Systematic chemical and molecular profiling of MLL-rearranged infant acute lymphoblastic leukemia reveals efficacy of romidepsin.

To address the poor prognosis of mixed lineage leukemia (MLL)-rearranged infant acute lymphoblastic leukemia (iALL), we generated a panel of cell lines from primary patient samples and investigated cytotoxic responses to contemporary and novel Food and Drug Administration-approved chemotherapeutics. To characterize representation of primary disease within cell lines, molecular features were compared using RNA-sequencing and cytogenetics. High-throughput screening revealed variable efficacy of currently used drugs, however identified consistent efficacy of three novel drug classes: proteasome inhibitors, histone deacetylase inhibitors and cyclin-dependent kinase inhibitors. Gene expression of drug targets was highly reproducible comparing iALL cell lines to matched primary specimens. Histone deacetylase inhibitors, including romidepsin (ROM), enhanced the activity of a key component of iALL therapy, cytarabine (ARAC) in vitro and combined administration of ROM and ARAC to xenografted mice further reduced leukemia burden. Molecular studies showed that ROM reduces expression of cytidine deaminase, an enzyme involved in ARAC deactivation, and enhances the DNA damage-response to ARAC. In conclusion, we present a valuable resource for drug discovery, including the first systematic analysis of transcriptome reproducibility in vitro, and have identified ROM as a promising therapeutic for MLL-rearranged iALL.

High expression of connective tissue growth factor accelerates dissemination of leukaemia.

To improve treatment of acute lymphoblastic leukaemia (ALL), a better understanding of disease development is needed to tailor new therapies. Connective tissue growth factor (CTGF/CCN2) is highly expressed in leukaemia cells from the majority of paediatric patients with B-lineage ALL (pre-B ALL). CTGF is a matricellular protein and plays a role in aggressive cancers. Here we have genetically engineered leukaemia cells to modulate CTGF expression levels. Elevated CTGF levels accelerated disease dissemination and reduced survival in NOD/SCID mice. In vitro studies showed that CTGF protein induces stromal cell proliferation, promotes adhesion of leukaemia cells to stromal cells and leads to overexpression of genes associated with cell cycle and synthesis of extracellular matrix (ECM). Corresponding data from our leukaemia xenograft models demonstrated that CTGF leads to increased proliferation of non-leukaemia cells and deposition of ECM in the bone marrow. We document for the first time a functional role of CTGF in altering disease progression in a lymphoid malignancy. The findings provide support for targeting the bone marrow microenvironment in aggressive forms of leukaemia.

A pre-clinical model of resistance to induction therapy in pediatric acute lymphoblastic leukemia.

Relapse and acquired drug resistance in T-cell acute lymphoblastic leukemia (T-ALL) remains a significant clinical problem. This study was designed to establish a preclinical model of resistance to induction therapy in childhood T-ALL to examine the emergence of drug resistance and identify novel therapies. Patient-derived T-ALL xenografts in immune-deficient (non-obese diabetic/severe combined immunodeficient) mice were exposed to a four-drug combination of vincristine, dexamethasone (DEX), L-asparaginase and daunorubicin (VXLD). 'Relapse' xenografts were characterized by responses to drugs, changes in gene expression profiles and Connectivity Map (CMap) prediction of strategies to reverse drug resistance. Two of four xenografts developed ex vivo and in vivo drug resistance. Both resistant lines showed altered lipid and cholesterol metabolism, yet they had a distinct drug resistance pattern. CMap analyses reinforced these features, identifying the cholesterol pathway inhibitor simvastatin (SVT) as a potential therapy to overcome resistance. Combined ex vivo with DEX, SVT was significantly synergistic, yet when administered in vivo with VXLD it did not delay leukemia progression. Synergy of SVT with established chemotherapy may depend on higher drug doses than are tolerable in this model. Taken together, we have developed a clinically relevant in vivo model of T-ALL suitable to examine the emergence of drug resistance and to identify novel therapies.

