A pilot clinical trial of oral tetrahydrouridine/decitabine for noncytotoxic epigenetic therapy of chemoresistant lymphoid malignancies

One mechanism by which lymphoid malignancies resist standard apoptosis-intending (cytotoxic) treatments is genetic attenuation of the p53/p16-CDKN2A apoptosis axis. Depletion of the epigenetic protein DNA methyltransferase 1 (DNMT1) using the deoxycytidine analog decitabine is a validated approach to cytoreduce malignancy independent of p53/p16. In vivo decitabine activity, however, is restricted by rapid catabolism by cytidine deaminase (CDA). We, therefore, combined decitabine with the CDA-inhibitor tetrahydrouridine and conducted a pilot clinical trial in patients with relapsed lymphoid malignancies: the doses of tetrahydrouridine/decitabine used (∼10/0.2 mg/kg orally (PO) 2×/week) were selected for the molecular pharmacodynamic objective of non-cytotoxic, S-phase dependent, DNMT1-depletion, guided by previous Phase 1 studies. Patients with relapsed/refractory B- or T-cell malignancies (n = 7) were treated for up to 18 weeks. Neutropenia without concurrent thrombocytopenia is an expected toxicity of DNMT1-depletion and occurred in all patients (Grade 3/4). Subjective and objective clinical improvements occurred in 4 of 7 patients, but these responses were lost upon treatment interruptions and reductions to manage neutropenia. We thus performed parallel experiments in a preclinical in vivo model of lymphoma to identify regimen refinements that might sustain DNMT1-targeting in malignant cells but limit neutropenia. We found that timed-alternation of decitabine with the related molecule 5-azacytidine, and combination with inhibitors of CDA and de novo pyrimidine synthesis could leverage feedback responses of pyrimidine metabolism to substantially increase lymphoma cytoreduction but with less neutropenia. In sum, regimen innovations beyond incorporation of a CDA-inhibitor are needed to sustain decitabine DNMT1-targeting and efficacy against chemo-resistant lymphoid malignancy. Such potential solutions were explored in preclinical in vivo studies.     

Effects of tetrahydrouridine on pharmacokinetics and pharmacodynamics of oral decitabine

The deoxycytidine analog decitabine (DAC) can deplete DNA methyl-transferase 1 (DNMT1) and thereby modify cellular epigenetics, gene expression, and differentiation. However, a barrier to efficacious and accessible DNMT1-targeted therapy is cytidine deaminase, an enzyme highly expressed in the intestine and liver that rapidly metabolizes DAC into inactive uridine counterparts, severely limiting exposure time and oral bioavailability. In the present study, the effects of tetrahydrouridine (THU), a competitive inhibitor of cytidine deaminase, on the pharmacokinetics and pharmacodynamics of oral DAC were evaluated in mice and nonhuman primates. Oral administration of THU before oral DAC extended DAC absorption time and widened the concentration-time profile, increasing the exposure time for S-phase-specific depletion of DNMT1 without the high peak DAC levels that can cause DNA damage and cytotoxicity. THU also decreased interindividual variability in pharmacokinetics seen with DAC alone. One potential clinical application of DNMT1-targeted therapy is to increase fetal hemoglobin and treat hemoglobinopathy. Oral THU-DAC at a dose that would produce peak DAC concentrations of less than 0.2μM administered 2×/wk for 8 weeks to nonhuman primates was not myelotoxic, hypomethylated DNA in the γ-globin gene promoter, and produced large cumulative increases in fetal hemoglobin. Combining oral THU with oral DAC changes DAC pharmacology in a manner that may facilitate accessible noncytotoxic DNMT1-targeted therapy.     

