Biallelic variants of ATP13A3 cause dose-dependent childhood-onset pulmonary arterial hypertension characterised by extreme morbidity and mortality.

BACKGROUND: The molecular genetic basis of pulmonary arterial hypertension (PAH) is heterogeneous, with at least 26 genes displaying putative evidence for disease causality. Heterozygous variants in the ATP13A3 gene were recently identified as a new cause of adult-onset PAH. However, the contribution of ATP13A3 risk alleles to child-onset PAH remains largely unexplored. METHODS AND RESULTS: We report three families with a novel, autosomal recessive form of childhood-onset PAH due to biallelic ATP13A3 variants. Disease onset ranged from birth to 2.5 years and was characterised by high mortality. Using genome sequencing of parent-offspring trios, we identified a homozygous missense variant in one case, which was subsequently confirmed to cosegregate with disease in an affected sibling. Independently, compound heterozygous variants in ATP13A3 were identified in two affected siblings and in an unrelated third family. The variants included three loss of function variants (two frameshift, one nonsense) and two highly conserved missense substitutions located in the catalytic phosphorylation domain. The children were largely refractory to treatment and four died in early childhood. All parents were heterozygous for the variants and asymptomatic. CONCLUSION: Our findings support biallelic predicted deleterious ATP13A3 variants in autosomal recessive, childhood-onset PAH, indicating likely semidominant dose-dependent inheritance for this gene.

Recent and Ongoing Research into Metastatic Osteosarcoma Treatments.

The survival rate for metastatic osteosarcoma has not improved for several decades, since the introduction and refinement of chemotherapy as a treatment in addition to surgery. Over two thirds of metastatic osteosarcoma patients, many of whom are children or adolescents, fail to exhibit durable responses and succumb to their disease. Concerted efforts have been made to increase survival rates through identification of candidate therapies via animal studies and early phase trials of novel treatments, but unfortunately, this work has produced negligible improvements to the survival rate for metastatic osteosarcoma patients. This review summarizes data from clinical trials of metastatic osteosarcoma therapies as well as pre-clinical studies that report efficacy of novel drugs against metastatic osteosarcoma in vivo. Considerations regarding the design of animal studies and clinical trials to improve survival outcomes for metastatic osteosarcoma patients are also discussed.

Tetrazolium reduction assays under-report cell death provoked by clinically relevant concentrations of proteasome inhibitors.

High throughput cell viability screening assays often capitalize on the ability of active enzymes or molecules within viable cells to catalyze a quantifiable chemical reaction. The tetrazolium reduction (MTT) assay relies on oxidoreductases to reduce tetrazolium into purple formazan crystals that are solubilized so absorbance reflects viability, while other assays use cellular ATP to catalyze a luminescence-emitting reaction. It is therefore important to know how accurately these assays report cellular responses, as cytotoxic anti-cancer agents promote cell death via a variety of signaling pathways, some of which may alter how these assays work. In this study, we compared the magnitude of cytotoxicity to different cell types provoked by currently used anti-cancer agents, using three different cell viability assays. We found the three assays were consistent in reporting the viability of cells treated with chemotherapy drugs or the BH3 mimetic navitoclax, but the MTT assay underreported the killing capacity of proteasome inhibitors. Additionally, the MTT assay failed to confirm the induction of caspase-mediated cell death by bortezomib at physiologically relevant concentrations, thereby mischaracterizing the mode of cell death. While the cell viability assays used allow for the rapid identification of novel cytotoxic compounds, our study emphasizes the importance for these screening assays to be complemented with a direct measure of cell death or another independent measure of cell viability. We caution researchers against using MTT assays for monitoring cytotoxicity induced by proteasome inhibitors.

The Role of Caregiver-Reported Risks in Predicting Adverse Pediatric Outcomes.