The evolution of clinical trials for infant acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) in infants has a significantly inferior outcome in comparison with older children. Despite initial improvements in survival of infants with ALL since establishment of the first pediatric cooperative group ALL trials, the poor outcome has plateaued in recent years. Historically, infants were treated on risk-adapted childhood ALL protocols. These studies were pivotal in identifying the need for infant-specific protocols, delineating prognostic categories and the requirement for a more unified approach between study groups to overcome limitations in accrual because of low incidence. This subsequently led to the development of collaborative infant-specific studies. Landmark outcomes have included the elimination of cranial radiotherapy following the discovery of intrathecal and high-dose systemic therapy as a superior and effective treatment strategy for central nervous system disease prophylaxis, with improved neurodevelopmental outcome. Universal prospective identification of independent adverse prognostic factors, including presence of a mixed lineage leukemia rearrangement and young age, has established the basis for risk stratification within current trials. The infant-specific trials have defined limits to which conventional chemotherapeutic agents can be intensified to optimize the balance between treatment efficacy and toxicity. Despite variations in therapeutic intensity, there has been no recent improvement in survival due to the equilibrium between relapse and toxicity. Ultimately, to improve the outcome for infants with ALL, key areas still to be addressed include identification and adaptation of novel prognostic markers and innovative therapies, establishing the role of hematopoietic stem cell transplantation in first complete remission, treatment strategies for relapsed/refractory disease and monitoring and timely intervention of late effects in survivors. This would be best achieved through a single unified international trial.

Comparative drug screening in NUT midline carcinoma.

BACKGROUND: The NUT midline carcinoma (NMC) is a rare but fatal cancer for which systematic testing of therapy options has never been performed. METHODS: On the basis of disease biology, we compared the efficacy of the CDK9 inhibitor flavopiridol (FP) with a panel of anticancer agents in NMC cell lines and mouse xenografts. RESULTS: In vitro anthracyclines, topoisomerase inhibitors, and microtubule poisons were among the most cytotoxic drug classes for NMC cells, while efficacy of the bromodomain inhibitor JQ1 varied considerably between lines carrying different BRD4 (bromodomain-containing protein 4)-NUT (nuclear protein in testis) translocations. Efficacy of FP was comparable to vincristine and doxorubicin, drugs that have been previously used in NMC patients. All three compounds showed significantly better activity than etoposide and vorinostat, agents that have also been used in NMC patients. Statins and antimetabolites demonstrated intermediate single-agent efficacy. In vivo, vincristine significantly inhibited tumour growth in two different NMC xenografts. Flavopiridol in vivo was significantly effective in one of the two NMC xenograft lines, demonstrating the biological heterogeneity of this disease. CONCLUSIONS: These results demonstrate that FP may be of benefit to a subset of patients with NMC, and warrant a continued emphasis on microtubule inhibitors, anthracyclines, and topoisomerase inhibitors as effective drug classes in this disease.

Cumulative illness severity and progression from moderate to severe retinopathy of prematurity.

OBJECTIVE: To test cumulative neonatal illness severity (IS) and IS fluctuation as predictors of progression from moderate to severe retinopathy of prematurity (ROP). METHODS: Data from research databases and medical record review were collected for infants from four neonatal intensive care unit (NICUs) admitted between 1995 and 2001 and diagnosed with prethreshold ROP. Cumulative neonatal IS measured using daily Scores for Neonatal Acute Physiology (SNAP) for the first 28 days of life, and IS fluctuation as assessed by summing changes between daily SNAP scores, were tested as predictors of progression to threshold ROP using logistic regression. RESULTS: Infants progressing to threshold (n=79), compared to those not progressing to threshold (n=130), had significantly (P<0.05) lower gestational ages (25.2+/-1.1 versus 25.8+/-1.4 weeks), higher cumulative neonatal SNAP (255+/-77 versus 224+/-63 weeks) and had more severe hospitalizations as indicated by diagnoses and medical management. In regression analysis, gestational age, chronological age and presence of plus disease at first diagnosis of prethreshold were associated with development of threshold. After adjusting for these factors, cumulative neonatal SNAP was significantly associated with progression to threshold. However, addition of cumulative SNAP to the model only increased receiver-operating characteristic curve area from 0.77 to 0.78 (NS). Other factors, including SNAP fluctuation, were not associated with progression to threshold after adjustment using this model. CONCLUSIONS: Cumulative neonatal IS, as measured by cumulative SNAP, is an independent risk factor for progression from moderate to severe ROP. However, cumulative IS does not enhance assessment of risk for ROP progression after adjusting for simpler clinical factors.

