dataesr/scientific-paragraphs-categorization huggingface dataset:
The analysis of the RAW files was performed using the MaxQuant software package (version 1. 6. 5. 0). The extracted spectra were matched against the mouse sequences from SwissProt (version 2019-04). Sequences of known contaminants were added to this database, and the reverse decoy was strictly set to FDR of 0. 01. Database searches were done with 20 ppm and 0. 5 Da mass tolerances for precursor ions and fragmented ions respectively. Trypsin (two missed cleavages allowed) was selected as the protease. Dynamic modifications included N-terminal acetylation and oxidation of methionine, while Cystein carbamidomethylation was set as a fixed modification. Protein abundances were determined by label-free quantitation using the composite iBAQ intensity values determined by MaxQuant and normalized within single or multiple migration profiles of individual proteins.
Normalized protein quantification values were then imported into the Perseus 70 software platform for log transformation, data imputation, and statistical analysis. For proteins missing values for 1 of the 3 experimental replicates in a particular band and genotype, the mean iBAQ value of the two other replicates was imputed. All proteins missing more than one value in a particular band and genotype were considered undetected. For each protein, gel migration profiles were created and normalized to the maximum abundance across all samples analyzed. The migration profile of proteins belonging to known multi-protein complexes were then manually extracted and heatmaps of relative abundance were generated using GraphPad Prism 8 software package (version 8. 4. 2) to visualize co-migration. The absolute abundance of selected protein complexes was also estimated by summing the iBAQ values of all detected protein components co-migrating in the same band.
For statistical analysis, one sample T tests comparing the two genotypes were performed for each band. A P value of less than 0. 05 was set as the arbitrary threshold for statistical significance. For each P value obtained, a corresponding False Discovery Rate (FDR) was calculated according to the Benjamini and Hochberg method and lists of significantly affected proteins were generated at FDR thresholds of < 5, < 10, and < 15% FDR values. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE  71  partner repository with the dataset identifier PXD021867. Quantitative RT-PCR. Chop transcript level from frozen liver tissue was assessed using an RNeasy kit (QIAGEN) for RNA extraction and reverse transcribed with the High-Capacity CDNA RT kit (Thermo Fisher Scientific) according to the manufacturer's recommendations.
Quantification was allowed using the ΔΔCT method and the TATA Binding Protein gene expression was used as a housekeeping gene. Primers used: Chop (NM_007837), sense 5′-TATCTCATCCCCAGGAAACG-3′, anti-sense 5′-CAGGGTCAAGAGTAGTGAAGGTTT-3′; Tbp (NM_013684), sense 5′-GGCCTCTCAGAAGCATCACTA-3′, anti-sense 5′-GCCAAGCCCTGAGCATAA-3′. Lipidomics. Lipid extraction and data analysis were done as previously described  31  and adapted for isolated mitochondria  14. In brief, lipids were extracted from isolated mitochondria or digitonin-extract and spiked with six internal standards: LPC 13:0, PC19:0/19:0, PC14:0/14:0, PS12:0/12:0, PG15:0/15:0, and PE17:0/17:0 (Avanti Polar Lipids Inc, Alabaster, USA). For both kinds of samples, protein concentration was determined using a colorimetric-based assay based on the Bradford dye-binding method to set the volume of injection.
5 or 0. 75. Statistical analysis was performed on the data using an unpaired student's t-test with Storey correction for multiple testing using the Q-value package from Bioconductor. Data are depicted as volcano plots and box plots, where the midline represents the median fold change vs. controls, the box represents the interquartile range (IQR) between the first and third quartile, and whiskers represent the lowest or highest values. Statistics and Reproducibility. For functional immunoblot and Q-PCR experiments, values are reported as mean ± sem for a minimum of 3 mice per experimental group and graphically represented as violin plots. Unpaired two-sided t-tests with Welch's correction (GraphPad Prism 8. 4. 3) were used to determine statistical difference when two means were compared, with a significance threshold set at p < 0.
