-bio]/Cancer, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, HGAIN, Medicine, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Digital polymerase chain reaction, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, 0303 health sciences, business.industry, medicine.disease, 3. Good health, [SDV] Life Sciences [q-bio], Oncology, chemistry, 030220 oncology & carcinogenesis, Cancer research, Droplet-based digital PCR, business, Metastatic anal cancer, DNA
Abstract

International audience; no abstract

Published
2019

33. Circulating tumor DNA is a prognostic marker of tumor recurrence in stage II and III colorectal cancer: multicentric, prospective cohort study (ALGECOLS)

Author

Delphine Le Corre, Juliette Djadi-Prat, Olivier Bouché, Leonor Benhaim, Bruno Landi, Julien Taieb, Jean-François Emile, Corinne Normand, Pierre Laurent-Puig, Karla Perez-Toralla, Valérie Taly, Hélène Blons, Gilles Chatellier, Geoffroy Poulet, Audrey Didelot, Marie Selvy, Thierry Lecomte, Claire Mulot, Sonia Garrigou, Deniz Pekin, Darren Roy Link, and Shu-Fang Wang-Renault

Subjects
Adult, Male, Cancer Research, medicine.medical_specialty, Colorectal cancer, Gastroenterology, Circulating Tumor DNA, Cohort Studies, Interquartile range, Internal medicine, Biomarkers, Tumor, Medicine, Humans, Longitudinal Studies, Prospective Studies, Liquid biopsy, Stage (cooking), Prospective cohort study, Aged, Aged, 80 and over, business.industry, Hazard ratio, Middle Aged, medicine.disease, Prognosis, Minimal residual disease, Confidence interval, Oncology, Female, Neoplasm Recurrence, Local, business, Colorectal Neoplasms
Abstract

Background In non-metastatic colorectal cancer (CRC), we evaluated prospectively the pertinence of longitudinal detection and quantification of circulating tumor DNA (ctDNA) as a prognostic marker of recurrence. Method The presence of ctDNA was assessed from plasma collected before and after surgery for 184 patients classified as stage II or III and at each visit during 3–4 years of follow-up. The ctDNA analysis was performed by droplet-based digital polymerase chain reaction, targeting mutation and methylation markers, blindly from the clinical outcomes. Multivariate analyses were adjusted on age, gender, stage, and adjuvant chemotherapy. Results Before surgery, 27.5% of patients were positive for ctDNA detection. The rate of recurrence was 32.7% and 11.6% in patients with or without detectable ctDNA respectively (P = 0.001). Time to recurrence (TTR) was significantly shorter in patients with detectable ctDNA before (adjusted hazard ratio [HR] = 3.58, 95% confidence interval [CI] 1.71–7.47) or immediately after surgery (adjusted HR = 3.22, 95% CI 1.32–7.89). The TTR was significantly shorter in patients with detectable ctDNA during the early postoperative follow-up (1–6 months) (adjusted HR = 5, 95% CI 1.9–12.9). Beyond this period, ctDNA remained a prognostic marker with a median anticipated diagnosis of recurrence of 13.1 weeks (interquartile range 28 weeks) when compared to imaging follow-up. The rate of ctDNA+ might be underestimated knowing that consensus pre-analytical conditions were not described at initiation of the study. Conclusion This prospective study confirms the relevance of ctDNA as a recurrence risk factor in stage II and III CRC before surgery and as a marker of minimal residual disease after surgery that may predict recurrence several months before imaging techniques.

Published
2021

34. Advances in multiplexed techniques for the detection and quantification of microRNAs

Author

Thomas Jet, Yannick Rondelez, Guillaume Gines, Valérie Taly, Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Gulliver (UMR 7083), Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), TALY, Valerie, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), and HAL-SU, Gestionnaire

Subjects
Computer science, [SDV]Life Sciences [q-bio], Computational biology, 010402 general chemistry, 01 natural sciences, Multiplexing, 03 medical and health sciences, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Neoplasms, microRNA, Humans, Multiplex, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], 030304 developmental biology, 0303 health sciences, Liquid Biopsy, High-Throughput Nucleotide Sequencing, Neurodegenerative Diseases, General Chemistry, Microarray Analysis, 3. Good health, 0104 chemical sciences, Diagnosis methods, [SDV] Life Sciences [q-bio], MicroRNAs, Potential biomarkers, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Nucleic Acid Amplification Techniques
Abstract

International audience; MicroRNA detection is currently a crucial analytical chemistry challenge: almost 2000 papers were referenced in PubMed in 2018 and 2019 for the keywords "miRNA detection method". MicroRNAs are potential biomarkers for multiple diseases including cancers, neurodegenerative and cardiovascular diseases. Since miRNAs are stably released in bodily fluids, they are of prime interest for the development of non-invasive diagnosis methods, such as liquid biopsies. Their detection is however challenging, as high levels of sensitivity, specificity and robustness are required. The analysis also needs to be quantitative, since the aim is to detect miRNA concentration changes. Moreover, a high multiplexing capability is also of crucial importance, since the clinical potential of miRNAs probably lays in our ability to perform parallel mapping of multiple miRNA concentrations and recognize typical disease signature from this profile. A plethora of biochemical innovative detection methods have been reported recently and some of them provide new solutions to the problem of sensitive multiplex detection. In this review, we propose to analyze in particular the new developments in multiplexed approaches to miRNA detection. The main aspects of these methods (including sensitivity and specificity) will be analyzed, with a particular focus on the demonstrated multiplexing capability and potential of each of these methods.

Published
2021

35. Usefulness of Plasma SARS-CoV-2 RNA Quantification by Droplet-based Digital PCR to Monitor Treatment Against COVID-19 in a B-cell Lymphoma Patient

