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Supplementary Figure from The Polarity and Specificity of Antiviral T Lymphocyte Responses Determine Susceptibility to SARS-CoV-2 Infection in Patients with Cancer and Healthy Individuals

Supplementary Data from The Polarity and Specificity of Antiviral T Lymphocyte Responses Determine Susceptibility to SARS-CoV-2 Infection in Patients with Cancer and Healthy Individuals

Vaccination against coronavirus disease 2019 (COVID-19) relies on the in-depth understanding of protective immune responses to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). We characterized the polarity and specificity of memory T cells directed against SARS-CoV-2 viral lysates and peptides to determine correlates with spontaneous, virus-elicited, or vaccine-induced protection against COVID-19 in disease-free and cancer-bearing individuals. A disbalance between type 1 and 2 cytokine release was associated with high susceptibility to COVID-19. Individuals susceptible to infection exhibited a specific deficit in the T helper 1/T cytotoxic 1 (Th1/Tc1) peptide repertoire affecting the receptor binding domain of the spike protein (S1-RBD), a hotspot of viral mutations. Current vaccines triggered Th1/Tc1 responses in only a fraction of all subject categories, more effectively against the original sequence of S1-RBD than that from viral variants. We speculate that the next generation of vaccines should elicit Th1/Tc1 T-cell responses against the S1-RBD domain of emerging viral variants.Significance:This study prospectively analyzed virus-specific T-cell correlates of protection against COVID-19 in healthy and cancer-bearing individuals. A disbalance between Th1/Th2 recall responses conferred susceptibility to COVID-19 in both populations, coinciding with selective defects in Th1 recognition of the receptor binding domain of spike.See related commentary by McGary and Vardhana, p. 892.This article is highlighted in the In This Issue feature, p. 873

Supplementary Figure from Immune Infiltrate and Tumor Microenvironment Transcriptional Programs Stratify Pediatric Osteosarcoma into Prognostic Groups at Diagnosis

Supplementary Table from Immune Infiltrate and Tumor Microenvironment Transcriptional Programs Stratify Pediatric Osteosarcoma into Prognostic Groups at Diagnosis

The outcomes of adolescents/young adults with osteosarcoma have not improved in decades. The chaotic karyotype of this rare tumor has precluded the identification of prognostic biomarkers and patient stratification. We reasoned that transcriptomic studies should overcome this genetic complexity. RNA sequencing (RNA-seq) of 79 osteosarcoma diagnostic biopsies identified stable independent components that recapitulate the tumor and microenvironment cell composition. Unsupervised classification of the independent components stratified this cohort into favorable (G1) and unfavorable (G2) prognostic tumors in terms of overall survival. Multivariate survival analysis ranked this stratification as the most influential variable. Functional characterization associated G1 tumors with innate immunity and G2 tumors with angiogenic, osteoclastic, and adipogenic activities as well as PPARγ pathway upregulation. A focused gene signature that predicted G1/G2 tumors from RNA-seq data was developed and validated within an independent cohort of 82 osteosarcomas. This signature was further validated with a custom NanoString panel in 96 additional osteosarcomas. This study thus proposes new biomarkers to detect high-risk patients and new therapeutic options for osteosarcoma.Significance:These findings indicate that the osteosarcoma microenvironment composition is a major feature to identify hard-to-treat patient tumors at diagnosis and define the biological pathways and potential actionable targets associated with these tumors.

The outcomes of adolescents/young adults with osteosarcoma have not improved in decades. The chaotic karyotype of this rare tumor has precluded the identification of prognostic biomarkers and patient stratification. We reasoned that transcriptomic studies should overcome this genetic complexity. RNA sequencing (RNA-seq) of 79 osteosarcoma diagnostic biopsies identified stable independent components that recapitulate the tumor and microenvironment cell composition. Unsupervised classification of the independent components stratified this cohort into favorable (G1) and unfavorable (G2) prognostic tumors in terms of overall survival. Multivariate survival analysis ranked this stratification as the most influential variable. Functional characterization associated G1 tumors with innate immunity and G2 tumors with angiogenic, osteoclastic, and adipogenic activities as well as PPARγ pathway upregulation. A focused gene signature that predicted G1/G2 tumors from RNA-seq data was developed and validated within an independent cohort of 82 osteosarcomas. This signature was further validated with a custom NanoString panel in 96 additional osteosarcomas. This study thus proposes new biomarkers to detect high-risk patients and new therapeutic options for osteosarcoma. Significance: These findings indicate that the osteosarcoma microenvironment composition is a major feature to identify hard-to-treat patient tumors at diagnosis and define the biological pathways and potential actionable targets associated with these tumors.