A pilot study of subcutaneous decitabine in β-thalassemia intermedia

Ineffective erythropoiesis, the hallmark of β-thalassemia, is a result of α/non-α globin chain imbalance. One strategy to redress globin-chain imbalance is to induce γ-globin gene (HBG) expression. Repression of HBG in adult erythroid cells involves DNA methylation and other epigenetic changes. Therefore, the cytosine analog decitabine, which can deplete DNA methyltransferase 1 (DNMT1), can potentially activate HBG. In 5 patients with β-thalassemia intermedia, a dose and schedule of decitabine intended to deplete DNMT1 without causing significant cytotoxicity (0.2 mg/kg subcutaneous 2 times per week for 12 weeks) increased total hemoglobin from 7.88 ± 0.88 g/dL to 9.04 ± 0.77 g/dL (P = .004) and absolute fetal hemoglobin from 3.64 ± 1.13 g/dL to 4.29 ± 1.13 g/dL (P = .003). Significant favorable changes also occurred in indices of hemolysis and red blood cell densitometry. Consistent with a noncytotoxic, differentiation altering mechanism of action, the major side effect was an asymptomatic increase in platelet counts without erythrocyte micronucleus or VDJ recombination assay evidence of genotoxicity. This study was registered at www.clinicaltrials.gov as #NCT00661726.     

In vitro and in vivo induction of fetal hemoglobin with a reversible and selective DNMT1 inhibitor

Pharmacological induction of fetal hemoglobin (HbF) expression is an effective therapeutic strategy for the management of β-hemoglobinopathies such as sickle cell disease. DNA methyltransferase (DNMT) inhibitors 5-azacytidine (5-aza) and 5-aza-2′-deoxycytidine (decitabine) have been shown to induce HbF expression in both preclinical models and clinical studies, but are not currently approved for the management of hemoglobinopathies. We report here the discovery of a novel class of orally bioavailable DNMT1-selective inhibitors as exemplified by GSK3482364. This molecule potently inhibits the methyltransferase activity of DNMT1, but not DNMT family members DNMT3A or DNMT3B. In contrast with cytidine analog DNMT inhibitors, the DNMT1 inhibitory mechanism of GSK3482364 does not require DNA incorporation and is reversible. In cultured human erythroid progenitor cells, GSK3482364 decreased overall DNA methylation resulting in derepression of the -globin genes HBG1 and HBG2 and increased HbF expression. In a transgenic mouse model of sickle cell disease, orally administered GSK3482364 caused significant increases in both HbF levels and in the percentage HbF-expressing erythrocytes, with good overall tolerability. We conclude that in these preclinical models, selective, reversible inhibition of DNMT1 is sufficient for the induction of HbF, and is well-tolerated. We anticipate that GSK3482364 will be a useful tool molecule for the further study of selective DNMT1 inhibition both in vitro and in vivo.     

Comparative clinical effectiveness of azacitidine versus decitabine in older patients with myelodysplastic syndromes

The hypomethylating agents (HMAs) azacitidine and decitabine are both approved for treatment of myelodysplastic syndromes (MDS) in the USA. In Europe, decitabine is not approved due to lack of survival advantage in randomized trials. The two drugs have not been compared in clinical trials. We identified patients diagnosed with MDS between 2004 and 2011 from the Surveillance, Epidemiology, and End Results (SEER)‐Medicare linked database in the USA who received ≥ 10 doses of either HMA. We estimated survival from HMA initiation with Kaplan–Meier methods and used multivariate Cox proportional hazards models to adjust for covariates. Analyses controlled for histological subtype and we conducted a subset analysis limited to patients with refractory anaemia with excess blasts (RAEB). In 2025 HMA‐treated patients, median survival was 15 months with no difference in survival based on the HMA received in adjusted analysis (decitabine versus azacitidine, hazard ratio = 1·06, 95% confidence interval: 0·94–1·19, P = 0·37). For RAEB patients (n = 523), median survival was 12 months, with no significant difference based on HMA received. No significant survival difference was found between azacitidine and decitabine in patients with MDS, including RAEB. Importantly, population‐based survival of azacitidine‐treated RAEB patients was substantially shorter than in the AZA‐001 clinical trial (11 versus 24·5 months).     

