dataesr/scientific-paragraphs-categorization huggingface dataset:
FACS sorting of LSK cells Bone marrow was flushed from donor femurs with sterile PBS containing 1% heat-inactivated fetal calf serum (PBS/FCS). Bone marrow cells were stained with a cocktail of antibodies from BD Biosciences: biotinylated lineage 1 antibodies (CD3, CD11b, CD19, CD49b, IgM, and Ter119), PE-conjugated lineage 2 antibodies (CD4, CD8, GR-1, and I-A b ), as well as, B220 PE-Cy5, c-kit APC, and Sca-1 PE-Cy7, followed by streptavidin APC-Cy7. For some experiments, cells were sorted using a FACS-Aria cell sorter and data analyzed using Diva Version 5. 1 software (both from BD Biosciences). After initial scatter-based gating to exclude doublets, the B220 -population was further gated to identify and sort the lineage -Sca-1 + c-Kit + (LSK) cell population that contains hematopoietic stem cells  [44].
Histology and immunofluorescence microscopy Imatinib-treated (66. 7mg/kg/day) or water control pumps were implanted one day prior to infection. Day 7 post-treatment, spleens were harvested, frozen in optimum cutting temperature (OCT) compound, cut into 6μm sections, mounted on slides, and then fixed with %100 acetone for 10 min at -20°C. Following rehydration in PBS, slides were permeabilized with PBS +0. 5% Triton X-100 +1% BSA, blocked with normal rat serum and anti-FcRIII/I antibodies and then incubated with anti-CD169-FITC (Serotec; Oxford, UK), CD11b-PE (eBioscience), and F4/ 80-biotin (eBioscience). Tissues were incubated with Streptavidin-APC and mounted with Prolong-GOLD (with DAPI) (Invitrogen). Images were captured using the x10 objective on a Zeiss Axioscope (Carl Zeiss, Germany) and analyzed using ImageJ (National Institute of Mental Health) and DoubleTake (Echo One, Denmark) software.
Ongoing Trials When this Trial Watch was being redacted (September 2014), official sources listed no less than 19 clinical trials initiated after September 1 st , 2013 to evaluate the safety and efficacy of recombinant/purified TAAs or peptides thereof as anticancer vaccines in patients (source http://www. clinicaltrials. gov). Of these studies, 13 rely on one or more TAA-derived short peptides injected as such (NCT01949688; NCT01949701; NCT01950156; NCT01961115; NCT01970358; NCT01989559; NCT01989572; NCT02019524; NCT02077114; NCT02126579; NCT02134925; NCT02149225; NCT02193347), 2 on TAA-derived short peptides encapsulated within liposomes (NCT01978964; NCT02065973), 1 on an SLP (NCT02128126), 1 on a CMP (NCT02229084), 1 on a full length, recombinant TAA (NCT02015416), and one on purified TAAs complexed with heat shock protein 90kDa b (Grp94), member 1 (HSP90B1, best known as gp96)  [190] [191] [192] [193] [194]  (NCT02122822) (Table  1 ).
In particular, (1) mucin 1 (MUC1)-derived peptides are being tested as standalone, Hiltonol Ò -adjuvanted or PET-Lipid Aadjuvanted and liposome-encapsulated interventions in patients with colorectal adenomas or other MUC1-expressing solid tumors (NCT02134925; NCT01978964); (2) HPV-16 E6-and E7-derived epitopes are being investigated as liposome-encapsulated adjuvanted interventions or combined with pegylated IFNa, carboplatin (an FDA-approved platinum derivative commonly used for the treatment of ovarian carcinoma)  [195] [196] [197]  and paclitaxel (an FDA-approved microtubular poison employed against various neoplasms),  198, 199  in women with histologically confirmed cervical intraepithelial neoplasia or cervical carcinoma (NCT02065973; NCT02128126); (3) peptides derived from folate receptor 1 (FOLR1, also known as FBP)  200, 201  are being tested as GM-CSF-adjuvanted standalone interventions in breast and ovarian carcinoma patients (NCT02019524); (4) peptides derived from HPV-16 E7 and premelanosome protein (PMEL, best known as gp100) are being assessed as Montanide ISA-51adjuvanted standalone interventions in melanoma patients (NCT01989559); (5) IDH1-derived epitopes are being tested as.