Background Currently, mammographic-based breast cancer screening (BCS) using age as the single criterion for population selection, apart from rare high-risk indications, is being questioned for its imperfect sensitivity (interval cancers) and specificity (false positive recalls), as well as the risk of over-diagnoses. BC risk scores incorporating personal and family history, breast mammographic density and genetic information based on a polygenic score (PRS) give a promisingly accurate likelihood of a woman developing invasive BC in the next 5 years. MyPeBS, a European Commission H2020-funded randomized clinical trial (NCT03672331) conducted in 6 countries (Belgium, France, Israel, Italy, Spain and UK) aims to demonstrate the usefulness of a risk-based screening approach to improve BCS in the general population. Methods MyPeBS’s primary objective is to show non-inferiority of the risk-stratified BCS arm in terms of incidence rate of breast cancer of stage 2 and higher, compared to the standard BCS arm. The key secondary objective, if non-inferiority is shown, is to demonstrate superiority of the risk-based screening arm. MyPeBS also assesses whether this strategy allows reduces morbidity (less false positives); is equally or more cost-effective, and impacts women’s understanding, awareness and emotional responses as compared to standard of care. Women aged 40 to 70 are eligible if they did not have prior BC or high risk-condition, and live in a participating country and area. Participants are randomized 1:1 between standard BCS according to country’s/region’s ongoing policy, or risk-stratified BCS, where screening frequency and method depend on their individual 5 year predicted risk of invasive BC (Table 1). Stratification factors include age, prior mammogram and country. Risk assessment uses a centralized genotyping on a saliva sample to assess PRS 313 (Mavaddat et al, Am J Hum Genet 2019), which is embedded in either a BCSC-derived score for women with at most one first-degree relative with BC; or Tyrer-Cuzick score for women with > 1 family member with breast cancer. Women participate for 4 years. Planned accrual is 85000. On June 30th, 2021, 13882 women have been randomized. Table 1.BC Screening schedule in MyPeBS’ Risk-based armRisk levelLow riskAverage riskHigh riskVery high riskNumerical definition (invasive breast cancer risk at 5 years) Citation Format: Suzette Delaloge, Paolo Giorgi Rossi, Michal Guindy, Fiona Gilbert, Jean-Benoit Burrion, Corinne Balleyguier, Marta Roman Exposito, Livia Giordano, Harry De Koning, Sandrine de Montgolfier, Stephane Ragusa, Damien Drubay, Marie-Eve Rouge-Bugat, Gareth D Evans, Debbie Keatley, Helene Blanche, Anne Boland, Emilien Gauthier, Aloys Dubois d'Aische, Cécile Vissac-Sabatier, Daniel Couch, Camille Baron, Jean-François Deleuze, Paul Pharoah, Stefan Michiels. Mypebs: An international randomized study comparing personalized, risk-stratified to standard breast cancer screening in women aged 40-70 [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr OT2-10-02.

The development of prognostic molecular signatures considering the inter-patient heterogeneity is a key challenge for the precision medicine. We propose a joint model of this heterogeneity and the patient survival, assuming that tumor expression results from a mixture of a subset of independent signatures. We deconvolute the omics data using a non-parametric independent component analysis with a double sparseness structure for the source and the weight matrices, corresponding to the gene-component and individual-component associations, respectively. In a simulation study, our approach identified the correct number of components and reconstructed with high accuracy the weight ([Formula: see text]0.85) and the source ([Formula: see text]0.75) matrices sparseness. The selection rate of components with high-to-moderate prognostic impacts was close to 95%, while the weak impacts were selected with a frequency close to the observed false positive rate ([Formula: see text]25%). When applied to the expression of 1063 genes from 614 breast cancer patients, our model identified 15 components, including six associated to patient survival, and related to three known prognostic pathways in early breast cancer (i.e. immune system, proliferation, and stromal invasion). The proposed algorithm provides a new insight into the individual molecular heterogeneity that is associated with patient prognosis to better understand the complex tumor mechanisms.

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is the consequence of platelet destruction by maternal alloantibodies against fetal human platelet antigens (HPA). This may result in intracranial haemorrhages (ICH) or even fetal death. Currently, fetal HPA genotyping is performed using invasive procedures. Here, we carried out a proof-of-concept study for non-invasive prenatal diagnosis of fetal platelet genotyping in four HPA systems (HPA-1, -3, -5 and-15) by droplet digital polymerase chain reaction (ddPCR) using cell-free DNA extracts from the plasma of 47 pregnant women with suspected, or history of, FNAIT. Results showed that 74% (35/47) of pregnant women presented incompatibility in at least one HPA system, and 38% (18/47) of cases presented HPA-1 incompatibility, including nine women with multiple incompatibilities. ICH occurred in one case of profound fetal thrombocytopenia with HPA-15 incompatibility, confirming the need for non-invasive prenatal genotyping in systems other than HPA-1. Fetal HPA genotypes predicted by ddPCR were confirmed in all FNAIT cases after amniocentesis or delivery. Fetal HPA genotyping on maternal plasma based on ddPCR is a fast, safe and reliable non-invasive method. This technique will be useful for the early identification of pregnancies at high risk of FNAIT requiring antenatal management to minimize the risk of fetal/neonatal haemorrhage.