Hydrofluoroalkane-beclomethasone versus chlorofluorocarbon-beclomethasone delivery in neonatal models.

Dose delivery of hydrofluoroalkane-beclomethasone and chlorofluorocarbon-beclomethasone was compared during in vitro neonatal simulations: mechanical ventilation with 40% and 100% relative humidity + Neonatal Chamber-Ventilator System/endotracheal tube; manual ventilation + Neonatal Chamber/endotracheal tube; "spontaneous breathing" + Neonatal Chamber/face mask without/with manual assistance. The delivery of hydrofluoroalkane-beclomethasone was significantly greater in each simulation.

Postnatal glucocorticoid therapy for prevention of bronchopulmonary dysplasia: routes of administration compared.

Postnatal systemic and inhaled glucocorticoid therapies continue to be used in the management of bronchopulmonary dysplasia (BPD). Systemic dexamethasone therapy has been studied longer than aerosolized glucocorticoid therapy. Several prophylactic trials of systemic dexamethasone therapy demonstrated a reduction in the incidence of BPD and neonatal mortality. However, evidence of potentially serious acute and long-term adverse effects has reduced enthusiasm for use of systemic dexamethasone therapy. Although no trial to date of inhaled glucocorticoid therapy showed a reduction in the incidence of BPD, some studies demonstrated secondary pulmonary benefits with few short-term adverse effects. This article compares the clinical efficacy and safety of these two routes of administration of glucocorticoid therapy.

Inhaled glucocorticoid therapy in infants at risk for neonatal chronic lung disease.

The primary impetus for the study of inhaled glucocorticoid therapy in the treatment and prevention of neonatal chronic lung disease (CLD) was to achieve effective anti-inflammatory therapy with few adverse effects. Initial reports of inhaled glucocorticoid therapy in infants with established CLD suggest modest improvement in neonatal respiratory outcomes. Recent randomized trials also indicate that inhaled glucocorticoid therapy may provide some benefit, but have not demonstrated a reduction in CLD. Some studies suggest that the pulmonary response to systemic glucocorticoid may be greater and faster than response to inhaled glucocorticoid therapy. Few adverse effects have been noted with inhaled glucocorticoid therapy. One limitation of studies of inhaled glucocorticoid therapy is the uncertainty of the dose delivered and deposited in peripheral airways and regions of the lungs. Experience with and systematic study of inhaled glucocorticoid therapy is still in its early stages. The role of inhaled glucocorticoid therapy in the treatment and prevention of CLD is evolving. Advances in delivery devices and new developments of drug formulations should improve aerosol delivery and deposition in infants. Given the clinical dilemma of systemic glucocorticoid therapy and potential benefits demonstrated by recent trials of inhaled glucocorticoid therapy, further study of inhaled glucocorticoid therapy for CLD is warranted.

Postnatal glucocorticosteroid therapy for treatment and prevention of neonatal chronic lung disease.

Neonatal chronic lung disease (CLD) is a persistent complication, primarily of premature infants. Postnatal glucocorticoid therapy is widely used in the treatment and prevention of CLD. Most studies reveal acute improvement in the pulmonary status of infants treated with postnatal glucocorticoid therapy. Recent studies of 'earlier' intervention (< 14 days of age) demonstrated a reduction in mortality and in the occurrence of CLD between 28 days of age and 36 weeks postmenstrual age. Great concern remains, however, regarding the potential adverse outcomes, including growth inhibition, infection, catastrophic GI complications and CNS injury. Therefore, the use of postnatal glucocorticoid therapy remains controversial with respect to the clinical indications for initiating therapy, the dose, duration, onset and route of administration, as well as potential benefits and risks. Inhaled glucocorticoid therapy is increasingly used to treat and prevent CLD in order to avoid adverse effects of high dose systemic glucocorticoid therapy. Recent studies with inhaled glucocorticoid therapy show promise. Further work, however, for improving aerosol delivery and deposition, will be needed to refine their role in the prevention and treatment of CLD. Future studies enabling early, accurate identification of infants at greatest risk for CLD, coupled with a more comprehensive understanding of the different pathogeneses, will provide information regarding appropriate timing of onset, dosing, route of therapy and duration of intervention.

