ABSTRACT
To study the mechanisms of relapse in acute lymphoblastic leukemia (ALL), we performed whole-genome sequencing of 103 diagnosis-relapse-germline trios and ultra-deep sequencing of 208 serial samples in 16 patients. Relapse-specific somatic alterations were enriched in 12 genes (NR3C1, NR3C2, TP53, NT5C2, FPGS, CREBBP, MSH2, MSH6, PMS2, WHSC1, PRPS1, and PRPS2) involved in drug response. Their prevalence was 17% in very early relapse (<9 months from diagnosis), 65% in early relapse (9-36 months), and 32% in late relapse (>36 months) groups. Convergent evolution, in which multiple subclones harbor mutations in the same drug resistance gene, was observed in 6 relapses and confirmed by single-cell sequencing in 1 case. Mathematical modeling and mutational signature analysis indicated that early relapse resistance acquisition was frequently a 2-step process in which a persistent clone survived initial therapy and later acquired bona fide resistance mutations during therapy. In contrast, very early relapses arose from preexisting resistant clone(s). Two novel relapse-specific mutational signatures, one of which was caused by thiopurine treatment based on in vitro drug exposure experiments, were identified in early and late relapses but were absent from 2540 pan-cancer diagnosis samples and 129 non-ALL relapses. The novel signatures were detected in 27% of relapsed ALLs and were responsible for 46% of acquired resistance mutations in NT5C2, PRPS1, NR3C1, and TP53. These results suggest that chemotherapy-induced drug resistance mutations facilitate a subset of pediatric ALL relapses.
Subject(s)
Biomarkers, Tumor/genetics , Methotrexate/therapeutic use , Mutagenesis/drug effects , Mutation , Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , 5'-Nucleotidase/genetics , Antimetabolites, Antineoplastic/therapeutic use , Child , DNA Mutational Analysis , Female , Follow-Up Studies , Genomics , High-Throughput Nucleotide Sequencing , Humans , Male , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Prognosis , Receptors, Glucocorticoid/genetics , Survival Rate , Tumor Suppressor Protein p53/geneticsABSTRACT
MOTIVATION: Recent single-cell DNA sequencing technologies enable whole-genome sequencing of hundreds to thousands of individual cells. However, these technologies have ultra-low sequencing coverage (<0.5× per cell) which has limited their use to the analysis of large copy-number aberrations (CNAs) in individual cells. While CNAs are useful markers in cancer studies, single-nucleotide mutations are equally important, both in cancer studies and in other applications. However, ultra-low coverage sequencing yields single-nucleotide mutation data that are too sparse for current single-cell analysis methods. RESULTS: We introduce SBMClone, a method to infer clusters of cells, or clones, that share groups of somatic single-nucleotide mutations. SBMClone uses a stochastic block model to overcome sparsity in ultra-low coverage single-cell sequencing data, and we show that SBMClone accurately infers the true clonal composition on simulated datasets with coverage at low as 0.2×. We applied SBMClone to single-cell whole-genome sequencing data from two breast cancer patients obtained using two different sequencing technologies. On the first patient, sequenced using the 10X Genomics CNV solution with sequencing coverage ≈0.03×, SBMClone recovers the major clonal composition when incorporating a small amount of additional information. On the second patient, where pre- and post-treatment tumor samples were sequenced using DOP-PCR with sequencing coverage ≈0.5×, SBMClone shows that tumor cells are present in the post-treatment sample, contrary to published analysis of this dataset. AVAILABILITY AND IMPLEMENTATION: SBMClone is available on the GitHub repository https://github.com/raphael-group/SBMClone. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Subject(s)
Genomics , Software , Algorithms , Clone Cells , High-Throughput Nucleotide Sequencing , Humans , Mutation , Whole Genome SequencingABSTRACT
BACKGROUND: The Wessex Modified Richmond Sedation Scale (WMRSS) has been developed with the aim of improving the early identification of patients requiring decompressive hemicraniectomy for malignant middle cerebral artery syndrome (MMS). The objective of this study was to evaluate the WMRSS against the Glasgow Coma Scale (GCS). METHODS: A retrospective study was conducted of patients admitted to our unit for observation of MMS. Data were obtained on WMRSS and GCS recordings from admission up to 120-h post-ictus. Patients' meeting inclusion criteria were recommended for theatre based on subsequent deteriorations in consciousness on either WMRSS or GCS from a 6-h post-stroke baseline, after ruling out non-neurological causes. RESULTS: Approximately, 60% of those eligible for monitoring were not recommended for theatre, and none died; however, these patients continued to demonstrate some variability in recorded conscious level. Patients requiring surgical intervention showed earlier drops in WMRSS compared to GCS. Neither the GCS nor the WMRSS on admission predicted the subsequent need for decompressive surgery. There was no increase in mortality with the introduction of WMRSS. CONCLUSIONS: WMRSS adds value to monitoring MMS by indicating need for surgery prior to GCS. Early reduction in consciousness may not be sufficient for proceeding to surgical intervention, but subsequent reduction in consciousness may be a more appropriate criterion for surgery.
