Single-cell multiomics reveals increased plasticity, resistant populations, and stem-cell-like blasts in KMT2A-rearranged leukemia.

KMT2A-rearranged (KMT2A-r) infant acute lymphoblastic leukemia (ALL) is a devastating malignancy with a dismal outcome, and younger age at diagnosis is associated with increased risk of relapse. To discover age-specific differences and critical drivers that mediate poor outcome in KMT2A-r ALL, we subjected KMT2A-r leukemias and normal hematopoietic cells from patients of different ages to single-cell multiomics analyses. We uncovered the following critical new insights: leukemia cells from patients <6 months have significantly increased lineage plasticity. Steroid response pathways are downregulated in the most immature blasts from younger patients. We identify a hematopoietic stem and progenitor-like (HSPC-like) population in the blood of younger patients that contains leukemic blasts and form an immunosuppressive signaling circuit with cytotoxic lymphocytes. These observations offer a compelling explanation for the ability of leukemias in young patients to evade chemotherapy and immune-mediated control. Our analysis also revealed preexisting lymphomyeloid primed progenitors and myeloid blasts at initial diagnosis of B-ALL. Tracking of leukemic clones in 2 patients whose leukemia underwent a lineage switch documented the evolution of such clones into frank acute myeloid leukemia (AML). These findings provide critical insights into KMT2A-r ALL and have clinical implications for molecularly targeted and immunotherapy approaches. Beyond infant ALL, our study demonstrates the power of single-cell multiomics to detect tumor intrinsic and extrinsic factors affecting rare but critical subpopulations within a malignant population that ultimately determines patient outcome.

KMT2A-MAML2 rearrangement emerged and regressed during neuroblastoma therapy without leukemia after 12.8-year follow-up.

Twelvepatients without therapy-related leukemia were studied after completing TOP2 poison chemotherapy in a high-risk neuroblastoma regimen. One patient harbored an inv(11) that was a KMT2A rearrangement. The KMT2A-MAML2 transcript was expressed at low level. The patient was prospectively followed. The inv(11) was undetectable in ensuing samples. Leukemia never developed after a 12.8-year follow-up period. Enriched etoposide-induced TOP2A cleavage in the relevant MAML2 genomic region supports a TOP2A DNA damage mechanism. After completing TOP2 poison chemotherapies, covert KMT2A-R clones may occur in a small minority of patients; however, not all KMT2A rearrangements herald a therapy-related leukemia diagnosis.

Genome-wide TOP2A DNA cleavage is biased toward translocated and highly transcribed loci.

Type II topoisomerases orchestrate proper DNA topology, and they are the targets of anti-cancer drugs that cause treatment-related leukemias with balanced translocations. Here, we develop a high-throughput sequencing technology to define TOP2 cleavage sites at single-base precision, and use the technology to characterize TOP2A cleavage genome-wide in the human K562 leukemia cell line. We find that TOP2A cleavage has functionally conserved local sequence preferences, occurs in cleavage cluster regions (CCRs), and is enriched in introns and lincRNA loci. TOP2A CCRs are biased toward the distal regions of gene bodies, and TOP2 poisons cause a proximal shift in their distribution. We find high TOP2A cleavage levels in genes involved in translocations in TOP2 poison-related leukemia. In addition, we find that a large proportion of genes involved in oncogenic translocations overall contain TOP2A CCRs. The TOP2A cleavage of coding and lincRNA genes is independently associated with both length and transcript abundance. Comparisons to ENCODE data reveal distinct TOP2A CCR clusters that overlap with marks of transcription, open chromatin, and enhancers. Our findings implicate TOP2A cleavage as a broad DNA damage mechanism in oncogenic translocations as well as a functional role of TOP2A cleavage in regulating transcription elongation and gene activation.

Clinical and patient-reported outcomes following Low Intensity Pulsed Ultrasound (LIPUS, Exogen) for established post-traumatic and post-surgical nonunion in the foot and ankle.