Author

Valérie Taly, Laure Izquierdo, Laurent Bélec, Hélène Péré, Solen Kernéis, Celia Azoulay, Thomas Serrano, Nicolas Robillard, Pierre Laurent-Puig, Karine Lacombe, Benjamin Terrier, Deborah Eshagh, Adrien Contejean, Marguerite Vignon, David Veyer, Tali Anne Szwebel, Thomas Hueso, Flore Rozenberg, Etienne Canouï, Thomas Bruneau, CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Unité de Virologie [Hôpital Européen Georges Pompidou - APHP], Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École Pratique des Hautes Études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Equipe Mobile d'Infectiologie [APHP Centre, Paris], Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Cité (UPCité), Labex OncoImmunology, Equipe labellisée Ligue Contre le Cancer, Université Paris Descartes - Paris 5 (UPD5), Département d'hématologie [Gustave Roussy], Institut Gustave Roussy (IGR), Service de Virologie [CHU Cochin], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Paris-Centre de Recherche Cardiovasculaire (PARCC (UMR_S 970/ U970)), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Epidémiologie et modélisation de la résistance aux antimicrobiens - Epidemiology and modelling of bacterial escape to antimicrobials (EMAE), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pierre Louis d'Epidémiologie et de Santé Publique (iPLESP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), CHU Saint-Antoine [AP-HP], This work was supported by the Assistance Publique Hôpitaux de Paris (AP-HP), Ministère de l’Enseignement Supérieur et de la Recherche, the Université de Paris, the Institut National de la Santé et de la Recherche Médicale (INSERM) and the Collecte Crise COVID/APHP Centre-Université de Paris., Centre National de Référence Maladies auto-immunes Systémiques Rares [CHU Pitié-Salpêtrière], Service de Département de médecine interne et immunologie clinique [CHU Pitié-Salpêtrière] (DMIIC), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), École pratique des hautes études (EPHE), TALY, Valerie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université de Paris (UP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), and Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
0301 basic medicine, Oncology, medicine.medical_specialty, medicine.drug_class, business.industry, [SDV]Life Sciences [q-bio], RNA, medicine.disease, Monoclonal antibody, Article, 3. Good health, Lymphoma, [SDV] Life Sciences [q-bio], 03 medical and health sciences, 030104 developmental biology, 0302 clinical medicine, 030220 oncology & carcinogenesis, Internal medicine, medicine, Biomarker (medicine), Digital polymerase chain reaction, Stem cell, business, B-cell lymphoma, Viral load
Abstract

International audience; We report the case of an HIV-1-infected patient, treated with anti-CD20 monoclonal antibody for a B-cell lymphoma previously treated by autologous stem cell transplant. He suffered from chronic COVID19 and we monitored by plasma SARS-CoV-2 RNA by highly sensitive droplet-based digital PCR technology (ddPCR). Under tocilizumab therapy and despite a first clinical improvement biologically associated with decreasing inflammatory markers, a slight increase of SARS-CoV-2 RNAaemia quantified by ddPCR was highlighted, confirming the absence of viral efficacy of this treatment and predicting the subsequent observed deterioration. As expected, his complete recovery, finally achieved after COVID-19 convalescent plasmatherapy, strictly paralleled plasma SARS-CoV-2 RNA clearance. With these results, we confirmed the interest of SARS-CoV-2 RNAaemia monitoring by ddPCR in COVID-19 patients, particularly during treatment, and firstly showed that this new and specific biomarker could be helpful to select eligible patient for anti-IL6 receptors therapy considering the variable levels of efficacy recently observed with such therapy.

Published
2021

36. Prognostic Value and Relation with Adjuvant Treatment Duration of ctDNA in Stage III Colon Cancer: a

Author

Julien, Taieb, Valérie, Taly, Julie, Henriques, Camille, Bourreau, Laurent, Mineur, Jaafar, Bennouna, Jérôme, Desrame, Christophe, Louvet, Céline, Lepere, May, Mabro, Joëlle, Egreteau, Olivier, Bouche, Claire, Mulot, Katia, Hormigos, Kariman, Chaba, Thibault, Mazard, Aimery, de Gramont, Dewi, Vernerey, Thierry, André, and Pierre, Laurent-Puig

Subjects
Male, Duration of Therapy, Chemotherapy, Adjuvant, Colonic Neoplasms, Humans, Female, Prospective Studies, Prognosis, Disease-Free Survival, Article, Aged, Circulating Tumor DNA, Neoplasm Staging
Abstract

Circulating tumor DNA (ctDNA) has been suggested as a major prognostic factor in resected stage-III colon cancer. We analyzed ctDNA of patients randomized in the phase III IDEA-France trial.ctDNA was tested forOf 2,010 randomized patients, 1,345 had available ctDNA samples (1,017 collected both post-surgery and pre-chemotherapy). More Eastern Cooperative Oncology Group performance status (ECOG PS) of 0 (78% versus 69%) and T4 and/or N2 (40% versus 36%) were observed in patients studied (In this first ctDNA assessment of a large series of patients with stage III colon cancer enrolled in phase III trial, post-surgery ctDNA was found in 13.8% of them and was confirmed as an independent prognostic marker.

Published
2021

37. Technological Advances in Tumor-On-Chip Technology: From Bench to Bedside

Author

Valérie Taly, Alexandra Harrison, Wenjin Xiao, Santa Bērziņa, Gestionnaire, HAL Sorbonne Université 5, Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP)

Subjects
Computer science, [SDV.CAN]Life Sciences [q-bio]/Cancer, 02 engineering and technology, Review, tumor model, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, on-chip analysis, RC254-282, 030304 developmental biology, 0303 health sciences, business.industry, Translational medicine, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, personalized medicine, 021001 nanoscience & nanotechnology, Bench to bedside, 3. Good health, ComputingMethodologies_PATTERNRECOGNITION, Oncology, tumor-on-chip, Systems engineering, cancer research, Personalized medicine, Analysis tools, 0210 nano-technology, business
Abstract

Simple Summary Various 3D in vitro tumor models are rapidly advancing cancer research. Unlike animal models, they can be produced quickly and are amenable to high-throughput studies. Growing tumor spheroids in microfluidic tumor-on-chip platforms has particularly elevated the capabilities of such models. Tumor-on-chip devices can mimic multiple aspects of the dynamic in vivo tumor microenvironment in a precisely controlled manner. Moreover, new technologies for the on- and off-chip analysis of these tumor mimics are continuously emerging. There is thus an urgent need to review the latest developments in this rapidly progressing field. Here, we present an overview of the technological advances in tumor-on-chip technology by reviewing state-of-the-art tools for on-chip analysis. In particular, we evaluate the potential for tumor-on-chip technology to guide personalized cancer therapies. We strive to appeal to cancer researchers and biomedical engineers alike, informing on current progress, while provoking thought on the outstanding developments needed to achieve clinical-stage research. Abstract Tumor-on-chip technology has cemented its importance as an in vitro tumor model for cancer research. Its ability to recapitulate different elements of the in vivo tumor microenvironment makes it promising for translational medicine, with potential application in enabling personalized anti-cancer therapies. Here, we provide an overview of the current technological advances for tumor-on-chip generation. To further elevate the functionalities of the technology, these approaches need to be coupled with effective analysis tools. This aspect of tumor-on-chip technology is often neglected in the current literature. We address this shortcoming by reviewing state-of-the-art on-chip analysis tools for microfluidic tumor models. Lastly, we focus on the current progress in tumor-on-chip devices using patient-derived samples and evaluate their potential for clinical research and personalized medicine applications.