05. For lipidomics, data are depicted as volcano plots and box plots, where the midline represents the median fold change vs. controls, the box represents the interquartile range (IQR) between the first and third quartile, and whiskers represent the lowest or highest values (n = 4 mice per group). All discriminant MS features were selected according to the following criteria of selection: Q-value < 0. 05 (as an estimation of false discovery rate (FDR) < 5%) and a relative fold change (FC) of 1. 5 or 0. 75. Statistical analysis was performed on the data using an unpaired student's t-test with Storey correction for multiple testing using the Qvalue package from Bioconductor. For proteomics, replicate samples from 6 mice (3 H-Lrpprc -/-and 3 H-Lrpprc +/+ ) were analyzed. The difference in the abundance of individual proteins between genotypes was determined in each band using unpaired two-tailed T tests.
Acknowledgements This work was funded by grants from the  Natural Sciences and Engineering Council of Canada  ( RGPIN 2016-09932 ), The  Canadian Institutes of Health Research  ( MOP 136999 ), the  Fondation du Grand Défi Pierre Lavoie , and the  Canadian Foundation for Innovation. The authors are grateful to the  Leigh Syndrome French Canadian Consortium (LSFC)  for the support in creating tools and reagents to support this research project. LSFC consortium members include.
• Some automation and remote controls will be needed during the inspection process. The inspection area is quite large as the whole tube bodies will be inspected. • Internal surfaces located between the nested tubes likely cannot be inspected for corrosion and therefore, sealing the space from oxygen and water will be likely be the best option. • Experimental reference is a critical next step of the work to assess and qualify the inspection methods. As the next step, CEA will continue discussions on requirements of the mock-ups: for eddy current inspection, a Zircaloy-4 planar specimen with artificial reflectors is needed. For ultrasonic reference, a representative mock-up of the cylinder with a weld and artificial defect(s) is needed. The mock-up testing will provide an experimental reference to CIVA calculations as simulations of this work have been performed with CIVA software, which is extensively used in the industry and research organizations in simulation of nondestructive control.
B. Ultrasonic simulations Ultrasonic testing is a family of non-destructive testing techniques based on the propagation of ultrasonic waves in the object or material tested. In most common applications, very short ultrasonic pulse-waves with center frequencies ranging from 0. 1-15 MHz are transmitted into materials to detect internal flaws or to characterize materials. Such ultrasonic simulation can be performed using CIVA module which simulates the beam-defect interaction and predicts the amplitude and the time of flight of various echoes: direct echo, corner effect, tip diffraction echo, etc. It can also calculate echoes scattered back by the geometry (backwall, entry surface and interior specular interfaces echoes) and takes into account mode conversions. Ultrasonic sensors chosen for this study are 2 MHz frequency, 7x7 mm and 10 MHz , 8x8 mm size.
GITR activation by its ligand in CD4 effector cells and CTLs increases cell proliferation and effector function. GITR stimulation in Treg cells leads to instability, Treg cell depletion, and decreases Treg cells suppressive function  18, 19. Anti-GITR agonistic antibody therapy has been shown to cause tumor regression in a number of preclinical non-CNS tumor models  18, 20  and is under evaluation as monotherapy (NCT01239134) and in combination with aPD1 therapy in a number of clinical trials (NCT04021043, NCT02740270, NCT02628574, NCT02598960, and NCT04225039). αGITR therapy combined with stereotactic radiation in an experimental model of glioma improves survival  21. While these studies are insightful, the mechanisms mediating therapeutic response in GBM for αGITR alone or in combination with aPD1 or radiation remain to be elucidated.
Finally, we used 005GSC, which is derived from a genetically-engineered mouse model and faithfully recapitulates the stem-like features of human GBM  30  , and thus it is likely to have a low mutational burden. Mice bearing orthotopic, sizematched murine GL261-MGH, CT2A, and 005GSC GBM tumors were treated with IgG2a control or αPD1 antibodies. In this treatment setting, we did not observe significant survival benefits (Fig. 1a ), consistent with the outcomes of randomized phase III clinical trials in patients with recurrent  2  or newly diagnosed GBM (NCT02617589)  31. There were no significant changes in the frequency of CD8 T cells (Supplementary Fig. 1a-c ), except for a modest reduction in PD1 + Lag3 + exhausted CD8 T cells (Supplementary Fig. 1d ). Interestingly, we observed that CD4 T cells   were consistently enriched in the GBM tissues with a CD4/CD8 ratio of 2 or higher, irrespective of treatment (Supplementary Fig.