Basal/squamous (Ba/Sq) subtype represents an intrinsic and robust group in the consensus molecular classification of muscle-invasive bladder cancer (MIBC), with poor outcome and controversial chemosensitivity. We aimed to investigate the spectrum of intratumor heterogeneity (ITH) in the Ba/Sq subtype. First, we validated a 29-gene NanoString CodeSet to predict the Ba/Sq subtype for FFPE samples. We identified heterogeneous Ba/Sq tumors in a series of 331 MIBC FFPE samples using dual GATA3/KRT5/6 immunohistochemistry (IHC). Heterogeneous regions with distinct immunostaining patterns were studied separately for gene expression using the 29-gene CodeSet, for mutations by targeted next-generation sequencing, and for copy number alteration (CNA) by microarray hybridization. Among 83 Ba/Sq tumors identified by GATA3/KRT5/6 dual staining, 19 tumors showed heterogeneity at the IHC level. In one third of the 19 cases, regions from the same tumor were classified in different distinct molecular subtypes. The mutational and CNA profiles confirmed the same clonal origin for IHC heterogeneous regions with possible subclonal evolution. Overall, two patterns of intratumoral heterogeneity (ITH) were observed in Ba/Sq tumors: low ITH (regions with distinct immunostaining, but common molecular subtype and shared CNA) or high ITH (regions with distinct immunostaining, molecular subtype, and CNA). These results showed multilayer heterogeneity in Ba/Sq MIBC. In view of personalized medicine, this heterogeneity adds complexity and should be taken into account for sampling procedures used for diagnosis and treatment choice. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Identifying individual mechanisms involved in complex diseases, such as cancer, is essential for precision medicine. Their characterization is particularly challenging due to the unknown relationships of high-dimensional omics data and their inter-patient heterogeneity. We propose to model individual gene expression as a combination of unobserved molecular mechanisms (molecular components) that may differ between the individuals. Considering a baseline molecular profile common to all individuals, these molecular components may represent molecular pathways differing from the population background. We defined an infinite sparse graphical independent component analysis (isgICA) to identify these molecular components. This model relies on double sparseness: the source matrix sparseness defines the subset of genes involved in each molecular component, whereas the weight matrix sparseness identifies the subset of molecular components associated with each patient. As the number of molecular components is unknown but likely high, we simultaneously inferred it and the weight matrix sparseness using the beta-Bernoulli process (BBP). We simulated data from a double sparse ICA with 10/30 components with specific sparseness structures for 100/500 individuals and 500/1000/5000 genes with different noise variance levels to evaluate the reconstruction of the latent structures by our model. For all simulations, the isgICA was able to reconstruct with higher accuracy than 2 state-of-the-art methods ( ica and fastICA) the number of components, the weight and source matrix sparsenesses (correlation simulated/estimated >.8). Applying our model to the expression of 1063 genes of 614 breast cancer patients, the isgICA identified 22 components. According to the source matrix, 7 of these 22 components seemed to be specifically related to 3 known molecular pathways with a prognostic effect in early breast cancer (immune system, proliferation, and stroma invasion). This proposed algorithm provides an insight into individual molecular heterogeneity to better understand complex disease mechanisms.

Vaccination against coronavirus disease 2019 (COVID-19) relies on the in-depth understanding of protective immune responses to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). We characterized the polarity and specificity of memory T cells directed against SARS-CoV-2 viral lysates and peptides to determine correlates with spontaneous, virus-elicited, or vaccine-induced protection against COVID-19 in disease-free and cancer-bearing individuals. A disbalance between type 1 and 2 cytokine release was associated with high susceptibility to COVID-19. Individuals susceptible to infection exhibited a specific deficit in the T helper 1/T cytotoxic 1 (Th1/Tc1) peptide repertoire affecting the receptor binding domain of the spike protein (S1-RBD), a hotspot of viral mutations. Current vaccines triggered Th1/Tc1 responses in only a fraction of all subject categories, more effectively against the original sequence of S1-RBD than that from viral variants. We speculate that the next generation of vaccines should elicit Th1/Tc1 T-cell responses against the S1-RBD domain of emerging viral variants. Significance: This study prospectively analyzed virus-specific T-cell correlates of protection against COVID-19 in healthy and cancer-bearing individuals. A disbalance between Th1/Th2 recall responses conferred susceptibility to COVID-19 in both populations, coinciding with selective defects in Th1 recognition of the receptor binding domain of spike. See related commentary by McGary and Vardhana, p. 892. This article is highlighted in the In This Issue feature, p. 873

Optimal vaccination and immunotherapy against coronavirus disease COVID-19 relies on the in-depth comprehension of immune responses determining the individual susceptibility to be infected by SARS-CoV-2 and to develop severe disease. We characterized the polarity and specificity of circulating SARS-CoV-2-specific T cell responses against whole virus lysates or 186 unique peptides derived from the SARS-CoV-2 or SARS-CoV-1 ORFeome on 296 cancer-bearing and 86 cancer-free individuals who were either from the pre-COVID-19 era (67 individuals) or contemporary COVID-19-free (237 individuals) or who developed COVID-19 (78 individuals) in 2020/21. The ratio between the prototypic T helper 1 (TH1) cytokine, interleukin-2, and the prototypic T helper 2 (TH2) cytokine, interleukin-5 (IL-5), released from SARS-CoV-2-specific memory T cells measured in early 2020, among SARS-CoV-2-negative persons, was associated with the susceptibility of these individuals to develop PCR-detectable SARS-CoV-2 infection in late 2020 or 2021. Of note, T cells from individuals who recovered after SARS-CoV-2 re-infection spontaneously produced elevated levels of IL-5 and secreted the immunosuppressive TH2 cytokine interleukin-10 in response to SARS-CoV-2 lysate, suggesting that TH2 responses to SARS-CoV-2 are inadequate. Moreover, individuals susceptible to SARS-CoV-2 infection exhibited a deficit in the TH1 peptide repertoire affecting the highly mutated receptor binding domain (RBD) amino acids (331-525) of the spike protein. Finally, current vaccines successfully triggered anti-RBD specific TH1 responses in 88% healthy subjects that were negative prior to immunization. These findings indicate that COVID-19 protection relies on TH1 cell immunity against SARS-CoV-2 S1-RBD which in turn likely drives the phylogenetic escape of the virus. The next generation of COVID-19 vaccines should elicit high-avidity TH1 (rather than TH2)-like T cell responses against the RBD domain of current and emerging viral variants.