Subject(s)
Glasgow Coma Scale , Infarction, Middle Cerebral Artery/pathology , Monitoring, Physiologic/methods , Adult , Aged , Consciousness , Decompression, Surgical , Female , Humans , Infarction, Middle Cerebral Artery/surgery , Male , Middle AgedABSTRACT
Bulk DNA sequencing of multiple samples from the same tumor is becoming common, yet most methods to infer copy-number aberrations (CNAs) from this data analyze individual samples independently. We introduce HATCHet2, an algorithm to identify haplotype- and clone-specific CNAs simultaneously from multiple bulk samples. HATCHet2 extends the earlier HATCHet method by improving identification of focal CNAs and introducing a novel statistic, the minor haplotype B-allele frequency (mhBAF), that enables identification of mirrored-subclonal CNAs. We demonstrate HATCHet2's improved accuracy using simulations and a single-cell sequencing dataset. HATCHet2 analysis of 10 prostate cancer patients reveals previously unreported mirrored-subclonal CNAs affecting cancer genes.
Subject(s)
Algorithms , DNA Copy Number Variations , Haplotypes , Prostatic Neoplasms , Humans , Prostatic Neoplasms/genetics , Male , Sequence Analysis, DNA/methods , Neoplasms/genetics , Gene Frequency , Single-Cell AnalysisABSTRACT
Whole-genome doubling (WGD) is a critical driver of tumor development and is linked to drug resistance and metastasis in solid malignancies. Here, we demonstrate that WGD is an ongoing mutational process in tumor evolution. Using single-cell whole-genome sequencing, we measured and modeled how WGD events are distributed across cellular populations within tumors and associated WGD dynamics with properties of genome diversification and phenotypic consequences of innate immunity. We studied WGD evolution in 65 high-grade serous ovarian cancer (HGSOC) tissue samples from 40 patients, yielding 29,481 tumor cell genomes. We found near-ubiquitous evidence of WGD as an ongoing mutational process promoting cell-cell diversity, high rates of chromosomal missegregation, and consequent micronucleation. Using a novel mutation-based WGD timing method, doubleTime , we delineated specific modes by which WGD can drive tumor evolution: (i) unitary evolutionary origin followed by significant diversification, (ii) independent WGD events on a pre-existing background of copy number diversity, and (iii) evolutionarily late clonal expansions of WGD populations. Additionally, through integrated single-cell RNA sequencing and high-resolution immunofluorescence microscopy, we found that inflammatory signaling and cGAS-STING pathway activation result from ongoing chromosomal instability and are restricted to tumors that remain predominantly diploid. This contrasted with predominantly WGD tumors, which exhibited significant quiescent and immunosuppressive phenotypic states. Together, these findings establish WGD as an evolutionarily 'active' mutational process that promotes evolvability and dysregulated immunity in late stage ovarian cancer.
ABSTRACT
Multi-region DNA sequencing of primary tumors and metastases from individual patients helps identify somatic aberrations driving cancer development. However, most methods to infer copy-number aberrations (CNAs) analyze individual samples. We introduce HATCHet2 to identify haplotype- and clone-specific CNAs simultaneously from multiple bulk samples. HATCHet2 introduces a novel statistic, the mirrored haplotype B-allele frequency (mhBAF), to identify mirrored-subclonal CNAs having different numbers of copies of parental haplotypes in different tumor clones. HATCHet2 also has high accuracy in identifying focal CNAs and extends the earlier HATCHet method in several directions. We demonstrate HATCHet2's improved accuracy using simulations and a single-cell sequencing dataset. HATCHet2 analysis of 50 prostate cancer samples from 10 patients reveals previously-unreported mirrored-subclonal CNAs affecting cancer genes.
