Genomic Characterization of Partial Tandem Duplication Involving the KMT2A Gene in Adult Acute Myeloid Leukemia.

BACKGROUND: Gene rearrangements affecting KMT2A are frequent in acute myeloid leukemia (AML) and are often associated with a poor prognosis. KMT2A gene fusions are often detected by chromosome banding analysis and confirmed by fluorescence in situ hybridization. However, small intragenic insertions, termed KMT2A partial tandem duplication (KMT2A-PTD), are particularly challenging to detect using standard molecular and cytogenetic approaches. METHODS: We have validated the use of a custom hybrid-capture-based next-generation sequencing (NGS) panel for comprehensive profiling of AML patients seen at our institution. This NGS panel targets the entire consensus coding DNA sequence of KMT2A. To deduce the presence of a KMT2A-PTD, we used the relative ratio of KMT2A exons coverage. We sought to corroborate the KMT2A-PTD NGS results using (1) multiplex-ligation probe amplification (MLPA) and (2) optical genome mapping (OGM). RESULTS: We analyzed 932 AML cases and identified 41 individuals harboring a KMT2A-PTD. MLPA, NGS, and OGM confirmed the presence of a KMT2A-PTD in 22 of the cases analyzed where orthogonal testing was possible. The two false-positive KMT2A-PTD calls by NGS could be explained by the presence of cryptic structural variants impacting KMT2A and interfering with KMT2A-PTD analysis. OGM revealed the nature of these previously undetected gene rearrangements in KMT2A, while MLPA yielded inconclusive results. MLPA analysis for KMT2A-PTD is limited to exon 4, whereas NGS and OGM resolved KMT2A-PTD sizes and copy number levels. CONCLUSIONS: KMT2A-PTDs are complex gene rearrangements that cannot be fully ascertained using a single genomic platform. MLPA, NGS panels, and OGM are complementary technologies applied in standard-of-care testing for AML patients. MLPA and NGS panels are designed for targeted copy number analysis; however, our results showed that integration of concurrent genomic alterations is needed for accurate KMT2A-PTD identification. Unbalanced chromosomal rearrangements overlapping with KMT2A can interfere with the diagnostic sensitivity and specificity of copy-number-based KMT2A-PTD detection methodologies.

Routine Use of a Standardized Mastectomy Diagram by Surgeons Improves Accuracy and Timeliness of the Final Pathological Report.

BACKGROUND: Accurate and timely assessment of pathology specimens is critical for patient care and oncologic management. This study aimed to determine whether a standardized mastectomy diagram would facilitate communication among surgeons and pathologists and improve pathologic processing. METHODS: A prospective quality improvement study was conducted over a continuous 12-month period. During the first 6 months, usual pathologic processing of mastectomy specimens was performed per standard department protocol. In the second 6 months, a standardized mastectomy diagram was completed at the time of surgery, noting the location and preoperative pathologic diagnosis of all benign and malignant lesions. An analysis of covariance was used to compare the number of breast lesions identified and the number of days between specimen receipt and the date of the final pathology report between each group. RESULTS: Time from specimen receipt to final pathologic report decreased from a mean (± SE) of 8.3 ± 0.7 days in the usual processing group to 6.1 ± 0.6 days with the use of the standardized mastectomy diagram, for a between-group difference of 2.1 days (95% confidence interval [CI] 0.3-4.0; p = 0.02). The number of lesions identified increased from 1.8 ± 0.2 to 2.6 ± 0.2, for a between-group difference of 0.8 (95% CI 0.1-1.5; p = 0.02). CONCLUSION: A standardized mastectomy diagram completed at the time of surgery improves the quality of pathologic processing. The diagram, which serves as a mastectomy lesion map, assists lesion localization, enhances accuracy, and reduces time to final pathology report.

Biallelic disruption of DDX41 activity is associated with distinct genomic and immunophenotypic hallmarks in acute leukemia.