Patients with cancer are at higher risk of severe coronavirus infectious disease 2019 (COVID-19), but the mechanisms underlying virus-host interactions during cancer therapies remain elusive. When comparing nasopharyngeal swabs from cancer and non-cancer patients for RT-qPCR cycle thresholds measuring acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in 1063 patients (58% with cancer, 89% COVID-19+), we found that malignant disease favors the magnitude and duration of viral RNA shedding concomitant with prolonged serum elevations of type 1 IFN that anticorrelated with anti-RBD IgG antibodies. Chronic viral RNA carriers exhibited the typical immunopathology of severe COVID-19 at the early phase of infection including circulation of immature neutrophils, depletion of non-conventional monocytes and a general lymphopenia that, however, was accompanied by a rise in plasmablasts, activated follicular T helper cells, and non-naive Granzyme B+ FasL+, EomehighTCF-1high, PD-1+CD8+ Tc1 cells. Virus-induced lymphopenia worsened cancer-associated lymphocyte loss, and low lymphocyte counts correlated with chronic SARS-CoV-2 RNA shedding, COVID-19 severity and a higher risk of cancer-related death in the first and second surge of the pandemic. Lymphocyte loss correlated with significant changes in metabolites from the polyamine and biliary salt pathways as well as increased blood DNA from Enterobacteriaceae and Micrococcaceae gut family members in long term viral carriers. We surmise that cancer therapies may exacerbate the paradoxical association between lymphopenia and COVID-19-related immunopathology, and that the prevention of COVID-19-induced lymphocyte loss may reduce cancer-associated death.

International audience; Limited experimental evidence bridges nutrition and cancer immunosurveillance. Here, we show that ketogenic diet (KD) - or its principal ketone body, 3-hydroxybutyrate (3HB), most specifically in intermittent scheduling - induced T cell-dependent tumor growth retardation of aggressive tumor models. In conditions in which anti-PD-1 alone or in combination with anti-CTLA-4 failed to reduce tumor growth in mice receiving a standard diet, KD, or oral supplementation of 3HB reestablished therapeutic responses. Supplementation of KD with sucrose (which breaks ketogenesis, abolishing 3HB production) or with a pharmacological antagonist of the 3HB receptor GPR109A abolished the antitumor effects. Mechanistically, 3HB prevented the immune checkpoint blockade-linked upregulation of PD-L1 on myeloid cells, while favoring the expansion of CXCR3+ T cells. KD induced compositional changes of the gut microbiota, with distinct species such as Eisenbergiella massiliensis commonly emerging in mice and humans subjected to carbohydrate-low diet interventions and highly correlating with serum concentrations of 3HB. Altogether, these results demonstrate that KD induces a 3HB-mediated antineoplastic effect that relies on T cell-mediated cancer immunosurveillance.

Aside from PD-L1 expression, biomarkers of response to immune checkpoint inhibitors (ICIs) in non-small-cell lung cancer (NSCLC) are needed. In a previous retrospective analysis, we documented that fecal Akkermansia muciniphila (Akk) was associated with clinical benefit of ICI in patients with NSCLC or kidney cancer. In the current study, we performed shotgun-metagenomics-based microbiome profiling in a large cohort of patients with advanced NSCLC (n = 338) treated with first- or second-line ICIs to prospectively validate the predictive value of fecal Akk. Baseline stool Akk was associated with increased objective response rates and overall survival in multivariate analyses, independent of PD-L1 expression, antibiotics, and performance status. Intestinal Akk was accompanied by a richer commensalism, including Eubacterium hallii and Bifidobacterium adolescentis, and a more inflamed tumor microenvironment in a subset of patients. However, antibiotic use (20% of cases) coincided with a relative dominance of Akk above 4.8% accompanied with the genus Clostridium, both associated with resistance to ICI. Our study shows significant differences in relative abundance of Akk that may represent potential biomarkers to refine patient stratification in future studies.

Summary Blood myeloid cells are known to be dysregulated in the coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2. It is unknown whether the innate myeloid response differs with disease severity, and whether markers of innate immunity discriminate high risk patients. Thus, we performed high dimensional flow cytometry and single cell RNA sequencing of COVID-19 patient peripheral blood cells and detected the disappearance of non-classical CD14LowCD16High monocytes, the accumulation of HLA-DRLow classical monocytes, and the release of massive amounts of calprotectin (S100A8/S100A9) in severe cases. Immature CD10LowCD101-CXCR4+/- neutrophils with an immuno-suppressive profile accumulated as well in blood and lungs, suggesting emergency myelopoiesis. We finally showed that calprotectin plasma level and a routine flow cytometry assay detecting decreased frequencies of non-classical monocytes could discriminate patients who develop a severe COVID-19 form, suggesting a predictive value that deserves prospective evaluation., Highlights • Severe patients accumulate HLA-DRLow monocytes and immature neutrophils in blood/lung. • Calprotectin level positively correlates with neutrophil count and disease severity. • Loss of non-classical monocytes could identify high risk of severe COVID-19.

