dataesr/scientific-paragraphs-categorization huggingface dataset:
PARP Inhibitors and Antiangiogenic Agents The rationale for the combination of PARP inhibitors with anti-angiogenic agents is based on the fact that PARP inhibition decreases angiogenesis, whereas hypoxic state and vascular endothelial growth factor receptor 3 (VEGFR3) inhibitors induce down-regulation of HR repair proteins, such as BRCA1/2 and RAD51. However, hypoxia is also associated with hypoxia inducible factor 1 alpha (HIF1α) up-regulation. Due to PARP1 involvement in HIF1α stabilization, inhibition of PARP may prevent HIF1α accumulation that leads to targeted hypoxic-induced apoptosis. The combination of the antiangiogenic agent cediranib and olaparib versus olaparib monotherapy has been evaluated in a randomized, open label, phase II study (NCT01116648)  [85]. Interim analysis revealed a significantly longer PFS of 17.
7 months in the experimental arm compared with 9. 0 months for patients treated with singleagent olaparib (HazR 0. 42; p = 0. 005). The purpose of the GY004 trial (NCT02446600) was to compare olaparib monotherapy versus doublet therapy of olaparib and cediranib versus standard platinum-based chemotherapy in patients with platinum-sensitive recurrent ovarian cancer  [86]. Similarly, the ICON 9 trial (NCT03278717) is examining maintenance therapy with the doublet of cediranib and olaparib, versus single agent olaparib  [87]. With regards to platinumresistant disease, three phase II/III trials are currently in progress  [88] [89] [90]. As previously mentioned, PAOLA-1/ENGOT-ov25 (NCT02477644) is the first phase III trial that evaluated efficacy and safety of a PARP inhibitor plus bevacizumab as firstline maintenance therapy in advanced EOC  [48].
2 months, compared with 17. 7 months with placebo (HazR 0. 33, 95% CI 0. 25-0. 45). HR-deficient BRCA wild-type patients treated with olaparib reached a median PFS of 28. 1 months versus 16. 6 months with placebo (HazR 0. 43, 95% CI 0. 28-0. 66). However, the benefit of olaparib in negative or unknown HR deficiency status was limited; median PFS was 16. 9 months versus 16. 0 months for olaparib and placebo, respectively (HazR 0. 92, 95% CI 0. 72-1. 17). Overall, PAOLA-1/ENGOT-ov25 met its primary objective, demonstrating a statistically significant prolongation of PFS in the intent-to-treat population. Patients with BRCA mutation and those with HR deficiency are most likely to benefit from this maintenance therapeutic strategy. Similarly, in the phase II OVARIO study (NCT03326193), niraparib plus bevacizumab is evaluated as a maintenance treatment in patients with advanced EOC who have recovered from primary debulking surgery and have responded to frontline platinum-based chemotherapy with bevacizumab  [91].
The starting dose of niraparib was based on baseline body weight and/or platelet count, whereas bevacizumab dosage was 15 mg/kg on a 3-weekly schedule up to 15 months, until disease progression or unacceptable toxicity. Finally, AVANOVA (NCT02354131) is a phase II superiority trial, comparing single-agent niraparib 300 mg with combination niraparib-bevacizumab (15 mg/kg once every 3 weeks), until disease progression  [92]. Ninety-seven enrolled women with high-grade serous or endometrioid platinum-sensitive EOC were assessed based on myChoice HR-deficient scores. Niraparib plus bevacizumab significantly improved PFS compared with niraparib alone [median PFS 11. 9 months (95% CI 8. 5-16. 7) vs 5. 5 months (95% CI 3. 8-6. 3), respectively; HazR 0. 35, 95% CI 0. 21-0. 57; p < 0. 0001].
