Osteopetrorickets due to Snx10 deficiency in mice results from both failed osteoclast activity and loss of gastric acid-dependent calcium absorption.

Mutations in sorting nexin 10 (Snx10) have recently been found to account for roughly 4% of all human malignant osteopetrosis, some of them fatal. To study the disease pathogenesis, we investigated the expression of Snx10 and created mouse models in which Snx10 was knocked down globally or knocked out in osteoclasts. Endocytosis is severely defective in Snx10-deficient osteoclasts, as is extracellular acidification, ruffled border formation, and bone resorption. We also discovered that Snx10 is highly expressed in stomach epithelium, with mutations leading to high stomach pH and low calcium solubilization. Global Snx10-deficiency in mice results in a combined phenotype: osteopetrosis (due to osteoclast defect) and rickets (due to high stomach pH and low calcium availability, resulting in impaired bone mineralization). Osteopetrorickets, the paradoxical association of insufficient mineralization in the context of a positive total body calcium balance, is thought to occur due to the inability of the osteoclasts to maintain normal calcium-phosphorus homeostasis. However, osteoclast-specific Snx10 knockout had no effect on calcium balance, and therefore led to severe osteopetrosis without rickets. Moreover, supplementation with calcium gluconate rescued mice from the rachitic phenotype and dramatically extended life span in global Snx10-deficient mice, suggesting that this may be a life-saving component of the clinical approach to Snx10-dependent human osteopetrosis that has previously gone unrecognized. We conclude that tissue-specific effects of Snx10 mutation need to be considered in clinical approaches to this disease entity. Reliance solely on hematopoietic stem cell transplantation can leave hypocalcemia uncorrected with sometimes fatal consequences. These studies established an essential role for Snx10 in bone homeostasis and underscore the importance of gastric acidification in calcium uptake.

A retrospective study of intravenous pentamidine for Pneumocystis jirovecii pneumonia prophylaxis in adult patients with hematologic malignancies-its utility during respiratory virus pandemics.

OBJECTIVES: In hematology, prophylaxis for Pneumocystis jirovecii pneumonia (PCP) is recommended for patients undergoing hematopoietic stem cell transplantation and in selected categories of intensive chemotherapy for hematologic malignancies. Trimethoprim-sulfamethoxazole (TMP-SMX) is the recommended first-line agent; however, its use is not straightforward. Inhaled pentamidine is the recommended second-line agent; however, aerosolized medications were discouraged during respiratory virus outbreaks, especially during the COVID-19 pandemic, in view of potential contamination risks. Intravenous (IV) pentamidine is a potential alternative agent. We evaluated the effectiveness and tolerability of IV pentamidine use for PCP prophylaxis in adult allogeneic hematopoietic stem cell transplantation recipients and patients with hematologic malignancies during COVID-19. RESULTS: A total of 202 unique patients who received 239 courses of IV pentamidine, with a median of three doses received (1-29). The largest group of the patients (49.5%) who received IV pentamidine were undergoing or had received a hematopoietic stem cell transplant. The most common reason for not using TMP-SMX prophylaxis was cytopenia (34.7%). We have no patients who had breakthrough PCP infection while on IV pentamidine. None of the patients developed an infusion reaction or experienced adverse effects from IV pentamidine. CONCLUSIONS: Pentamidine administered IV monthly is safe and effective.

Management of classical Philadelphia chromosome-negative myeloproliferative neoplasms in Asia: consensus of the Asian Myeloid Working Group.

Myeloproliferative neoplasms (MPN) are a heterogeneous group of clonal hematopoietic stem cell disorders characterized clinically by the proliferation of one or more hematopoietic lineage(s). The classical Philadelphia-chromosome (Ph)-negative MPNs include polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF). The Asian Myeloid Working Group (AMWG) comprises representatives from fifteen Asian centers experienced in the management of MPN. This consensus from the AMWG aims to review the current evidence in the risk stratification and treatment of Ph-negative MPN, to identify management gaps for future improvement, and to offer pragmatic approaches for treatment commensurate with different levels of resources, drug availabilities and reimbursement policies in its constituent regions. The management of MPN should be patient-specific and based on accurate diagnostic and prognostic tools. In patients with PV, ET and early/prefibrotic PMF, symptoms and risk stratification will guide the need for early cytoreduction. In younger patients requiring cytoreduction and in those experiencing resistance or intolerance to hydroxyurea, recombinant interferon-α preparations (pegylated interferon-α 2A or ropeginterferon-α 2b) should be considered. In myelofibrosis, continuous risk assessment and symptom burden assessment are essential in guiding treatment selection. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) in MF should always be based on accurate risk stratification for disease-risk and post-HSCT outcome. Management of classical Ph-negative MPN entails accurate diagnosis, cytogenetic and molecular evaluation, risk stratification, and treatment strategies that are outcome-oriented (curative, disease modification, improvement of quality-of-life).