ABSTRACT
Methods of assessment in anatomy vary across medical schools in the United Kingdom (UK) and beyond; common methods include written, spotter, and oral assessment. However, there is limited research evaluating these methods in regards to student performance and perception. The National Undergraduate Neuroanatomy Competition (NUNC) is held annually for medical students throughout the UK. Prior to 2017, the competition asked open-ended questions (OEQ) in the anatomy spotter examination, and in subsequent years also asked single best answer (SBA) questions. The aim of this study is to assess medical students' performance on, and perception of, SBA and OEQ methods of assessment in a spotter style anatomy examination. Student examination performance was compared between OEQ (2013-2016) and SBA (2017-2020) for overall score and each neuroanatomical subtopic. Additionally, a questionnaire explored students' perceptions of SBAs. A total of 631 students attended the NUNC in the studied period. The average mark was significantly higher in SBAs compared to OEQs (60.6% vs. 43.1%, P < 0.0001)-this was true for all neuroanatomical subtopics except the cerebellum. Students felt that they performed better on SBA than OEQs, and diencephalon was felt to be the most difficult neuroanatomical subtopic (n = 38, 34.8%). Students perceived SBA questions to be easier than OEQs and performed significantly better on them in a neuroanatomical spotter examination. Further work is needed to ascertain whether this result is replicable throughout anatomy education.
Subject(s)
Anatomy , Education, Medical, Undergraduate , Students, Medical , Anatomy/education , Curriculum , Educational Measurement , Humans , Neuroanatomy/education , Schools, Medical , Surveys and QuestionnairesABSTRACT
BACKGROUND: Neurosurgery is a notoriously difficult career to enter and requires medical students to engage in extracurricular activities to demonstrate their commitment to the specialty. The National Undergraduate Neuroanatomy Competition (NUNC) was established in 2013 as a means for students to display this commitment as well as academic ability. METHODS: A bespoke 22-item questionnaire was designed to determine career outcomes and the role of competition attendance in job applications. It was distributed using the SurveyMonkey website to the 87 attendees at the 2013 and 2014 competitions. RESULTS: Responses were received by 40 competitors (response rate, 46.0%). Twenty-four responders (60.0%) intended to pursue a career in either neurosurgery (n = 18) or neurology (n = 6). This included 10 responders (25.0%) who had successfully entered either neurosurgery (n = 9) or neurology (n = 1). The performance of these 10 was significantly better than the other responders (57.0 ± 13.6% vs. 46.5 ± 13.5% [n = 30]; P = 0.036). Seventeen responders (42.5%) either included their attendance at NUNC in a post-Foundation job application or intend to. CONCLUSIONS: The NUNC provides the opportunity for medical students to demonstrate their interest in neurosurgery. It has the potential to be used as a tool for recognizing medical students suitable for neurosurgery training.
Subject(s)
Career Choice , Neurosurgery , Professional Competence , Students, Medical , Awards and Prizes , Humans , NeuroanatomyABSTRACT
Longitudinal DNA sequencing of cancer patients yields insight into how tumors evolve over time or in response to treatment. However, sequencing data from bulk tumor samples often have considerable ambiguity in clonal composition, complicating the inference of ancestral relationships between clones. We introduce Cancer Analysis of Longitudinal Data through Evolutionary Reconstruction (CALDER), an algorithm to infer phylogenetic trees from longitudinal bulk DNA sequencing data. CALDER explicitly models a longitudinally observed phylogeny incorporating constraints that longitudinal sampling imposes on phylogeny reconstruction. We show on simulated bulk tumor data that longitudinal constraints substantially reduce ambiguity in phylogeny reconstruction and that CALDER outperforms existing methods that do not leverage this longitudinal information. On real data from two chronic lymphocytic leukemia patients, we find that CALDER reconstructs more plausible and parsimonious phylogenies than existing methods, with CALDER phylogenies containing fewer tumor clones per sample. CALDER's use of longitudinal information will be advantageous in further studies of tumor heterogeneity and evolution.