NCT03426254 Active, not recruiting display a higher neo-antigen load than HR-proficient cancers  [93]. The targeted blockade of the PD-L1 pathway can restore antitumor immunity and potentiate the antitumor activity of PARP inhibitors. Currently, there are available data from olaparib-durvalumab, and niraparib-pembrolizumab combinations  [94, 95]. The reported toxicities are compatible with those that were observed for the relevant agents in monotherapy settings. The phase I/II basket MEDIOLA trial (NCT02734004), evaluated the combination of olaparib and durvalumab in selected advanced solid cancers  [94]. The phase I trial in patients with triple-negative breast and gynecological cancers demonstrated early evidence of efficacy, without overlapping toxicities  [95]. In the phase II study among 32 patients with germline BRCA1/2 mutant platinum-sensitive ovarian cancer, disease control rate at 12 weeks and ORR were 81% and 63%, respectively.
The most common reported adverse events of grade 3 or more were anemia (12%) and increased lipase (9%), along with any-grade hypothyroidism (15%) and rash (12%)  [94]. The phase I/II TOPACIO trial (NCT02657889) enrolled heavily pretreated patients with platinum-resistant, or secondarily platinum-refractory EOC  [96]. Based on dose finding in phase I, the recommended phase II dose of niraparib and pembrolizumab was 200 mg orally once daily and 200 mg intravenously 3-weekly, respectively. Among 60 patients evaluable for initial response assessment, 64% had platinum-resistant, 19% had platinum-refractory, and 17% had platinum-sensitive ovarian cancer, respectively  [97]. The overall ORR and disease control rate were 25% and 68%, respectively, whilst in the BRCA1/2 mutant cohort they were higher (45% and 73%, respectively).
Interestingly enough, the ORR in BRCA wild-type and HR deficiency negative subgroups was 24% and 27%, respectively. This is suggestive of treatment efficacy even in populations not typically responsive to single-agent PARP inhibitors. Preliminary data revealed adverse events compatible with treatment with single agents. Grade 3 or more toxicities included anemia (17%), fatigue (6%), and thrombocytopenia (3%)  [98]. Currently, there are five ongoing phase III studies in the maintenance setting. FIRST (NCT03602859) was designed to assess platinum and the PD-L1 inhibitor TSR-042, followed by niraparib and TSR-042 maintenance therapy, versus standard platinum-based treatment followed by maintenance niraparib or placebo, as first-line treatment of advanced EOC  [99]. The ENGOT-ov46/AGO/DUO-O study (NCT03737643) evaluates the efficacy and safety of the standard platinum-based chemotherapy and bevacizumab followed by maintenance bevacizumab as monotherapy, or in combination with durvalumab, either alone or combined with olaparib  [100].
ATHENA is a four-arm study (NCT03522246) that is currently evaluating the combination of rucaparib with nivolumab for those responded to front-line platinum-based chemotherapy  [101]. Pembrolizumab in combination with chemotherapy followed by olaparib maintenance therapy is being investigated in the phase III, randomized, double-blind ENGOT-ov43/ KEYLYNK-001 study (NCT03740165) in first-line treatment of patients with BRCA1/2 wild-type advanced EOC  [102]. Randomization is in a 1:1:1 ratio to pembrolizumab + chemotherapy followed by olaparib maintenance; pembrolizumab + chemotherapy followed by placebo; or placebo + chemotherapy followed by placebo. Bevacizumab use is determined at the investigator's discretion before randomization. Finally, JAVELIN Ovarian PARP 100 (  B9991030 ) is an open-label, international, multi-center, randomized study designed to evaluate the efficacy and safety of the combination of avelumab and chemotherapy followed by avelumab and talazoparib maintenance compared with chemotherapy and bevacizumab followed by bevacizumab maintenance in treatment-naïve patients with locally advanced or metastatic EOC (NCT03642132)  [103].