Published
2021

38. Circadian rhythm and circulating cell-free DNA release on healthy subjects

Author

Geoffroy Poulet, Jean-Sébastien Hulot, Anne Blanchard, Damien Bergerot, Wenjin Xiao, Frederic Ginot, Audrey Boutonnet-Rodat, Abdelli Justine, Guillaume Beinse, Vanna Geromel, Laurence Pellegrina, Michel Azizi, Pierre Laurent-Puig, Leonor Benhaim, and Valerie Taly

Subjects
Medicine, Science
Abstract

Abstract In the last decade, clinical studies have investigated the clinical relevance of circulating cell-free-DNA (ccfDNA) as a diagnostic and prognosis tool in various diseases including cancers. However, limited knowledge on ccfDNA biology restrains its full development in the clinical practice. To improve our understanding, we evaluated the impact of the circadian rhythm on ccfDNA release in healthy subjects over a 24-h period. 10 healthy female subjects underwent blood sampling at 8am and 20 healthy male subjects underwent serial blood sampling (8:00 AM, 9:00 AM, 12:00 PM, 4:00 PM, 8:00 PM, 12:00 AM, 4 AM (+ 1 Day) and 8 AM (+ 1 Day)). We performed digital droplet-based PCR (ddPCR) assays to target 2 DNA fragments (69 & 243 bp) located in the KRAS gene to determine the ccfDNA concentration and fragmentation profile. As control, half of the samples were re-analyzed by capillary miniaturized electrophoresis (BIAbooster system). Overall, we did not detect any influence of the circadian rhythm on ccfDNA release. Instead, we observed a decrease in the ccfDNA concentration after meal ingestion, suggesting either a post-prandial effect or a technical detection bias due to a higher plasma load in lipids and triglycerides. We also noticed a potential effect of gender, weight and creatinine levels on ccfDNA concentration.

Published
2023
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39. Highly sensitive quantification of plasma SARS-CoV-2 RNA shelds light on its potential clinical value

Author

Benjamin Terrier, Hélène Péré, Nicolas Robillard, Laurent Bélec, Flore Rozenberg, Jérôme Hadjadj, Solen Kernéis, Valérie Taly, Maxime Wack, Pierre Laurent-Puig, Anne-Sophie L’Honneur, David Veyer, Geoffroy Poulet, Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Infection, Anti-microbiens, Modélisation, Evolution (IAME (UMR_S_1137 / U1137)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP)-Université Sorbonne Paris Nord, Eurofins Biomnis, Institut Pasteur [Paris], Imagine - Institut des maladies génétiques (IHU) (Imagine - U1163), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Paris-Centre de Recherche Cardiovasculaire (PARCC (UMR_S 970/ U970)), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), École Pratique des Hautes Études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité)-Université Sorbonne Paris Nord, Institut Pasteur [Paris] (IP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), and TALY, Valerie

Subjects
0301 basic medicine, Microbiology (medical), medicine.medical_specialty, Critical Illness, [SDV]Life Sciences [q-bio], Severity of Illness Index, Gastroenterology, law.invention, 03 medical and health sciences, 0302 clinical medicine, law, Internal medicine, Severity of illness, medicine, Major Article, Humans, Digital polymerase chain reaction, 030212 general & internal medicine, Respiratory system, Polymerase chain reaction, Liver injury, business.industry, Diagnostic Tests, Routine, SARS-CoV-2, COVID-19, medicine.disease, 3. Good health, [SDV] Life Sciences [q-bio], Editorial Commentary, Pneumonia, 030104 developmental biology, Infectious Diseases, AcademicSubjects/MED00290, RNA, Viral, Biomarker (medicine), RNA, business, Viral load
Abstract

Background Coronavirus disease 2019 (COVID-19) is a global public health problem that has already caused more than 662 000 deaths worldwide. Although the clinical manifestations of COVID-19 are dominated by respiratory symptoms, some patients present other severe damage such as cardiovascular, renal and liver injury, and/or multiple organ failure, suggesting a spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in blood. Recent ultrasensitive polymerase chain reaction (PCR) technology now allows absolute quantification of nucleic acids in plasma. We intend to use the droplet-based digital PCR technology to obtain sensitive detection and precise quantification of plasma SARS-CoV-2 viral load (SARS-CoV-2 RNAemia) in hospitalized COVID-19 patients. Methods Fifty-eight consecutive COVID-19 patients with pneumonia 8 to 12 days after onset of symptoms and 12 healthy controls were analyzed. Disease severity was categorized as mild to moderate in 17 patients, severe in 16, and critical in 26. Plasma SARS-CoV-2 RNAemia was quantified by droplet digital Crystal Digital PCR next-generation technology (Stilla Technologies, Villejuif, France). Results Overall, SARS-CoV-2 RNAemia was detected in 43 (74.1%) patients. Prevalence of positive SARS-CoV-2 RNAemia correlated with disease severity, ranging from 53% in mild-to-moderate patients to 88% in critically ill patients (P = .036). Levels of SARS-CoV-2 RNAemia were associated with severity (P = .035). Among 9 patients who experienced clinical deterioration during follow-up, 8 had positive SARS-CoV-2 RNAemia at baseline, whereas only 1 critical patient with undetectable SARS-CoV-2 RNAemia at the time of analysis died at day 27. Conclusion SARS-CoV-2 RNAemia measured by droplet-based digital PCR constitutes a promising prognosis biomarker in COVID-19 patients.

Published
2020

40. Decision for adjuvant treatment in stage II colon cancer based on circulating tumor DNA:The CIRCULATE-PRODIGE 70 trial

Author

Thierry André, Karine Le Malicot, Flore Geillon, Julien Taieb, Jean-François Emile, Marion Chauvenet, Sylvain Manfredi, Côme Lepage, Léonor Benhaim, David Tougeron, Pierre Laurent Puig, Valérie Taly, Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Institut Gustave Roussy (IGR), Département de chirurgie viscérale [Gustave Roussy], Lipides - Nutrition - Cancer [Dijon - U1231] (LNC), Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Institut National de la Santé et de la Recherche Médicale (INSERM), Biomarqueurs et essais cliniques en Cancérologie et Onco-Hématologie (BECCOH), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Université Paris-Saclay, Centre hospitalier universitaire de Poitiers (CHU Poitiers), Hospices Civils de Lyon (HCL), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Sorbonne Université (SU), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), and TALY, Valerie