Elevated fetal haemoglobin is a predictor of better outcome in MDS/AML patients receiving 5‐aza‐2′‐deoxycytidine (Decitabine)

Although azanucleoside DNA‐hypomethylating agents (HMAs) are routinely used for the treatment of myelodysplastic syndrome/acute myeloid leukaemia (MDS/AML), very few outcome predictors have been established. Expression of the β‐like globin gene locus is tightly regulated by DNA methylation, is HMA‐sensitive in vitro, and fetal haemoglobin (HbF) expression is under study as a potential biomarker for response of MDS patients to azacitidine. We determined HbF expression in 16 MDS and 36 AML patients receiving decitabine (DAC). Pre‐treatment HbF was already elevated (>1·0% of total haemoglobin) in 7/16 and 12/36 patients, and HbF was induced by DAC in 81%/54% of MDS/AML patients, respectively. Elevated pre‐treatment HbF was associated with longer median overall survival (OS): 26·6 vs. 8·6 months for MDS (hazard ratio [HR] 8·56, 95% confidence interval [CI] 1·74–42·49, P = 0·008, with similarly longer progression‐free and AML‐free survival), and 10·0 vs. 2·9 months OS for AML (HR 3·01, 95% CI 1·26–7·22, P = 0·014). In a multivariate analysis, the prognostic value of HbF was retained. Time‐dependent Cox models revealed that the prognostic value of treatment‐induced HbF induction was inferior to that of pre‐treatment HbF. In conclusion, we provide first evidence for in vivo HbF induction by DAC in MDS/AML, and demonstrate prognostic value of elevated pre‐treatment HbF, warranting prospective, randomized studies.     

Phase II study of tosedostat with cytarabine or decitabine in newly diagnosed older patients with acute myeloid leukaemia or high‐risk MDS

Tosedostat, an oral aminopeptidase inhibitor, has synergy with cytarabine and hypomethylating agents. We performed a Phase II trial to determine rates of complete remission (CR) and survival using tosedostat with cytarabine or decitabine in older patients with untreated acute myeloid leukaemia (AML) or high‐risk myelodysplastic syndrome (MDS). Thirty‐four patients ≥60 years old (median age 70 years; range, 60–83) were randomized to receive tosedostat (120 mg on days 1–21 or 180 mg continuously) with 5 d of either cytarabine (1 g/m²/d) or decitabine (20 mg/m²/d) every 35 d. Twenty‐nine patients (85%) had AML, including 15 (44%) with secondary AML/MDS, and 5 (15%) had MDS‐refractory anaemia with excess blasts type 2. The CR/CR with incomplete count recovery (CRi) rate was 53% [9 in each arm; 14 CR (41%) and 4 CRi (12%)], attained in 6 of 14 patients with adverse cytogenetics and 4 of 7 with FLT3‐internal tandem duplication mutations. Median follow‐up was 11·2 months (range, 0·5–22·3), and median survival was 11·5 months (95% confidence interval, 5·2–16·7). Twenty‐three patients (67·6%) were treated as outpatients and 10 of these patients required hospitalization for febrile neutropenia. No Grade 3–4 non‐haematological toxicities required withdrawal from study. Tosedostat with cytarabine or decitabine is tolerated in older patients with untreated AML/MDS, results in a CR/CRi rate of >50%, and warrants further study in larger trials.     

A Meta‐analysis of intradermal versus intramuscular influenza vaccines: Immunogenicity and Adverse Events