Furthermore, intermittent dosing schedules and supportive treatment with granulocyte colonystimulating factors are also reasonable strategies to improve the tolerability of chemotherapy in combination with PARP inhibitors. A phase III neoadjuvant Br ighTNess tr ial (NCT02032277) in patients with stage II-III, operable triple-negative breast cancer is a placebo-controlled study, where patients were randomized 2:1:1 to veliparib 50 mg twice daily in combination with the standard chemotherapy doublet of carboplatin and paclitaxel, versus paclitaxel plus carboplatin plus veliparib placebo (twice daily), versus paclitaxel plus carboplatin placebo plus veliparib placebo  [105]. Pathologic CR for the experimental arm was not significantly better than standard chemotherapy. As such, subtherapeutic tolerated doses of PARP inhibitors with chemotherapy may not potentiate therapeutic efficacy.
Among studies that investigate the utility of a combination of immune checkpoint blockade with PARP inhibitors, a phase Ib/II JAVELIN PARP MEDLEY (NCT03330405) trial is currently recruiting patients with germline BRCA1/2 mutations or ATM-deficient advanced solid tumors including breast and ovarian cancers, for evaluation of talazoparib in combination with PD-L1 inhibitor avelumab  [106]. Given the non-overlapping toxicities of immune checkpoint therapies and PARP inhibitors, the combination is expected to be well tolerated. Α study demonstrated that combination treatment of talazoparib and the next-generation DNA methyltransferase inhibitor guadecitabine optimized PARP inhibitors' response in breast and high-grade serous EOC cell lines, both in vitro and in vivo, regardless of BRCA status  [107].
Talazoparib was generally well tolerated; nevertheless, grade 3/4 myelosuppression was higher with talazoparib than chemotherapy (55 vs 39%), but still manageable by dose modifications or treatment delays. This grade 3/4 hematological toxicity is more prominent in EMBRACA as compared with the previously reported NCT01286987 dose-escalation study of talazoparib. Nevertheless, the combination of talazoparib with carboplatin in the study of Dhawan et al. [72].
Talazoparib was generally well tolerated; nevertheless, grade 3/4 myelosuppression was higher with talazoparib than chemotherapy (55 vs 39%), but still manageable by dose modifications or treatment delays. This grade 3/4 hematological toxicity is more prominent in EMBRACA as compared with the previously reported NCT01286987 dose-escalation study of talazoparib. Nevertheless, the combination of talazoparib with carboplatin in the study of Dhawan et al. [72] Study Phase Patient s (n) Description Population Outcome Trial, status de Bono et al. [13] I 113 Talazoparib 1 mg daily 1. Solid tumors (34/113 1. ORR: 41. 7% NCT01286987 Com- platinum-treated 2. gBRCAm: ORR: pleted EOC, primary perito- 55% in platinum- neal or fallopian tube sensitive cancer) ORR: 20% in platinum- 2. gBRCAm (25/34 resistant EOC) 3.
PFS: 36. 4 mo Dhawan et al. [72] I 24 Talazoparib + carbo- 1. Solid tumors (2/24 1. 14% ORR Completed platin EOC) 2. 52% SD Talazoparib starting 2. 14/24 (58%) of 3. Dose reduction: 50% dose of 0. 75 mg daily patients received prior 4. Dose interruptions: One cycle equaled platinum CTH 75% 21 days 3. gBRCAm (7/24, 5. Pharmacokinetics 29%) 4. sBRCAm (3/24, 12. 5%) POSITION [73] I 30 Talazoparib 1 mg daily 1. EOC, primary perito- Basal levels and effects NCT02316834 neal or fallopian tube of talazoparib on Ongoing cancer DNA copy number, 2. Neoadjuvant setting LOH, and mutation, and level of RNA and protein expres- sion in HRD-related pathways before and after treatment NCT02326844 [74] II 3 Talazoparib 1 mg daily 1. Recurrent and/or 1. Objective response NCT02326844 metastatic EOC (CR + PR) Terminated (closed by 2.