Subjects
Oncology, Adult, Male, medicine.medical_specialty, Methylated biomarker, Adolescent, Colorectal cancer, medicine.medical_treatment, [SDV]Life Sciences [q-bio], Leucovorin, Rectum, Disease-Free Survival, 03 medical and health sciences, Young Adult, 0302 clinical medicine, Internal medicine, medicine, Adjuvant therapy, Biomarkers, Tumor, Humans, Adjuvant, Aged, Neoplasm Staging, Circulating tumor DNA, Predictive marker, Hepatology, business.industry, Gastroenterology, Middle Aged, medicine.disease, 3. Good health, Oxaliplatin, Colon cancer, [SDV] Life Sciences [q-bio], medicine.anatomical_structure, Treatment Outcome, Chemotherapy, Adjuvant, 030220 oncology & carcinogenesis, Colonic Neoplasms, Adenocarcinoma, 030211 gastroenterology & hepatology, Female, Fluorouracil, France, Bolus (digestion), business, medicine.drug
Abstract

International audience; Background: Adjuvant treatment for stage II colon cancer remains debated. Finding a tool to select patients at risk for disease recurrence may help the clinical decision. Circulating tumor DNA (ctDNA) has been reported recently as a potential predictive marker for disease recurrence. We thus aim to test its ability to better select stage II colon cancer patients for adjuvant therapy.Methods: This national, phase III trial (NCT00002019-000935-15) conducted in more than 100 centers in France, plans to screen around 2640 patients in order to randomize (2:1; minimization method) 198 ctDNA positive patients. Patients aged 18 to 75 years with ECOG performance status ≤1 with R0 surgical resection of a pT3-T4aN0 colon or high rectum adenocarcinoma will be randomized within 63 days after curative-intent surgery, to adjuvant mFOLFOX6 (oxaliplatin 85 mg/m², leucovorin 400 mg/m², and 5-FU bolus 400 mg/m2 then 5FU Continuous infusion 2.4 g/m²) every two weeks for 12 cycles or observation. Patients will be followed for maximum 7 years. A gain of 17.5% in 3-yr disease free survival (DFS) is expected (42.5% in the experimental arm vs. 25% in the control arm; HR:0.62; α, 5% [two-sided log-rank test]; 1-β, 80%). Secondary endpoints include 2-yr DFS, overall survival, and toxicity. Recruitement began End of January 2020.

Published
2020

41. Author Correction: High throughput single cell counting in droplet-based microfluidics

Author

Jeremy Vrignon, Valérie Taly, Zakaria El Harrak, Ouriel Caen, Eleonora Zonta, Philippe Nizard, Jean-Christophe Baret, and Heng Lu

Subjects
Multidisciplinary, Cell Survival, Computer science, lcsh:R, Microfluidics, lcsh:Medicine, HL-60 Cells, Nanotechnology, Microfluidic Analytical Techniques, Cell counting, High-Throughput Screening Assays, Lab-On-A-Chip Devices, Escherichia coli, Humans, lcsh:Q, Droplet-based microfluidics, Single-Cell Analysis, lcsh:Science, Author Correction, Throughput (business), Cell Proliferation
Abstract

Droplet-based microfluidics is extensively and increasingly used for high-throughput single-cell studies. However, the accuracy of the cell counting method directly impacts the robustness of such studies. We describe here a simple and precise method to accurately count a large number of adherent and non-adherent human cells as well as bacteria. Our microfluidic hemocytometer provides statistically relevant data on large populations of cells at a high-throughput, used to characterize cell encapsulation and cell viability during incubation in droplets.

Published
2020

42. Streamlined digital bioassays with a 3D printed sample changer

Author

Adèle Dramé-Maigné, Valérie Taly, Yannick Rondelez, Guillaume Gines, Roberta Menezes, Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP), Centre National de la Recherche Scientifique (CNRS), Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Gulliver (UMR 7083), Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), TALY, Valerie, and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité)

Subjects
Sample (material), Femto, [SDV]Life Sciences [q-bio], Microfluidics, Dispersity, 02 engineering and technology, 01 natural sciences, Biochemistry, Polymerase Chain Reaction, Analytical Chemistry, Robustness (computer science), Electrochemistry, Environmental Chemistry, Humans, Digital polymerase chain reaction, Deoxyribonucleases, Type II Site-Specific, Throughput (business), Spectroscopy, ComputingMilieux_MISCELLANEOUS, chemistry.chemical_classification, Biomolecule, 010401 analytical chemistry, DNA, Equipment Design, Microfluidic Analytical Techniques, 021001 nanoscience & nanotechnology, [SDV.BIBS]Life Sciences [q-bio]/Quantitative Methods [q-bio.QM], 0104 chemical sciences, 3. Good health, [SDV] Life Sciences [q-bio], MicroRNAs, chemistry, Printing, Three-Dimensional, Biological Assay, Emulsions, 0210 nano-technology, Biological system
Abstract

International audience; Droplet-based microfluidics has permeated many areas of life sciences including biochemistry, biology and medicine. Water-in-oil droplets act as independent femto- to nano-liter reservoirs, enabling the parallelization of (bio)chemical reactions with a minimum sample input. Among the range of applications spanned by droplet microfluidics, digital detection of biomolecules, using Poissonian isolation of single molecules in compartments, has gained considerable attention due to the high accuracy, sensitivity and robustness of these methods. However, while the droplet throughput can be very high, the sample throughput of these methods is poor in comparison to well plate-based assays. This limitation comes from the necessity to convert independently each sample into a monodisperse emulsion. In this paper, we report a versatile device that performs the quick sequential partitioning of up to 15 samples using a single microfluidic chip. A 3D printed sample rotor is loaded with all samples and connected to a pressure source. Simple magnetic actuation is then used to inject the samples in the microfluidic chip without pressure disruption. This procedure generates monodisperse droplets with high sample-to-sample consistency. We also describe a fluorescent barcoding strategy that allows all samples to be collected, incubated, imaged and analyzed simultaneously, thus decreasing significantly the time of the assay. As an example of application, we perform a droplet digital PCR assay for the quantification of a DNA amplicon from 8 samples in less than 2 hours. We further validate our approach demonstrating the parallel quantification of 11 microRNAs from a human sample using an isothermal nucleic acid amplification chemistry. As an off-chip device, the sample changer can be connected to a variety of microfluidic geometries and therefore, used for a wide range of applications.