Introduction: Inherited DDX41 mutations cause familial predisposition to hematologic malignancies including acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS), with the majority of DDX41 mutated MDS/AMLs described to date harboring germline DDX41 and co-occurring somatic DDX41 variants. DDX41-AMLs were shown to share distinguishing clinical features such as a late AML onset and an indolent disease associated with a favorable outcome. However, genotype-phenotype correlation in DDX41-MDS/AMLs remain poorly understood. Methods: Here, we studied the genetic profile, bone marrow morphology and immunophenotype of 51 patients with DDX41 mutations. We further assessed the functional impact of ten previously uncharacterized DDX41 variants of uncertain significance. Results: Our results demonstrate that MDS/AML cases harboring two DDX41 variants share specific clinicopathologic hallmarks that are not seen in other patients with monoallelic DDX41 related hematologic malignancies. We further showed that the features seen in these individuals with two DDX41 variants were concordant with biallelic DDX41 disruption. Discussion: Here, we expand on previous clinicopathologic findings on DDX41 mutated hematologic malignancies. Functional analyses conducted in this study unraveled previously uncharacterized DDX41 alleles and further illustrate the implication of biallelic disruption in the pathophysiology of this distinct AML entity.

The Long Noncoding RNA NEAT1 Promotes Sarcoma Metastasis by Regulating RNA Splicing Pathways.

Soft-tissue sarcomas (STS) are rare malignancies showing lineage differentiation toward diverse mesenchymal tissues. Half of all high-grade STSs develop lung metastasis with a median survival of 15 months. Here, we used a genetically engineered mouse model that mimics undifferentiated pleomorphic sarcoma (UPS) to study the molecular mechanisms driving metastasis. High-grade sarcomas were generated with Cre recombinase technology using mice with conditional mutations in Kras and Trp53 (KP) genes. After amputation of the limb bearing the primary tumor, mice were followed for the development of lung metastasis. Using RNA-sequencing of matched primary KP tumors and lung metastases, we found that the long noncoding RNA (lncRNA) Nuclear Enriched Abundant Transcript 1 (Neat1) is significantly upregulated in lung metastases. Furthermore, NEAT1 RNA ISH of human UPS showed that NEAT1 is upregulated within a subset of lung metastases compared with paired primary UPS. Remarkably, CRISPR/Cas9-mediated knockout of Neat1 suppressed the ability of KP tumor cells to colonize the lungs. To gain insight into the underlying mechanisms by which the lncRNA Neat1 promotes sarcoma metastasis, we pulled down Neat1 RNA and used mass spectrometry to identify interacting proteins. Interestingly, most Neat1 interacting proteins are involved in RNA splicing regulation. In particular, KH-Type Splicing Regulatory Protein (KHSRP) interacts with Neat1 and is associated with poor prognosis of human STS. Moreover, depletion of KHSRP suppressed the ability of KP tumor cells to colonize the lungs. Collectively, these results suggest that Neat1 and its interacting proteins, which regulate RNA splicing, are involved in mediating sarcoma metastasis. IMPLICATIONS: Understanding that lncRNA NEAT1 promotes sarcoma metastasis, at least in part, through interacting with the RNA splicing regulator KHSRP may translate into new therapeutic approaches for sarcoma.

Osteosarcoma and soft-tissue sarcomas with an immune infiltrate express PD-L1: relation to clinical outcome and Th1 pathway activation.

Immune checkpoint proteins, such as PD-L1 and PD-1, are important in several cancers; however, their role in osteosarcoma (OSA) and soft tissue sarcoma (STS) remains unclear. Our aims were to determine whether subsets of OSA/STS harbor tumor-infiltrating lymphocytes (TILs) and express PD-L1, and how PD-L1 expression is related to clinical outcome. Tissue sections of 25 cases each of untreated undifferentiated pleomorphic sarcoma (UPS), myxofibrosarcoma (MFS), liposarcoma (LPS) and 24 of leiomyosarcoma (LMS) were subjected to immunohistochemistry (IHC) for immune cells, PD-L1 and PD-1. RT-qPCR was utilized to quantify levels of PD-L1 mRNA from 33 UPS, 57 MFS and 79 OSA primary-untreated specimens. PD-L1 mRNA levels were tested for their correlation with overall survival in patients presenting without metastases. Transcriptome analysis evaluated biological pathway differences between high and low PD-L1 expressers. A subset of UPS and MFS contained TILs and expressed PD-L1 and PD-1; LMS and LPS did not. PD-L1 levels by IHC and RT-qPCR were positively correlated. PD-L1 over-expression was associated with better survival for UPS and OSA, but not MFS. The Th1 pathway was significantly activated in UPS with high levels of PD-L1 and improved survival. Some sarcoma subtypes harbor TILs and express PD-L1. Patients with UPS and OSA with high levels of PD-L1 had better overall survival than those with low expression levels. Important biological pathways distinguish PD-L1 high and low groups. The stratification of patients with OSA/STS with respect to potential immune therapies may be improved through investigation of the expression of immune cells and checkpoint proteins.