International audience; Objective: Health risk behaviors (HRB) of childhood cancer survivors (CCS) are generally studied separately, despite the evidence suggesting that HRB are not independent. To our knowledge, few studies have examined HRB profiles in the former pediatric cancer patients. In this study, we identified HRB profiles and examined predictors engaging in unhealthy behaviors in CCS. Methods: We used data from a French cohort of CCS that includes five-year survivors diagnosed between 1945 and 2000 and treated before reaching age 18, in five centers in France. A total of 2961 adult CCS answered a self-reported questionnaire pertaining to HRB. Latent class analysis was used to identify HRB profiles combining physical activity, smoking, cannabis use, and alcohol drinking. Multinomial logistic analyses examined predictors for engaging in unhealthy behaviors. Results: Three HRB patterns emerged: “Low-risk” (n = 1846, 62.3%) included CCS who exhibited the highest frequency for usual physical activity and the lowest probabilities for current smoking or cannabis use, but most drank at least moderately; “Moderate-risk behaviors” (n = 291, 9.8%), and “High-risk behaviors” (n = 824, 27.8%) for CCS who exhibited the highest frequencies for current smoking, cannabis use, and heavy drinking. The multivariable regression revealed that male CCS, less educated or not married were significantly more likely to be in the high-risk behaviors group than the low-risk group. Conclusions: As CCS remain a vulnerable population, screening for HRB should be routinized in long-term follow-up care and interventions targeting multiple HRB simultaneously among survivors should be developed.

Background LY2780301, a dual inhibitor of protein kinase B (AKT) and the downstream effector p70 ribosomal protein S6 kinase (p70S6K), may inhibit progression in tumours relying on phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signalling pathway activation. This phase IB trial investigated the maximum tolerated dose (MTD), dose-limiting toxicities (DLTs), safety, pharmacokinetics (PK) and antitumour activity of LY2780301 plus gemcitabine in patients with advanced/metastatic solid tumours. Methods This was a non-randomised, open-label, dose escalation and dose expansion trial. Patients harbouring molecular alterations of the PI3K/AKT/mTOR pathway received once daily (QD) oral LY2780301 (400 or 500 mg) in combination with intravenous gemcitabine (750 or 1000 mg/m 2 ) on days 1, 8 and 15 of a 28-d cycle. Dose escalation followed a 3 + 3 design. Assessments included adverse events (AEs), PK and preliminary antitumour activity. Results Fifty patients (median age, 53 years; 74% female) predominantly with mutations/amplifications of PI3K (60%) and phosphatase and tensin homologue (PTEN) gene/protein inactivation (42%) were treated for up to 14 cycles. The MTD was LY2780301 500 mg QD with gemcitabine 750 mg/m 2 . DLTs during cycle 1 were grade IV thrombocytopenia, grade III skin rash and grade III increase in alkaline phosphatase, gamma glutamyltransferase and alanine aminotransferase, occurring in one patient each. Most common AEs were anaemia (84%), fatigue (84%), transaminase increase (74%), thrombocytopenia (74%), nausea/vomiting (70%), neutropenia (68%) and lymphopenia (56%). Among the efficacy-evaluable population, two patients (5%) had a partial response; the disease control rate was 74% at cycle 2. Conclusions Addition of LY2780301 to gemcitabine showed manageable toxicity and encouraging antitumour activity in patients with molecular alterations of the PI3K/AKT/mTOR pathway. Clinical trial registration number NCT02018874.

The aim of the current study was to conduct a pooled analysis of studies that have investigated the prognostic value of tumor-infiltrating lymphocytes (TILs) in early-stage triple negative breast cancer (TNBC)., SCOPUS: ar.j, info:eu-repo/semantics/published

International audience; Background and purpose The dose effect-effect relationship for cardiac diseases following radiotherapy suffers from uncertainties. Three dimensional coronary artery (CA) dose calculation after mediastinal Hodgkin lymphoma radiotherapy was performed, using the patient's coronary CT angiography (CCTA), and the relationship between the coronary arteries’ radiation doses and the risk of stenosis was estimated. Materials and methods Radiotherapy simulation CT scans and CCTAs of patients treated for a mediastinal Hodgkin lymphoma were used to merge thoracic and detailed cardiovascular anatomies. Radiation treatment parameters were used to estimate CA radiation doses. Twenty-one patients without coronary stenosis (controls) were matched with twelve patients with stenosis (cases). CA segments were considered as sub-volumes of interest. Radiation doses to stenotic segments were compared with those received by normal segments (from cases and controls) using a logistic regression. Results In eleven cases out of twelve, the highest of the coronary dose distribution was on a damaged segment. Logistic regression with CA segments yielded an odds ratio associated with the risk of coronary stenosis of 1.049 per additional gray with the CA segment median dose (95% confidence interval, 1.004–1.095; p-value

The most commonly used method for dose finding, the 3 + 3, has poor performance. New adaptive designs are more efficient. Nevertheless, they have reached a maximum performance level, and further improvement requires either larger sample sizes or outcomes measures richer than the simplistic severe toxicity measured at cycle 1. Clin Cancer Res; 23(15); 3977–9. ©2017 AACR. See related article by Yan et al., p. 3994