BACKGROUND: Biophysical methods including Low Intensity Pulsed Ultrasound (LIPUS) are emerging as potential alternatives to revision surgery for treating established nonunions. We aim to prospectively review the clinical and patient-reported outcomes of patients treated with LIPUS following post-traumatic and post-surgical nonunions in the foot and ankle. METHODS: Forty-seven consecutive patients underwent Exogen treatment. Patient-reported outcome scores included MOXFQ, EQ-5D and VAS. Patients were divided in to 3 groups: fractures (A), hindfoot procedures (B) and midfoot/forefoot procedures (C). RESULTS: Thirty-seven patients (78.7%) clinically united, 4 patients (8.5%) noticed no significant improvement but did not want further intervention and 6 patients (12.8%) underwent revision surgery. The mean duration of Exogen treatment was 6 months. Union rates of 93%, 67% and 78% were noted in the three groups. Significant improvement in functional outcomes and potential cost savings were observed. CONCLUSIONS: Exogen for established nonunion in the foot and ankle is a safe, valuable and economically viable clinical option as an alternative to revision surgery. We observed better results in the fracture and midfoot/forefoot groups and relatively poorer results in the hindfoot fusion group.

Modifiable lifestyle factors associated with risk of sessile serrated polyps, conventional adenomas and hyperplastic polyps.

OBJECTIVE: To identify modifiable factors associated with sessile serrated polyps (SSPs) and compare the association of these factors with conventional adenomas (ADs) and hyperplastic polyps (HPs). DESIGN: We used data from the Tennessee Colorectal Polyp Study, a colonoscopy-based case-control study. Included were 214 SSP cases, 1779 AD cases, 560 HP cases and 3851 polyp-free controls. RESULTS: Cigarette smoking was associated with increased risk for all polyps and was stronger for SSPs than for ADs (OR 1.74, 95% CI 1.16 to 2.62, for current vs never, ptrend=0.008). Current regular use of non-steroidal anti-inflammatory drugs was associated with a 40% reduction in SSP risk in comparison with never users (OR 0.68, 95% CI 0.48 to 0.96, ptrend=0.03), similar to the association with AD. Red meat intake was strongly associated with SSP risk (OR 2.59, 95% CI 1.41 to 4.74 for highest vs lowest intake, ptrend<0.001) and the association with SSP was stronger than with AD (ptrend=0.003). Obesity, folate intake, fibre intake and fat intake were not associated with SSP risk after adjustment for other factors. Exercise, alcohol use and calcium intake were not associated with risk for SSPs. CONCLUSIONS: SSPs share some modifiable risk factors for ADs, some of which are more strongly associated with SSPs than ADs. Thus, preventive efforts to reduce risk for ADs may also be applicable to SSPs. Additionally, SSPs have some distinctive risk factors. Future studies should evaluate the preventive strategies for these factors. The findings from this study also contribute to an understanding of the aetiology and biology of SSPs.

Evaluation of pro-inflammatory markers plasma C-reactive protein and urinary prostaglandin-E2 metabolite in colorectal adenoma risk.

C-reactive protein (CRP) is a pro-inflammatory protein with potential as a biomarker in predicting colon cancer risk. However, little is known regarding its association with risk of colorectal adenomas, particularly by subtypes. We conducted a colonoscopy-based matched case-control study to assess whether elevated plasma CRP levels may be associated with colorectal adenoma risk and further whether this association may be modified by urinary prostaglandin E2 metabolite (PGE-M), a biomarker of systemic prostaglandin E2 production. Included in the study were 226 cases with a single small tubular adenoma, 198 cases with multiple small tubular adenomas, 283 cases with at least one advanced adenoma, and 395 polyp-free controls. No apparent association between CRP level and risk of single small tubular adenomas was found (ptrend = 0.59). A dose-response relationship with CRP level was observed for risk of either multiple small tubular adenomas (OR = 2.01, 95%CI = 1.10-3.68 for the highest versus lowest tertile comparison; ptrend = 0.03) or advanced adenomas (OR = 1.81, 95%CI = 1.10-2.96 for the highest versus lowest tertile comparison; ptrend = 0.02). In a joint analysis of CRP level and PGE-M, risk of multiple or advanced adenoma was greatest among those with highest levels of both CRP and PGE-M in comparison to those with low CRP and low PGE-M (OR = 3.72, 95%CI = 1.49-9.72). Our results suggest that elevated CRP, particularly in the context of concurrent elevated PGE-M, may be a biomarker of multiple or advanced adenoma risk in a screening age population. © 2015 Wiley Periodicals, Inc.