Progression on PARP 2. Safety the Cancer Therapy inhibitors mono- 3. Duration of response Evaluation Program) therapy 4. PFS 3. gBRCAm NCT02836028 [75] II N/A Arm 1: talazoparib 1. Recurrent EOC, ORR NCT02836028 1 mg daily primary peritoneal or Withdrawn Arm 2: talazoparib fallopian tube cancer 1 mg daily + temo-zolomide 37. 5 mg/m 2 2. <3 prior lines of CTH on days 1-5 3. gBRCAm, or sBR- CAm, or HRD(+) BRCA breast cancer susceptibility genes, CR complete response, CTH chemotherapy, EOC epithelial ovarian cancer, HRD homologous recombination deficiency, LOH loss of heterozygosis, mo months, ORR objective response rate, PARP poly(ADP-ribose) polymerase, PFS progressionfree survival, PR partial response, SD stable disease.
Using a hybrid assembly approach incorporating short-and long-read sequencing followed by manual finishing, we demonstrate how a highly contiguous assembly overcomes some of the critical limitations imposed by draft assemblies in interpreting large-scale genetic and functional genomic datasets. We provide insight into patterns of transcriptional change and putative co-regulation using a significantly improved genome annotation, derived de novo from both shortand long-read cDNA sequencing, and describe within-and between-population genetic diversity that shapes the genome. This chromosome-scale assembly now offers insight into genome evolution among a broad group of important parasite species and a robust scaffold for genome-wide analyses of important parasite traits such as anthelmintic resistance.
The overall annotation strategy is presented in Supplementary Fig. 6  and consisted of four key stages: (i) RNA-seq evidence was used to train ab initio gene finding using Braker 37 ; (ii) full-length high-quality isoforms generated using Iso-Seq were used to identify candidate gene models using the annotation tool Program to Assemble Spliced Alignments (PASA  38  ); (iii) evidence from steps (i) and (ii), together with 110 manually curated genes and orthologs from the C. elegans and H. contortus V1 genomes, were incorporated using EvidenceModeller; and finally (iv) UTRs and multievidence gene models were updated using the full-length isoforms once again using PASA. Together, this strategy achieved a high level of sensitivity and precision, and represented substantial improvement over the V1 annotation (Supplementary Table  4 ).
contortus and H. placei 39 , a gastrointestinal pathogen commonly associated with cattle. The increased orthology between close (H. placei) and more distantly (C. elegans) related species provides support for a more complete and biologically representative gene set in the V4 annotation than the previously described V1 and McMaster annotations. Transcriptional dynamics throughout development and between sexes. Both 2013 publications describing draft assemblies of H. contortus provide extensive descriptions of the key developmental transitions between sequential pairs of life stages throughout the life cycle  23, 24. We revisited this using the current annotation and, for visual comparison, present the top 1,000 most variable transcripts across all life stage samples in the MHco3 (ISE). N1 datasets (Fig.
This approach was initially difficult and timeconsuming and was further confounded by a large number of repetitive sequences present in each haplotype. Significant improvements in scaffold length were gained by the integration of OpGen (http://www. opgen. com/) optical mapping data. Optical mapping was performed following methods described previously  11  with the following exceptions: high molecular weight DNA was prepared by proteinase K lysis of pools of ~500 H. contortus L3 embedded in agarose plugs, after which one of three restriction enzymes (KpnI, AflII and KpnI) were used to generate three separate restriction map datasets. Initial attempts to generate a de novo assembly using optical molecules alone was unsuccessful, and therefore, optical contigs were generated using DNA sequence seeds from the genome assembly using GenomeBuilder (OpGen) and visualised and manually curated using AssemblyViewer (OpGen).