The importance of aspirin, catheterization accuracy, and catheter design in external ventricular drainage-related hemorrhage: a multicenter study of 1002 procedures.

BACKGROUND: External ventricular drainage (EVD) is the commonest neurosurgical procedure performed in daily neurosurgical practice, but relatively few studies have investigated the incidence and risk factors of its related hemorrhagic complications. METHODS: This was a multicenter retrospective review of consecutive EVD procedures. Patients 18 years or older who underwent EVD and had a routine postoperative computed tomography (CT) scan performed within 24 hours were included. EVD-related hemorrhage was defined as new intracranial hemorrhage immediately adjacent or within the ventricular catheter trajectory. The volume of hemorrhage and the position of the catheter tip were assessed. A review of patient-, disease-, and surgery-related factors including the ventricular catheter design utilized was conducted. The Bonferroni correction was applied to the alpha level of significance (0.05) for multivariable analysis. RESULTS: Nine hundred sixty-two patients underwent 1002 EVD performed by neurosurgeons in the operating theater. Sixteen percent (154) of patients were on aspirin before the procedure. Thirty-four percent (333) of patients had intracerebral hemorrhage, 25% (251) had aneurysmal subarachnoid hemorrhage and 16% (158) had traumatic brain injury. The mean duration from EVD to the first postoperative CT scan was 20 ± 4 h. EVD-related hematomas were detected after 81 procedures with a per-catheter risk of 8.1%. Mean hematoma volume was 1.2 ± 3.3 ml. Most were less than 1 ml (grade I, 79%, 64), 1 to 15 ml (grade II) in 20% (16) and a single clot larger than 15 ml (grade III, 1%) were detected. Clinically significant hemorrhage that resulted in catheter occlusion occurred in 1.7% (17) of procedures. Most catheters (62%, 625) were optimally placed, i.e., its tip being within the ipsilateral frontal horn or third ventricle. Three non-antibiotic-impregnated ventricular catheter designs were used with 55% (550) being the 2.2-mm Integra™ catheter, 14% (137) being the 2.8-mm Medtronic™ catheter, and 31% (315) being the 3.1-mm Codman™ catheter. Independent significant predictors for EVD-related hemorrhage were the preoperative prescription of aspirin (adjusted OR 1.94; 95% CI 1.10-3.44), catheter malposition (aOR 1.99; 95% CI 1.22-3.23), and use of the 2.8-mm Medtronic™ catheter (aOR 4.22; 95% CI 2.39-7.41). CONCLUSIONS: The per-catheter risk of hemorrhage was 8.1%, but the incidence of symptomatic hemorrhage was low. The only patient risk factor was aspirin intake. This is the first study to evaluate and establish an association between catheter malposition and catheter design with EVD-related hemorrhage.

A Systematic Review of Utility Score Assessments in the Breast Surgery Cost-Analysis Literature.

BACKGROUND: Surgery for breast cancer can have significant impact on patient quality-of-life. Cost-utility analysis provides a way to analyze the economic impact of a surgical procedure with the change in a patient's quality of life. Utility scores are used in these analyses to quantify the impact on quality of life. We undertook a systematic review of the literature on breast cancer surgical procedures to compile a repository of utility scores and to assess gaps in the current literature. METHODS: Following PRISMA guidelines, a systematic review was performed for studies reporting utility scores for breast surgery and breast reconstruction. The health states and utility scores were extracted and grouped into seven procedural categories based on oncologic and reconstructive methods. Mean utility score and ranges were calculated and reported for each procedural category. RESULTS: Nineteen articles met the inclusion criteria assessing 118 health states. Most utility scores were obtained from healthcare professionals. Breast-conserving therapy yielded the highest mean utility score at 0.79, whereas mastectomy yielded a mean utility score of 0.75. Among reconstruction health states, implant reconstruction had a lower score than autologous reconstruction (0.64 implant vs. latissimus dorsi 0.69 and TRAM/DIEP 0.71). No utility scores were found associated with oncoplasty or nipple-sparing mastectomy procedures. CONCLUSIONS: A reliable body of utility scores is important in enabling future cost-utility and value-based analysis comparisons for breast surgical oncology. Additional work is needed to obtain health state assessments from the patient perspective, as well as assessment of more modern surgical and reconstructive approaches.