Stent thrombogenicity early in high-risk interventional settings is driven by stent design and deployment and protected by polymer-drug coatings.

BACKGROUND: Stent thrombosis is a lethal complication of endovascular intervention. Concern has been raised about the inherent risk associated with specific stent designs and drug-eluting coatings, yet clinical and animal support is equivocal. METHODS AND RESULTS: We examined whether drug-eluting coatings are inherently thrombogenic and if the response to these materials was determined to a greater degree by stent design and deployment with custom-built stents. Drug/polymer coatings uniformly reduce rather than increase thrombogenicity relative to matched bare metal counterparts (0.65-fold; P=0.011). Thick-strutted (162 µm) stents were 1.5-fold more thrombogenic than otherwise identical thin-strutted (81 µm) devices in ex vivo flow loops (P<0.001), commensurate with 1.6-fold greater thrombus coverage 3 days after implantation in porcine coronary arteries (P=0.004). When bare metal stents were deployed in malapposed or overlapping configurations, thrombogenicity increased compared with apposed, length-matched controls (1.58-fold, P=0.001; and 2.32-fold, P<0.001). The thrombogenicity of polymer-coated stents with thin struts was lowest in all configurations and remained insensitive to incomplete deployment. Computational modeling-based predictions of stent-induced flow derangements correlated with spatial distribution of formed clots. CONCLUSIONS: Contrary to popular perception, drug/polymer coatings do not inherently increase acute stent clotting; they reduce thrombosis. However, strut dimensions and positioning relative to the vessel wall are critical factors in modulating stent thrombogenicity. Optimal stent geometries and surfaces, as demonstrated with thin stent struts, help reduce the potential for thrombosis despite complex stent configurations and variability in deployment.

Sun exposure and risk of lymphoid neoplasms in Singapore.

BACKGROUND: Epidemiologic studies have reported an inverse association between sun exposure and non-Hodgkin lymphoma (NHL), but these have been almost exclusively conducted in Western populations residing in temperate locations. We evaluated the association between personal outdoor sun exposure and risk of malignant lymphomas in Singapore. METHODS: A hospital-based case-control study of 541 incident cases of lymphoid neoplasms and 830 controls were recruited during 2004-2008. Participants were interviewed regarding recreational or occupational outdoor activities during childhood and in adulthood. Basic demographics and potential confounders were also collected. Odds ratios (OR) and 95 % confidence intervals (CI) were calculated using unconditional logistic regression analysis. RESULTS: Compared with individuals who did not have regular sun exposure, a lower risk of NHL was observed for those who reported regular exposure on non-school days during childhood [OR, 0.62; 95 % CI, 0.46-0.83] and non-working days in adulthood [OR, 0.70; 95 % CI, 0.51-0.97]. The protective effect was more evident among women. CONCLUSION: Our findings support an inverse relationship between intermittent sun exposure and the risk of NHL. These findings are consistent with the growing evidence from various countries, but further studies, especially prospective studies, are needed in Asian populations.

Yeast arginine methyltransferase Hmt1p regulates transcription elongation and termination by methylating Npl3p.

The heterogeneous nuclear ribonucleoprotein Npl3p of budding yeast is a substrate of arginine methyltransferase Hmt1p, but the role of Hmt1p in regulating Npl3p's functions in transcription antitermination and elongation were unknown. We found that mutants lacking Hmt1p methyltransferase activity exhibit reduced recruitment of Npl3p, but elevated recruitment of a component of mRNA cleavage/termination factor CFI, to the activated GAL10-GAL7 locus. Consistent with this, hmt1 mutants displayed increased termination at the defective gal10-Delta56 terminator. Remarkably, hmt1Delta cells also exhibit diminished recruitment of elongation factor Tho2p and a reduced rate of transcription elongation in vivo. Importantly, the defects in Npl3p and Tho2p recruitment, antitermination and elongation in hmt1Delta cells all were mitigated by substitutions in Npl3p RGG repeats that functionally mimic arginine methylation by Hmt1p. Thus, Hmt1p promotes elongation and suppresses termination at cryptic terminators by methylating RGG repeats in Npl3p. As Hmt1p stimulates dissociation of Tho2p from an Npl3p-mRNP complex, it could act to recycle these elongation and antitermination factors back to sites of ongoing transcription.