OBJECTIVE: Certain social risk factors (e.g., housing instability, food insecurity) have been shown to directly and indirectly influence pediatric health outcomes; however, there is limited understanding of which social factors are most salient for children admitted to the hospital. This study examines how caregiver-reported social and medical characteristics of children experiencing an inpatient admission are associated with the presence of future health complications. METHODS: Caregivers of children experiencing an inpatient admission (N = 249) completed a predischarge questionnaire designed to capture medical and social risk factors across systems (e.g., patient, caregiver, family, community, healthcare environment). Electronic health record (EHR) data were reviewed for child demographic data, chronic disease status, and subsequent emergency department visits or readmissions (i.e., acute events) 90 days postindex hospitalization. Associations between risk factors and event presence were estimated using odds ratios (ORs) and confidence intervals (CI), both unadjusted and adjusted OR (aOR) for chronic disease and age. RESULTS: Thirty-three percent (N = 82) of children experienced at least one event. After accounting for child age and chronic disease status, caregiver perceptions of child's health being generally "poor" or "not good" prior to discharge (aOR = 4.7, 95% CI = 2.3, 9.7), having high care coordination needs (aOR = 3.2, 95% CI = 1.6, 6.1), and experiencing difficulty accessing care coordination (aOR = 2.5, 95% CI = 1.4, 4.7) were significantly associated with return events. CONCLUSIONS: Caregiver report of risks may provide valuable information above and beyond EHR records to both determine risk of future health problems and inform intervention development.

Proteasome inhibitors trigger mutations via activation of caspases and CAD, but mutagenesis provoked by the HDAC inhibitors vorinostat and romidepsin is caspase/CAD-independent.

Genotoxic anti-cancer therapies such as chemotherapy and radiotherapy can contribute to an increase in second malignancies in cancer survivors due to their oncogenic effects on non-cancerous cells. Inhibition of histone deacetylase (HDAC) proteins or the proteasome differ from chemotherapy in that they eliminate cancer cells by regulating gene expression or cellular protein equilibrium, respectively. As members of these drug classes have been approved for clinical use in recent times, we investigated whether these two drug classes exhibit similar mutagenic capabilities as chemotherapy. The HDAC inhibitors vorinostat/SAHA and romidepsin/FK288 were found to induce DNA damage, and mis-repair of this damage manifested into mutations in clonogenically viable surviving cells. DNA damage and mutations were also detected in cells treated with the proteasome inhibitor bortezomib. Exposure to both drug classes stimulated caspase activation consistent with apoptotic cell death. Inhibition of caspases protected cells from bortezomib-induced acute (but not clonogenic) death and mutagenesis, implying caspases were required for the mutagenic action of bortezomib. This was also observed for second generation proteasome inhibitors. Cells deficient in caspase-activated DNase (CAD) also failed to acquire DNA damage or mutations following treatment with bortezomib. Surprisingly, vorinostat and romidepsin maintained an equivalent level of killing and mutagenic ability regardless of caspase or CAD activity. Our findings indicate that both drug classes harbour mutagenic potential in vitro. If recapitulated in vivo, the mutagenicity of these agents may influence the treatment of cancer patients who are more susceptible to oncogenic mutations due to dysfunctional DNA repair pathways.

Smac mimetics LCL161 and GDC-0152 inhibit osteosarcoma growth and metastasis in mice.

