Low-temperature (< 200 °C) degradation of electronic nicotine delivery system liquids generates toxic aldehydes.

Electronic cigarette usage has spiked in popularity over recent years. The enhanced prevalence has consequently resulted in new health concerns associated with the use of these devices. Degradation of the liquids used in vaping have been identified as a concern due to the presence of toxic compounds such as aldehydes in the aerosols. Typically, such thermochemical conversions are reported to occur between 300 and 400 °C. Herein, the low-temperature thermal degradation of propylene glycol and glycerol constituents of e-cigarette vapors are explored for the first time by natural abundance 13C NMR and 1H NMR, enabling in situ detection of intact molecules from decomposition. The results demonstrate that the degradation of electronic nicotine delivery system (ENDS) liquids is strongly reliant upon the oxygen availability, both in the presence and absence of a material surface. When oxygen is available, propylene glycol and glycerol readily decompose at temperatures between 133 and 175 °C over an extended time period. Among the generated chemical species, formic and acrylic acids are observed which can negatively affect the kidneys and lungs of those who inhale the toxin during ENDS vapor inhalation. Further, the formation of hemi- and formal acetals is noted from both glycerol and propylene glycol, signifying the generation of both formaldehyde and acetaldehyde, highly toxic compounds, which, as a biocide, can lead to numerous health ailments. The results also reveal a retardation in decomposition rate when material surfaces are prevalent with no directly observed unique surface spectator or intermediate species as well as potentially slower conversions in mixtures of the two components. The generation of toxic species in ENDS liquids at low temperatures highlights the dangers of low-temperature ENDS use.

Burosumab for the Treatment of Tumor-Induced Osteomalacia.

Tumor-induced osteomalacia (TIO) is caused by phosphaturic mesenchymal tumors producing fibroblast growth factor 23 (FGF23) and is characterized by impaired phosphate metabolism, skeletal health, and quality of life. UX023T-CL201 is an ongoing, open-label, phase 2 study investigating the safety and efficacy of burosumab, a fully human monoclonal antibody that inhibits FGF23, in adults with TIO or cutaneous skeletal hypophosphatemia syndrome (CSHS). Key endpoints were changes in serum phosphorus and osteomalacia assessed by transiliac bone biopsies at week 48. This report focuses on 14 patients with TIO, excluding two diagnosed with X-linked hypophosphatemia post-enrollment and one with CSHS. Serum phosphorus increased from baseline (0.52 mmol/L) and was maintained after dose titration from week 22 (0.91 mmol/L) to week 144 (0.82 mmol/L, p < 0.0001). Most measures of osteomalacia were improved at week 48: osteoid volume/bone, osteoid thickness, and mineralization lag time decreased; osteoid surface/bone surface showed no change. Of 249 fractures/pseudofractures detected across 14 patients at baseline, 33% were fully healed and 13% were partially healed at week 144. Patients reported a reduction in pain and fatigue and an increase in physical health. Two patients discontinued: one to treat an adverse event (AE) of neoplasm progression and one failed to meet dosing criteria (receiving minimal burosumab). Sixteen serious AEs occurred in seven patients, and there was one death; all serious AEs were considered unrelated to treatment. Nine patients had 16 treatment-related AEs; all were mild to moderate in severity. In adults with TIO, burosumab exhibited an acceptable safety profile and was associated with improvements in phosphate metabolism and osteomalacia. © 2020 The Authors. Journal of Bone and Mineral Research published by American Society for Bone and Mineral Research..

Molecular Control of Phosphorus Homeostasis and Precision Treatment of Hypophosphatemic Disorders.