Please cite this paper as: Marra et al. (2012) A Meta‐analysis of intradermal versus intramuscular influenza vaccines: immunogenicity and adverse events. Influenza and Other Respiratory Viruses 7(4), 584–603. Objective To determine immunogenicity and safety of intradermal (ID) influenza vaccines compared with intramuscular (IM) administration and effect of dose and age. Design Meta‐anlysis. Setting Systematic review and meta‐analysis of randomized controlled trials on influenza vaccines. Sample Randomized, controlled trials comparing ID seasonal split‐virus influenza vaccines with 15 μg IM control in subjects 18 years of age or older and assessed antibody response at 21–28 days post‐vaccination were considered for inclusion. Results A total of 13 trials were included. The pooled immunogenicity outcomes did not differ significantly between the IM and ID vaccine groups for the H1N1 (ratio of GMTR: 0·92, 95% confidence interval 0·77–1·09; seroconversion: 0·94, 0·86–1·02; seroprotection: 0·97, 0·94–1·00) and B strains (GMTR: 0·93, 0·80–1·08; seroconversion: 0·91, 0·80–1·04; seroprotection: 0·97, 0·91–1·03). For the H3N2 strain, there was no significant difference in GMTR (0·97, 0·80–1·18); however, there was a lower pooled seroconversion (0·89, 0·80–0·99) and seroprotection rate (0·98, 0·96–0·99) for ID recipients. There was a statistically significant association between increasing doses of the ID vaccination with increasing immunogenicity response (P = 0·01). There were no differences in adverse event rates within 3 days post‐vaccination for ID versus IM. But for adverse events occurring 7 days post‐vaccination, ID vaccination was associated with a greater incidence of local events but not systemic events. Conclusions There was no significant difference in immunologic response when comparing ID with IM administration of the influenza vaccination in the overall population, but higher doses of ID vaccine in the older adult population produced a better response.     

Plasma aldosterone response to the low-dose adrenocorticotrophin (ACTH 1-24) stimulation test

Background Endocrine tests for adrenal insufficiency use pharmacological doses of stimulant such as ACTH. More physiological tests have often used high‐dose protocols for sampling frequency. Aims To evaluate the response of plasma aldosterone concentration to low doses (125, 250 and 500 ng/m² body surface area) of synthetic ACTH. Design A randomised trial in six normal adult males aged 18–27 years. Materials and methods Aldosterone concentration was measured by radioimmunoassay in serum from blood samples taken at 10 min intervals for 90 min. Results All three doses produced a significant rise in plasma aldosterone concentration (125 ng/m², P = 0·003; 250 ng/m², P < 0·001; 500 ng/m², P < 0·001) but there was no effect of dose on either the peak or incremental plasma aldosterone concentration. Mean time to peak was similar between the doses and the two higher doses were associated with a longer secretory profile (125 ng/m² 56 (26 SD) mins, 250 ng/m² 74 (19) mins, 500 ng/m² 77 (21) mins; F = 3·39; P = 0·04). Peaks of 100% were detected within 30 min of drug administration and peak response was associated with the prestimulation plasma aldosterone concentration (r = 0·45; P = 0·003). The between‐ and within‐individual coefficients of variation for prestimulation concentrations were 37·0% and 32·8%, and for the peak response were 27·2% and 27·2%, respectively. Conclusions The response of plasma aldosterone concentrations to low‐dose ACTH administration requires a blood sampling protocol of 0, 10, 20 and 30 min to capture concentrations near the peak response. The high‐dose protocol would have missed the response. Over the dose range studied no dose–response was observed so the selection of dose should be based on the dose effective to release steroids in the glucocorticoid pathway if this study is to be used in conjunction with such evaluation.     

Phase 1 study of low-dose prolonged exposure schedules of the hypomethylating agent 5-aza-2'-deoxycytidine (decitabine) in hematopoietic malignancies

Decitabine (5-aza-2'-deoxycytidine) inhibits DNA methylation and has dual effects on neoplastic cells, including the reactivation of silenced genes and differentiation at low doses and cytotoxicity at high doses. We evaluated, in a phase 1 study, low-dose prolonged exposure schedules of decitabine in relapsed/refractory leukemias. Patient cohorts received decitabine at 5, 10, 15, or 20 mg/m2 intravenously over one hour daily, 5 days a week for 2 consecutive weeks, doses 5- to approximately 30-fold lower than the maximum tolerated dose (MTD). There were 2 groups that also received 15 mg/m2 daily for 15 or 20 days. A total of 50 patients were treated (44 with acute myelogenous leukemia [AML]/myelodysplasia [MDS], 5 with chronic myelogenous leukemia [CML], and 1 with acute lymphocytic leukemia [ALL]), and the drug was well tolerated at all dose levels, with myelosuppression being the major side effect. Responses were seen at all dose levels. However, the dose of 15 mg/m2 for 10 days appeared to induce the most responses (11 of 17 or 65%), with fewer responses seen when the dose was escalated or prolonged (2 of 19 or 11%). There was no correlation between P15 methylation at baseline or after therapy and response to decitabine. We conclude that decitabine is effective in myeloid malignancies, and low doses are as or more effective than higher doses.     