1e ). These data are consistent with the hypothesis that Treg cells may mediate resistance to αPD1 therapy in GBM. Despite confinement of GBMs in the intracranial space, systemic T cells are also dysfunctional in GBM patients  41. To evaluate this dysfunction, including induction of Treg cell anergy, we analyzed tumor-draining lymph nodes and spleens of GBM tumor-bearing mice. While the frequency and numbers of Treg cells were not altered in these lymphoid organs during tumor growth (Supplementary Fig. 3a ), the ratio of FR4 hi CD73 hi / FR4 lo CD73 lo was higher compared to non-tumor-bearing mice (Supplementary Fig. 3b ). These data indicate that Treg cells outside the brain can also acquire a suppressive phenotype and might induce systemic immunosuppression during tumor progression.
Given that αGITR antibodies are currently being evaluated in a number of clinical trials (NCT04021043, NCT02740270, NCT02628574, NCT02598960, and NCT04225039), our study provides mechanistic insights and compelling preclinical data in support of testing the combination of αGITR with αPD1/PDL1with or without the SoC therapy-in GBM patients with a high intratumoral accumulation of Treg cells.
Methods Antibodies. For ease of identification, full details of antibodies are shown under the registry of publicly available data base (antibodyregistry. org) that provides a comprehensive list of suppliers, and appropriate references for usage. This information is summarized under "antibody registry number" in the Supplementary Table  1  included in the supplementary information file. Mice. C57BL/6, FoxP3 DTR , and Rag1 -/-mice were obtained from the Cox-7 gnotobiotic animal facility operated by the Edwin L. Steele Laboratories, Department of Radiation Oncology at the MGH. All animal experiments were performed in the Cox-7 animal facility, accredited by the Association for Assessment and Accreditation of Laboratory Animal Care International. Female and male mice were used. Animal protocols were approved by the Institutional Animal Care and Use Committees at MGH.
For cytokine expression analysis, cells were activated with Leukocyte Activation Cocktail (BD Biosciences, Cat no: 554656) in RPMI containing 10% FBS and 1% penicillin-streptomycin (10,000 U/mL) for 4. 5 h. For the flowcytometric analyses, cells were first stained with surface markers in FACS buffer (2% BSA in PBS) for 20 min on ice. Then cells were fixed in Fix/Perm buffer (eBioscience) for 15 min, washed in permeabilization buffer (eBioscience), and stained for intracellular factors in permeabilization buffer for 20 min on ice. Live cells for transfer experiments and in vitro assays were sorted using BD FACSAria ™ II cell sorter (BD Biosciences). Cells were analyzed on Aria II (BD Biosciences) or LSRII (BD Biosciences), and data analysis was performed on FlowJo (Tree Star 10. 7v). Tumor models.
Patient records were reviewed for age, sex, medical history, surgical intervention, therapeutic regimens, IDH1 mutation status, MGMT promotor methylation, cytogenetics and overall survival. All patients (except 1 patient which opted for no treatment) received SoC therapy ± immune checkpoint blockade ± Bevacizumab. PBMCs were collected from healthy volunteers and not cancer patients. The data were anonymized and categorized under MGH general informed consent protocol. RNA-seq analysis of Treg cells. Treg cells were sorted from tumor tissues or spleens of treated mice. Next-generation sequencing libraries were prepared according to the Smart-Seq2 protocol as previously described  [78] [79] [80]. Briefly, RNA from cell lysates were captured using Agencourt RNA Clean beads then converted to cDNA.