Published
2020

43. Plasma circulating tumor DNA in pancreatic adenocarcinoma for screening, diagnosis, prognosis, treatment and follow-up: A systematic review

Author

Pierre Laurent-Puig, Debora Basile, Aziz Zaanan, Raëf Abdallah, Jean-Baptiste Bachet, Daniel Pietrasz, Valérie Taly, Julien Taieb, Shulin Zhao, Léo Mas, TALY, Valerie, Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université Paris Cité (UPCité), Università degli Studi di Udine - University of Udine [Italie], and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Université de Paris (UP)

Subjects
0301 basic medicine, Oncology, medicine.medical_specialty, Poor prognosis, [SDV]Life Sciences [q-bio], Disease, 03 medical and health sciences, 0302 clinical medicine, Internal medicine, Diagnosis, medicine, Advanced disease, Biomarkers, Tumor, Humans, Radiology, Nuclear Medicine and imaging, Radical surgery, Circulating tumor DNA, business.industry, Treatment monitoring, General Medicine, medicine.disease, Prognosis, Minimal residual disease, 3. Good health, Clinical trial, Pancreatic Neoplasms, [SDV] Life Sciences [q-bio], 030104 developmental biology, 030220 oncology & carcinogenesis, Screening, Adenocarcinoma, business, Pancreatic adenocarcinoma, Carcinoma, Pancreatic Ductal, Follow-Up Studies
Abstract

International audience; While no biomarker is currently recommended for the management of pancreatic adenocarcinoma (PA), circulating tumor DNA (ctDNA) seems promising but little is known on how it may help to manage our patients in the near future. This systematic review of literature was designed to explore the current knowledge on ctDNA as a screening, diagnostic, prognostic, predictive and theranostic biomarker in the management of PA. We retrieved 62 full-text articles, 3 meta-analyses, 2 clinical trials, 1 abstract and 13 ongoing trials. Results were categorized into sections about screening, diagnosis, prognosis and follow-up of localized and advanced PA together with possible theranostics applications. Although its specificity is excellent, the current sensitivity of ctDNA remains a limitation especially in patients without metastatic disease. Therefore, this biomarker cannot be currently used as a screening or diagnostic tool. Increasing evidence suggests that ctDNA is a relevant candidate biomarker to assess minimal residual disease after radical surgery, but also a strong independent biomarker linked to a poor prognosis in advanced PA. Some recent data also indicates that ctDNA is an attractive biomarker for longitudinal follow-up and possibly early treatment adaptation. Its role in tumor profiling in advanced disease to decide targeted treatments remains to be explored. Altogether, ctDNA appears to be a reliable prognostic tool. Though promising results have been reported, further studies are still needed to define exactly how ctDNA can help physicians in the screening, diagnosis and treatment, as PA is expected to become a major cause of cancer-related deaths in the forthcoming decade.

Published
2020

44. Beyond the on/off chip trade-off: A reversibly sealed microfluidic platform for 3D tumor microtissue analysis

Author

Valérie Taly, Antoine Riaud, Gabriele Pitingolo, Philippe Nizard, Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Equipe labellisée Ligue contre le Cancer, This work was supported by the Ministère de l’Enseignement Supérieur et de la Recherche, the Université Paris-Descartes, the Centre National de la Recherche Scientifique (CNRS), the Institut National de la Santé et de la Recherche Médicale (INSERM), the Institut National du Cancer (INCA) and canceropole Ile-de-France (no. 2016-1-EMERG-54-UP 5-1), the SIRIC CARPEM, the Fondation de la Recherche Medicale (SPF20160936257) and the ligue nationale contre le cancer (LNCC, Program 'Equipe labelisée LIGUE', no. EL2016.LNCC/VaT). The authors acknowledges the financial support of HTE Program-HetColi network, INSERM Physicancer program (no. PC201423) and the SIRIC CARPEM. Financial supports from CAMPUS FRANCE (n° 39525QJ) and Università Italo-Francese (n° G18_208) are gratefully acknowledged. The authors would like to thank Saksham Sharma for his important contribution to the development of the numerical models. This work has benefited from the facility and expertise of the electron microscopy facility of Institut de Biologie Paris-Seine (IBPS/FR3631 - UPMC/CNRS- Paris, France). This work was carried out with the support of the Pierre-Gilles de Gennes Institute equipment ('Investissements d’Avenir' program, reference: ANR 10-NANO 0207)., and TALY, Valerie

Subjects
0301 basic medicine, Materials science, Growth kinetics, [SDV]Life Sciences [q-bio], Microfluidics, Nanotechnology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 02 engineering and technology, Fluorescence, SEM analysis, 03 medical and health sciences, 3D cell culture, Tumor microtissue, Materials Chemistry, Drug response, Electrical and Electronic Engineering, Modular chip, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Instrumentation, Fluorescence staining, [SDV.IB] Life Sciences [q-bio]/Bioengineering, Metals and Alloys, 021001 nanoscience & nanotechnology, Condensed Matter Physics, Chip, Surfaces, Coatings and Films, Electronic, Optical and Magnetic Materials, [SDV] Life Sciences [q-bio], Spheroid, 030104 developmental biology, Microfluidic chip, [SDV.IB]Life Sciences [q-bio]/Bioengineering, 0210 nano-technology
Abstract

International audience; Nowadays, microfluidic 3D cell culture is widely used to mimic complex microtissue and dynamic environment, performing more realistic in vitro assays for drug testing. Herein, we developed a novel microfluidic platform for tumor microtissue culture, drug response analysis and versatile microscopic characterization. By reversibly bonding the chip, we go beyond the on/off chip tradeoff, which allows us to perform both fluorescence and SEM characterization of tumor microtissues on a simple platform. The microfluidic chip consists of spherical microwells connected via microchannels, bonded through a magnetic system. Colorectal cancer HT-29 cells were cultured as spherical microtissues on chip and their growth kinetics monitored. The cytotoxic activity of Camptothecin was evaluated by in situ live/dead fluorescence staining and quantification of morphology parameters. Finally, we demonstrated the possibility to collect the 3D tumor microtissues and characterize their surface damaged by the drug using scanning electron microscopy. This reversibly sealed microfluidic platform thus enables to grow sets of 3D tumor microtissues in a controlled dynamic microenvinroment, and subsequently to retrieve the 3D tumor microtissues after chemotherapeutic treatment for in-depth analysis.