A novel network pharmacology approach for leukaemia differentiation therapy using Mogrify®.

Acute myeloid leukaemia (AML) is a rapidly fatal blood cancer that is characterised by the accumulation of immature myeloid cells in the blood and bone marrow as a result of blocked differentiation. Methods which identify master transcriptional regulators of AML subtype-specific leukaemia cell states and their combinations could be critical for discovering novel differentiation-inducing therapies. In this proof-of-concept study, we demonstrate a novel utility of the Mogrify® algorithm in identifying combinations of transcription factors (TFs) and drugs, which recapitulate granulocytic differentiation of the NB4 acute promyelocytic leukaemia (APL) cell line, using two different approaches. In the first approach, Connectivity Map (CMAP) analysis of these TFs and their target networks outperformed standard approaches, retrieving ATRA as the top hit. We identify dimaprit and mebendazole as a drug combination which induces myeloid differentiation. In the second approach, we show that genetic manipulation of specific Mogrify®-identified TFs (MYC and IRF1) leads to co-operative induction of APL differentiation, as does pharmacological targeting of these TFs using currently available compounds. We also show that loss of IRF1 blunts ATRA-mediated differentiation, and that MYC represses IRF1 expression through recruitment of PML-RARα, the driver fusion oncoprotein in APL, to the IRF1 promoter. Finally, we demonstrate that these drug combinations can also induce differentiation of primary patient-derived APL cells, and highlight the potential of targeting MYC and IRF1 in high-risk APL. Thus, these results suggest that Mogrify® could be used for drug discovery or repositioning in leukaemia differentiation therapy for other subtypes of leukaemia or cancers.

Real-World Experience of Measurable Residual Disease Response and Prognosis in Acute Myeloid Leukemia Treated with Venetoclax and Azacitidine.

The prognostic value of measurable residual disease (MRD) by flow cytometry in acute myeloid leukemia (AML) patients treated with non-intensive therapy is relatively unexplored. The clinical value of MRD threshold below 0.1% is also unknown after non-intensive therapy. In this study, MRD to a sensitivity of 0.01% was analyzed in sixty-three patients in remission after azacitidine/venetoclax treatment. Multivariable cox regression analysis identified prognostic factors associated with cumulative incidence of relapse (CIR), progression-free survival (PFS) and overall survival (OS). Patients who achieved MRD < 0.1% had a lower relapse rate than those who were MRD ≥ 0.1% at 18 months (13% versus 57%, p = 0.006). Patients who achieved an MRD-negative CR had longer median PFS and OS (not reached and 26.5 months) than those who were MRD-positive (12.6 and 10.3 months, respectively). MRD < 0.1% was an independent predictor for CIR, PFS, and OS, after adjusting for European Leukemia Net (ELN) risk, complex karyotype, and transplant (HR 5.92, 95% CI 1.34−26.09, p = 0.019 for PFS; HR 2.60, 95% CI 1.02−6.63, p = 0.046 for OS). Only an MRD threshold of 0.1%, and not 0.01%, was predictive for OS. Our results validate the recommended ELN MRD cut-off of 0.1% to discriminate between patients with improved CIR, PFS, and OS after azacitidine/venetoclax therapy.

Midostaurin in acute myeloid leukemia: current evidence and practical considerations in routine clinical use.

Mutation within the FMS-like tyrosine kinase 3 (FLT3) gene are one of the most frequent genetic alterations in acute myeloid leukemia. A high mutation fraction of FLT3-ITD molecules on the surface of leukemia cells is associated with short remissions and overall adverse outcomes in AML. In this article we summarize the clinical trial data of midostaurin - one of the FLT3 inhibitors. We review its use in various combinations both in relapsed/refractory acute myeloid leukemia as well as in the newly diagnosed patients and recollect the evidence of its use as maintenance therapy post allogenic stem cell transplantation. We enumerate the practical issues faced in the use of midostaurin like antifungal prophylaxis, dosage of concomitant chemotherapy agents as well as available data on sequencing of the FLT3 inhibitors. Lastly, we provide our perspective of the future directions for FLT3 inhibition especially midostaurin, the underlying resistance mechanisms and the need for standardization of the FLT3 tests.