Oral decitabine reactivates expression of the methylated gamma-globin gene in Papio anubis

The silencing of tumor suppressor genes associated with increased DNA methylation of the promoter regions is a frequent observation in many forms of cancer. Reactivation of these genes using pharmacological inhibitors of DNA methyltransferase such as 5-aza-2'-deoxycytidine (decitabine) is a worthwhile therapeutic goal. The effectiveness and tolerability of low-dose intravenous and subcutaneous decitabine regimens to demethylate and reactivate expression of the methylated gamma-globin gene in baboons and in patients with sickle cell disease led to successful trials of low-dose regimens of this drug in patients with myelodysplastic syndrome. Since these low-dose regimens are well-tolerated with minimal toxicity, they are suitable for chronic dosing to maintain promoter hypomethylation and expression of target genes. The development of an orally administered therapy using DNA methyltransferase inhibitors would facilitate such chronic approaches to therapy. We tested the ability of decitabine and a new salt derivative, decitabine mesylate, to reactivate the methylated gamma-globin gene in baboons when administered orally. Our results demonstrate that oral administration of these drugs at doses 17-34 times optimal subcutaneous doses of decitabine reactivates fetal hemoglobin, demethylates the epsilon- and gamma-globin gene promoters, and increases histone acetylation of these promoters in baboons (Papio anubis).     

Impact of vitamin D-related serum PTH reference values on the diagnosis of mild primary hyperparathyroidism, using bivariate calcium/PTH reference regions

Background, objective An international consensus conference underlined the importance of defining upper parathyroid hormone (PTH) reference values based on 25‐OH‐vitamin D [25(OH)D] to diagnose mild primary hyperparathyroidism. We determined the importance of this factor in a Belgian population. Design, patients, methods Intact PTH and 25(OH)D were measured in 261 healthy controls (18–65 years, winter/summer). They were classified as 25(OH)D replete (50–153 nmol/l; n = 129) or deplete (8–50 nmol/l; n = 132). PTH was determined in 49 patients with surgically proven primary hyperparathyroidism. PTH thresholds for 95% specificities and corresponding sensitivities were computed from both 25(OH)D replete and deplete receiver operating characteristic (ROC) curves. The 95% bivariate reference ellipses, relating PTH to calcium for 25(OH)D replete and deplete controls, were compared to the PTH/calcium pairs of patients with primary hyperparathyroidism. Results Parathyroid hormone correlated with 25(OH)D (r = ?0·3232; P < 0·0001). PTH normative values were 20% lower in 25(OH)D replete than deplete controls (P < 0·0001). PTH thresholds, providing 95% specificities for primary hyperparathyroidism diagnosis, were 7·6 pmol/l and 5·8 pmol/l, using ROC curves derived from 25(OH)D deplete or replete controls, respectively. Corresponding sensitivities were of 56%vs 88%, respectively (P < 0·05). The 95% PTH/calcium bivariate reference ellipses for?deplete and replete 25(OH)D controls differed, but the PTH/calcium pairs of patients with primary hyperparathyroidism did not overlap these ellipses. Conclusion For a given specificity, primary hyperparathyroidism diagnostic parathyroid hormone thresholds were lower and sensitivities higher using ROC curves, derived from 25(OH)D replete vs deplete controls. The 25(OH)D status does not affect the efficiency of primary hyperparathyroidism diagnosis, using bivariate PTH/calcium reference density ellipses.     