Two to Five-Year Outcomes of Total Ankle Arthroplasty with the Infinity Fixed-Bearing Implant: A Concise Follow-up of a Previous Report.

LEVEL OF EVIDENCE: Therapeutic Level II. See Instructions for Authors for a complete description of levels of evidence.

Prosthetic Mitral Valve Thrombosis: A Complication Following Mitral Valve Surgery and Coronary Artery Bypass Graft Surgery.

Mitral valve regurgitation is a common valvular defect that can lead to severe complications, requiring surgical intervention, often in the form of either mitral valve repair or replacement. This case report follows a 63-year-old male with multivessel coronary artery disease, who initially presented to the emergency department (ED) with a non-ST elevation myocardial infarction (NSTEMI) secondary to multivessel coronary artery disease with severe mitral regurgitation, and subsequently underwent coronary artery bypass grafting (CABG) with repair of the mitral valve. He was readmitted a month later with endocarditis of the mitral valve and underwent a reoperation with a bioprosthetic mitral valve replacement and massive reconstruction of the right ventricle, after which he failed to recover postoperatively. A repeat transesophageal echocardiogram (TEE) during his final chest washout procedure revealed echodensities suspicious for thrombi and, despite the team's best efforts, the patient expired. This report demonstrates that even with appropriate medical decision-making, poor outcomes still result, especially in patients with comorbidities including multivessel disease, respiratory illness, and endocarditis. This study suggests that continuing to characterize repairs or replacements of the mitral valve is essential. Additionally, aggressive and newly emerging procedures, such as percutaneous approaches to mitral valve repair or replacement, may be considered for use to mitigate negative outcomes, especially with an aging population.

Recurrent Hiatal Hernia Resulting in Rightward Mediastinal Shift: Diagnostics in Cardiology and Clinical Pearls.

On radiographic imaging, the finding of a right-sided heart location can be due to multiple etiologies and may be congenital or acquired. We present the case of a 71-year-old male with a self-reported past medical history of hiatal hernia and previously diagnosed dextrocardia. The patient experienced cardiovascular intervention following an ST-elevation myocardial infarction. In the cardiac workup, a low-voltage normal electrocardiogram confirmed dextroposition of the heart due to significant herniation of gastric contents into the thoracic cavity. This gentleman had presumably been diagnosed with dextrocardia, a right-left reversal of heart anatomy and electrophysiology, based on imaging and incomplete workup. Dextroposition refers to a rightward shift of the mediastinum with no changes in orientation of cardiac anatomy, and therefore unchanged directional orientation of conduction. This is an important distinction from dextrocardia, a mirror-image reversal of the cardiac chambers and heart location in the chest wall, such as that due to congenital ciliary dysfunction. A sliding hernia is an uncommon cause of the rightward mediastinal shift, with few such cases documented in the literature, and cardiovascular manifestations of hiatal hernias are discussed. This case exemplifies the role of an electrocardiogram in distinguishing between dextrocardia and dextroposition for accurate diagnosis and management.

Missed Lisfranc injuries-surgical vs conservative treatment.

INTRODUCTION: Lisfranc injuries form a distinct group of rare but severe injury. Literature suggests a low incidence, but failure to diagnose these injuries early and its subsequent delay in management will affect the patient's mobility and quality of life significantly. The preferred mode of management is said to be surgical. Conversely, the method of intervention for patients not suitable for surgery is less clear. AIM: This study aims to evaluate the effect of delayed diagnosis and the treatment provided on the overall functional outcome for the patients with missed Lisfranc injury. METHODOLOGY: The study was conducted at a specialist centre in the North-West of UK between January 2011 and November 2016. All patients with acute Lisfranc injuries were included in this study. Patient data was collected through electronic notes and analysed to ascertain missed diagnosis. It was also used to evaluate functional and radiological outcome. RESULTS: In our series, 58.8% of Lisfranc injuries were missed on their initial presentation. We report better results for the surgical group when compared with the non-operated group, in spite of the delay in diagnosis. CONCLUSION: We believe that definitive treatment in the form of surgical fixation and anatomical reduction has more influence on the functional outcome than the timing of the surgical fixation in case of subtle Lisfranc injuries.

