A single droplet digital PCR for ESR1 activating mutations detection in plasma.

Activating mutations in the estrogen receptor 1 (ESR1) gene confer resistance to aromatase inhibitors (AI), and may be targeted by selective estrogen receptor downregulators. We designed a multiplex droplet digital PCR (ddPCR), which combines a drop-off assay, targeting the clustered hotspot mutations found in exon 8, with an unconventional assay interrogating the E380Q mutation in exon 5. We assessed its sensitivity in vitro using synthetic oligonucleotides, harboring E380Q, L536R, Y537C, Y537N, Y537S, or D538G mutations. Further validation was performed on plasma samples from a prospective study and compared with next generation sequencing (NGS) data. The multiplex ESR1-ddPCR showed a high sensitivity with a limit of detection ranging from 0.07 to 0.19% in mutant allele frequency. The screening of plasma samples from patients with AI-resistant metastatic breast cancer identified ESR1 mutations in 29% of them, all mutations being confirmed by NGS. In addition, this test identifies patients harboring polyclonal alterations. Furthermore, the monitoring of circulating tumor DNA using this technique during treatment follow-up predicts the clinical benefit of palbociclib-fulvestrant. The multiplex ESR1-ddPCR detects, in a single reaction, the most frequent ESR1 activating mutations with good sensitivity. This method allows real-time liquid biopsy for ESR1 mutation monitoring in large cohorts of patients.

Toronto Workshop on Late Recurrence in Estrogen Receptor-Positive Breast Cancer: Part 2: Approaches

Late disease recurrence (more than 5 years after initial diagnosis) represents a clinical challenge in the treatment and management of estrogen receptor-positive breast cancer (BC). An international workshop was convened in Toronto, Canada, in February 2018 to review the current understanding of late recurrence and to identify critical issues that require future study. The underlying biological causes of late recurrence are complex, with the processes governing cancer cell dormancy, including immunosurveillance, cell proliferation, angiogenesis, and cellular stemness, being integral to disease progression. These critical processes are described herein as well as their role in influencing risk of recurrence. Moreover, observational and interventional clinical trials are proposed, with a focus on methods to identify patients at risk of recurrence and possible strategies to combat this in patients with estrogen receptor-positive BC. Because the problem of late BC recurrence of great importance, recent advances in disease detection and patient monitoring should be incorporated into novel clinical trials to evaluate approaches to enhance patient management. Indeed, future research on these issues is planned and will offer new options for effective late recurrence treatment and prevention strategies.

Toronto Workshop on Late Recurrence in Estrogen Receptor-Positive Breast Cancer: Part 1: Late

Disease recurrence (locoregional, distant) exerts a significant clinical impact on the survival of estrogen receptor-positive breast cancer patients. Many of these recurrences occur late, more than 5 years after original diagnosis, and represent a major obstacle to the effective treatment of this disease. Indeed, methods to identify patients at risk of late recurrence and therapeutic strategies designed to avert or treat these recurrences are lacking. Therefore, an international workshop was convened in Toronto, Canada, in February 2018 to review the current understanding of late recurrence and to identify critical issues that require future study. In this article, the major issues surrounding late recurrence are defined and current approaches that may be applicable to this challenge are discussed. Specifically, diagnostic tests with potential utility in late-recurrence prediction are described as well as a variety of patient-related factors that may influence recurrence risk. Clinical and therapeutic approaches are also reviewed, with a focus on patient surveillance and the implementation of extended endocrine therapy in the context of late-recurrence prevention. Understanding and treating late recurrence in estrogen receptor-positive breast cancer is a major unmet clinical need. A concerted effort of basic and clinical research is required to confront late recurrence and improve disease management and patient survival.