PURPOSE OF REVIEW: Serum phosphorus is maintained in a narrow range by balancing dietary phosphate absorption, influx and efflux of phosphorus from bone and intracellular stores, and renal reabsorption of filtered phosphate. Acute hypophosphatemia, typically caused by transient increases in cellular uptake, can lead to severe complications such as cardiopulmonary dysfunction and rhabdomyolysis that can warrant parenteral phosphate repletion. Chronic hypophosphatemia, however, generally represents true phosphate deficiency and may result in long-term metabolic and skeletal complications, particularly in children due to the critical importance of phosphorus to skeletal mineralization and longitudinal growth. RECENT FINDINGS: In addition to the well characterized roles of vitamin D and parathyroid hormone (PTH), a new bone-kidney axis has been discovered that regulates phosphate homeostasis through the bone-derived hormone Fibroblast Growth Factor 23 (FGF23) and its phosphaturic actions that are mediated by activation of fibroblast growth factor receptors (FGFRs) complexed with α-Klotho in renal tubules. Chronic hypophosphatemia can now be classified as FGF23 dependent or independent. SUMMARY: In cases of FGF23 dependent hypophosphatemia, traditional non-specific treatments with elemental phosphorus and 1,25(OH)2 vitamin D (calcitriol) can now be replaced with a targeted approach by using an FGF-23 blocking antibody (Burosumab).

Continued Beneficial Effects of Burosumab in Adults with X-Linked Hypophosphatemia: Results from a 24-Week Treatment Continuation Period After a 24-Week Double-Blind Placebo-Controlled Period.

Burosumab, a fully human monoclonal antibody to FGF23, is the only approved treatment for X-linked hypophosphatemia (XLH), a rare genetic disorder characterized by renal phosphate wasting and substantial cumulative musculoskeletal morbidity. During an initial 24-week randomized, controlled trial, 134 adults with XLH received burosumab 1 mg/kg (n = 68) or placebo (n = 66) every 4 weeks. After 24 weeks, all subjects received open-label burosumab until week 48. This report describes the efficacy and safety of burosumab during the open-label treatment period. From weeks 24-48, serum phosphorus concentrations remained normal in 83.8% of participants who received burosumab throughout and were normalized in 89.4% who received burosumab after placebo. By week 48, 63.1% of baseline fractures/pseudofractures healed fully with burosumab, compared with 35.2% with burosumab after placebo. In both groups, burosumab was associated with clinically significant and sustained improvement from baseline to week 48 in scores for patient-reported outcomes of stiffness, pain, physical function, and total distance walked in 6 min. Rates of adverse events were similar for burosumab and placebo. There were no fatal adverse events or treatment-related serious adverse events. Nephrocalcinosis scores did not change from baseline by more than one grade at either week 24 or 48. These data demonstrate that in participants with XLH, continued treatment with burosumab is well tolerated and leads to sustained correction of serum phosphorus levels, continued healing of fractures and pseudofractures, and sustained improvement in key musculoskeletal impairments.

Tenofovir Has Minimal Effect on Biomarkers of Bone Health in Youth with HIV Receiving Initial Antiretroviral Therapy.

Both HIV infection and tenofovir disoproxil fumarate (TDF) treatment adversely impact bone metabolism and may lead to osteopenia, which has critical implications for youth with HIV (YWH). This study evaluates changes in the biomarkers of bone metabolism and inflammation among YWH receiving initial treatment with TDF- and non-TDF-containing antiretroviral therapies (ARTs). YWH [n = 23, median age 21 years (range 18-24), 87% male, 61% African American] were assessed for inflammatory and bone metabolism biomarkers at enrollment, after 48 weeks of TDF-containing ART, and 96 weeks of ART without TDF with continued viral suppression. Spearman's rank correlation evaluated biomarker associations. Bone alkaline phosphatase, parathyroid hormone, and osteopontin increased after TDF treatment. All fell after TDF was discontinued. Levels of RANKL and osteoprotegerin did not change throughout the study. There was little correlation between biomarkers of bone metabolism and either macrophage or lymphocyte activation at any time point. Our results establish baseline associations between bone metabolism and immune biomarkers for this population, and find that before CD4 T cell decline chronic inflammation does not perturb biomarkers of bone metabolism among YWH. The adverse effects of TDF on bone health may be marginal for YWH at the early stages of disease.

Stable Acinar Progenitor Cell Model Identifies Treacle-Dependent Radioresistance.