Surgical management of invasive fungal infections in adult leukemia patients: experience from a large tertiary center in Southeast-Asia.

Objectives: Invasive fungal infections (IFIs) are a major cause of morbidity and mortality in acute leukemia patients undergoing chemotherapy or hematopoietic stem cell transplantation (HSCT). Surgical interventions may be necessary to improve the survival outcomes of these patients. The aim of this study is to report a single-center experience using surgical intervention as adjunctive treatment for IFI in adult leukemia patients. Methods: A retrospective review was conducted to obtain clinical characteristics and outcomes of surgically managed IFI patients diagnosed between January 2005 and December 2015 in our center. Results: Nineteen acute leukemia patients, median age 46 years (range 19-65), underwent 20 surgical procedures as management for IFI. Three patients had proven IFI diagnoses prior to surgery. Sixteen patients underwent surgery for both diagnostic and therapeutic purposes. Post-surgery, the diagnostic yield for proven IFI increased by a factor of 5, and 15 patients had definitive IFI diagnoses. Surgical complications included 2 pleural effusions, 4 pneumothoraxes, and 1 hydropneumothorax. The median duration of hospitalization for patients with complications was 9 days (range 3-64). Thirteen patients benefited overall from the procedure, 3 had temporary clinical benefits, and 2 had progression of IFI. After surgery, the 3-month and 2-year overall survival rates were 89.5% and 57.9%, respectively. The median time from surgery to resumption of chemotherapy or HSCT was 25 days. Conclusions: Surgical interventions for IFI are feasible in selected leukemia patients, as they yield valuable information to guide antifungal therapy or enable therapeutic outcomes with acceptable risk, thereby allowing patients to proceed with curative chemotherapy and HSCT.

The utility of voriconazole therapeutic drug monitoring in a multi-racial cohort in Southeast Asia.

OBJECTIVES: Voriconazole serum concentration, which is affected by several factors, is associated with treatment response and toxicity. There is paucity of data on voriconazole therapeutic drug monitoring (TDM) among Southeast Asians, who exhibit a higher prevalence of CYP2C19-poor metabolisers compared with Caucasians and East Asians. Hence, there are concerns for higher risk of voriconazole accumulation and toxicity. We aim to determine the utility of voriconazole TDM through establishing: (1) proportion of patients achieving therapeutic troughs without dose adjustments; (2) characterisation of patients with sub-therapeutic, therapeutic and supra-therapeutic levels; (3) appropriate dose titrations/dose required for therapeutic troughs; (4) correlation between troughs and adverse events, treatment response/fungal breakthrough. PATIENTS AND METHODS: A single-centre retrospective analysis of data from adults (≥21 years old) with ≥1 voriconazole trough measured at Singapore General Hospital from 2015 to 2017 was performed. RESULTS: Thirty-two patients (45.7%) among 70 patients achieved therapeutic troughs (defined as 2.0-5.5 mg/L) without dose adjustments. Eleven patients (15.7%) experienced hepatotoxicity (troughs 0.5 to >7.5 mg/L). Neurotoxicity occurred in three patients (4.3%) (troughs ≥6.7 mg/L) and all patients had symptom resolution upon dose reduction. Treatment failure of invasive fungal infection appeared less in patients with therapeutic troughs compared with sub-therapeutic troughs (11.4% vs. 14.2%). Two patients experienced treatment failure despite supra-therapeutic voriconazole troughs. CONCLUSIONS: TDM should be implemented due to significant unpredictability in dose exposure. TDM can reduce unnecessary switches to alternatives due to intolerability and rule in the possibility of resistant organisms in the event of treatment failure despite therapeutic troughs, alerting clinicians to switch to alternatives promptly.

Procedural and Anatomical Determinants of Multielectrode Renal Denervation Efficacy.

Radiofrequency renal denervation is under investigation for treatment of hypertension with variable success. We developed preclinical models to examine the dependence of ablation biomarkers on renal denervation treatment parameters and anatomic variables. One hundred twenty-nine porcine renal arteries were denervated with an irrigated radiofrequency catheter with multiple helically arrayed electrodes. Nerve effects and ablation geometries at 7 days were characterized histomorphometrically and correlated with associated renal norepinephrine levels. Norepinephrine exhibited a threshold dependence on the percentage of affected nerves across the range of treatment durations (30-60 s) and power set points (6-20 W). For 15 W/30 s treatments, norepinephrine reduction and percentage of affected nerves tracked with number of electrode treatments, confirming additive effects of helically staggered ablations. Threshold effects were only attained when ≥4 electrodes were powered. Histomorphometry and computational modeling both illustrated that radiofrequency treatments directed at large neighboring veins resulted in subaverage ablation areas and, therefore, contributed suboptimally to efficacy. Account for measured nerve distribution patterns and the annular geometry of the artery revealed that, regardless of treatment variables, total ablation area and circumferential coverage were the prime determinants of renal denervation efficacy, with increased efficacy at smaller diameters.