GM-CSF-adjuvanted standalone interventions in subjects bearing IDH1 C recurrent Grade II gliomas (NCT02193347); (  6 ) the safety and efficacy of peptides derived from indoleamine 2,3dioxygenase 1 (IDO1)  [202] [203] [204] [205] [206]  combined with the FDA-approved immune checkpoint blocker ipilimumab (also known as Yervoy TM )  [207] [208] [209] [210] [211] [212]  and vemurafenib (an FDA-approved inhibitor of mutant BRAF)  [213] [214] [215] [216] [217]  are being evaluated in melanoma patients (NCT02077114); (7) the therapeutic profile of a CMP targeting ganglioside GD2 and the so-called Lewis Y antigen (2 non-peptide TAAs),  [218] [219] [220]  employed as a PADRE-and Montanide ISA-51-adjuvanted intervention in combination with standard chemotherapy, is being assessed in breast carcinoma patients (NCT02229084); (8) full-length, recombinant cancer/ testis antigen 1B (CTAG1B, best known as NY-ESO-1)  107, [221] [222] [223] [224]  is being tested as a standalone intervention adjuvanted with GLA-SE, a synthetic TLR4 agonist,  [225] [226] [227] [228]  in subjects with solid tumors (NCT02015416); (  9 ) the safety and efficacy of peptides derived from lymphocyte antigen 6 complex, locus K (LY6K, also known as URLC10)  105, 108, [229] [230] [231]  administered as a standalone Montanide ISA-51-adjuvanted intervention are being evaluated in non-small cell lung carcinoma (NSCLC) patients (NCT01949701); (10) epitopes derived from fms-related tyrosine kinase 1 (FLT1, best known as VEGFR1) and kinase insert domain receptor (KDR, best known as VEGFR2) are being tested as standalone Montanide ISA-51-adjuvanted intervention in subjects affected by various solid tumors (NCT01949688); (11) the clinical profile of a not-better defined peptide-based vaccine (LPV7) adjuvanted with Montanide ISA-51, Hiltonol Ò , the tetanus toxoid (an FDA-approved adjuvant),  232  and resiquimod (a hitherto experimental TLR7 agonist)  [233] [234] [235] [236] [237]  is being assessed in melanoma patients (NCT02126579); (12) PPV approaches are being tested as Hiltonol Ò -adjuvanted standalone interventions in combination with standard chemotherapy in patients with melanoma, glioblastoma or glioma (NCT01970358; NCT02122822; NCT02149225); and (13) the clinical profile of multipeptidebased vaccines employed as standalone Montanide ISA-51-adjuvanted intervention, as standalone Montanide ISA-51-and GM-CSF-adjuvanted intervention, or combined with the IDO1 inhibitor INCB024360,  202, 238, 239  is being tested in melanoma or NSCLC patients (NCT01950156; NCT01961115; NCT 01989572).
Of note, both NCT01989572 and NCT01989559 are listed as "Completed" on http://www. clinicaltrials. gov. However, to the best of our knowledge, the results of neither of these studies have already been disseminated. As for the clinical trials listed in our 2 previous Trial Watches dealing with this topic,  47, 48  the following studies have changed status since the publication of the latter: NCT01854099, now listed as "Withdrawn;" NCT00624182 and NCT01264731, now listed as "Suspended;" NCT00455572, NCT00977145 and NCT01058850, now listed as "Terminated;" NCT00480025, NCT00643097, NCT00694551, NCT00819806, NCT00821652, NCT01149343, NCT01222221, NCT01250470, NCT01355393, NCT01425749, NCT01437605, NCT01462838, NCT01507103, NCT01580696, NCT01592617, NCT01606241 and NCT01632332, and now listed as "Active, not recruiting;" NCT00911560, NCT00960752, NCT01220128, NCT01307618, NCT01479244, NCT01526473, NCT01532960, NCT01570036 and NCT01922921, now listed as "Recruiting;" NCT00626015, NCT00655161, NCT00773097, NCT00841399, NCT00854789, NCT01639885 and NCT01673217, now listed as "Completed" (source http://www.