The role of personality in patients with fibromyalgia.

Previous studies suggest personality, the multifaceted characteristics underlying a person's affect, cognition, and behavior, may influence fibromyalgia. We examined associations among personality, fibromyalgia impact, and health-related outcomes in patients with fibromyalgia. We further tested whether anxiety and depression mediated the effect of personality on fibromyalgia impact. We performed a secondary analysis using baseline data from a randomized trial on fibromyalgia. Personality was assessed using the NEO-Five Factor Inventory 3. Fibromyalgia impact was evaluated using the revised Fibromyalgia Impact Questionnaire (FIQR). We also measured symptom severity, anxiety, depression, stress, quality of life, social support, self-efficacy, outcome expectations, and mindfulness. Multivariable linear regression was performed to evaluate each association. Mediation analysis assessed whether anxiety and depression mediated the relationship between personality and FIQR. There were 92 participants, 95% female, mean age 52 years, body mass index (BMI) 30 kg/m2, 52% white, and mean duration of body pain 14 years. Higher neuroticism was significantly associated with higher FIQR (P = 0.002) and symptom severity (P = 0.008), as well as higher levels of anxiety, depression and stress, worse mental component quality of life, and lower self-efficacy, mindfulness, and social support. Higher conscientiousness and extraversion were associated with better psychological health and health-related outcomes. The effect of neuroticism on fibromyalgia impact was mediated by anxiety and depression. Personality was associated with fibromyalgia impact and a variety of health outcomes. Identifying the factors that influence fibromyalgia will help us better understand the condition and provide insight for more effective treatment.

Intraventricular urokinase to treat a blocked ventriculoperitoneal shunt in a glioblastoma patient with leptomeningeal spread.

Leptomeningeal spread and hydrocephalus are increasingly recognized as late disease complications of glioblastoma with almost a quarter of patients requiring early cerebrospinal fluid shunting. The neurosurgeon is challenged with maintaining shunt patency when tumor disease progression is rapid and adjuvant oncologic therapy has yet to be initiated. We describe our experience in treating a young female with diffuse glioblastoma leptomeningeal spread and communicating hydrocephalus who had several episodes of shunt obstruction due to intraluminal tumor cell-fibrin deposits. Regular intraventricular instillations of urokinase fibrinolytic therapy not only re-established shunt patency but also contributed to the resolution of her hydrocephalus.

Gene Expression Profiling of Endoplasmic Reticulum Stress in Hepatitis C Virus-Containing Cells Treated with an Inhibitor of Protein Disulfide Isomerases.

Protein disulfide isomerases (PDIs) catalyze disulfide bond formation between protein cysteine residues during protein folding in the endoplasmic reticulum (ER) lumen and are essential for maintaining ER homoeostasis. The life cycle of the hepatitis C virus (HCV) is closely associated with the ER. Synthesis and maturation of HCV proteins occur in the ER membrane and are mediated by multiple host cell factors that include also PDI. Here, we present a study investigating the effect of PDI inhibition on Huh7 human hepatoma cells harboring an HCV subgenomic replicon using the abscisic acid-derived PDI inhibitor origamicin. Transcriptional profiling shows that origamicin changed the expression levels of genes involved in the oxidative and ER stress responses and the unfolded protein response, as indicated by the upregulation of antioxidant enzymes and chaperone proteins, the downregulation of cell-cycle proteins, and induction of apoptosis-associated genes. Our data suggest that origamicin negatively impacts HCV replication by causing an imbalance in cellular homoeostasis and induction of stress responses. These insights suggest that inhibition of PDIs by low-molecular-weight inhibitors could be a promising approach to the discovery of novel antiviral compounds.