Published
2018

45. Development of molecular analyzes by digital PCR for clinical practice: positioning, current applications and perspectives

Author

Nasrin Sarafan-Vasseur, Daniel Henaff, Valérie Taly, Pauline Romanet, Frédéric Fina, Pierre-Jean Lamy, Romain Appay, Jérôme Alexandre Denis, Benoit Busser, Alexandre Perrier, Anne-Sophie Alary, Juliette Nectoux, Pascal Pedini, Centre de Recherche Saint-Antoine (CR Saint-Antoine), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Service de biochimie et de génétique moléculaire [CHU Cochin], Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institut médical d'anayse génomique (IMAGENOME), Labosud, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Génomique et Médecine Personnalisée du Cancer et des Maladies Neuropsychiatriques (GPMCND), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM), ID Solutions [Grabels, France], Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Assistance Publique - Hôpitaux de Marseille (APHM), Anthropologie bio-culturelle, Droit, Ethique et Santé (ADES), Aix Marseille Université (AMU)-EFS ALPES MEDITERRANEE-Centre National de la Recherche Scientifique (CNRS), Marseille medical genetics - Centre de génétique médicale de Marseille (MMG), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), TALY, Valerie, Centre de Recherche Saint-Antoine (CRSA), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Cochin [AP-HP]

Subjects
[SDV]Life Sciences [q-bio], Medical laboratory, Fibrous Dysplasia, Polyostotic, Polymerase Chain Reaction, 03 medical and health sciences, 0302 clinical medicine, Pregnancy, Neoplasms, Prenatal Diagnosis, RNA analysis, Humans, Digital polymerase chain reaction, Practice Patterns, Physicians', Precision Medicine, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, 0303 health sciences, business.industry, Hematopoietic Stem Cell Transplantation, General Medicine, Precision medicine, Data science, 3. Good health, Clinical Practice, [SDV] Life Sciences [q-bio], Molecular Diagnostic Techniques, 030220 oncology & carcinogenesis, Female, business
Abstract

This review is the second part of the workshop on digital PCR (dPCR) proposed by the working group of the French society of clinical biology. The first part of the paper discusses the advantages and limitations of dPCR for the search of different molecular abnormalities such as point mutations, copy number variants, DNA methylation, RNA analysis and a more innovative application, the single-cell dPCR. This synthesis makes it possible to propose a positioning of the dPCR compared to the other available technologies in a medical laboratory. In a second part, the main current applications of the dPCR will be addressed including the oncology of solid tumors and liquid biopsies, oncohematology and the follow-up of hemopathies treatments by hematopoietic stem cell transplantation. We will also detail non-invasive prenatal diagnosis and diagnosis of mosaic genetic disease, using the example of McCune-Albright syndrome. Several French specialists in the field who have implemented these techniques in their laboratory have written these different examples of applications jointly. In summary, this manuscript offers an up-to-date view of the positioning of dPCR in relation to other existing technologies in order to best meet the expectations of precision medicine.

Published
2019

46. BIABooster: Online DNA Concentration and Size Profiling with a Limit of Detection of 10 fg/μL and Application to High-Sensitivity Characterization of Circulating Cell-Free DNA

Author

Valérie Taly, Aurélien Bancaud, Audrey Didelot, Fanny Garlan, Sonia Garrigou, Comtet-Louis Andriamanampisoa, Frederic Ginot, Audrey Boutonnet-Rodat, Daniel Henaff, Jacques Fabre, Laure Saias, Arnaud Morin, Frédéric Fina, Vincent Picot, Aziz Zaanan, Pierre Laurent-Puig, Caroline Gaudy, Pascale Tomasini, Picometrics Technologies, Équipe Micro-Nanofluidique pour les sciences de la vie et de l’environnement (LAAS-MILE), Laboratoire d'analyse et d'architecture des systèmes [Toulouse] (LAAS), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UPS), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UPS), Université Fédérale Toulouse Midi-Pyrénées, Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Descartes - Paris 5 (UPD5), Centre de Recherches en Oncologie biologique et Oncopharmacologie (CRO2), Aix Marseille Université (AMU)- Hôpital de la Timone [CHU - APHM] (TIMONE)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire des Sciences de l'Information et des Systèmes (LSIS), Aix Marseille Université (AMU)-Université de Toulon (UTLN)-Arts et Métiers Paristech ENSAM Aix-en-Provence-Centre National de la Recherche Scientifique (CNRS), HEMODIA, Institut de Recherche en Infectiologie de Montpellier (IRIM), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Laboratoire d'Innovation pour les Technologies des Energies Nouvelles et les nanomatériaux (LITEN), Institut National de L'Energie Solaire (INES), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Centre National de la Recherche Scientifique (CNRS), Physico-Chimie-Curie (PCC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie-Université Pierre et Marie Curie - Paris 6 (UPMC), Service d'Oncologie Multidisciplinaire et d'Innovations Thérapeutiques, Nord University Hospital of Marseille, Laboratoire d'analyse et d'architecture des systèmes (LAAS), Université Toulouse Capitole (UT Capitole), Institut National des Sciences Appliquées (INSA)-Université Fédérale Toulouse Midi-Pyrénées-Institut National des Sciences Appliquées (INSA)-Université Toulouse - Jean Jaurès (UT2J)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Toulouse Capitole (UT Capitole), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL), Université de Picardie Jules Verne (UPJV), Laboratoire de Transfert d'Oncologie Biologique [Hôpital Nord - APHM], Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)- Hôpital Nord [CHU - APHM], Institut de Génétique Moléculaire de Montpellier (IGMM), Unicancer, Institut Curie [Paris], Laboratoire des Composants PEM (LCPEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), TALY, Valerie, Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université Toulouse - Jean Jaurès (UT2J), Université de Toulouse (UT)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Université Toulouse Capitole (UT Capitole), Université de Toulouse (UT), École Pratique des Hautes Études (EPHE), Université Toulouse - Jean Jaurès (UT2J)-Université Toulouse 1 Capitole (UT1), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Université Fédérale Toulouse Midi-Pyrénées-Université Toulouse - Jean Jaurès (UT2J)-Université Toulouse 1 Capitole (UT1), Hôpital Nord [CHU - APHM]-Assistance Publique - Hôpitaux de Marseille (APHM)-Aix Marseille Université (AMU), Centre National de la Recherche Scientifique (CNRS)-Arts et Métiers Paristech ENSAM Aix-en-Provence-Université de Toulon (UTLN)-Aix Marseille Université (AMU), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)

Subjects
0301 basic medicine, Accuracy and precision, Capillary action, [SDV]Life Sciences [q-bio], [PHYS.PHYS.PHYS-BIO-PH]Physics [physics]/Physics [physics]/Biological Physics [physics.bio-ph], Polymerase Chain Reaction, Analytical Chemistry, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Capillary electrophoresis, Limit of Detection, Neoplasms, Biomarkers, Tumor, Humans, Digital polymerase chain reaction, Detection limit, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, Chromatography, Dna concentration, Electrophoresis, Capillary, Equipment Design, Circulating Cell-Free DNA, [SDV] Life Sciences [q-bio], 030104 developmental biology, chemistry, 030220 oncology & carcinogenesis, Hydrodynamics, Cell-Free Nucleic Acids, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, DNA
Abstract