clinicaltrials. gov). NCT01854099 has been withdrawn prior to enrollment following the decision of the investigator, while NCT01264731 has been suspended because the study staff is out on medical leave. NCT00455572, NCT00977145 and NCT01058850 were all terminated owing to issues with patient accrual (source http://www. clinicaltrials. gov). NCT00626015 (a Phase I trial) tested the safety and efficacy of a peptide vaccine targeting the mutant epidermal growth factor receptor (EGFR) in combination with daclizumab (a monoclonal antibody specific for the IL-2 receptor a chain, CD25),  240, 241  temozolomide (an immunogenic alkylating agent)  70, 71, 242  and radiation therapy  72, 243, 244  in glioblastoma patients. 245 In this patient cohort, daclizumab provoked a decline in the circulating levels of CD4 C CD25 C FOXP3 C regulatory T cells  246  that correlated with the elicitation of humoral EGFR-specific immune responses.
245 NCT00773097 (a Phase II trial) tested the safety and immunological activity of a MUC1-derived peptide administered in combination with a TLR3 ligand in 46 advanced colorectal carcinoma patients. In this setting, all patients that completed the study (39 individuals) developed anti-MUC1 antibodies. No severe (Grade III-IV) side effects were recorded, but all patients experienced mild (Grade I-II) toxicities, notably skin reactions at the injection site (source http:// www. clinicaltrials. gov). NCT00841399 (a Phase I-II study) investigated the clinical profile of an HER2-derived peptide combined with GM-CSF in node-positive breast carcinoma patients. Of 195 enrolled patients, 187 were assessable at the end of the study, 108 of which were vaccinated as per compatible MHC profile. Vaccination was well tolerated and associated with a disease-free survival rate of 89.
7%, even though many individual received suboptimal vaccine doses (80. 2% among non-vaccinated patients, 94. 6% among optimally dosed patients). 134 A randomized, Phase III clinical trial has been initiated to confirm these results in a large patient cohort (NCT01479244, see above). To the best of our knowledge, the results of NCT00655161, NCT00854789, NCT01639885 and NCT01673217 have not been released yet (source http://www. clinicaltrials. gov).
Authors are supported by Ligue contre le Cancer ( equipe labelis ee); Agence National de la Recherche (ANR); Association pour la recherche sur le cancer (ARC); Canc eropôle Ile-de-France; AXA Chair for Longevity Research; Institut National du Cancer (INCa); Fondation Bettencourt-Schueller; Fondation de France; Fondation pour la Recherche M edicale (FRM); the European Commission (ArtForce); the European Research Council (ERC); the LabEx Immuno-Oncology; the SIRIC Stratified Oncology Cell DNA Repair and Tumor Immune Elimination (SOCRATE); the SIRIC Cancer Research and Personalized Medicine (CARPEM); and the Paris Alliance of Cancer Research Institutes (PACRI).
Funding Authors are supported by Ligue contre le Cancer ( equipe labelis ee); Agence National de la Recherche (ANR); Association pour la recherche sur le cancer (ARC); Canc eropôle Ile-de-France; AXA Chair for Longevity Research; Institut National du Cancer (INCa); Fondation Bettencourt-Schueller; Fondation de France; Fondation pour la Recherche M edicale (FRM); the European Commission (ArtForce); the European Research Council (ERC); the LabEx Immuno-Oncology; the SIRIC Stratified Oncology Cell DNA Repair and Tumor Immune Elimination (SOCRATE); the SIRIC Cancer Research and Personalized Medicine (CARPEM); and the Paris Alliance of Cancer Research Institutes (PACRI).