Mutation Detection and Dynamics in Meningeal Carcinomatosis in Breast Cancer.

mutation is frequently encountered in hormone receptor (HR)-positive, human epidermal growth factor receptor 2 (HER2)-negative metastatic breast cancer (MBC), especially after aromatase inhibitor (AI) therapy, as a mechanism of resistance to endocrine therapy. Circulating tumor DNA-based detection of mutation in plasma has been demonstrated as a prognostic and predictive factor for poor outcomes in subsequent AI therapy. In this case report, for the first time, we describe the detection of mutation (p.Tyr537Ser) only in the cerebrospinal fluid (CSF) and not in the plasma of a patient with isolated leptomeningeal progression who was treated with AI for HR-positive, HER2-negative MBC (bone metastasis only). Circulating tumor DNA levels also appeared to be correlated with clinical evolution. We suggest that in the presence of isolated leptomeningeal metastasis and when tamoxifen or AI has been prescribed for HR-positive MBC, CSF should be screened for mutations to potentially adjust systemic treatment.

Clinical Correlations of Programmed Cell Death Ligand 1 Status in Liquid and Standard Biopsies in

Data regarding the prognostic value of programmed cell death ligand 1 (PD-L1) expression on circulating tumor cells (CTCs) are lacking. However, CTCs could represent an alternative approach to serial biopsies, allowing real-time monitoring of cancer phenotype.

Serial analysis of circulating tumor cells in metastatic breast cancer receiving first-line

We examined the prognostic significance of circulating tumor cell (CTC) dynamics during treatment in metastatic breast cancer (MBC) patients receiving first-line chemotherapy.

HER2 expression on tumor-derived extracellular vesicles and circulating tumor cells in metastatic

Tumor-derived extracellular vesicles (tdEVs) and circulating tumor cells (CTCs) in the blood of metastatic cancer patients associate with poor outcomes. In this study, we explored the human epidermal growth factor receptor 2 (HER2) expression on CTCs and tdEVs of metastatic breast cancer patients.

Plasma thymidine kinase 1 activity and outcome of ER+ HER2- metastatic breast cancer patients

Previous cohort studies have reported plasma TK1 activity (pTKa) as a potential prognostic biomarker in estrogen receptor-positive (ER+) HER2-negative (HER2-) metastatic breast cancer (MBC). In this prospective study, we report here the prognostic impact of pTKa in ER+/HER2- MBC patients treated with endocrine therapy and CDK4/6 inhibitor.

Efficacy of Circulating Tumor Cell Count-Driven vs Clinician-Driven First-line Therapy Choice in

The choice between chemotherapy and endocrine therapy as first-line treatment for hormone receptor-positive, ERBB2 (also known as HER2)-negative metastatic breast cancer is usually based on the presence of clinical features associated with a poor prognosis. In this setting, a high circulating tumor cell (CTC) count (≥5 CTCs/7.5 mL) is a strong adverse prognostic factor for overall survival and progression-free survival (PFS).

Clinical utility of circulating tumor cells: an update.

The prognostic role of circulating tumor cells (CTCs) has been clearly demonstrated in many types of cancer. However, their roles in diagnostic and treatment strategies remain to be defined. In this review, we present an overview of the current clinical validity of CTCs in non-metastatic and metastatic cancer, and the main studies or concepts investigating the clinical utility of CTCs. In particular, we focus on breast-, lung-, colorectal- and prostate cancer. Two major topics concerning the clinical utility of CTC are discussed: treatment based on CTC count or CTC variations; and treatment based on the molecular characteristics of CTCs. Although some of these studies are inconclusive, many are still ongoing, and their results could help to define the role of CTCs in the management of cancers. A summary of published or ongoing phase II-III trials is also presented.

Genuage

Genuage for multidimensional Point-cloud data visualization and analysis

Single-molecule localization microscopy (SMLM) is becoming the method of choice in several biological studies. SMLM explores the cellular structures and composition at the nanometric scale. Single-molecule tracking provides also valuable information about the molecular interaction.

New optical methods are now being developed and the complexity of the recorded data is increasing. This raises many concerns regarding data representation, interaction and analysis.

Genuage is an open-source software dedicated for multidimensional point-cloud data visualization and analysis.

Genuage features two visualization modes: (i) a desktop mode for a simple visualization of the data sets on a 2D screen and (ii) a virtual reality mode for an immersive experience inside the point cloud.