Subject(s)
Algorithms , Computational Biology/methods , Neoplasms/genetics , Phylogeny , Software , Base Sequence , Cell Lineage , Computer Simulation , DNA, Neoplasm , Data Analysis , Humans , Leukemia, Lymphoid/geneticsABSTRACT
OBJECTIVE: Falls from standing are common, particularly amongst the aging population, due to declining mobility, proprioception and vision. They are often complicated by fragility fractures, including vertebral fractures, that are associated with significant morbidity and may represent a pre-terminal condition with high one-year mortality rates. PATIENTS AND METHODS: A retrospective review of the Trauma Audit and Research Network database for a major trauma centre was conducted for all patients admitted between January 2011 and December 2016. Patients with a spinal fracture and a confirmed fall from standing height were eligible for inclusion. Case notes were reviewed for demographics, Injury Severity Score, Charlson co-morbidity score, treatment, complications and outcomes. RESULTS: Of 1408 patients with a spine fracture admitted during the study period, 229 (16.3%) were confirmed to be secondary to a fall from standing height. The average age of this cohort was 76.6 ± 14.5 years and 134 (58.5%) cases were female. The average ISS score was 9.7 ± 5.4. The 229 patients sustained 283 fractures with a distribution of: cervical (n = 140), thoracic (n = 65) and lumbar (n = 78) spine. Fifty-six (24.5%) patients underwent surgical intervention. Forty-three patients (18.7%) died within 6 months of admission and all-cause mortality was significantly higher in patients with increasing age and Charlson co-morbidity score. CONCLUSION: Spinal fractures due to a fall from standing height represent one sixth of the fracture workload of the emergency spinal service at a major trauma centre. Whilst the majority of patients can be managed conservatively there are still considerable implications for hospital bed usage and patient mortality.
Subject(s)
Accidental Falls/statistics & numerical data , Cervical Vertebrae/surgery , Spinal Cord Injuries/surgery , Spinal Fractures/surgery , Adolescent , Adult , Aged , Aged, 80 and over , Cervical Vertebrae/injuries , Cohort Studies , Female , Hospitalization/statistics & numerical data , Humans , Male , Middle Aged , Retrospective Studies , Spinal Cord Injuries/complications , Spinal Fractures/epidemiology , Trauma Centers/statistics & numerical dataABSTRACT
Within medical education a reduction in curriculum time for subjects, such as anatomy puts pressure on educators to ensure the same learning outcomes are conveyed in less time. This has the potential to impact negatively on student experience. Near-peer teaching (NPT) is often praised as an effective revision tool, but its use as a frontline teaching resource remains unreported. The study explores the potential for NPT to promote delivery of learning outcomes and maximize student experience within a neuroanatomy module for second year medical students. The study occurred in three educational settings, (1) frontline NPT of cranial nerves, (2) revision session NPT of cranial nerves, and (3) NPT alongside faculty staff in laboratory-based neuroanatomy practical exercises. For the first and second components, knowledge was measured using a pre- and post-session test and student perception was ascertained with a questionnaire. For the third component, student perception was assessed with an end-of-module survey. The results show that overall, NPT was well received by learners. A significant knowledge gain was seen between the pre- and post-session test of the frontline NPT session. The study presents evidence in favor of using NPTs to supplement the delivery of learning outcomes in a time and resource constrained curriculum. In particular, for the effective delivery of frontline material. Anat Sci Educ 0: 1-9. © 2018 American Association of Anatomists.
Subject(s)
Anatomy/education , Education, Medical, Undergraduate/organization & administration , Peer Group , Students, Medical/statistics & numerical data , Teaching/organization & administration , Cross-Sectional Studies , Curriculum/statistics & numerical data , Curriculum/trends , Education, Medical, Undergraduate/methods , Education, Medical, Undergraduate/trends , Educational Measurement , Humans , Learning , Program Evaluation , Schools, Medical/organization & administration , Schools, Medical/statistics & numerical data , Schools, Medical/trends , Teaching/trendsABSTRACT
Undergraduates often perceive neuroscience to be a challenging discipline. As the scope of neuroscience continues to expand, it is important to provide undergraduates with sufficient opportunities to develop their knowledge and skills with the aim of encouraging the future generation of basic and clinical neuroscientists. Through our experience of developing the National Undergraduate Neuroanatomy Competition (NUNC), we have accrued an extensive volume of performance data and subjective insight into the delivery of undergraduate neuroanatomy education, which has the potential to inform how to better engage students within this field. More broadly, our group has implemented a technology enhanced learning platform alongside a peer-assisted teaching program. These achieve the dual purpose of compensating for the reduction in dedicated neuroanatomy teaching hours and encouraging undergraduates to develop an interest in the neurosciences. Here, we consider how improving the learning experience at an undergraduate level encourages further engagement in the neurosciences and the importance of this within the wider neuroscience community.