The whole transcriptome was amplified before quantification and dual-index barcoding with an Illumina Nextera XT kit. The cDNA libraries were enriched then pooled in equal volumes before final bead clean-up and sequencing on a NextSeq500 instrument. Reads were aligned to mm10 mouse reference genome using Tophat, and RSEM  81  was used to calculate transcripts per million. Normalized read counts were filtered for robust expression (>10) and low CV and processed in the Multiplot suite and Morpheus (https:// software. broadinstitute. org/morpheus/). In vitro co-culture assays. In preclinical models, Treg cells or CD8 T cells were isolated from tumors, spleen, or LN of tumor-bearing mice or naive C57BL/6 mice. Cell sorting was done post enrichment using Treg cell isolation or CD8 isolation kits.
Acknowledgements We would like to thank  Dr. Thomas N. Seyfried  at  Boston College  for providing the murine CT2A cell line,  Dr. Martine F. Roussel  at  St. Jude Children's Research Hospital  for providing the Myc-MB cell line,  Dr. Joan Massagué  at  Memorial Sloan Kettering Institute (NY)  for providing ErbB2-P cell line, and  Dr. Mark Cobbold  for providing human PBMC cells. We thank  Drs. Rebecca Betensky  and  Hang Lee  for the statistical analyses,  Mark Duquette  at the  Steele laboratories, and the staff of the flow cytometry Core facility at Simches research center, MGH , for their technical assistance. We also thank  Drs. Patrik Andersson ,  Meenal Datta ,  Dan Duda ,  Lance Munn ,  Timothy Padera ,  Mikael Pittet , and  Nilesh Talele  for their helpful input on this paper. We thank  Dr.
Data availability All data generated by this study are available within the Article, Supplementary Information or available from the authors upon request. Source data are provided with this paper. Our RNA sequencing data from GBM Tregs vs. spleen Tregs are available in Gene Expression Omnibus (GEO) under accession code GSE167746. Materials are available under a material transfer agreement (contact person R. K. J. ). Source data are provided with this paper.
DISCUSSION The study succession documents the evolution of a salt-marsh depositional environment over a time period of 650 years, thus including the two historically documented time-spans of active fluvial input by the Brenta River (i. e. , BR1 and BR2 periods). The dataset presented here shows that, despite the large volume of sediments delivered by the Brenta River during these stages -which determined during BR2 an expansion of the total marsh area of 24 km 2 according to the available historical maps  (Roner et al. , 2017; Tommasini et al. , 2019)  the ratio between organic and inorganic sedimentation, as well as the inorganic grain size, remained almost constant. Moreover, an initially stable upper salt-marsh environment is also documented by the foraminifera assemblage, whereas a middlemarsh environment was suggested to occur in the early 20th century (Figure  4 ).
ACKNOWLEDGMENTS We are indebted to  Stéphane Augros ,  Johannes Berger ,  Boris Brückmann ,  Rémy Eudeline ,  Ludovic Montfort , and  Cynthia Y. H. Wang-Claypool  for their support in the field surveys, to  Athena W. Lam  for help in the molecular lab, and to  Michael Franzen  for providing photographs of the preserved holotype. Furthermore, we thank  Guillaume Decalf  and the authorities at the  Direction de l'Environnement, de l'Aménagement et du Logement (DEAL), Mayotte (France)  for issuing research and export permits and the  BOLD team  for handling the barcode database entries. Finally, we thank  Rebecca Laver ,  Juan Diego Daza , and  Scott Miller  for organizing this special issue and inviting us to contribute to it, and two anonymous referees for providing highly valuable comments.
Delays Related to Neoadjuvant Treatment Data from large, neoadjuvant CT trials in resectable NSCLC (Table  1 ) serve as historical benchmarks for comparing perioperative outcomes in neoadjuvant ICI trials. A National Cancer Database study revealed that 43% of 2185 patients with stage IIIA NSCLC treated with neoadjuvant CT had long delays before surgery (>114 d), 37% had medium delays (77-114 d), and 30% had short delays (<77 d). 9  The delay to surgery did not influence 30-or 90-day mortality rates. Nevertheless, 1and 3-year survival analyses revealed that long delays to surgery increased mortality compared with short delays (hazard ratio ¼ 1. 25, p ¼ 0. 0005), suggesting overall oncologic outcome is influenced by the timing of surgery after neoadjuvant treatment. The interval between neoadjuvant CT and resection has rarely been reported in large multicenter studies.