Radiotherapy for head and neck cancers can result in extensive damage to the salivary glands, significantly affecting patient quality of life. However, the salivary gland can recover in patients receiving lower doses of radiation. In addition, there is considerable interest in delineating the mechanisms by which stem cells survive radiation exposure and promote tissue regeneration. In this study, we isolated stable radioresistant acinar progenitor cells from the submaxillary gland of the Sprague Dawley rat. Progenitor cells are characterized as c-Kithigh/alpha-amylase+ and are resistant to X rays (≤5 Gy).We further isolated a radiosensitive acinar counterpart, characterized as c-Kitlow/alpha-amylase+, which is effectively killed by exposure to 2 Gy X ray of radiation. Phosphopeptides with homology to the treacle protein (TCOF1) were disproportionately increased in progenitor cells, compared to their radiosensitive counterparts. Silencing of TCOF1 expression (shRNA) radiosensitized progenitor cells, a response conserved in human cells with TCOF1 knockdown. Collectively, these observations indicate that radiation resistance is an intrinsic property of c-Kithigh salivary gland progenitor cells. Since human salivary gland stem cells with c-Kit expression are believed to have enhanced regenerative potencies, our model system provides a stable platform to investigate molecular features associated with c-Kit expression that may contribute to protection or stabilization of the stem cell niche.

Evaluation of non-invasive biomonitoring of 2,4-Dichlorophenoxyacetic acid (2,4-D) in saliva.

The objective of this study was to evaluate the potential for non-invasive biomonitoring of 2,4-Dichlorophenoxyacetic acid (2,4-D) in saliva. Using an in vitro rat salivary gland epithelial cell (SGEC) system, a collection of experiments investigating chemical protein binding, temporal and directional transport, as well as competitive transport with para-aminohippuric acid (PAH), a substrate for renal organic anion transporters, was conducted to identify cellular transport parameters required to computationally model salivary transport of 2,4-D. Additionally, a physiological protein gradient was implemented to mimic physiologically relevant concentrations of protein in rat plasma and saliva, and under these conditions the transfer of 2,4-D was markedly slower, driven by increased protein binding (i.e. reduced free 2,4-D species available to cross salivary barrier). The rate of transfer was directly proportional to the amount of unbound 2,4-D and demonstrated no indication of active transport. An in vivo assessment of 2,4-D exposure in rats revealed non-linear protein binding in plasma, indicating saturated protein binding and increased levels of unbound 2,4-D species at higher doses. A strong correlation between 2,4-D concentrations in saliva and unbound 2,4-D in plasma was observed (Pearson correlation coefficient = 0.95). Saliva:plasma 2,4-D ratios measured in vivo (0.0079) were consistent within the linear protein binding range and expected 2,4-D levels from occupational exposures but were significantly different than ratios measured in vitro (physiological conditions) (0.034), possibly due to 2,4-D concentrations in saliva not being at equilibrium with 2,4-D concentrations in blood, as well as physiological features absent in in vitro settings (e.g. blood flow). We demonstrated that 2,4-D is consistently transported into saliva using both in vitro and in vivo models, making 2,4-D a potential candidate for human non-invasive salivary biomonitoring. Further work is needed to understand whether current sensor limits of detection are sufficient to measure occupationally relevant exposures.

Impaired calcium sensing distinguishes primary hyperparathyroidism (PHPT) patients with low bone mineral density.