Motivation Detailed knowledge of coding sequences has led to different candidate models for pathogenic variant prioritization. Several deleteriousness scores have been proposed for the non-coding part of the genome, but no large-scale comparison has been realized to date to assess their performance. Results We compared the leading scoring tools (CADD, FATHMM-MKL, Funseq2 and GWAVA) and some recent competitors (DANN, SNP and SOM scores) for their ability to discriminate assumed pathogenic variants from assumed benign variants (using the ClinVar, COSMIC and 1000 genomes project databases). Using the ClinVar benchmark, CADD was the best tool for detecting the pathogenic variants that are mainly located in protein coding gene regions. Using the COSMIC benchmark, FATHMM-MKL, GWAVA and SOMliver outperformed the other tools for pathogenic variants that are typically located in lincRNAs, pseudogenes and other parts of the non-coding genome. However, all tools had low precision, which could potentially be improved by future non-coding genome feature discoveries. These results may have been influenced by the presence of potential benign variants in the COSMIC database. The development of a gold standard as consistent as ClinVar for these regions will be necessary to confirm our tool ranking. Availability and implementation The Snakemake, C++ and R codes are freely available from https://github.com/Oncostat/BenchmarkNCVTools and supported on Linux. Supplementary information Supplementary data are available at Bioinformatics online.

Introduction L’agence europeenne du medicament recommande que « les toxicites de faible grade sur de longues periodes qui affectent la tolerance (…) soient considerees dans la definition de la toxicite dose-limitantes (DLT) et de la dose recommandee pour les phase II (RP2D) » dans les essais de phase precoce. En particulier, les toxicites repetees de faible grade de severite peuvent fortement impacter l’adherence au traitement. Neanmoins, l’analyse de ces donnees longitudinales est limitee par le faible nombre de patients, rendant utiles les hypotheses fonctionnelles. Dans ce travail, nous explorons la structure de mesures repetees de toxicites gradees et en particulier si l’hypothese des odds proportionnels (OP) sur les grades de differentes toxicites en fonction de la dose et des cycles de traitement est plausible. Methodes Dans une meta-analyse sur donnees individuelles de 53 etudes de phase I de monotherapies ciblees nous caracterisons les relations entre la dose et les grades de toxicite cutanee, digestive, hematologique, troubles generaux et « autres » toxicites. Les paliers de doses ont ete standardises sur la dose maximum toleree DMT de chaque essai. Nous avons employe un modele dit « continuation ratio » (CR) multivarie pour modeliser conjointement le risque de chacun de ces cinq types de toxicite ordinale (grade 1, 2 ou ≥ 3) sur les six premiers cycles de traitement. L’hypothese OP correspond a la nullite de certains parametres. Le caractere longitudinal et multivarie des donnees a ete pris en compte a l’aide d’effets aleatoires toxicite-specifiques distribues selon une loi Gaussienne multidimensionnelle. Pour estimer ce modele complexe, l’inference bayesienne avec l’algorithme de Monte-Carlo hamiltonien a ete privilegiee. Le modele a ete regularise en utilisant les distributions a priori horseshoe pour les effets fixes et LKJ pour la matrice de correlation des effets aleatoires. Afin d’obtenir des conclusions generalisables a de futurs essais, nous avons compare ces modeles a l’aide du critere WAIC. Resultats Les 2048 patients ont presente un total de 9904 toxicites, dont 5563 de grade 1, 3128 de grade 2 et 1213 de grade ≥ 3. Pour le modele CR, la dose augmentait le risque de chaque type de toxicite avec un odd ratio (OR) superieur a 2. L’estimation la plus faible concernait l’OR pour le risque de trouble general (2,16 intervalle de credibilite a 95 % [1,77 ; 2,65]). L’hypothese OP ne semblait pas respectee ni pour les troubles generaux, ni pour les toxicites digestives, ou l’OR augmentait avec le grade de toxicite. Elle ne semblait pas respectee pour le cycle qui etait associe a une augmentation des risques de toxicites cutanees, troubles generaux et « autres » avec un OR plus faible pour les grades ≥ 2. Ce modele presentait le meilleur WAIC (30911), qui n’est que 1,7 % inferieur a celui du modele OP (31432), qui donne des conclusions similaires pour les effets dose et cycle. Conclusion Notre etude montre que les donnees generees par les essais de phase I sont plus riches que le critere classique binaire de toxicite au 1er cycle. Meme si le modele CR presentait le meilleur WAIC, la faible difference avec le modele OP suggere que de nouveaux designs pourraient etre elabores selon cette hypothese pour selectionner la dose optimale de facon plus fiable.