Effects of early inhaled beclomethasone therapy on tracheal aspirate inflammatory mediators IL-8 and IL-1ra in ventilated preterm infants at risk for bronchopulmonary dysplasia.

We tested the hypothesis that inhaled beclomethasone therapy for prevention of bronchopulmonary dysplasia (BPD) reduces pulmonary inflammation. As part of a randomized, placebo-controlled trial, interleukin-8 (IL-8) and interleukin-1 receptor antagonist (IL-1ra) concentrations in tracheal aspirates were measured as markers of pulmonary inflammation. On study days 1 (baseline), 8, 15, and day 28 of age, samples were obtained from enrolled infants (birth weights <1,251 g, gestational age <33 week, 3 to 14 days of age) who remained ventilated and had not received systemic glucocorticoid therapy. Cytokine levels (pg/microg of free secretory component of immunoglobulin A) were compared between groups. We determined whether baseline cytokine levels modified treatment effect regarding subsequent need for systemic glucocorticoid therapy or occurrence of BPD (age 28 days). Tracheal aspirates were obtained from 161 infants (77 receiving beclomethasone, 84 receiving placebo). Median IL-8 levels were lower in beclomethasone versus placebo infants on study days 8 (82.9 vs. 209.2, P < 0.01) and 15 (37.4 vs. 77.4, P < 0.03) after controlling for antenatal glucocorticoid therapy and maternal race. Median IL-1ra levels were lower in beclomethasone versus placebo infants only on study day 8 (86.5 vs. 153.3, P < 0.01). Fewer beclomethasone infants with baseline IL-8 levels in the interquartile range required systemic glucocorticoid therapy (beclomethasone 30.6% vs. placebo 65.8%, P < 0.01) or developed BPD (beclomethasone 42.4% vs. placebo 69.4%, P < 0.03). We conclude that early-inhaled beclomethasone therapy was associated with a reduction in pulmonary inflammation after 1 week of therapy. Beclomethasone-treated infants with moderately elevated baseline IL-8 levels received less subsequent systemic glucocorticoid therapy and had a lower incidence of BPD than nontreated infants.

Special problems in aerosol delivery: neonatal and pediatric considerations.

Identification of the determinants of efficient aerosol delivery and the specific challenges of aerosol delivery to infants and children can facilitate a systematic approach to optimize aerosol delivery to this population. There are inherent anatomical, physiologic, pathophysiologic, and technical limitations of aerosol efficiency in infants and young children. Nevertheless, one can enhance aerosol efficiency through application of sound principles of aerosol delivery and by exerting control over factors that are amenable to intervention. Improvements in aerosol formulations and delivery systems are being made that will enhance efficiency, decrease risk, and reduce waste and cost. Attention to aerosol particle size (1-3 microm mass median aerodynamic diameter and geometric standard deviation < 2 microm), and the concentration of this respirable particle fraction produced by an aerosol system may enhance delivery through endotracheal tubes and to the lower respiratory tract in infants and children with low V(T) and low inspiratory rates. Attention to the choice of delivery system and to details of proper MDI technique (shaking, priming, immediate actuation, and avoiding multiple actuations prior to inhalation), choice of the aerosol spacer and patient interface (type of face mask, endotracheal tube, mouthpiece), spacer cleaning, and consideration of the medicine to be aerosolized (solution or suspension, viscosity) permit adjustment of the aerosol regimen to optimize delivery. All the patient-related, system-related, and operator-dependent considerations combined can greatly impact aerosol delivery efficacy and improve therapeutic response. Therefore, education and motivation of medical personnel, parents and caregivers, and patients regarding factors that influence aerosol efficiency and teaching of proper technique must be prioritized in order to improve aerosol delivery. Aerosol therapy to all patients, especially infants and young children, would be well served if we had a clear understanding of the efficiency and functional differences among the various drugs and devices. These are substantive issues with daily therapeutic impact that have received increasingly outspoken concern over the past decade by aerosol scientists and clinicians. These issues must be given due attention by drug and device manufacturers as well as by regulatory agencies. The medication, the device, and the conditions under which they are tested must be considered together and studied as thoroughly as the medications themselves with respect to total output and particle size distribution. As noted by Bisgaard, medication dose recommendations are useless unless the device and technique used are specified. Medication dose recommendation could be facilitated by setting equivalent standards for generic and brand-name medications and devices. In addition, standardization of in vitro models with better replicas of infants' and children's anatomy (oropharynx, upper airways), and better in vitro lung models, plus utilization of realistic breathing patterns of infants and children will improve in vitro prediction of the in vivo dose delivered to lower airways. This would greatly facilitate selection of delivery systems under specific circumstances for infants and children of various ages). Safety profile, therapeutic efficacy, and efficiency of aerosolized medications delivered to infants and children need to be rigorously studied. This is particularly true for medications with potentially great benefit but possible adverse effects, such as inhaled glucocorticoid therapy in extremely premature infants. Common sense, ethics, and due respect for the same high standard of approval requirements of adults and older children should motivate further research in understanding and improving aerosol delivery in infants and young children.