International audience; We describe a technology to perform sizing and concentration analysis of double stranded DNA with a sensitivity of 10 fg/μL in an operating time of 20 min. The technology is operated automatically on a commercial capillary electrophoresis instrument using electro-hydrodynamic actuation. It relies on a new capillary device that achieves online concentration of DNA at the junction between two capillaries of different diameters, thanks to viscoelastic lift forces. Using a set of DNA ladders in the range of 100–1500 bp, we report a sizing accuracy and precision better than 3% and a concentration quantification precision of ∼20%. When the technology is applied to the analysis of clinical samples of circulating cell-free DNA (cfDNA), the measured cfDNA concentrations are in good correlation with those measured by digital PCR. Furthermore, the cfDNA size profiles indicate that the fraction of low molecular weight cfDNA in the range of 75–240 bp is a candidate biomarker to discriminate between healthy subjects and cancer patients. We conclude that our technology is efficient in analyzing highly diluted DNA samples and suggest that it will be helpful in translational and clinical research involving cfDNA.

Published
2018

47. High throughput single cell counting in droplet-based microfluidics

Author

Jeremy Vrignon, Valérie Taly, Heng Lu, Ouriel Caen, Zakaria El Harrak, Eleonora Zonta, Jean-Christophe Baret, Philippe Nizard, Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique (MEPPOT - U1147), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche Paul Pascal (CRPP), Université de Bordeaux (UB)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), SIRIC CARPEMERC (FP7/2007-2013/ERC Starting Grant 306385-SoftRégion Aquitaine, ANR-15-CE11-0008,CHIPSeT,Caracterisation Structurale des Couplages entre Chromatine et Homeostasie Protéique par Combinaison d'Analyses de Coevolution et de Perturbation des Interfaces de Complexes a Haut-Debit(2015), ANR-10-IDEX-0003,IDEX BORDEAUX,Initiative d'excellence de l'Université de Bordeaux(2010), Teulet, Nadine, Caracterisation Structurale des Couplages entre Chromatine et Homeostasie Protéique par Combinaison d'Analyses de Coevolution et de Perturbation des Interfaces de Complexes a Haut-Debit - - CHIPSeT2015 - ANR-15-CE11-0008 - AAPG2015 - VALID, and Initiative d'excellence de l'Université de Bordeaux - - IDEX BORDEAUX2010 - ANR-10-IDEX-0003 - IDEX - VALID

Subjects
Multidisciplinary, Cell growth, Science, 010401 analytical chemistry, Microfluidics, 02 engineering and technology, Biology, 021001 nanoscience & nanotechnology, Bioinformatics, Cell counting, 01 natural sciences, Article, 0104 chemical sciences, Hemocytometer, Medicine, Droplet-based microfluidics, Viability assay, [PHYS.COND]Physics [physics]/Condensed Matter [cond-mat], 0210 nano-technology, Cell encapsulation, Biological system, Throughput (business), [PHYS.COND] Physics [physics]/Condensed Matter [cond-mat]
Abstract

Droplet-based microfluidics is extensively and increasingly used for high-throughput single-cell studies. However, the accuracy of the cell counting method directly impacts the robustness of such studies. We describe here a simple and precise method to accurately count a large number of adherent and non-adherent human cells as well as bacteria. Our microfluidic hemocytometer provides statistically relevant data on large populations of cells at a high-throughput, used to characterize cell encapsulation and cell viability during incubation in droplets.

Published
2017

48. EP364 Prognostic value of ddPCR HPV in lymph nodes of early cervical cancer

Author

Hélène Péré, Valérie Taly, D Veyer, M Wack, B Rance, Meriem Koual, M Deloménie, Vincent Balaya, A S Bats, Charlotte Ngo, H Bonsang-Kitzis, M-A Lefrère-Belda, H T Nguyen-Xuan, Patrice Mathevet, R Montero Macias, Marion Mandavit, and Fabrice Lecuru

Subjects
Oncology, Cervical cancer, medicine.medical_specialty, business.industry, medicine.disease, DNA extraction, medicine.anatomical_structure, Internal medicine, Nodal status, medicine, Biomarker (medicine), Digital polymerase chain reaction, Lymph, business, Pathological, Lymph node
Abstract

Introduction/Background In early stages (Ia to IIa1) of cervical cancer (CC), nodal status is the cornerstone of treatment algorithm and patients with nodal metastasis (N+) present worse survival. However, 10–15% of patients without nodal metastasis (N0) present the same survival to patients N+. No clinical, imaging or pathological risk-factor is today effective to select these patients for different management. In HPV-related cancers, like CC, HPV DNA could be assimilated to tumoral DNA and represent an interesting marker of node metastasis. In this study, we evaluate the presence of tumoral HPV DNA in pelvic lymph nodes (PLN) by new ultrasensitive droplet-based digital polymerase chain reaction (ddPCR) to explore it as a new additional biomarker to classify early stages of CC. Methodology This study was conducted on 30 patients from Senticol prospective trials. Formalin-fixed paraffin (FFPE) embedded tumours and associated pelvic lymph nodes samples were available for only 25 patients in our institution. In HPV16-positive tumour patients (n=18; 3 patients were N+ and 15 N0), we performed the specific HPV16 ddPCR in associated pelvic lymph node samples after a manual DNA extraction. Simultaneously, albumin was detected as internal control. Results Unfortunately, 22.2% of PLN were not interpretable (NI) with a very low level of albumin suggesting that no sufficient amount of DNA was extracted for HPV16 tumoral DNA interpretation. It could be explained by DNA destruction over time considering that FFPE biopsies were collected from 2005 to 2011. Insufficient follow-up data is Conclusion As expected, HPV16 tumoral DNA was detected in N+ patient PLN. In the N0 group, 40% (2/5) of patients with HPV16-positive PLN have relapsed regarding 12,5% (1/8) relapsing patients in the group of HPV16-negative PLN. These interesting preliminary results invite to test this new molecular biomarker in largest cohorts to evaluate its interest in early CC management. Disclosure Nothing to disclose.