Background The NIVOREN GETUG-AFU 26 study reported safety and efficacy of nivolumab in patients with RCC in a “real world setting”. A translational research program was launched to characterize immune cell populations in fresh whole blood at baseline and after 2 months of treatment (C3) to determine association with high-grade toxicity and clinical outcome. Methods Absolute number of 106 immune cell populations were analysed in patients treated at a single institution as part of the NIVOREN GETUG-AFU 26 in fresh whole blood using dry formulation panels for multicolor flow cytometry. Predefined clinical endpoints were 6-month grade 3-4 toxicity (treatment related adverse event, TRAE) occurrence and 6-month disease progression occurrence. Missing values were imputed using multivariate imputation by chained equations. Multivariate differential count analysis was done using the DESeq2 R package. Results Overall 44 patients were included in this fresh whole blood immune-monitoring. Higher occurrence of grade 3-4 TRAE at 6 months was observed in patients with lower number of CD4+PD-1neg4.1BB+ T cell (non-exhausted activated CD4 T cells) counts at baseline (log2 fold change (LFC) = -1.993 95% confidence interval (95%CI)[-2.172; -1.815] ) and with lower number of CD56+ T cells (NKT cells) at C3 (LFC = -1.464 95%CI[-1.597; -1.331]). Higher occurrence of disease progression at 6-month was observed in patients with lower number of CD4+PD-1posCD69+ T cells (exhausted activated CD4 T cells) (LFC = -1.186 95%CI[ -1.308; -1.064 ]) and with higher number of CD4+CD244+ T cells (exhausted CD4 T cells) (LFC = 1.686 95%CI[ 1.533; 1.840]) at baseline. No immune cell populations were associated with the occurrence of a 6-month progression during the course of nivolumab (C3). Conclusions Fresh whole blood monitoring at baseline and after 2 months of nivolumab identified immune cell populations associated with grade 3-4 TRAE (CD4+PD-1neg4.1BB+ T cells and CD56+ T cells) and disease progression (CD4+PD-1posCD69+ T cells and CD4+CD244+ T cells). Functional analyses and external validation are ongoing. Clinical trial identification NCT03013335. Legal entity responsible for the study UNICANCER. Funding Bristol-Myers Squibb. Disclosure B. Beuselinck: Honoraria (self), Advisory / Consultancy, Research grant / Funding (institution): BMS. B. Escudier: Advisory / Consultancy, Research grant / Funding (institution), Travel / Accommodation / Expenses: BMS; Advisory / Consultancy, Research grant / Funding (institution), Travel / Accommodation / Expenses: Pfizer; Advisory / Consultancy, Research grant / Funding (institution): Novartis; Advisory / Consultancy: Roche; Advisory / Consultancy: Ipsen; Advisory / Consultancy: EUSA; Advisory / Consultancy, Research grant / Funding (institution): Aveo. L. Albiges: Advisory / Consultancy, compensated to institution: Pfizer; Advisory / Consultancy, compensated to institution: Novartis; Advisory / Consultancy, compensated to institution: Roche; Advisory / Consultancy, compensated to institution: Bristol-Myers Squibb; Advisory / Consultancy, compensated to institution: IPSEN; Advisory / Consultancy, compensated to institution: MSD; Advisory / Consultancy, compensated to institution: AstraZeneca. All other authors have declared no conflicts of interest.

3034Background: In the context of the checkpoint inhibitors (CPI) era and the strong demand for identification of predictive biomarkers, the role of the host must be considered. We developed a cust...

467 Background: Recent and independent muscle-invasive bladder cancer (MIBC) molecular classifications identified the basal / squamous-like (BASQ) tumours as an intrinsic and robust subtype, with a poor outcome and possible chemosensitivity to cisplatin based regimen, making mandatory the development of a diagnostic tool for their identification in routine samples. Our study aimed to evaluate the diagnostic accuracy of a Nanostring classifier for tumor subtype prediction on FPPE specimens. Methods: Two series of MIBC were used (CIT n = 51 & Stransky n = 22) for which BASQ tumours were identified previously using Affymetrix transcriptome data obtained from frozen samples (Rebouissou, Science Trans Med 2014). 29 genes were selected to predict the basal subtype, RNA expression of matched frozen and FFPE samples was studied using Nanostring technology. To define the classifiers for Affymetrix, frozen and FFPE Nanostring expression matrix on CIT samples the centroid of each cluster was calculated using the expression of 29 genes. Internal validation used leave-one-out cross-validation to train and test the prediction accuracy of the new classifier. For external validation, the CIT samples were used as training set and the Stransky samples as validation set. Predictive accuracy expressed as percentage of correctly classified samples is provided. Results: Correlations between Affymetrix, frozen and FFPE Nanostring data set were checked for gene expression and samples. Using CIT samples as train and test set, the predictive accuracy for BASQ tumour identification was for Affymetrix, frozen Nanostring and FFPE Nanostring classifiers, 90.20% [78.59%; 96.74%], 88.24% [76.13%; 95.56%] and 92.16% [81.12%; 97.82%], respectively. Using training on CIT samples and test on Stransky samples, this predictive accuracy was 90.91% [70.84%; 98.88%] both for Affymetrix, frozen Nanostring and FFPE Nanostring classifiers. Conclusions: The 29 gene expression Nanostring codeset was able to identify reliably basal / squamous like tumours on FFPE samples from MIBC in comparison with the gold standard approach based on transcriptomic profile, appearing as a promising diagnostic tool.

International audience; Purpose The objectives are to analyze mortality risks in the extended follow-up of the French uranium miners’ cohort and to examine their potential relation to occupational exposure to ionizing radiation (IR). Methods The total cohort includes 5,086 uranium miners employed in the CEA-COGEMA group and followed up from 1946 to 2007. Vital status, causes of death, and cumulative radon exposures were recorded. The post-55 subcohort includes 3,377 miners first employed after 1955, for whom long-lived radionuclides (LLR) and external gamma-ray exposure were also recorded. External mortality analyses were performed by computing standardized mortality ratios (SMR). Excess relative risks (ERRs) due to IR exposures were estimated from Poisson regression models. Results The miners included in the total cohort were followed up for 35.4 years and exposed to 36.6 working level months (WLM) on average. There was no evidence of a difference in overall mortality between miners and the general French male population. Miners had a statistically significant excess mortality rate from lung cancer (SMR = 1.34 [95 % CI 1.16–1.53]) and from kidney cancer (SMR = 1.60 [1.03–2.39]). Cumulative radon exposure was significantly associated with lung cancer risk (ERR/100 WLM = 0.71 [0.31–1.30]) and cerebrovascular risk (ERR/100 WLM = 0.41 [0.04–1.03]). In the post-55 subcohort, this excess mortality from lung cancer remained associated with exposure to radon, and also with exposure to LLR and external gamma rays. Conclusions The analyses in the extended follow-up strengthen the results previously observed among French uranium miners about their excess risk of mortality and its association with their occupational IR exposure. © 2014, Springer-Verlag Berlin Heidelberg.