Disruption of intraflagellar transport in adult mice leads to obesity and slow-onset cystic kidney disease.

The assembly of primary cilia is dependent on intraflagellar transport (IFT), which mediates the bidirectional movement of proteins between the base and tip of the cilium. In mice, congenic mutations disrupting genes required for IFT (e.g., Tg737 or the IFT kinesin Kif3a) are embryonic lethal, whereas kidney-specific disruption of IFT results in severe, rapidly progressing cystic pathology. Although the function of primary cilia in most tissues is unknown, in the kidney they are mechanosenstive organelles that detect fluid flow through the tubule lumen. The loss of this flow-induced signaling pathway is thought to be a major contributing factor to cyst formation. Recent data also suggest that there is a connection between ciliary dysfunction and obesity as evidenced by the discovery that proteins associated with human obesity syndromes such as Alström and Bardet-Biedl localize to this organelle. To more directly assess the importance of cilia in postnatal life, we utilized conditional alleles of two ciliogenic genes (Tg737 and Kif3a) to systemically induce cilia loss in adults. Surprisingly, the cystic kidney pathology in these mutants is dependent on the time at which cilia loss was induced, suggesting that cyst formation is not simply caused by impaired mechanosensation. In addition to the cystic pathology, the conditional cilia mutant mice become obese, are hyperphagic, and have elevated levels of serum insulin, glucose, and leptin. We further defined where in the body cilia are required for normal energy homeostasis by disrupting cilia on neurons throughout the central nervous system and on pro-opiomelanocortin-expressing cells in the hypothalamus, both of which resulted in obesity. These data establish that neuronal cilia function in a pathway regulating satiety responses.

Quantifying Y Balance Test performance with multiple and single inertial sensors.

A growing body of evidence has highlighted that inertial sensor data can increase the sensitivity and clinical utility of the Y Balance Test, a commonly used clinical dynamic balance assessment. While early work has demonstrated the value of a single lumbar worn inertial sensor in quantifying dynamic balance control, no research has investigated if alternative (shank) or combined (lumbar and shank) sensor mounting locations may improve the assessments discriminant capabilities. Determining the optimal sensor set-up is crucial to ensuring minimal cost and maximal utility for clinical users The aim of this cross-sectional study was to investigate if single or multiple inertial sensors, mounted on the lumbar spine and/or shank could differentiate young (18-40 years [n = 41]) and middle-aged (40-65 years [n = 42]) adults, based on dynamic balance performance. Random-forest classification highlighted that a single lumbar sensor could classify age-related differences in performance with an accuracy of 79% (sensitivity = 81%; specificity = 78%). The amalgamation of shank and lumbar data did not significantly improve the classification performance (accuracy = 73-77%; sensitivity = 71-76%; specificity = 73-78%). Jerk magnitude root-mean-square consistently demonstrated predictor importance across the three reach directions: posteromedial (rank 1), anterior (rank 3) and posterolateral (rank 6).

An evaluation of the training determinants of marathon performance: A meta-analysis with meta-regression.

OBJECTIVES: Marathoners rely on expert-opinion and the anecdotal advice of their peers when devising their training plans for an upcoming race. The accumulation of results from multiple scientific studies has the potential to clarify the precise training requirements for the marathon. The purpose of the present study was to perform a systematic review, meta-analysis and meta-regression of available literature to determine if a dose-response relationship exists between a series of training behaviours and marathon performance. DESIGN: Systematic review, meta-analysis and meta-regression. METHODS: A systematic search of multiple literature sources was undertaken to identify observational and interventional studies of elite and recreational marathon (42.2km) runners. RESULTS: Eighty-five studies which included 137 cohorts of runners (25% female) were included in the meta-regression, with average weekly running distance, number of weekly runs, maximum running distance completed in a single week, number of runs ≥32km completed in the pre-marathon training block, average running pace during training, distance of the longest run and hours of running per week used as covariates. Separately conducted univariate random effects meta-regression models identified a negative statistical association between each of the above listed training behaviours and marathon performance (R2 0.38-0.81, p<0.001), whereby increases in a given training parameter coincided with faster marathon finish times. Meta-analysis revealed the rate of non-finishers in the marathon was 7.27% (95% CI 6.09%-8.65%). CONCLUSIONS: These data can be used by athletes and coaches to inform the development of marathon training regimes that are specific to a given target finish time.