CONTEXT: A subset of PHPT patients exhibit a more severe disease phenotype characterized by bone loss, fractures, recurrent nephrolithiasis, and other dysfunctions, but the underlying reasons for this disparity in clinical presentation remain unknown. OBJECTIVE: We sought to identify new mechanistic indices that could inform more personalized management of PHPT. DESIGN: Pre-, peri-, and postoperative data and demographic, clinical, and pathological information from patients undergoing parathyroidectomy for PHPT were collected. Univariate and partial Spearman correlation was used to estimate the association of parathyroid tumor calcium sensing capacity with select variables. PATIENTS OR OTHER PARTICIPANTS: An unselected series of 237 patients aged >18years and undergoing parathyroidectomy for PHPT were enrolled. MAIN OUTCOME MEASURES: Calcium sensing capacity, expressed as the concentration required for half-maximal biochemical response (EC50), was evaluated in parathyroid tumors from an unselected series of 74 patients and assessed for association with clinical parameters. The hypothesis was that greater disease severity would be associated with attenuated calcium sensitivity and biochemically autonomous parathyroid tumor behavior. RESULTS: Parathyroid tumors segregated into two distinct groups of calcium responsiveness (EC50<3.0 and ≥3.0mM). The low EC50 group (n=27) demonstrated a mean calcium EC50 value of 2.49mM [95% confidence interval (CI): 2.43-2.54mM], consistent with reference normal activity. In contrast, the high EC50 group (n=47) displayed attenuated calcium sensitivity with a mean EC50 value of 3.48mM [95% CI: 3.41-3.55mM]. Retrospective analysis of the clinical registry data suggested that high calcium EC50 patients presented with a more significant preoperative bone mineral density (BMD) deficit with a t-score of -2.7, (95% CI: -3.4 to -1.9) versus 0.9, (95% CI: -2.1 to -0.4) in low EC50 patients (p<0.001). After adjusting for gender, age, BMI, 25 OH vitamin D level and preoperative iPTH, lowest t-score and calcium EC50 were inversely correlated, with a partial Spearman correlation coefficient of -0.35 (p=0.02). CONCLUSIONS: Impaired calcium sensing in parathyroid tumors is selectively observed in a subset of patients with more severe bone mineral density deficit. Assessment of parathyroid tumor biochemical behavior may be a useful predictor of disease severity as measured by bone mineral density in patients with PHPT.

Non-invasive saliva human biomonitoring: development of an in vitro platform.

Direct measurements of exposure represent the most accurate assessment of a subject's true exposure. The clearance of many drugs and chemicals, including pesticides such as chlorpyrifos (CPF), can be detected non-invasively in saliva. Here we have developed a serous-acinar transwell model system as an in vitro screening platform to prioritize chemicals for non-invasive biomonitoring through salivary clearance mechanisms. Rat primary serous-acinar cells express both α-amylase and aquaporin-5 proteins and develop significant tight junctions at postconfluence - a feature necessary for chemical transport studies in vitro. CPF exhibited bidirectional passage across the serous-acinar barrier that was disproportional to the passage of a cell impermeable chemical (lucifer yellow), consistent with a hypothesized passive diffusion process. CPF was metabolized to trichlorpyridinol (TCPy) by serous-acinar cells, and TCPy also displayed bidirectional diffusion in the transwell assay. This model system should prove useful as an in vitro screening platform to support the non-invasive monitoring of toxicons and pharmacons in human saliva and provide guidance for development of advanced in vitro screening platforms utilizing primary human salivary gland epithelial cells.

Burden of disease in adult patients with hypophosphatasia: Results from two patient-reported surveys.

BACKGROUND: Hypophosphatasia (HPP) is a rare metabolic bone disease caused by loss-of-function mutation(s) in the tissue-nonspecific alkaline (TNSALP) phosphatase gene, which manifests as rickets and/or osteomalacia with systemic complications and affects patients of all ages. The burden of disease is poorly characterized in adult patients. AIMS: We assessed patient-reported burden of disease using two surveys reasonably specific for HPP symptomatology, the Hypophosphatasia Impact Patient Survey (HIPS) and the Hypophosphatasia Outcomes Study Telephone interview (HOST). METHODS: Patients with HPP were invited to participate via patient advocacy groups or their medical provider. Survey questions captured demography, HPP-related medical history, mobility, and health-related quality of life (using Short Form 12 [version 2] Health Survey [SF-12v2]) via internet report (HIPS) or telephone interview (HOST). RESULTS: One hundred twenty-five adults responded (mean [standard deviation, SD] age: 45 [14.3] years). Eighty-four patients (67%) reported pediatric-onset of their symptoms. Common clinical features in the study population included pain (95% of patients), fractures (86% of patients) muscle weakness (62%) and unusual gait (52%). Use of assistive devices for mobility (60%) was also prevalent. Twenty-six percent of patients reported more than 10 fractures. Seventy-four percent of patients had undergone orthopedic/dental surgical procedures. The health profile of patients responding on the SF-12 showed a broad and substantial impact of HPP on health-related quality of life, with domains related to physical ability showing the greatest decrement compared to normative data. CONCLUSIONS: In aggregate, these data indicate that HPP can confer a high burden of illness in adulthood.