5× in single male sequencing libraries. We resolved the X chromosome using linked read long-range sequencing using the 10X Genomics Chromium platform (www. 10xgenomics. com). DNA prepared from pooled worms was used to generate 10X sequencing libraries, which were subsequently sequenced on a HiSeq 2500 using 250 bp PE reads. Reads were mapped using bwa mem (v0. 7. 17-r1188), followed by scaffolding using ARCS 78 and LINKS  79. A single scaffold was generated using this approach, merging the two major X-linked scaffolds as well as two short sequences putatively identified as being X-linked but previously unplaced (Supplementary Fig. 2 ). The completion of the X chromosome, together with the polishing of the genome first with arrow (https://github. com/PacificBiosciences/ GenomicConsensus) followed by Pilon v1.
22  80  , finalised the genome to produce the V4 version presented here. Genome completeness. Assembly completeness was determined throughout the improvement process using Core Eukaryotic Genes Mapping Approach (CEGMA; v2. 5)  81  and Benchmarking Universal Single-Copy Orthologs (BUSCO; v3. 0; -mode genome -lineage metazoa_odb9 -long) 82 gene completeness metrics. For comparison, these metrics were also determined for the complete set of nematode genomes from the WormBase ParaSite version 12 release. Haplotype analyses. The identification and discrimination of the major and minor haplotypes in our assembly offered an opportunity to characterise the distribution of haplotypes throughout the genome, and the relative diversity between them. To visualise the broad-scale variation between the chromosome and haplotypes, we generated an approximate genome graph using minigraph (https:// github.
com/lh3/minigraph; parameters: -xggs). The output GFA file was visualised using Bandage 83 ; a representative set of haplotypes were chosen for further comparison, and was compared using nucmer and visualised using Genome Ribbon 84. To quantify the distribution and diversity of haplotypes, we used minimap to align haplotypes per chromosome. Haplotype density was calculated in 1 Mbp windows throughout the genome using bedtools coverage  85  and visualised using the R package circlize  86. We determined the relative usage of detected haplotypes in the MHco3(ISE). N1 population by using Illumina sequencing of DNA from 11 individual worms and competitively mapping the sequencing data to both the reference and haplotype assemblies using bwa mem  87. Relative sequence coverage was determined using bedtools coverage  85  , which was normalised per sample to enable equivalent comparison between samples.
Transcriptome library preparation and sequencing. We made use of the RNAseq data generated for each of the life stages as previously described  23  together with additional full-length cDNA (PacBio Iso-Seq) sequencing performed with the specific aim of generating evidence for the annotation. Three life stages were used in the Iso-Seq analyses: adult female, adult male, and L3. RNA was extracted by homogenising the samples in Trizol in Fast prep (MP bio) followed by chloroform extraction and ethanol precipitation of the RNA, after which 1. 2 μg of total RNA was converted to cDNA using SmartSeq2 protocol  88. Size fractionation of equimolar pooled cDNA of the different stages was performed using a SageELF electrophoresis system (Sage Sciences), from which cDNA size fraction pools spanning 400 to 6,500 bp were collected.
Generation of a high-quality non-redundant Iso-Seq dataset. Full-length Iso-Seq subreads were mapped to the V4 genome using minimap2 (version 2. 6-r639dirty  89  ) with splice-aware mapping enabled (parameter: -ax splice). The generation of high-quality Iso-Seq reads was performed following the Iso-Seq3 pipeline (https://github. com/PacificBiosciences/IsoSeq3), which consisted of four steps: (i) the generation of circular consensus sequencing (CCS) reads from raw subreads (ccs subreads. bam subreads. ccs. bam -noPolish -minPasses 1), (ii) the removal of library primers, and reorientation of reads (lima subreads. ccs. bam primers. fa subreads. demux. bam -isoseq -no-pbi), (iii) clustering of transcripts, removal of concatemers and trimming of polyA tails (isoseq3 cluster subreads. demux. primer_5p-primer_3p.