a. Adjuvanted with Montanide ISA-51, NCT02126579 Hiltonol Ò , tetanus toxoid and resiquimod II Recruiting Multiple Combined with an IDO1 inhibitor NCT01961115 III Completed Multiple As Montanide ISA-51-and GM-CSF- NCT01989572 adjuvanted intervention n. a. Completed E7 Adjuvanted with Montanide ISA-51 NCT01989559 PMEL NSCLC I/II Recruiting LY6K Adjuvanted with Montanide ISA-51 NCT01949701 I/II Recruiting Multiple Adjuvanted with Montanide ISA-51 NCT01950156 Solid tumors I/II Recruiting FLT1 Adjuvanted with Montanide ISA-51 NCT01949688 KDR SLPs Cervical carcinoma I/II Recruiting E6 As IFNa-adjuvanted intervention, NCT02128126 E7 combined with carboplatin and paclitaxel Abbreviations: CIN, cervical intraepithelial neoplasia; CMP, carbohydrate-mimetic peptide; FLT1, fms-related tyrosine kinase 1; FOLR1, folate receptor 1; GM- CSF, granulocyte macrophage colony-stimulating factor; IDH1, isocitrate dehydrogenase 1 (NADPC), soluble; IDO1, indoleamine 2,3-dioxygenase 1; IFNa, interferon a; KDR, kinase insert domain receptor; LeY, Lewis Y; LY6K, lymphocyte antigen 6 complex, locus K; MUC1, mucin 1; n.
a. , not available; NSCLC, non- small cell lung carcinoma; PADRE, pan-DR binding peptide epitope; PMEL, premelanosome protein; PPV, personalized peptide vaccination; SLP, synthetic long peptide; TAA, tumor-associated antigen. *Based on clinical trials initiated after September 2014, 1 st (source www. clinicaltrials. gov). e974411-4 OncoImmunology Volume 4 Issue 4.
Conclusions The therapeutic potential of Tregs in organ transplantation is wide and very promising. A number of clinical trials are currently taking place and will evaluate the safety and efficacy of FOXP3 Treg cellular therapy in kidney transplant recipients (https://clinicaltrials. gov/ct2/show/NCT02088931; https://clinicaltrials. gov/ct2/show/NCT02129881). However, the basic biology of Tregs has to be thoroughly studied to decipher the mechanisms that influence their stability and functions if we want to effectively and safely exploit these cells and propose them as a suitable therapeutic alternative for kidney transplant patients. A better understanding of how different promoting or inhibiting factors regulate Treg stability, under homeostatic and inflammatory settings, could lead to the emergence of alternative strategies for enhancing Treg stability and inducing long-term graft survival.
Thus, more sensitive genotyping techniques are needed to assess HIV tropism prior to initiating maraviroc therapy  [8]. Next-generation sequencing methods, such as ultra-deep pyrosequencing (UDPS), have been developed to increase sequencing capacity while generating clonal sequences. They have been shown to be as sensitive as phenotypic methods  [9, 10]. An important challenge with this technology is the very large number of sequences generated, that requires complex dataset analyses in order that the information becomes clinically meaningful. Bioinformatics algorithms that differentiate CCR5 from CXCR4 viral variants classically use rules based on the presence of substitutions at positions 11 and 25 and the global charge of the V3 loop  [11]  or comparisons with phenotypic test databases.
UDPS sequence analyses First, the quality of the experiments was analyzed and demultiplexing was performed with our in-house software Pyroclass  [15]. Four methods analyzing and reporting UDPS data were then compared. Method 1: consensus sequences were generated from UDPS data at D0 and used as a surrogate of population sequencing; they were interpreted by means of the geno2pheno[coreceptor] algorithm (Max-Planck-Institute), based on recommendations from the European Consensus Group on Clinical Management of HIV-1 Tropism Testing (false-positive rate: 5%)  [16]. Method 2: the consensus sequences were interpreted by means of the 11/25/charge rule   cp/mL]   4. 2 (3. 4-4. 9)  2. 0 (1. 6-2. 8)  1. 8 (1. 0-2. 5)  1. 8 (1. 0-2. 4) HIV-1 subtype B [%] 100.