PictureGenuage

The source codes can be found here:

https://github.com/Genuage/Genuage

the different releases:

https://github.com/Genuage/Genuage/releases

Check out our YouTube channel : https://www.youtube.com/channel/UC2Z1xaGfhUNkEqdHXiJwkAg/videos 

Genuage is continuously being updates with increasing functionalities.

Current functionalities:

  • Desktop and Virtual reality modes
  • Can represent: 2D and 3D super-resolution data sets, single-particle trajectories, 3D molecular orientations, multicolor
  • Easy visual parameter reconfiguration : Column reassignment, scaling, point size, color map…
  • Data thresholding
  • 3D selection tools
  • Clipping plane at arbitrary angle
  • Trajectory analysis
  • Density calculation
  • Interfaceable with MATLAB and Python
  • Compatible with user defined dlls

People actively involved in this project:

Thomas Blanc (Institut Curie), Mohamed El-Beheiry (Institut Curie and Institut Pasteur), Jean-Baptiste Masson (Institut Pasteur) and Bassam Hajj (Institut Curie)

 

In the news:

https://institut-curie.org/actualite/innovation/genuage-4-dimensional-visualization-and-analysis-super-resolution-microscopy

https://www.pasteur.fr/en/research-journal/news/genuage-virtual-reality-supporting-super-resolution-microscopy 

Genuage

Genuage for multidimensional Point-cloud data visualization and analysis

Single-molecule localization microscopy (SMLM) is becoming the method of choice in several biological studies. SMLM explores the cellular structures and composition at the nanometric scale. Single-molecule tracking provides also valuable information about the molecular interaction.

New optical methods are now being developed and the complexity of the recorded data is increasing. This raises many concerns regarding data representation, interaction and analysis.

Genuage is an open-source software dedicated for multidimensional point-cloud data visualization and analysis.

Genuage features two visualization modes: (i) a desktop mode for a simple visualization of the data sets on a 2D screen and (ii) a virtual reality mode for an immersive experience inside the point cloud.

PictureGenuage

The source codes can be found here:

https://github.com/Genuage/Genuage

the different releases:

https://github.com/Genuage/Genuage/releases

Check out our YouTube channel : https://www.youtube.com/channel/UC2Z1xaGfhUNkEqdHXiJwkAg/videos 

Genuage is continuously being updates with increasing functionalities.

Current functionalities:

  • Desktop and Virtual reality modes
  • Can represent: 2D and 3D super-resolution data sets, single-particle trajectories, 3D molecular orientations, multicolor
  • Easy visual parameter reconfiguration : Column reassignment, scaling, point size, color map…
  • Data thresholding
  • 3D selection tools
  • Clipping plane at arbitrary angle
  • Trajectory analysis
  • Density calculation
  • Interfaceable with MATLAB and Python
  • Compatible with user defined dlls

People actively involved in this project:

Thomas Blanc (Institut Curie), Mohamed El-Beheiry (Institut Curie and Institut Pasteur), Jean-Baptiste Masson (Institut Pasteur) and Bassam Hajj (Institut Curie)

In the news:

https://institut-curie.org/actualite/innovation/genuage-4-dimensional-visualization-and-analysis-super-resolution-microscopy

https://www.pasteur.fr/en/research-journal/news/genuage-virtual-reality-supporting-super-resolution-microscopy 

Apprentissage statistique et modélisation des systèmes biologiques

De nombreuses technologies récentes permettent d’observer les organismes vivants à une échelle jusqu’alors inexplorée. Par exemple, les techniques de biopuces ou de séquençage massivement parallèle permettent de lire l’information génétique complète d’un échantillon biologique ; la spectrométrie de masse permet de quantifier le protéome ; la vidéo-microscopie à haute résolution permet de suivre les évolutions de populations de cellules ; et les méthodes de criblage à haut débit sont utilisées pour caractériser les effets biologiques de grandes quantités de molécules. Enfin, les dossiers patient électroniques contiennent de grands volumes de texte, d’images, ou de résultats d’analyses qui décrivent la dynamique du diagnostic et de la réponse au traitement.