BACKGROUND: Current therapies fail to cure over a third of osteosarcoma patients and around three quarters of those with metastatic disease. "Smac mimetics" (also known as "IAP antagonists") are a new class of anti-cancer agents. Previous work revealed that cells from murine osteosarcomas were efficiently sensitized by physiologically achievable concentrations of some Smac mimetics (including GDC-0152 and LCL161) to killing by the inflammatory cytokine TNFα in vitro, but survived exposure to Smac mimetics as sole agents. METHODS: Nude mice were subcutaneously or intramuscularly implanted with luciferase-expressing murine 1029H or human KRIB osteosarcoma cells. The impacts of treatment with GDC-0152, LCL161 and/or doxorubicin were assessed by caliper measurements, bioluminescence, 18FDG-PET and MRI imaging, and by weighing resected tumors at the experimental endpoint. Metastatic burden was examined by quantitative PCR, through amplification of a region of the luciferase gene from lung DNA. ATP levels in treated and untreated osteosarcoma cells were compared to assess in vitro sensitivity. Immunophenotyping of cells within treated and untreated tumors was performed by flow cytometry, and TNFα levels in blood and tumors were measured using cytokine bead arrays. RESULTS: Treatment with GDC-0152 or LCL161 suppressed the growth of subcutaneously or intramuscularly implanted osteosarcomas. In both models, co-treatment with doxorubicin and Smac mimetics impeded average osteosarcoma growth to a greater extent than either drug alone, although these differences were not statistically significant. Co-treatments were also more toxic. Co-treatment with LCL161 and doxorubicin was particularly effective in the KRIB intramuscular model, impeding primary tumor growth and delaying or preventing metastasis. Although the Smac mimetics were effective in vivo, in vitro they only efficiently killed osteosarcoma cells when TNFα was supplied. Implanted tumors contained high levels of TNFα, produced by infiltrating immune cells. Spontaneous osteosarcomas that arose in genetically-engineered immunocompetent mice also contained abundant TNFα. CONCLUSIONS: These data imply that Smac mimetics can cooperate with TNFα secreted by tumor-associated immune cells to kill osteosarcoma cells in vivo. Smac mimetics may therefore benefit osteosarcoma patients whose tumors contain Smac mimetic-responsive cancer cells and TNFα-producing infiltrating cells.

Aptamer micelles targeting fractalkine-expressing cancer cells in vitro and in vivo.

In this work we hypothesized that the chemokine fractalkine can serve as a cancer molecular target. We engineered aptamer micelles functionalized with an outer poly(ethylene glycol) (PEG) corona, and investigated the extent and efficacy of using them as a targeting tool against fractalkine-expressing colon adenocarcinoma cells. In vitro cell binding results showed that aptamer micelles bound and internalized to fractalkine-expressing cancer cells with the majority of the micelles found free in the cytoplasm. Minimal surface binding was observed by healthy cells. Even though partial PEGylation did not prevent serum adsorption, micelles were highly resistant to endonuclease and exonuclease degradation. In vivo biodistribution studies and confocal studies demonstrated that even though both aptamer and control micelles showed tumor accumulation, only the aptamer micelles internalized into fractalkine-expressing cancer cells, thus demonstrating the potential of the approach and showing that fractalkine may serve as a specific target for nanoparticle delivery to cancer cells.

Your Exclusion, My Inclusion: Reflections on a Career Working With the Most Challenging and Vulnerable in Diabetes.

EDITOR'S NOTE: This article is adapted from the address Dr. Harris delivered as the recipient of the American Diabetes Association's Richard R. Rubin Award for 2017. This award recognizes a behavioral researcher who has made outstanding, innovative contributions to the study and understanding of the behavioral aspects of diabetes in diverse populations. Dr. Harris delivered the address in June 2017 at the Association's 77th Scientific Sessions in San Diego, Calif.

Partnering with Insurers in Caring for the Most Vulnerable Youth with Diabetes: NICH as an Integrator.

PURPOSE OF REVIEW: In this review, we outline barriers to appropriately caring for high-risk youth with diabetes and discuss efforts in partnering with insurers through Alternative Payment Models to achieve the Triple Aim (improved health, improved care, and reduced costs) for this population. RECENT FINDINGS: Current approaches in caring for youth with diabetes who evidence a high degree of social complexity are woefully ineffective. These youth are vulnerable to repeat diabetic ketoacidosis episodes, poor glycemic control, and excessive utilization of healthcare resources. To effectively pursue the Triple Aim, an "integrator" (i.e., an entity that accepts responsibility for all components of the Triple Aim for a specified population) must be identified; however, this does not fit into current fee-for-service models. Integrators for youth with diabetes are limited, but early examples of integrator efforts are promising. We present one successful "integrator," Novel Interventions in Children's Healthcare (NICH), and detail this program's efforts in partnering with insurers to serve high-risk youth with diabetes.

Phosphatidylcholine Coatings Deliver Local Antimicrobials and Reduce Infection in a Murine Model: A Preliminary Study.