Gene model prediction and genome annotation. A schematic of the strategy used for genome annotation is presented in Supplementary Fig. 6. Existing RNA-seq data were individually mapped to the V4 genome using STAR v2. 5. 2  90  before combined into a single bam using samtools merge. The merged bam was used as input to Braker (version 2. 0)  37, 91  , which can exploit mapped RNA-seq information as hints to train the gene prediction tools GeneMark-EX and Augustus, substantially improving genome annotation. The use of exon, intron, and other hints derived from RNA-seq or proteins mapped to the genome can be used to train Augustus directly; initial attempts to incorporate mapped Iso-Seq transcripts as an input to Braker, or by training Augustus using exon and intron hints, were successful and produced superior gene models compared with Braker using RNA-seq alone.
However, as Iso-Seq data encode full-length transcripts that include UTRs in a single sequence, and that in this data there was little discrimination between the depth of coverage between UTR and exon, many UTRs were incorrectly annotated as 5' or 3' coding exons. Accordingly, only short-read RNA-seq data were used as input to Braker, resulting in a primary annotation (GFF 1 ; Supplementary Fig. 6 ). To incorporate Iso-Seq data into the genome annotation, we used the PASA (v2. 2. 0) genome annotation tool  92  to map the high-quality full-length transcripts to the genome using both blat and gmap, akin to the use of a de novo transcriptome assembly produced from, for example, trinity, to generate a transcript database (https://github. com/PASApipeline/PASApipeline/wiki/PASA_comprehensive_db).
The Braker GFF 1 was subsequently loaded into PASA and integrated with the mapped Iso-Seq transcripts to produce the first round merged annotation (GFF 2 ; Supplementary Fig. 6 ). To incorporate further coding evidence into the annotation, we used EvidenceModeller v1. 1. 1  38  to apply a differential weighting schema to multiple evidence sources before merging them into a single genome annotation. Evidence used included the GFF 1 and GFF 2 annotations from Braker and PASA as ab initio and transcript evidence, respectively, and protein sequences from C. elegans and H. contortus V1 genome assemblies (obtained from Wormbase ParaSite release 9 caenorhabditis_elegans. PRJNA13758. WBPS9. protein. fa. gz and haemonchus_contortus. PRJEB506. WBPS9. protein. fa. gz, respectively), as well as 110 curated genes; each protein dataset was mapped to the genome using exonerate v2.
2. 0 (parameters: -model protein2genome -percent 50 -showtargetgff  93  ) followed by GFF extraction using Exonerate_to_evm_gff3. pl from EvidenceModeller. The weighting was applied as follows: PROTEIN (hc_v1 * curated models) = 2; PROTEIN (Ce) = 1; ABINITIO_PREDICTION (GFF 1 from Braker) = 2; TRANSCRIPT (GFF 2 from PASA) = 5. The merged output of EvidenceModeller (GFF 3 ) was imported back into PASA to update gene models with the full-length transcript information, including updates to UTRs and evidence for alternative splicing. Consistent with the author's recommendations, two rounds of iterative update and comparison were performed to maximise the use of the full-length transcript, i. e. , Iso-Seq, data (GFF 4 ; Supplementary Fig. 6 ). Gene IDs were subsequently renamed with the prefix HCON and numbered with an 8 digit identifier that incremented by a value of 10 to allow identifiers to be subsequently added for new features if required (GFF 5 ; Supplementary Fig.
elegans reference; no comparable annotation was generated for the intermediate assemblies generated before V4 was finalised, including assembly versions V2 and V3. Annotation comparisons to a curated gene set to determine the sensitivity and specificity of the approaches used were performed using gffcompare v0. 10. 1 (http://ccb. jhu. edu/ software/stringtie/gffcompare. shtml), and were further analysed for completeness using BUSCO (-mode proteins -lineage metazoa_odb9). The repetitive content of the genome was analysed using RepeatModeller (v1. 0. 11) followed by RepeatMasker (v4. 0. 7). Further analysis of LTRs was performed using LTRharvest and LTRdigest 96 from the GenomeTools v1. 5. 9  97  package. Transcriptome analyses. Quantitative transcriptome analyses were primarily performed using kallisto v0.