Prior antiretroviral treatments Median number of NRTIs (IQR) 6  (5) (6) (7)  Median number of NNRTIs (IQR) using a module included into PyroTrop, an in-house software developed for this application. Method 3: the full set of UDPS sequences was interpreted with geno2pheno[454], an adaptation of the geno2pheno algorithm to multiple sequence files, such as those obtained with UDPS  [7]. This algorithm uses a cutoff value of 2% for the risk of maraviroc failure  [17]. Method 4: the full set of UDPS sequences was interpreted with our in-house software PyroTrop. Briefly, the previously described PyroMute software  [15]  was modified to assess each validated V3 sequence by means of the 11/25/charge algorithm. PyroTrop uses three quality filters to eliminate unreliable sequences, including sequences with a PHRED quality score <20  [18] , sequences not spanning the full-length HIV V3 loop, and sequences that are too divergent from the HXB2 strain reference HIV sequence (GenBank accession number K03455).
The latter filter was used to eliminate eventual contaminations by non-HIV sequences. Then, PyroTrop ascribes each valid V3 sequence to either CCR5 or CXCR4 by means of the 11/25/charge algorithm  [19]  and provides the respective percentages of these two populations within the patient's quasispecies (2% of CXCR4 variants was set as the detection threshold)  [20]. The results are provided as the respective proportions of CCR5 and CXCR4 populations in the patient's viral quasispecies, with a predefined lower limit of detection of 2%  [7]. Additional modules were included in PyroTrop to assess the frequency of each amino acid at each position, allowing for "machine learning" on the full data set.
Results HIV RNA could not be amplified in 11 patients at D0. Ten of them had an HIV RNA level below 1000 copies/ml; the remaining one had an HIV RNA level of 4. 6 log copies/ml but could not be amplified on three repeat tests. Pyrosequencing data of good quality were obtained in the remaining 102 patients at D0; 85, 79 and 73 of them had samples available for HIV RNA level determination at M1, M3 and M6, respectively. Overall, 16%, 13% and 12% of patients were considered as non-responders (<1 log HIV RNA level decrease) at M1, M3 and M6, respectively. The HIV RNA kinetics in responders is shown in S1 Fig. UDPS generated a total of 626,874 sequences (mean coverage 6146±298 per sample), among which 539,096 were valid for geno2pheno  [454]  analysis (mean coverage 5285±290 per sample; rejection rate: 14.
3%) and 461,044 for PyroTrop analysis (mean coverage 4520±306 per sample; rejection rate: 26,6%). Most of the removed sequences were rejected because the coverage of the V3 loop was partial (criterion used by all methods). In addition, PyroTrop rejected poor quality sequences or sequences which acceptable quality on only a portion of the V3 loop. Concordance between the four methods is shown in Table  2  for the 102 patients with available UDPS sequences at D0. Best concordance (94. 1%) was obtained with the two methods (consensus and UDPS) using Pyrotrop software. The performance of the four methods in predicting the virological response to maraviroc was compared. As shown in Table  3 , sensitivity was slightly higher for methods based on analysis of the full set of UDPS sequences, whereas the best specificity was achieved with UDPS data analyzed by means of Pyrotrop.
13 and p = 0. 08, respectively) (Fig  1 ). H34Y/S was present in 23% of the maraviroc responders, in whom it represented on average 79. 6% of the viral quasispecies population. In contrast, it was not detectable at significant levels in any of the non-responders to maraviroc (p = 0. 01). The PPV was defined as the capacity of the test (CCR5 tropism) to correctly predict a significant viral level decrease; the NPV was defined as the capacity of the test (CXCR4 tropism) to predict the lack of significant viral level decrease. Consensus means that the consensus of UDPS sequences has been used as a surrogate of population sequencing; UDPS means that the full set of pyrosequencing sequences has been used; g2p means that the geno2pheno algorithm has been used for analysis; Pyrotrop means that our in-house software has been used for analysis.