Pharmacokinetics and pharmacodynamics of a human monoclonal anti-FGF23 antibody (KRN23) in the first multiple ascending-dose trial treating adults with X-linked hypophosphatemia.

In X-linked hypophosphatemia (XLH), serum fibroblast growth factor 23 (FGF23) is increased and results in reduced renal maximum threshold for phosphate reabsorption (TmP), reduced serum inorganic phosphorus (Pi), and inappropriately low normal serum 1,25 dihydroxyvitamin D (1,25[OH]2 D) concentration, with subsequent development of rickets or osteomalacia. KRN23 is a recombinant human IgG1 monoclonal antibody that binds to FGF23 and blocks its activity. Up to 4 doses of KRN23 were administered subcutaneously every 28 days to 28 adults with XLH. Mean ± standard deviation KRN23 doses administered were 0.05, 0.10 ± 0.01, 0.28 ± 0.06, and 0.48 ± 0.16 mg/kg. The mean time to reach maximum serum KRN23 levels was 7.0 to 8.5 days. The mean KRN23 half-life was 16.4 days. The mean area under the concentration-time curve (AUCn ) for each dosing interval increased proportionally with increases in KRN23 dose. The mean intersubject variability in AUCn ranged from 30% to 37%. The area under the effect concentration-time curve (AUECn ) for change from baseline in TmP per glomerular filtration rate, serum Pi, 1,25(OH)2 D, and bone markers for each dosing interval increased linearly with increases in KRN23 AUCn . Linear correlation between serum KRN23 concentrations and increase in serum Pi support KRN23 dose adjustments based on predose serum Pi concentration.

Prolonged Correction of Serum Phosphorus in Adults With X-Linked Hypophosphatemia Using Monthly Doses of KRN23.

CONTEXT: In X-linked hypophosphatemia (XLH), elevated fibroblast growth factor 23 (FGF23) decreases the renal tubular maximum reabsorption rate of phosphate/glomerular filtration rate (TmP/GFR) and serum inorganic phosphorus (Pi), resulting in rickets and/or osteomalacia. OBJECTIVE: The objective was to test the hypothesis that monthly KRN23 (anti-FGF23 antibody) would safely improve serum Pi in adults with XLH. DESIGN: Two sequential open-label phase 1/2 studies were done. SETTING: Six academic medical centers were used. PARTICIPANTS: Twenty-eight adults with XLH participated in a 4-month dose-escalation study (0.05-0.6 mg/kg); 22 entered a 12-month extension study (0.1-1 mg/kg). INTERVENTION: KRN23 was injected sc every 28 days. MAIN OUTCOME MEASURE: The main outcome measure was the proportion of subjects attaining normal serum Pi and safety. RESULTS: At baseline, mean TmP/GFR, serum Pi, and 1,25-dihydroxyvitamin D [1,25(OH)2D] were 1.6 ± 0.4 mg/dL, 1.9 ± 0.3 mg/dL, and 36.6 ± 14.3 pg/mL, respectively. During dose escalation, TmP/GFR, Pi, and 1,25(OH)2D increased, peaking at 7 days for TmP/GFR and Pi and at 3-7 days for 1,25(OH)2D, remaining above (TmP/GFR, Pi) or near [1,25(OH)2D] pre-dose levels at trough. After each of the four escalating doses, peak Pi was between 2.5 and 4.5 mg/dL in 14.8, 37.0, 74.1, and 88.5% of subjects, respectively. During the 12-month extension, peak Pi was in the normal range for 57.9-85.0% of subjects, and ≥25% maintained trough Pi levels within the normal range. Serum Pi did not exceed 4.5 mg/dL in any subject. Although 1,25(OH)2D levels increased transiently, mean serum and urinary calcium remained normal. KRN23 treatment increased biomarkers of skeletal turnover and had a favorable safety profile. CONCLUSIONS: Monthly KRN23 significantly increased serum Pi, TmP/GFR, and 1,25(OH)2D in all subjects. KRN23 has potential for effectively treating XLH.