Modeling Repeated M Dwarf Flaring at an Earth-like Planet in the Habitable Zone: Atmospheric Effects for an Unmagnetized Planet.

Understanding the impact of active M dwarf stars on the atmospheric equilibrium and surface conditions of a habitable zone Earth-like planet is key to assessing M dwarf planet habitability. Previous modeling of the impact of electromagnetic (EM) radiation and protons from a single large flare on an Earth-like atmosphere indicated that significant and long-term reductions in ozone were possible, but the atmosphere recovered. However, these stars more realistically exhibit frequent flaring with a distribution of different total energies and cadences. Here, we use a coupled 1D photochemical and radiative-convective model to investigate the effects of repeated flaring on the photochemistry and surface UV of an Earth-like planet unprotected by an intrinsic magnetic field. As input, we use time-resolved flare spectra obtained for the dM3 star AD Leonis, combined with flare occurrence frequencies and total energies (typically 1030.5 to 1034 erg) from the 4-year Kepler light curve for the dM4 flare star GJ1243, with varied proton event impact frequency. Our model results show that repeated EM-only flares have little effect on the ozone column depth but that multiple proton events can rapidly destroy the ozone column. Combining the realistic flare and proton event frequencies with nominal CME/SEP geometries, we find the ozone column for an Earth-like planet can be depleted by 94% in 10 years, with a downward trend that makes recovery unlikely and suggests further destruction. For more extreme stellar inputs, O3 depletion allows a constant ∼0.1-1 W m-2 of UVC at the planet's surface, which is likely detrimental to organic complexity. Our results suggest that active M dwarf hosts may comprehensively destroy ozone shields and subject the surface of magnetically unprotected Earth-like planets to long-term radiation that can damage complex organic structures. However, this does not preclude habitability, as a safe haven for life could still exist below an ocean surface.

Targeting EIF4E signaling with ribavirin in infant acute lymphoblastic leukemia.

The poor outcomes in infant acute lymphoblastic leukemia (ALL) necessitate new treatments. Here we discover that EIF4E protein is elevated in most cases of infant ALL and test EIF4E targeting by the repurposed antiviral agent ribavirin, which has anticancer properties through EIF4E inhibition, as a potential treatment. We find that ribavirin treatment of actively dividing infant ALL cells on bone marrow stromal cells (BMSCs) at clinically achievable concentrations causes robust proliferation inhibition in proportion with EIF4E expression. Further, we find that ribavirin treatment of KMT2A-rearranged (KMT2A-R) infant ALL cells and the KMT2A-AFF1 cell line RS4:11 inhibits EIF4E, leading to decreases in oncogenic EIF4E-regulated cell growth and survival proteins. In ribavirin-sensitive KMT2A-R infant ALL cells and RS4:11 cells, EIF4E-regulated proteins with reduced levels of expression following ribavirin treatment include MYC, MCL1, NBN, BCL2 and BIRC5. Ribavirin-treated RS4:11 cells exhibit impaired EIF4E-dependent nuclear to cytoplasmic export and/or translation of the corresponding mRNAs, as well as reduced phosphorylation of the p-AKT1, p-EIF4EBP1, p-RPS6 and p-EIF4E signaling proteins. This leads to an S-phase cell cycle arrest in RS4:11 cells corresponding to the decreased proliferation. Ribavirin causes nuclear EIF4E to re-localize to the cytoplasm in KMT2A-AFF1 infant ALL and RS4:11 cells, providing further evidence for EIF4E inhibition. Ribavirin slows increases in peripheral blasts in KMT2A-R infant ALL xenograft-bearing mice. Ribavirin cooperates with chemotherapy, particularly L-asparaginase, in reducing live KMT2A-AFF1 infant ALL cells in BMSC co-cultures. This work establishes that EIF4E is broadly elevated across infant ALL and that clinically relevant ribavirin exposures have preclinical activity and effectively inhibit EIF4E in KMT2A-R cases, suggesting promise in EIF4E targeting using ribavirin as a means of treatment.

Cilia proteins control cerebellar morphogenesis by promoting expansion of the granule progenitor pool.