Chromosomal microarray analysis is superior in identifying cryptic aberrations in patients with acute lymphoblastic leukemia at diagnosis/relapse as a single assay.

INTRODUCTION: Conventional cytogenetics (CC) is important in diagnosis, therapy, monitoring of post-transplant bone marrow, and prognosis assessment of acute lymphoblastic leukemia (ALL). However, due to the nature of ALL, CC often encounters difficulties of complex karyotype, poor chromosome morphology, low mitotic index, or normal cells dividing only. In contrast, chromosomal microarray analysis (CMA) showed a specificity >99% and a sensitivity of 100% in chronic lymphocytic leukemia (CLL) patients. Here, we report our experience with CMA on adult ALL patients. METHODS: Thirty-three bone marrow/blood samples from ALL patients (aged 18-79 years, median 44) at diagnosis/relapse, analyzed by CC and/or fluorescence in situ hybridization (FISH), were recruited. Chromosomal microarray analysis results were compared with CC. Fluorescence in situ hybridization analysis, if available, was applied when there was a discrepancy. RESULTS: Copy-neutral loss-of-heterozygosity (CN-LOH) was found in 8 cases (24.2%). Only CN-LOH at 9p was recurrent (3 cases, 9.1%). Copy number alterations (CNAs) were detected in 6 of 9 cases (66.7%) with normal karyotypes, in 3 of 5 cases (60.0%) with sole "balanced" translocations, and in 18 of 19 cases (94.7%) with complex karyotypes. Common CNAs involved CDKN2A/2B (30.3%), IKZF1 (27.3%), PAX5 (9.1%), RB1 (9.1%), BTG1 (6.7%), and ETV6 (6.7%), which regulate cell cycle, B lymphopoiesis, or act as tumor suppressors in ALL. Copy number alteration detection rate by CMA was 81.8% (27 of 33 cases) as compared to 57.6% (19 of 33 cases) by CC. CONCLUSION: Incorporation of CMA as a routine clinical test at the time of diagnosis/relapse, in conjunction with CC and/or FISH, is highly recommended.

Use of antibiotics in a haematology ward--an audit.

INTRODUCTION: Rising rates of antibiotic resistance prompted a review of antibiotic use policies hospitalwide. The Department of Haematology established a new set of consensus guidelines in 2002 for antibiotic use in febrile neutropenia. The aim of our study was to audit adherence to the guidelines established for febrile neutropenia in patients treated for haematologic malignancies. MATERIALS AND METHODS: An antibiotic escalation pathway was developed by haematologists and infectious disease physicians. Adherence to the guidelines was audited. Patients with acute myeloid leukaemia (AML) or acute lymphocytic leukaemia (ALL) who had febrile neutropenia after chemotherapy were reviewed. The audit was performed by a retrospective review of casenotes. RESULTS: Forty patients with 100 episodes of febrile neutropenia were surveyed. Thirty-two had AML, 7 had ALL and 1 had undifferentiated leukaemia. In 76% of episodes, fever developed within the first 14 days of neutropenia. In 31 episodes, cefepime was started as the first-line agent; hence, compliance with the first-line agent was 31%. Fever defervesced in 13 episodes. The most common reason for switching antibiotics was persistent fever. There were clinical indications for non-compliance with the use of the first-line agent in all cases. There were 3 deaths - none related to non-compliance with or strict adherence to the guidelines. Four patients had proven fungal infections. CONCLUSIONS: Given the complex nature of the cases, compliance was reasonable, as there were valid reasons in all cases where the guidelines were not adhered to. Based on our findings, the guidelines could be simplified.

A patient with extramedullary acute myeloid leukaemia involving the brachial plexus: Case report and review of the literature.

Extramedullary leukaemic involvement is not uncommon as part of the presentation in acute myeloid leukaemia (AML). However, infiltrative peripheral neuropathy due to AML was rarely reported. We report a case of extramedullary leukaemic infiltration of the brachial plexus in a patient with relapsed AML.