Harmine,a Novel DNA Methyltransferase 1 Inhibitor in the Leukemia Cell Line

DNA methylation followed by tumor suppressor gene repression plays a critical role in the leukemia development. So, DNA methyl transferase inhibitors have great importance in treatment of theses malignancies. Harmine, A beta carboline alkaloid derivative of Peganum harmala, had shown anti- proliferative effects on leukemic cell line. This study aimed to evaluate the effect of Harmine on DNMT1 (DNA methyl transferase 1) expression in a leukemic cell line. Cell proliferation and cell cycle analysis were studied in NB4 cell line after treatment with Harmine for 72 h. DNMT1 expression in treated cells was analyzed by real time PCR. Tumor suppressor gene hypometylation and reactivation was evaluated via MSP analysis and also real time PCR. Harmine reduced cell proliferation in NB4 cell line in a time and dose-dependent manner. 102 µg/ml of Harmine was increased amount of cells in G1 Phase of cell cycle (p < 0.05). Anti proliferative doses of Harmine, has suppressed DNMT1 gene in NB4 cell line. Down-regulated DNMT1 induced p15 tumor suppressor promoter hypomethylation and reactivation. Our data indicate that Harmine can be considered as a potential treatment for AML (Acute Myeloid Leukemia), and future studies are required to test the clinical efficacy of Harmine—whether used as a single agent or as an adjuvant—for AML treatment.     

Comparative analysis between azacitidine and decitabine for the treatment of myelodysplastic syndromes

The present study aimed to directly compare the efficacy and safety of azacitidine and decitabine in patients with myelodysplastic syndromes (MDS). We compared the overall response rate (ORR) (complete responses, partial responses, marrow complete responses, and haematological improvements), overall survival (OS), event‐free survival (EFS), time to leukaemic transformation, and adverse outcomes between azacitidine and decitabine. To minimize the effects of treatment selection bias in this observational study, adjustments were made using the propensity‐score matching method. Among 300 patients, 203 were treated with azacitidine and 97 with decitabine. Propensity‐score matching yielded 97 patient pairs. In the propensity‐matched cohort, there were no significant differences between the azacitidine and decitabine groups regarding ORR (44% vs. 52%), OS (26 vs. 22·9 months), EFS (7·7 vs. 7·0 months), and rate of leukaemic transformation (16% vs. 22% at 1 year). In patients ≥65 years of age, survival was significantly better in the azacitidine group (P = 0·017). Patients who received decitabine experienced more frequent episodes of grade 3 or 4 cytopenia and infectious episodes. We found that azacitidine and decitabine showed comparable efficacy. Among patients ≥65 years of age, survival was significantly better in the azacitidine group (ClinicalTrials.gov Identifier: NCT01409070).     

A CARBOXY-TERMINAL SPECIFIC ASSAY FOR HUMAN PARATHYROID HORMONE

An homologous carboxy-terminal specific immunoradiometric assay for human parathyroid hormone (PTH) has been developed which uses an antiserum raised in a goat against intact human PTH. This assay has been applied to the measurement of circulating PTH in man; the reference preparation was a 53–84 fragment prepared from native human parathyroid hormone. PTH was detectable in all the normal subjects studied (range 0·1–0·79 ng/ml) and elevated concentrations were found in a group of patients with primary hyperparathyroidism (range 0·82–25·5 ng/ml). Carboxy-terminal hormone concentrations were particularly raised in uraemic subjects (range 1–02–52–5 ng/ml) but some patients with vitamin D deficient osteomalacia had concentrations within the normal range (0·5–2·4 ng/ml). All patients with pseudohypoparathyroidism had elevated concentrations (range 1·0–2·8 ng/ml) and in patients with idiopathic or surgical hypoparathyroidism the concentration ranged from undetectable to 0·4 ng/ml. These results have been compared with those obtained using an amino-terminal specific assay. In normal subjects carboxy-terminal immuno-reactive hormone concentrations were, on average, three times higher than those measured in the amino-terminal assay (C:N ratio 3·28). In patients with secondary hyperparathyroidism this ratio was dependent on renal function and the plasma calcium concentration. The disappearance of endogenous carboxy-and amino-terminal PTH from the circulation of a patient with primary hyperparathyroidism was studied. Amino-terminal reactive PTH disappeared much more rapidly (t½= 1·5 min) than carboxy-terminal hormone (t½= 2·75 h). It is hoped that the carboxy-terminal specific assay described here will be of value in the further study of the secretion and metabolism of PTH.     