Published
2019

49. Isothermal digital detection of microRNA using background-free molecular circuit

Author

Anne-Sophie Kirstetter, Valérie Taly, Yannick Rondelez, Kaori Nara, Guillaume Gines, and Roberta Menezes

Subjects
Regulation of gene expression, 0303 health sciences, 03 medical and health sciences, 010405 organic chemistry, Chemistry, microRNA, Loop-mediated isothermal amplification, Molecular programming, Computational biology, 01 natural sciences, Isothermal process, 030304 developmental biology, 0104 chemical sciences
Abstract

MicroRNA, a class of transcripts involved in the regulation of gene expression, are emerging as promising disease-specific biomarkers accessible from tissues or bodily fluids. However, their accurate quantification from biological samples remains challenging. We report a sensitive and quantitative microRNA method using an isothermal amplification chemistry adapted to a droplet digital readout. Building on molecular programming concepts, we design DNA circuit that converts, threshold, amplifies and report the presence of a specific microRNA, down to the femtomolar concentration. Using a leak-absorption mechanism, we were able to suppress non-specific amplification, classically encountered in other exponential amplification reactions. As a result, we demonstrate that this isothermal amplification scheme is adapted to digital counting of microRNA: by partitioning the reaction mixture into water-in-oil droplets, resulting in single microRNA encapsulation and amplification, the method provides absolute target quantification. The modularity of our approach enables to repurpose the assay for various microRNA sequences.

Published
2019
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50. BRAF Mutation Status in Circulating Tumor DNA from Patients with Metastatic Colorectal Cancer: Extended Mutation Analysis from the AGEO RASANC Study

Author

Anne Thirot-Bidault, Olivier Bouché, Romain Coriat, Jean-Marc Gornet, Julien Taieb, Lysiane Marthey, Pascal Artru, I. Baumgaertner, David Tougeron, Thierry Lecomte, Corinne Normand, Valérie Taly, Dewi Vernerey, Florence Mary, Hélène Blons, Jean-Baptiste Bachet, Romain Cohen, Aurélia Meurisse, Samy Louafi, Thomas Aparicio, Pierre Laurent-Puig, Léo Mas, Service d'Hépato-Gastro-Entérologie [CHU Pitié-Salpêtrière], CHU Pitié-Salpêtrière [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Association des Gastroentérologues Oncologues [Paris] (AGEO), Sorbonne Université (SU), Université Pierre et Marie Curie - Paris 6 (UPMC), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Service d'Hépato-gastroentérologie [HU Robert Debré, Reims], Hôpital universitaire Robert Debré [Reims], Service de gastroenterologie [CHU HEGP], Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Service d'Oncologie Médicale [CHU Saint -Antoine], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Interactions hôte-greffon-tumeur, ingénierie cellulaire et génique - UFC (UMR INSERM 1098) (RIGHT), Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC), Service d'hépato-gastro-entérologie [Hôpital Saint-Louis, APHP], Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Service de Gastroentérologie [Hôpital Privé Jean Mermoz, Lyon], Hôpital privé Jean-Mermoz [Lyon] (Ramsay-GDS), Service de Gastroenterologie [CH Sud Francilien, Corbeil-Essonnes], Centre Hospitalier Sud Francilien [Corbeil-Essonnes] (CH Sud Francilien), Service de Gastroenterologie [GH Nord Essonne, Longjumeau], Groupe Hospitalier Nord Essonne [Longjumeau], Service d'Hépato-gastro-entérologie [APHP Kremlin-Bicêtre], AP-HP Hôpital Bicêtre (Le Kremlin-Bicêtre), Service d'Oncologie médicale [CHU Henri Mondor], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Henri Mondor, Service de Gastro-entérologie [CHU Cochin], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Service d’hépato-gastro-entérologie et assistance nutritive [CHU de Poitiers], Centre hospitalier universitaire de Poitiers (CHU Poitiers), Service de Gastroentérologie [CHRU Tours], Centre Hospitalier Régional Universitaire de Tours (CHRU TOURS), Service de Gastroenterologie [Hôpital Avicenne, APHP], Hôpital Avicenne [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance Publique - Hôpitaux de Paris, Service d'hépato-gastroentérologie [Hôpital Antoine Béclère, APHP], AP-HP - Hôpital Antoine Béclère [Clamart], Service de biochimie [AP-HP Hôpital Européen Georges Pompidou], Merck Serono S.A.S. (France), an aliate of Merck KGaA (Darmstadt, Germany), supported this trial through grants to AGEO, but had no role in study design, data collection, data analysis, data interpretation, or writing of the report. The AGEO Group sponsored the trial and investigators were responsible for study design,data collection, data analysis, and data interpretation., Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Service d'Oncologie Médicale [CHU Saint-Antoine], CHU Saint-Antoine [AP-HP], Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), TALY, Valerie, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Franche-Comté (UFC), and Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])

Subjects
0301 basic medicine, Oncology, Cancer Research, medicine.medical_specialty, Colorectal cancer, Concordance, [SDV]Life Sciences [q-bio], colorectal cancer, [SDV.CAN]Life Sciences [q-bio]/Cancer, lcsh:RC254-282, Article, Metastasis, methylated biomarker, 03 medical and health sciences, 0302 clinical medicine, [SDV.CAN] Life Sciences [q-bio]/Cancer, Internal medicine, Medicine, Prospective cohort study, circulating tumor DNA, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, accuracy, business.industry, lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens, medicine.disease, digestive system diseases, 3. Good health, [SDV] Life Sciences [q-bio], 030104 developmental biology, Circulating tumor DNA, 030220 oncology & carcinogenesis, NGS, Mutation (genetic algorithm), Mutation testing, Biomarker (medicine), business, liver metastases, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract

In patients with metastatic colorectal cancer (mCRC), RAS and BRAF mutations are currently determined by tumor sample analysis. Here, we report BRAF mutation status analysis in paired tumor tissue and plasma samples of mCRC patients included in the AGEO RASANC prospective cohort study. Four hundred and twenty-five patients were enrolled. Plasma samples were analyzed by next-generation sequencing (NGS). When no mutation was identified, we used two methylated specific biomarkers (digital droplet PCR) to determine the presence or absence of circulating tumor DNA (ctDNA). Patients with conclusive ctDNA results were defined as those with at least one mutation or one methylated biomarker. The kappa coefficient and accuracy were 0.79 (95% CI: 0.67&ndash, 0.91) and 97.3% (95% CI: 95.2&ndash, 98.6%) between the BRAF status in plasma and tissue for patients with available paired samples (n = 405), and 0.89 (95% CI: 0.80&ndash, 0.99) and 98.5% (95% CI: 96.4&ndash, 99.5%) for those with conclusive ctDNA (n = 323). The absence of liver metastasis was the main factor associated to inconclusive ctDNA results. In patients with liver metastasis, the kappa coefficient was 0.91 (95% CI, 0.81&ndash, 1.00) and accuracy was 98.6% (95% CI, 96.5&ndash, 99.6%). We demonstrate satisfying concordance between tissue and plasma BRAF mutation detection, especially in patients with liver metastasis, arguing for plasma ctDNA testing for routine BRAF mutation analysis in these patients.

Published
2019

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