Toutes ces technologies génèrent de très grandes quantités de données, qu’il est souvent difficile d’interpréter directement. Afin de mieux exploiter ces masses de données et d’en extraire des informations biologiques et médicales pertinentes, notre équipe développe des méthodes mathématiques et des algorithmes innovants, basés sur notre expertise en modélisation mathématique, statistique, apprentissage automatique, bioimage informatics et biologie structurale.

Nous développons de nouveaux outils pour examiner différentes questions biologiques et médicales, notamment:

  • Biologie et biologie des systèmes in silico : Nous développons des approches innovantes pour la reconstruction statistique de réseaux biologiques, l’analyse et la modélisation de la progression tumorale au niveau génomique, transcriptomique et épigénétique, l’annotation automatique de gènes et d’éléments fonctionnels par intégration de données hétérogènes et complexes, en particulier les données provenant de séquençage à haut débit ou de vidéo-microscopie.
  • Médecine prédictive et de précision : Nous développons des méthodes pour la classification de tumeurs et l’identification de biomarqueurs diagnostiques, pronostiques et prédictifs de réponse à un traitement. Ces classifieurs utilisent un grand nombre de paramètres mesurables comme les données cliniques du patient, l’ensemble des mutations somatiques de la tumeur, l’expression de gènes et de transcrits alternatifs, ou les modifications structurelles de l’ADN, et sont construits grâce à des méthodes d’apprentissage statistique en grande dimension.
  • Développement de médicaments : nous développons de nouvelles méthodes pour le criblage virtuel et la chémogénomique in silico, afin d’aider à identifier ou créer de petites molécules capables de se lier spécifiquement à des cibles thérapeutiques par modélisation informatique. Ces méthodes combinent des modèles basées sur la représentation sous forme de séquence, de graphe, ou d’information 3D des molécules et des procédures statistiques innovantes, et peuvent par exemple aider à prédire les cibles thérapeutiques, les profils d’activité ou les effets indésirables de candidats médicaments.
Data, machine learning, applications.

Le machine learning permet d’utiliser de nombreux types de données pour des applications en imagerie biomédicale, biologie des systèmes, conception de médicaments, et médecine de précision.

Shreyangi Chakraborty

Lack of evidence for CDK12 as an ovarian cancer predisposing gene.

CDK12 variants were investigated as a genetic susceptibility to ovarian cancer in a series of 416 unrelated and consecutive patients with ovarian carcinoma and who carry neither germline BRCA1 nor BRCA2 pathogenic variant. The presence of CDK12 variants was searched in germline DNA by massive parallel sequencing on pooled DNAs. The lack of detection of deleterious variants and the observed proportion of missense variants in the series of ovarian carcinoma patients as compared with all human populations strongly suggests that CDK12 is not an ovarian cancer predisposing gene.

DNA repair functional analyses of NBN hypomorphic variants associated with NBN-related infertility.

Nijmegen breakage syndrome caused by biallelic pathogenic variants of the DNA-damage response gene NBN, is characterized by severe microcephaly, cancer proneness, infertility, and karyotype abnormalities. We previously reported NBN variants in siblings suffering from fertility defects. Here, we identify a new founder NBN variant (c.442A>G, p.(Thr148Ala)) in Lebanese patients associated with isolated infertility. Functional analyses explored preserved or altered functions correlated with their remarkably mild phenotype. Transcript and protein analyses supported the use of an alternative transcript with in-frame skipping of exons 4-5, leading to p84-NBN protein with a preserved forkhead-associated (FHA) domain. The level of NBN was dramatically reduced and the MRN complex delocalized to the cytoplasm. Interestingly, ataxia-elangiectasia mutated (ATM) also shifted from the nucleus to the cytoplasm, suggesting some interaction between ATM and the MRN complex at a steady state. The ATM pathway activation, attenuated in typical patients with NBS, appeared normal under camptothecin treatment in these new NBN-related infertile patients. Cell cycle checkpoint defect was present in these atypical patients, although to a lesser extent than in typical patients with NBS. In conclusion, we report three new NBN-related infertile patients and we suggest that preserved FHA domain could be responsible for the mild phenotype and intermediate DNA-damage response defects.