Subject(s)
Education, Medical, Undergraduate/methods , Neuroanatomy/education , Students , Education, Distance , Humans , Teaching/trends , United Kingdom , UniversitiesABSTRACT
BACKGROUND: Near-peer teaching is used in anatomy education because of its benefits to the learner, teacher and faculty members. Despite the range of reports focusing on the learner, the advantages for the teacher, which are thought to include communication skills, subject knowledge and employability, are only beginning to be explored. METHOD: A questionnaire was distributed to the teachers involved in anatomy near-peer teaching at the University of Southampton and Brighton and Sussex Medical School (BSMS). This questionnaire was designed using a rating scale of 0-10 to assess teacher perspectives on their level of knowledge, teaching skills and enjoyment of teaching. Free-text responses determined the teachers' motivation and perceived benefits from the teaching. RESULTS: Twenty-eight questionnaires were gathered (54.9% response rate), including 20 from Southampton and eight from BSMS. Long-term knowledge retention and better understanding of the material were rated 8.1 and 7.9 out of 10, respectively. Eight responses were from currently practising doctors, who rated how much they now use their teaching skills as doctors as 8.9 out of 10. Of the eight doctors, seven gained points for their foundation programme applications as a direct result of near-peer teaching. The most common motivator for engaging in teaching was to improve subject matter knowledge and the most common benefit was improved communication skills. There are numerous advantages to being a near-peer teacher in medical school DISCUSSION: There are numerous advantages to being a near-peer teacher in medical school, which include knowledge improvement, transferrable professional skills and employability. These initial results support the hypothesised benefits to the teachers and provide a foundation for further longitudinal studies.
Subject(s)
Education, Medical, Undergraduate/methods , Peer Group , Students, Medical/psychology , Teaching/organization & administration , Clinical Competence , Communication , Curriculum , Humans , Knowledge , Motivation , Teaching/standards , United KingdomABSTRACT
Rats exhibit 'empathy' making them a model to understand the neural underpinnings of such behavior. We show data consistent with these findings, but also that behavior and dopamine (DA) release reflects subjective rather than objective evaluation of appetitive and aversive events that occur to another. We recorded DA release in two paradigms: one that involved cues predictive of unavoidable shock to the conspecific and another that allowed the rat to refrain from reward when there were harmful consequences to the conspecific. Behavior and DA reflected pro-social interactions in that DA suppression was reduced during cues that predicted shock in the presence of the conspecific and that DA release observed on self-avoidance trials was present when the conspecific was spared. However, DA also increased when the conspecific was shocked instead of the recording rat and DA release during conspecific avoidance trials was lower than when the rat avoided shock for itself.
Subject(s)
Behavior, Animal , Dopamine/metabolism , Social Behavior , Animals , Electroshock , Male , Rats, Sprague-Dawley , RewardABSTRACT
Neurological conditions are common so a knowledge of neuroanatomy is necessary for junior doctors. Additionally, some students have a particular interest in neuroscience. However, little time is dedicated to neuroanatomy in the medical curriculum, and many students struggle with neuroanatomy. The National Undergraduate Neuroanatomy Competition (NUNC) aims to support the development of neuroanatomical knowledge among medical students and promote interest in neurosciences. Students who attended the NUNC completed a series of neuroanatomy-based examinations and a questionnaire investigating aspects of neuroanatomy teaching and resources at their home university. 387 students attended the NUNC between 2013 and 2017, of which 382 had a complete data set (response rate 98.7%). Male students significantly outperformed female students (p<0.0001) and clinical students outperformed pre-clinical students (p<0.05). Best answered questions were on the spine (average score 53.9%), and the most poorly answered questions were on the vasculature (average score 44.7%). Students felt that the neuroanatomy teaching, time spent on neuroanatomy and dissection/prosection resources were all reasonable (6-7/10) at their home institution. Elearning resources were rated more poorly (5.4/10). We conclude that the NUNC gives students the opportunity to enhance their neuroanatomical knowledge and gives keen students the chance to develop their interest
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