Strategies for prevention of neonatal chronic lung disease.

Chronic lung disease (CLD) of prematurity remains a substantial problem despite modern perinatal and neonatal care. CLD remains related to gestational age and lung immaturity, although it has become clear that severe initial lung disease is not a prerequisite for CLD to develop. Attempts to prevent CLD to date have not adequately addressed the multifactorial nature of the complex pathophysiology that leads to CLD. Thus, results have been modest at best. Prevention of CLD will require a multifaceted approach with specific interventions and care practices focused on different aspects of the pathway that leads to CLD. This review considers new information related to causation of CLD and the magnitude of the effect of prevention strategies tested to date. This article also advances the hypothesis that CLD is preventable with a global strategy of minimizing inciting events, optimizing management, and specific therapies aimed at intrinsic vulnerabilities.

Meta-analysis of dexamethasone therapy started in the first 15 days of life for prevention of chronic lung disease in premature infants.

We performed a systematic review of randomized controlled trials to determine whether the use of dexamethasone therapy in the first 15 days of life is beneficial for prevention of chronic lung disease is prematurely born infants. Studies were identified by conducting a literature search using the Medline database (1970-1997) and supplemented by a search of the Cochrane Library (1998, issue 4). Inclusion criteria were: 1) prospective randomized design with initiation of dexamethasone therapy within the first 15 days of life; 2) report of outcome of interest; and 3) less than 20% cross-over between treatment and control group during the study period. Our primary outcomes were mortality at hospital discharge and the development of chronic lung disease at 28 days of life and 36 weeks postconceptional age. The secondary outcomes were the presence of a patent ductus arteriosus and treatment side effects. The overall baseline event rate in the control group and pooled risk ratio (RR) of event reduction with 95% confidence interval (CI) were calculated. With dexamethasone therapy, chronic lung disease was decreased by 26% at 28 days (RR, 0.74; 95% CI, 0.57-0.96) and 48% at 36 weeks postconceptional age (RR, 0.52; 95% CI, 0.33-0.81). These reductions were more significant when dexamethasone was started in the first 72 h of life. The relative risk reduction of 24% in deaths was marginally significant (RR, 0.76; 95% CI, 0.56-1.04). The 27% decrease in patent ductus arteriosus and 11% increase in infection were not statistically significant, nor were any other changes. We conclude from this meta-analysis that systemic dexamethasone given to at-risk infants soon after birth may have a beneficial effect in reducing the incidence of chronic lung disease. We did not find evidence of significant short-term adverse side effects. New studies are needed to clarify long-term outcomes in prematurely born infants treated with dexamethasone.

Adrenal function in premature infants during inhaled beclomethasone therapy.

OBJECTIVE: We tested the hypothesis that inhaled beclomethasone therapy for prevention of bronchopulmonary dysplasia does not cause adrenal suppression. STUDY DESIGN: Infants receiving ventilatory support with birth weights

The total care unit for pediatric hematology and oncology, Princess Margaret Hospital for Children, Perth, Australia.

The total care unit for the treatment of pediatric hematology/oncology in Perth, Australia is so named to embody the philosophy of multidisciplinary care of children and their families. Where possible, patients are treated according to randomized controlled trials of the large cooperative Children's Cancer Group. There is a seamless association of clinical and laboratory research. Hemopoietic stem cell transplantation is managed within the unit, as well as treatment of a range of non-malignant hematological disorders. Long-term follow-up of survivors of childhood cancer is coordinated from the unit.