International audience; Progress in next-generation sequencing provides unprecedented opportunities to fully characterize the spectrum of somatic mutations of cancer genomes. Given the large number of somatic mutations identified by such technologies, the prioritization of cancer-driving events is a consistent bottleneck. Most bioinformatics tools concentrate on driver mutations in the coding fraction of the genome, those causing changes in protein products. As more non-coding pathogenic variants are identified and characterized, the development of computational approaches to effectively prioritize cancer-driving variants within the non-coding fraction of human genome is becoming critical. After a short summary of methods for coding variant prioritization, we here review the highly diverse non-coding elements that may act as cancer drivers and describe recent methods that attempt to evaluate the deleteriousness of sequence variation in these elements. With such tools, the prioritization and identification of cancer-implicated regulatory elements and non-coding RNAs is becoming a reality.

We address here the issue of prioritizing non-coding mutations in the tumoral genome. To this aim, we created two independent computational models. The first (germline) model estimates purifying selection based on population SNP data. The second (somatic) model estimates tumor mutation density based on whole genome tumor sequencing. We show that each model reflects a different set of constraints acting either on the normal or tumor genome, and we identify the specific genome features that most contribute to these constraints. Importantly, we show that the somatic mutation model carries independent functional information that can be used to narrow down the non-coding regions that may be relevant to cancer progression. On this basis, we identify positions in non-coding RNAs and the non-coding parts of mRNAs that are both under purifying selection in the germline and protected from mutation in tumors, thus introducing a new strategy for future detection of cancer driver elements in the expressed non-coding genome., Author Summary Cancer cells undergo a mutation/selection process that resembles that of any living cell. Most mutations in cancer cell DNA occur in the so-called "non-coding" regions that represent 98.5% of the genome length. Pinning down which of these mutations contribute to the fitness of cancer cells would be important for identifying new "cancer drivers", which may in turn lead to future treatments. Unfortunately, predicting the impact of a non-coding DNA alteration remains extremely difficult. In this study, we analyze millions of non-coding cancer mutations and show cancer-specific mutational patterns can be used to predict non-coding regions that are preserved from mutations and may thus be important for cancer cell survival. Combining this information with population data, we propose a new scoring system that should help prioritize important non-coding mutations in future studies.

International audience; The investigation of potential adverse health effects of occupational exposures to ionizing radiation, on uranium miners, is an important area of research. Radon is a well-known carcinogen for lung, but the link between radiation exposure and other diseases remains controversial, particularly for kidney cancer. The aims of this study were therefore to perform external kidney cancer mortality analyses and to assess the relationship between occupational radiation exposure and kidney cancer mortality, using competing risks methodology, from two uranium miners cohorts. The French (n = 3,377) and German (n = 58,986) cohorts of uranium miners included 11 and 174 deaths from kidney cancer. For each cohort, the excess of kidney cancer mortality has been assessed by standardized mortality ratio (SMR) corrected for the probability of known causes of death. The associations between cumulative occupational radiation exposures (radon, external gamma radiation and long-lived radionuclides) or kidney equivalent doses and both the cause-specific hazard and the probability of occurrence of kidney cancer death have been estimated with Cox and Fine and Gray models adjusted to date of birth and considering the attained age as the timescale. No significant excess of kidney cancer mortality has been observed neither in the French cohort (SMR = 1.49, 95 % confidence interval [0.73; 2.67]) nor in the German cohort (SMR = 0.91 [0.77; 1.06]). Moreover, no significant association between kidney cancer mortality and any type of occupational radiation exposure or kidney equivalent dose has been observed. Future analyses based on further follow-up updates and/or large pooled cohorts should allow us to confirm or not the absence of association. © Springer-Verlag Berlin Heidelberg 2014.

Objectives There is currently no consensus for an association between risk of Circulatory System Diseases (CSD) and low exposure to ionising radiation. The aim is to study the relationship between CSD mortality and radon exposure in the French cohort of uranium miners considering classical CSD risk factors. Method The French cohort includes 5086 uranium miners followed from 1946 through 2007. Among the 1935 deaths, 442 were from CSD, including 167 cases of IHD and 105 cases of CVD. Annual radon exposures were assessed individually. A nested case-control study was performed to collect individual information concerning classical CSD risk factors (weight, height, smoking status, blood pressure, blood glucose and cholesterolemia) from medical files for 76 cases of death from CSD (including 26 from IHD and 16 from CVD) and 237 controls, matched for attained age and birth date and counter-matched for cumulative radon exposure. Results In the whole cohort, a significant association with cumulative radon exposure was observed for CVD mortality, but not for CSD or IHD mortality. In the case-control study, no significant unadjusted Odd-Ratio for cumulative radon exposure was observed for any endpoint. Analyses adjusted on CSD risk factors, for which missing data do not exceed 25%, are ongoing. Conclusions The issue of CSD associated to ionising radiation is crucial for radiation protection. The present study, allowing to consider individual data on major classical CSD risk factors, will contribute to improve knowledge on the effects of low dose exposure.