Insulin-like growth factor 1 and risk of later depression in older people: prospective evidence from the English Longitudinal Study of Ageing

BackgroundDepressive disorders are a leading cause of mortality and morbidity. Whereas the role of psychosocial and behavioural predictors has been well examined, little is known about the biological origins of depression. In mice in which insulin-like growth factor 1 (IGF-1) was knocked out with viral vectors, there was evidence of increased signs of depression. In other studies, IGF-1 infusions in rodents had antidepressant properties. We tested the association of depression with IGF-1 in a large population-based study.MethodsIGF-1 concentrations were measured from serum taken at a nurse visit in 2008 from adults participating in the English Longitudinal Study of Ageing. Symptoms of depression were assessed in 2008 and 2012 with the eight item Center for Epidemiological Studies Depression Scale (CES-D) and diagnosis was based on self-report. Cross-sectional analyses were carried out on measures collected at baseline, in 2008. Outcome measures in longitudinal analyses were new reports of depression symptoms and physician-diagnosed depression in 2012. We used logistic regression to test both the cross-sectional and longitudinal associations, adjusted in a stepwise manner for age, anthropometric measures, comorbidities, psychosocial factors, and health behaviours.FindingsSamples from 6017 adults were included (mean age 65·7 years [SD 9·3], range 50–99; mean IGF-1 15·9 nmol/L [SD 5·7]). Mean CES-D score at baseline was 1·2 (SD 1·8). In cross-sectional and longitudinal analyses, the association between IGF-1 and symptoms of depression followed a U-shaped pattern—ie, lower and higher concentrations of IGF-1 were associated with an increased risk of depression, whereas the lowest risk was seen around median concentrations. In women, with the lowest quintile of IGF-1 as the referent, the age-adjusted odds ratios for increasing quintiles of IGF-1 in longitudinal analysis were 0·88 (95% CI 0·60–1·29), 0·77 (0·51–1·16), 0·84 (0·53–1·35), and 0·95 (0·60–1·50) (p for curve=0·027). Corresponding results in men were 0·51 (0·28–0·91), 0·50 (0·27–0·92), 0·63 (0·35–1·15), and 0·63 (0·35–1·13) (p for curve=0·002). These associations were partly attributable to comorbidities, adverse socioeconomic circumstances, and psychosocial and behavioural factors. Similar results were observed for the association between IGF-1 and physician-diagnosed depression.InterpretationIn the present study, which is the first to our knowledge to use validated measures of depression, there was some evidence that low and high concentrations of IGF-1 were associated with a slightly elevated risk of diagnosed depression and symptoms of depression. Further studies are needed to examine whether the observed association is causal and whether normalising IGF-1 concentrations with drugs or IGF-1 infusions could be useful and safe in the treatment of depression in older people.FundingThis work was supported by an Economic and Social Research Council–Medical Research Council studentship (awarded to SC).     

Bone mineral density and bone turnover in Romanian children and young adults with classical 21-hydroxylase deficiency are influenced by glucocorticoid replacement therapy