Although human congenital cerebellar malformations are common, their molecular and developmental basis is still poorly understood. Recently, cilia-related gene deficiencies have been implicated in several congenital disorders that exhibit cerebellar abnormalities such as Joubert syndrome, Meckel-Gruber syndrome, Bardet-Biedl syndrome, and Orofaciodigital syndrome. The association of cilia gene mutations with these syndromes suggests that cilia may be important for cerebellar development, but the nature of cilia involvement has not been elucidated. To assess the importance of cilia-related proteins during cerebellar development, we studied the effects of CNS-specific inactivation of two mouse genes whose protein products are critical for cilia formation and maintenance, IFT88, (also known as polaris or Tg737), which encodes intraflagellar transport 88 homolog, and Kif3a, which encodes kinesin family member 3a. We showed that loss of either of these genes caused severe cerebellar hypoplasia and foliation abnormalities, primarily attributable to a failure of expansion of the neonatal granule cell progenitor population. In addition, granule cell progenitor proliferation was sensitive to partial loss of IFT function in a hypomorphic mutant of IFT88 (IFT88(orpk)), an effect that was modified by genetic background. IFT88 and Kif3a were not required for the specification and differentiation of most other cerebellar cell types, including Purkinje cells. Together, our observations constitute the first demonstration that cilia proteins are essential for normal cerebellar development and suggest that granule cell proliferation defects may be central to the cerebellar pathology in human cilia-related disorders.

The genomic landscape of childhood and adolescent melanoma.

Despite remarkable advances in the genomic characterization of adult melanoma, the molecular pathogenesis of pediatric melanoma remains largely unknown. We analyzed 15 conventional melanomas (CMs), 3 melanomas arising in congenital nevi (CNMs), and 5 spitzoid melanomas (SMs), using various platforms, including whole genome or exome sequencing, the molecular inversion probe assay, and/or targeted sequencing. CMs demonstrated a high burden of somatic single-nucleotide variations (SNVs), with each case containing a TERT promoter (TERT-p) mutation, 13/15 containing an activating BRAF V600 mutation, and >80% of the identified SNVs consistent with UV damage. In contrast, the three CNMs contained an activating NRAS Q61 mutation and no TERT-p mutations. SMs were characterized by chromosomal rearrangements resulting in activated kinase signaling in 40%, and an absence of TERT-p mutations, except for the one SM that succumbed to hematogenous metastasis. We conclude that pediatric CM has a very similar UV-induced mutational spectrum to that found in the adult counterpart, emphasizing the need to promote sun protection practices in early life and to improve access to therapeutic agents being explored in adults in young patients. In contrast, the pathogenesis of CNM appears to be distinct. TERT-p mutations may identify the rare subset of spitzoid melanocytic lesions prone to disseminate.

Viscous flow simulation in a stenosis model using discrete particle dynamics: a comparison between DPD and CFD.

Flow and stresses induced by blood flow acting on the blood cellular constituents can be represented to a certain extent by a continuum mechanics approach down to the order of the µm level. However, the molecular effects of, e.g., adhesion/aggregation bonds of blood clotting can be on the order of nm. The coupling of the disparate length and timescales between such molecular levels and macroscopic transport represents a major computational challenge. To address this challenge, a multiscale numerical approach based on discrete particle dynamics (DPD) methodology derived from molecular dynamics (MD) principles is proposed. The feasibility of the approach was firstly tested for its ability to simulate viscous flow conditions. Simulations were conducted in low Reynolds numbers flows (Re = 25-33) through constricted tubes representing blood vessels with various degrees of stenosis. Multiple discrete particles interacting with each other were simulated, with 1.24-1.36 million particles representing the flow domain and 0.4 million particles representing the vessel wall. The computation was carried out on the massive parallel supercomputer NY BlueGene/L employing NAMD-a parallel MD package for high performance computing (HPC). Typical recirculation zones were formed distal to the stenoses. The velocity profiles and recirculation zones were in excellent agreement with computational fluid dynamics (CFD) 3D Navier-Stokes viscous fluid flow simulations and with classic numerical and experimental results by YC Fung in constricted tubes. This feasibility analysis demonstrates the potential of a methodology that widely departs from a continuum approach to simulate multiscale phenomena such as flow induced blood clotting.