BACKGROUND: Phosphatidylcholine coatings have been shown to elute antibiotics for several days. A recently developed biofilm inhibitor, cis-2-decenoic acid (C2DA), has been shown to exhibit synergistic activity with several common antibiotics. This study aims to evaluate the effectiveness of C2DA and amikacin dual drug delivery from a phosphatidylcholine coating. QUESTIONS/PURPOSES: (1) What are the in vitro elution profiles of amikacin and C2DA from phosphatidylcholine-coated coupons in incubated phosphate-buffered saline? (2) Does the presence of C2DA in eluate samples lower the amount of amikacin needed for bacterial inhibition in overnight bacterial turbidity assays? (3) Does addition of amikacin and C2DA result in decreased colony-forming units (CFUs) on wire implants and bone when compared with phosphatidylcholine coatings alone in a mouse model of periprosthetic joint infection? METHODS: Effects of loading concentrations were assessed during 7-day in vitro elution studies for coatings containing all mixtures of 0%, 5%, 15%, and 25% wt of amikacin and C2DA (n = 4) through quantitative high-performance liquid chromatography concentration determination and plotting concentration eluted over time. Antimicrobial activity was assessed by overnight turbidity testing of elution study samples against Staphylococcus aureus or Pseudomonas aeruginosa. In vivo efficacy was assessed using phosphatidylcholine-coated wire implants in a murine (mouse) model of infection (n = 3). Wire implants were coated with phosphatidylcholine containing no antimicrobials, amikacin alone, C2DA alone, or amikacin and C2DA and then inserted into the intramedullary femur of each mouse and inoculated with S aureus. The number of viable bacterial colonies on the implant surface and in the surrounding bone was determined after 1 week with the goal of achieving complete bacterial clearance. Total viable CFU count and proportion of samples achieving complete clearance were compared between groups. RESULTS: Elution samples showed a burst response of amikacin and C2DA for 1 to 2 days with C2DA release continuing at low levels through Day 4. All tested eluate samples inhibited P aeruginosa. Samples from coatings containing 25% amikacin or 15% amikacin and any amount of C2DA were able to inhibit S aureus formation, but all coatings with 5% amikacin or 15% amikacin but no C2DA were not inhibitory. All in vivo treatment groups achieved complete bacterial clearance on the wire implant, and the C2DA alone and amikacin alone coatings cleared all CFUs in bone (pin: phosphatidylcholine only one of three; amikacin three of three, C2DA three of three, amikacin + C2DA three of three, p = 0.04 [Fisher's exact test]; bone: coating only: zero of three; amikacin: three of three; C2DA; three of three; C2DA + amikacin: one of three; p = 0.03 [Fisher's exact test]). CONCLUSIONS: Phosphatidylcholine coatings elute antimicrobials in vitro under infinite sink conditions for up to 4 days in phosphate-buffered saline and were able to reduce bacterial colonies in a preliminary in vivo model. Turbidity testing with eluate samples containing varying amounts of C2DA and amikacin agrees with previous studies showing synergy between them. CLINICAL RELEVANCE: Used as an adjunctive to systemic therapy, C2DA-loaded phosphatidylcholine coatings have potential value as a prophylactic infection prevention measure. Future studies may include different antibiotics, animal studies with larger sample sizes and more controls, and advanced coating delivery methods.

G-DOC Plus - an integrative bioinformatics platform for precision medicine.

BACKGROUND: G-DOC Plus is a data integration and bioinformatics platform that uses cloud computing and other advanced computational tools to handle a variety of biomedical BIG DATA including gene expression arrays, NGS and medical images so that they can be analyzed in the full context of other omics and clinical information. RESULTS: G-DOC Plus currently holds data from over 10,000 patients selected from private and public resources including Gene Expression Omnibus (GEO), The Cancer Genome Atlas (TCGA) and the recently added datasets from REpository for Molecular BRAin Neoplasia DaTa (REMBRANDT), caArray studies of lung and colon cancer, ImmPort and the 1000 genomes data sets. The system allows researchers to explore clinical-omic data one sample at a time, as a cohort of samples; or at the level of population, providing the user with a comprehensive view of the data. G-DOC Plus tools have been leveraged in cancer and non-cancer studies for hypothesis generation and validation; biomarker discovery and multi-omics analysis, to explore somatic mutations and cancer MRI images; as well as for training and graduate education in bioinformatics, data and computational sciences. Several of these use cases are described in this paper to demonstrate its multifaceted usability. CONCLUSION: G-DOC Plus can be used to support a variety of user groups in multiple domains to enable hypothesis generation for precision medicine research. The long-term vision of G-DOC Plus is to extend this translational bioinformatics platform to stay current with emerging omics technologies and analysis methods to continue supporting novel hypothesis generation, analysis and validation for integrative biomedical research. By integrating several aspects of the disease and exposing various data elements, such as outpatient lab workup, pathology, radiology, current treatments, molecular signatures and expected outcomes over a web interface, G-DOC Plus will continue to strengthen precision medicine research. G-DOC Plus is available at: https://gdoc.georgetown.edu .