Objective It remains controversial if glucocorticoid replacement therapy impairs bone mineral density (BMD) in young patients with 21‐hydroxylase deficiency. We aimed to analyze the impact of treatment variables, phenotype and genotype on BMD and bone metabolism in these patients. Design Cross‐sectional study. Measurements Twenty‐eight Caucasian patients with classical 21‐hydroxylase deficiency (5–39 years). Clinical parameters, hormonal status, osteocalcin (OC), C‐terminal telopeptide of type I collagen (CTX), genotype and lumbar BMD (Z‐scores) were assessed. Cumulative and mean hydrocortisone equivalent doses were calculated for the entire treatment period. Results Patients with severely reduced BMD Z‐scores (≤–2·5 SD) had significantly higher mean/cumulative glucocorticoid doses compared to patients with moderately reduced (P = 0·003/P = 0·026) and normal Z‐scores (> –1 SD) (P = 0·005/P = 0·011). Mean hydrocortisone equivalent doses > 20 mg/m²/day led to significantly lower lumbar BMD Z‐scores (–2·16 ± 1·4 SD) vs. doses ≤ 20 mg/m²/day (–0·59 ± 1·25 SD) (P = 0·008). BMD correlated negatively with mean/cumulative glucocorticoid doses and treatment duration. OC (86·45 ± 37·45 ng/ml) and CTX (1·45 ± 0·43 ng/ml) were significantly increased compared to an age‐ and sex‐matched control group in patients with active growth; only CTX was slightly increased in patients who completed growth. Conclusions High cumulative and mean glucocorticoid doses negatively impact on BMD in children and young adults with classical 21‐hydroxylase deficiency. Substitution therapy should be adapted particularly at this life period to prevent bone loss.     

Impact of MLL5 expression on decitabine efficacy and DNA methylation in acute myeloid leukemia

Hypomethylating agents are widely used in patients with myelodysplastic syndromes and unfit patients with acute myeloid leukemia. However, it is not well understood why only some patients respond to hypomethylating agents. We found previously that the effect of decitabine on hematopoietic stem cell viability differed between Mll5 wild-type and null cells. We, therefore, investigated the role of MLL5 expression levels on outcome of acute myeloid leukemia patients who were treated with decitabine. MLL5 above the median expression level predicted longer overall survival independent of DNMT3A mutation status in bivariate analysis (median overall survival for high vs. low MLL5 expression 292 vs. 167 days; P=0.026). In patients who received three or more courses decitabine, high MLL5 expression and wild-type DNMT3A independently predicted improved overall survival (median overall survival for high vs. low MLL5 expression 468 vs. 243 days; P=0.012). In transformed murine cells, loss of Mll5 was associated with resistance to low-dose decitabine, less global DNA methylation in promoter regions, and reduced DNA demethylation upon decitabine treatment. Together, these data support our clinical observation of improved outcome in decitabine-treated patients who express MLL5 at high levels, and suggest a mechanistic role of MLL5 in the regulation of DNA methylation.     

Low‐dose pembrolizumab induced remission in patients with refractory classical Hodgkin lymphoma

We aimed to compare hypofibrinogenaemia prevalence in major bleeding patients across all clinical contexts, fibrinogen supplementation practice, and explore the relationship between fibrinogen concentrations and mortality. This cohort study included all adult patients from 20 hospitals across Australia and New Zealand who received massive transfusion between April 2011 and October 2015. Of 3566 patients, 2829 (79%) had fibrinogen concentration recorded, with a median first and lowest concentration of 2·0 g/l (interquartile range [IQR] 1·5–2·7) and 1·8 g/l (IQR 1·3–2·4), respectively. Liver transplant (1·7 g/l, IQR 1·2–2·1), trauma (1·8, IQR 1·3–2·5) and vascular surgery (1·9 g/l, IQR 1·4–2·5) had lower concentrations. Total median fibrinogen dose administered from all products was 7·3 g (IQR 3·3–13·0). Overall, 1732 (61%) received cryoprecipitate and 9 (<1%) fibrinogen concentrate. Time to cryoprecipitate issue in those with initial fibrinogen concentration <1 g/l was 2·5 h (IQR 1·2–4·3 h). After adjustment, initial fibrinogen concentration had a U‐shaped association with in‐hospital mortality [adjusted odds ratios: fibrinogen <1 g/l, 2·31 (95% confidence interval (CI) 1·48–3·60); 1–1·9 g/l, 1·29 (95% CI 0·99–1·67) and >4 g/l, 2·03 (95% CI 1·35–3·04), 2–4 g/l reference category]. The findings indicate areas for practice improvement including timely administration of cryoprecipitate, which is the most common source of concentrated fibrinogen in Australia and New Zealand.