Neoadjuvant Therapy End Points Although the intent of neoadjuvant therapy is not to achieve downstaging to render the disease operable, tumor downstaging after neoadjuvant therapy predicts better survival. 34 Appropriate candidates for neoadjuvant therapy should have R0 resectable disease on the basis of baseline scans, confirmed by a multidisciplinary tumor board or consilium. Nodal downstaging predicted survival after neoadjuvant CT for stage IIIA (N2) NSCLC in CALGB protocol 893: patients with resected and no residual N2 disease had superior EFS compared with patients with persistent N2 disease (47. 8 versus 8. 2 mo, respectively). 35 It is therefore imperative to collect data on pretreatment clinical and posttreatment clinical and pathologic stages to allow assessment of down-and upstaging after neoadjuvant therapy (downstaging end points; Fig.
1 ). For right-sided resections, this involves lymph nodes from at least levels 4R, 7, 10R, and 11R. For leftsided resections, this involves lymph nodes from at least levels 5/6, 7, 10L, and 11L. Level 8 and 9 lymph nodes should be assessed by dissection or sampling, particularly for lower lobe cancers. In addition, prospective data on intraoperative complications for bronchial or vascular injuries should be collected (operative end points; Fig. 1 ). As in neoadjuvant CT studies (Table  1 ), lobectomy was the most common type of resection performed in phase 2 neoadjuvant ICI studies (65%-93% after monotherapy or dual immunotherapy and 73%-93% after ICI plus CT; Table  4 ). R0 rates ranged from 87% to 100%. In CheckMate 159, a total of 50% of conversions to thoracotomy owing to adhesions (two of four) were in patients with stage I or IIA disease.
1 ). NEOSTAR provided data on surgical complexity. 33 n a subjective 4-point complexity scale that assumed a score of 2 as "normal dissection," representing standard lobectomy for stage I NSCLC without neoadjuvant therapy, 40% of 37 surgeries were judged to be more difficult than usual (i. e. , score !3). 33  The median operative time was 147 minutes (range: 71-315 min); median blood loss was 100 mL (range: 50-1000 mL). 33.
More data on the clinical characteristics of patients found pre-or intraoperatively to have unresectable disease would help define the patient subgroups unlikely to respond to neoadjuvant ICI therapy, independent of pathologic response. Although controversial, patients at higher risk for unresectability, such as those with T4 tumors with direct mediastinal invasion or multistation N2 disease, may be considered for exclusion from neoadjuvant trials. Data from intraoperative end points that allow quantitation of surgical technical difficulty and quality of oncologic resection should be collected. Standardized grading of complexity with respect to lymphadenopathy and fibrosis would address any subjectivity underlying perceptions that surgery after ICI therapy is more difficult than after CT. Demonstration of high R0 rates in phase 3 trials would further increase confidence in neoadjuvant immunotherapy.
within <15 d of (2020) 12 that window NCT02259621 I-IIIA Nivolumab Approximately 4 wk 20 (95%)/21 Median, 18 (range: 0% 0% 1/21 (5%) CheckMate 159 (3 cycles on d after the first 11-29) d (tracheal Bott et al. À42, À28, À14 neoadjuvant dose No treatment-related invasion; (2019) 41 Forde et al. (2018) 14 NCT03158129 NEOSTAR Cascone et al. (2019) 13 Sepesi et al. (2019) 33 NCT02938624 MK3475-223 Bar et al. (2019) 24 I-IIIA I/II [±2 d] before surgery on d 0) Nivolumab (3 cycles) vs. nivolumab (3 cycles) þ ipilimumab (1 cycle) Pembrolizumab (2 cycles) Within 3-6 wk after last neoadjuvant dose 1-3 wk 37 (84%)/44 13 (87%)/15 delays Median, 31 (range: 21-87) d (n ¼ 8 [22%] delayed beyond 42 d) NR 1/13 (8%) had delay owing to treatment-related grade 3 myositis 5/44 (11%) N: n ¼ 1 (2%) SAE (grade 3 hypoxia) and high surgical risk NI: n ¼ 4 (9%) PD (1), lack of resectability (1), high surgical risk (1), declined surgery (1) 2/15 (13%) Treatment-related grade 3 myositis (7%); grade 3 myocardial infarction (7%; not treatment-related) 1/44 (2%) (pneumonitis and BPF) NR patient had stage IIIA) NR NR (continued) Neoadjuvant Immunotherapy End Points in NSCLC.