Treating the most vulnerable and costly in diabetes.

Diabetic ketoacidosis (DKA) is associated with negative health outcomes and high costs for patients, families, and communities. Interventions developed to effectively reduce DKA and related costs should target the multiple risk factors associated with DKA and adherence difficulties. Certain demographic, psychological, and family factors are associated with increased risk for adherence problems and DKA. Individuals with a combination of risk factors (e.g., mental health problems, low socioeconomic status, high family conflict) may be particularly vulnerable to DKA. Although several different interventions have demonstrated promise in improving adherence and/or decreasing the risk of DKA, the generalizability of treatment results to those individuals most vulnerable to DKA is limited. Approaches which include multiple evidence-based components of care, are flexible in treatment delivery (e.g., home- and community-based, utilize technology), and target the multiple risk factors across relevant systems (e.g., individual, family, school, medical) are warranted to effectively reduce DKA in vulnerable populations.

Executive function, adherence, and glycemic control in adolescents with type 1 diabetes: a literature review.

The aim of the present review was to examine and report findings from published research to date that has examined associations between executive function (EF), adherence, and glycemic control in youth with type 1 diabetes. A review of the published research is presented with the objectives of reporting the following: (1) the associations between EF and adherence, (2) the associations between EF and glycemic control, (3) proposed methodological considerations needed to advance related research, (4) recommendations for future research, and (5) clinical recommendations. The major conclusions of this review support the presence of an association between EF, adherence, and glycemic control. Additional prospective and controlled studies are necessary to fully understand the impact of EF on the ability of youth to independently manage type 1 diabetes.

When helping hurts: miscarried helping in families of youth with chronic pain.

OBJECTIVE: To examine "miscarried helping" as a maladaptive dyadic process in families of youth with chronic pain using the Actor-Partner Interdependence Model. METHODS: 210 adolescents with chronic pain (mean = 14.23 years; 73.9% female) and their parents participating in a multicenter study completed measures assessing pain characteristics, miscarried helping, family functioning, parental protectiveness, and child depressive symptoms. RESULTS: Multilevel modeling revealed significant actor effects of miscarried helping on family functioning for both parents and teens, but not partner effects. Individual-level factors, including child pain characteristics, depressive symptoms, and parental protectiveness, uniquely contributed to miscarried helping. CONCLUSIONS: Higher perceptions of miscarried helping contribute to worse family functioning and may be a useful target for psychological intervention in parents of children with chronic pain. Parents who exhibit more protective responses to pain and youth with more depressive symptoms may be at increased risk for a miscarried helping process to develop.

Percutaneous peripheral nerve stimulation for chronic pain in subacromial impingement syndrome: a case series.