Uveal melanoma.

Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. UMs are usually initiated by a mutation in GNAQ or GNA11, unlike cutaneous melanomas, which usually harbour a BRAF or NRAS mutation. The annual incidence in Europe and the USA is ~6 per million population per year. Risk factors include fair skin, light-coloured eyes, congenital ocular melanocytosis, ocular melanocytoma and the BAP1-tumour predisposition syndrome. Ocular treatment aims at preserving the eye and useful vision and, if possible, preventing metastases. Enucleation has largely been superseded by various forms of radiotherapy, phototherapy and local tumour resection, often administered in combination. Ocular outcomes are best with small tumours not extending close to the optic disc and/or fovea. Almost 50% of patients develop metastatic disease, which usually involves the liver, and is usually fatal within 1 year. Although UM metastases are less responsive than cutaneous melanoma to chemotherapy or immune checkpoint inhibitors, encouraging results have been reported with partial hepatectomy for solitary metastases, with percutaneous hepatic perfusion with melphalan or with tebentafusp. Better insight into tumour immunology and metabolism may lead to new treatments.

High-Accuracy Determination of Microsatellite Instability Compatible with Liquid Biopsies.

Microsatellite instability (MSI) has recently emerged as a predictive pan-tumor biomarker of immunotherapy efficacy, stimulating the development of diagnostic tools compatible with large-scale screening of patients. In this context, noninvasive detection of MSI from circulating tumor DNA stands as a promising diagnostic and posttreatment monitoring tool.

Three new cases of ataxia-telangiectasia-like disorder: No impairment of the ATM pathway, but

Ataxia-telangiectasia-like disorder (ATLD) is a rare genomic instability syndrome caused by biallelic variants of MRE11 (meiotic recombination 11) characterized by progressive cerebellar ataxia and typical karyotype abnormalities. These symptoms are common to those of ataxia-telangiectasia, which is consistent with the key role of MRE11 in ataxia-telangiectasia mutated (ATM) activation after DNA double-strand breaks. Three unrelated French patients were referred with ataxia. Only one had typical karyotype abnormalities. Unreported biallelic MRE11 variants were found in these three cases. Interestingly, one variant (c.424G>A) was present in two cases and haplotype analysis strongly suggested a French founder variant. Variants c.544G>A and c.314+4_314+7del lead to splice defects. The level of MRE11 in lymphoblastoid cell lines was consistently and dramatically reduced. Functional consequences were evaluated on activation of the ATM pathway via phosphorylation of ATM targets (KAP1 and CHK2), but no consistent defect was observed. However, an S-phase checkpoint activation defect after camptothecin was observed in these patients with ATLD. In conclusion, we report the first three French ATLD patients and a French founder variant, and propose an S-phase checkpoint activation study to evaluate the pathogenicity of MRE11 variants.

Prospective validation in epithelial tumors of a gene expression predictor of liver metastasis

Predicting the risk of liver metastasis can have important prognostic and therapeutic implications, given the availability of liver-directed therapy. Uveal melanoma has a striking predisposition for liver metastasis despite the absence of anatomical proximity. Understanding its biology may uncover factors promoting liver metastasis in other malignancies. We quantified gene expression by RNAseq in 76 uveal melanomas and combined with public data in a meta-analysis of 196 patients. The meta-analysis of uveal melanoma gene expression identified 63 genes which remained prognostic after adjustment for chromosome 3 status. Two genes, PTP4A3 and JPH1, were selected by L1-penalized regression and combined in a prognostic score. The score predicted liver-specific relapse in a public pan-cancer dataset and in two public colorectal cancer datasets. The score varied between colorectal consensus molecular subtypes (CMS), as did the risk of liver relapse, which was lowest in CMS1. Additional prospective validation was done by real-time PCR in 463 breast cancer patients. The score was significantly correlated with liver relapse in hormone receptor positive tumors. In conclusion, the expression of PTP4A3 and JPH1 correlates with risk of liver metastasis in colorectal cancer and breast cancer. The underlying biological mechanism is an interesting area for further research.