Discussion In this study, we assessed the performance of methods based on consensus (mimicking population sequencing) and UDPS sequence analysis, followed by geno2pheno or Pyrotrop data analysis, to predict the response to maraviroc in HIV-1 subtype B-infected patients under HAART therapy. Our results show that using UDPS instead of population sequencing improved specificity and the positive and negative predictive values, while the use of our in-house software Pyrotrop for UDPS sequence analysis further improved specificity and the positive predictive value as compared to geno2pheno. Overall, the rates of correct prediction of maraviroc response were significantly improved when using UDPS and analyzing the data with Pyrotrop, versus consensus sequence and the geno2pheno algorithm. It is currently possible to assess the distribution of HIV entry coreceptors and predict the response to maraviroc by means of sequencing methods, most often based on Sanger sequencing, now based on UDPS, using various software/algorithms.
Recent studies have suggested that UDPS improved the prediction of maraviroc response when used instead of Sanger sequencing  [8, 24]. Nevertheless, it has also been shown that different algorithms/software may yield different results, emphasizing the need for standardization and the combined use of complementary approaches  [10]. In this study, we confirmed the better performance of UDPS as compared to Sanger sequencing and validated our in-house software Pyrotrop as an alternative to geno2pheno to optimize analysis of UDPS data. Interestingly, the negative predictive value of UDPS sequencing was lower in our study than in a previous one comparing Sanger sequencing and UDPS  [8]. This might be due to the lower proportion of nonresponders in our study. Also, other reasons than tropism such as poor observance, pharmacological factors, etc, may, at least in part, explain the lack of response to maraviroc.
It is interesting to note that the H34S/Y substitution has been recently reported to play an important role in V3 affinity for both CCR5 and CXCR4, in particular when combined with a Q32K substitution  [25]. In our study, only half of the patients with an H34Y/S substitution also had a Q32K substitution and no relationship with the response was found. The predictive role of this substitution, alone or combined with H34S/Y will need to be further explored in larger series of patients. We looked at the frequency of these substitutions in the Los Alamos Database of HIV sequences (www. hiv. lanl. gov/content/sequence/HIV/mainpage. html). As shown in Table  5 , 60. 4% of the HIV sequences available in the database were wild-type, 25. 0% had one (17. 3%) or two (7. 7%) of these substitutions. When restricting the analysis to HIV-1 subtype B sequences, 69.
2% had the wild-type sequence, 16. 6% had one or two of these substitutions. These findings suggest that using this additional sequence parameter in algorithms for the determination of HIV tropism could benefit to a substantial number of patients. Nevertheless, the database is essentially based on population sequencing and does not provide information on the respective frequencies of the different viral populations in the quasispecies. In conclusion, UDPS is a powerful tool that can be used with confidence to predict maraviroc response in HIV-1-infected patients. Different data analysis tools can be used and we validated here the performance of our in-house Pyrotrop software that may represent a performing alternative to geno2pheno. Improvement of the predictive value of the algorithm used is possible and our results suggest that adding the H34S/Y substitution would substantially improve the performance of the 11/25 charge rule with these new approaches.
This research has been funded as part of the project "Collaborative HIV and Anti-HIV Drug Resistance Network (CHAIN)"within the 7th Framework Program of the European Community (FP7) and by the National Agency for Research on AIDS and Viral Hepatitis (ANRS). C. R. is the recipient of a pre-doctoral fellowship from ANRS. Dr Bernard Masquelier sadly passed away in March 2013. The authors are grateful to Dr Richard Harrigan for helpful discussion of the data and manuscript. We would like to acknowledge the contribution of the ANRS AC11 Resistance Study Group members: Aix en Provence (Dr E Lagier); Amiens (Dr C. Roussel); Angers (Dr H. Le Guillou); Avicenne (Dr C. Alloui); Besançon (Dr D. Bettinger); Bicêtre (Dr C. Pallier); Bordeaux (Pr H.
This research has been funded as part of the project " Collaborative HIV and Anti-HIV Drug Resistance Network (CHAIN) " within the  7th Framework Program  of the  European Community  ( FP7 ) and by the  National Agency for Research on AIDS and Viral Hepatitis (ANRS). C. R. is the recipient of a pre-doctoral fellowship from  ANRS. possible and our results suggest that adding the H34S/Y substitution would substantially improve the performance of the 11/25/charge rule.