OBJECTIVE: The objective of this study was to determine the effect of peripheral nerve stimulation (PNS) on pain reduction for those with refractory subacromial impingement syndrome (SIS) and to evaluate the association with reduced disability, impairment, and safety. Our hypotheses are that PNS will be associated with a reduction in pain, impairment and disability, and improvement in quality of life while demonstrating safety. MATERIAL AND METHODS: Adults with shoulder pain of at least six months duration were recruited for a three-week treatment of percutaneous PNS applied through a percutaneous electrode to the axillary motor points of the deltoid muscle. Subjects were followed for 12 weeks after treatment. The primary outcome was the worst pain in the last week, and secondary outcomes included pain interference, the Disabilities of the Arm, Shoulder, and Hand questionnaire, shoulder abduction range of motion, and safety. Analysis was with a linear mixed model. RESULTS: Ten subjects were recruited. Longitudinal analysis demonstrated significant reduction in pain relative to baseline (F(1, 66) = 12.9, p < 0.01). After correcting for multiple comparisons, there were significant reductions at explantation and all follow-up time points when compared with baseline. There were also significant improvements in pain interference (F(1,65) = 15.0, p < 0.01), the Disabilities of the Arm, Shoulder, and Hand questionnaire (F(1,35) = 7.0, p = 0.01), and shoulder abduction range of motion (F(1,35) = 6.3, p = 0.02). CONCLUSION: Intramuscular PNS for chronic shoulder pain due to SIS is a safe treatment associated with pain reduction, lower pain interference with activities of daily living, reduced disability, and improved shoulder abduction. Pain reduction is maintained for at least 12 weeks after treatment.

The anti-cancer immune response in breast cancer: current and emerging biomarkers and treatments.

Triple-negative breast cancers (TNBCs) exhibit heightened T cell infiltration, contributing to an enhanced response to immune checkpoint blockade (ICB) compared with other subtypes. An immune-rich immune microenvironment correlates with improved prognosis in early and advanced TNBC. Combination chemotherapy and ICB is now the standard of care in early- and late-stage TNBC. Although programmed death ligand-1 (PD-L1) positivity predicts ICB response in advanced stages, its role in early-stage disease remains uncertain. Despite neoadjuvant ICB becoming common in early-stage TNBC, the necessity of adjuvant ICB after surgery remains unclear. Understanding the molecular basis of the immune response in breast cancer is vital for precise biomarkers for ICB and effective combination therapy strategies.

Towards targeting the breast cancer immune microenvironment.

The tumour immune microenvironment is shaped by the crosstalk between cancer cells, immune cells, fibroblasts, endothelial cells and other stromal components. Although the immune tumour microenvironment (TME) serves as a source of therapeutic targets, it is also considered a friend or foe to tumour-directed therapies. This is readily illustrated by the importance of T cells in triple-negative breast cancer (TNBC), culminating in the advent of immune checkpoint therapy in combination with cytotoxic chemotherapy as standard of care for both early and advanced-stage TNBC, as well as recent promising signs of efficacy in a subset of hormone receptor-positive disease. In this Review, we discuss the various components of the immune TME in breast cancer and therapies that target or impact the immune TME, as well as the complexity of host physiology.

Social Determinants of Health Screening in Type 1 Diabetes Management.

Type 1 diabetes management is intricately influenced by social determinants of health. Economic status impacts access to vital resources like insulin and diabetes technology. Racism, social injustice, and implicit biases affect equitable delivery of care. Education levels affect understanding of self-care, leading to disparities in glycemic outcomes. Geographic location can limit access to health care facilities. Stressors from discrimination or financial strain can disrupt disease management. Addressing these social factors is crucial for equitable diabetes care, emphasizing the need for comprehensive strategies that go beyond medical interventions to ensure optimal health outcomes for all individuals with type 1 diabetes.

Diabetes self-management profile short form: a preliminary report.

Effective family management of type 1 diabetes in childhood is critical to maintaining optimal glycemic control. The purpose of this study was to provide preliminary evidence for a reduced form of the Diabetes Self-Management Profile (DSMP) using Rasch modeling techniques. The study was a secondary analysis of DSMP data drawn from a previous study on patterns of self-management from 239 preadolescents with type 1 diabetes. Rasch modeling strategies were used to identify the most informative items and then a reduced score composite was correlated with hemoglobin A1c (A1c) and blood glucose monitoring (BGM) frequency. A short form of the DSMP was obtained using seven items that comprised all five subscales of the DSMP. The DSMP Short-Form (DSMP-SF) composite score correlated significantly with child's HbA1c and BGM frequency. The DSMP-SF may be considered a valid and effective screening interview alternative to the longer, original, DSMP, particularly when attempting to identify high-risk patients.