cisplatin/ failure) (2020) 50 docetaxel (3 cycles) NCT01820754 IB-IIIA Ipilimumab þ Within 12 wk of 13 (54%)/24 <12 wk (2 patients 11/24 (46%) 0% 0% TOP1201 chemotherapy completing [15%] had delay in (persistent N2 Yang et al. neoadjuvant surgery of 4 and 5 cancer: 5/24 (2018) 22 treatment wk, respectively, [21%]; inadequate owing to pulmonary ipilimumab- function: 2/24 related diarrhea) [8%]; PD: 2/24 [8%]; location of tumor: 1/24 [4%]; immune-related AE: 1/24 [4%]) NCT03081689 NADIM Provencio et al. (2019) 51.
5%), blood creatinine phosphokinase increased: 1/40 (2. 5%), lung infection: 1/40 (2. 5%) Combination immunotherapy plus chemotherapy NCT02716038 30 Atezolizumab þ 28/30 (93%) Possible irAEs were arthralgia or myalgia (grade 1/2; 5/30 Columbia 7 carboplatin þ nab- Increased ALT: 2/30 (7%), increased AST: 2/30 (7%), diarrhea: [17%]), diarrhea (grade 1/2; 8/30 [30%]; grade 3; 1/30 paclitaxel 1/30 (3%), anemia: 1/30 (3%), fatigue: 1/30 (3%), febrile [3%]), increased ALT (grade 1/2; 4/30 [13%]; grade 3; neutropenia: 1/30 (3%), hyperglycemia: 1/30 (3%), 2/30 [7%]), increased AST (grade 1/2; 3/30 [10%]; hyponatremia: 1/30 (3%), neutropenia: 15/30 (50%), grade 3; 2/30 [7%]), hypothyroidism (grade 1/2; 3/30 thrombocytopenia: 2/30 (7%), weight loss: 1/30 (3%) [10%]), hyperglycemia (grade 4; 1/30 [3%]) NCT02572843 67 Durvalumab þ Grade !3 all-cause AEs during neoadjuvant treatment: NR SAKK 16/14 50 cisplatin/docetaxel Chemotherapy: 45/67 (67%) Durvalumab: 8/62 (13%) NCT01820754 13 Ipilimumab þ Grade 3/4: 46% Grade 2 pneumonitis: 1/24 (4%); grade 3 adrenal TOP1201 22 chemotherapy insufficiency: 4/24 (17%); diarrhea/colitis (grade 1 or 2; 6/24 [25%]; grade 3; 3/24 [13%]) NCT03081689 46 Nivolumab þ Grade 3-5: 11/46 (24%) NR NADIM 51 paclitaxel þ Neutropenia: 3/46 (7%), febrile neutropenia: 2/46 (2%), carboplatin peripheral sensory neuropathy: 2/46 (4%), anorexia: 1/46 (2%), fatigue: 1/46 (2%), alopecia: 1/46 (2%), nephritis: 1/46 (2%) NCT03366766 52 13 Nivolumab þ Grade 3: 2/13 (15%) NR cisplatin þ Neutropenia: 2/13 (15%), anemia: 1/13 (8%), renal pemetrexed or dysfunction: 1/13 (8%) gemcitabine a Also termed "AESIs.