Data integration reveals key homeostatic mechanisms following low dose radiation exposure.

The goal of this study was to define pathways regulated by low dose radiation to understand how biological systems respond to subtle perturbations in their environment and prioritize pathways for human health assessment. Using an in vitro 3-D human full thickness skin model, we have examined the temporal response of dermal and epidermal layers to 10 cGy X-ray using transcriptomic, proteomic, phosphoproteomic and metabolomic platforms. Bioinformatics analysis of each dataset independently revealed potential signaling mechanisms affected by low dose radiation, and integrating data shed additional insight into the mechanisms regulating low dose responses in human tissue. We examined direct interactions among datasets (top down approach) and defined several hubs as significant regulators, including transcription factors (YY1, MYC and CREB1), kinases (CDK2, PLK1) and a protease (MMP2). These data indicate a shift in response across time - with an increase in DNA repair, tissue remodeling and repression of cell proliferation acutely (24-72h). Pathway-based integration (bottom up approach) identified common molecular and pathway responses to low dose radiation, including oxidative stress, nitric oxide signaling and transcriptional regulation through the SP1 factor that would not have been identified by the individual data sets. Significant regulation of key downstream metabolites of nitrative stress was measured within these pathways. Among the features identified in our study, the regulation of MMP2 and SP1 was experimentally validated. Our results demonstrate the advantage of data integration to broadly define the pathways and networks that represent the mechanisms by which complex biological systems respond to perturbation.

Deficient expression of aldehyde dehydrogenase 1A1 is consistent with increased sensitivity of Gorlin syndrome patients to radiation carcinogenesis.

Human phenotypes that are highly susceptible to radiation carcinogenesis have been identified. Sensitive phenotypes often display robust regulation of molecular features that modify biological response, which can facilitate identification of the pathways/networks that contribute to pathophysiological outcomes. Here we interrogate primary dermal fibroblasts isolated from Gorlin syndrome patients (GDFs), who display a pronounced inducible tumorigenic response to radiation, in comparison to normal human dermal fibroblasts (NHDFs). Our approach exploits newly developed thiol reactive probes to define changes in protein thiol profiles in live cell studies, which minimizes artifacts associated with cell lysis. Redox probes revealed deficient expression of an apparent 55 kDa protein thiol in GDFs from independent Gorlin syndrome patients, compared with NHDFs. Proteomics tentatively identified this protein as aldehyde dehydrogenase 1A1 (ALDH1A1), a key enzyme regulating retinoic acid synthesis, and ALDH1A1 protein deficiency in GDFs was confirmed by Western blot. A number of additional protein thiol differences in GDFs were identified, including radiation responsive annexin family members and lamin A/C. Collectively, candidates identified in our study have plausible implications for radiation health effects and cancer susceptibility.

Impaired osteoblast differentiation in annexin A2- and -A5-deficient cells.

Annexins are a class of calcium-binding proteins with diverse functions in the regulation of lipid rafts, inflammation, fibrinolysis, transcriptional programming and ion transport. Within bone, they are well-characterized as components of mineralizing matrix vesicles, although little else is known as to their function during osteogenesis. We employed shRNA to generate annexin A2 (AnxA2)- or annexin A5 (AnxA5)-knockdown pre-osteoblasts, and determined whether proliferation or osteogenic differentiation was altered in knockdown cells, compared to pSiren (Si) controls. We report that DNA content, a marker of proliferation, was significantly reduced in both AnxA2 and AnxA5 knockdown cells. Alkaline phosphatase expression and activity were also suppressed in AnxA2- or AnxA5-knockdown after 14 days of culture. The pattern of osteogenic gene expression was altered in knockdown cells, with Col1a1 expressed more rapidly in knock-down cells, compared to pSiren. In contrast, Runx2, Ibsp, and Bglap all revealed decreased expression after 14 days of culture. In both AnxA2- and AnxA5-knockdown, interleukin-induced STAT6 signaling was markedly attenuated compared to pSiren controls. These data suggest that AnxA2 and AnxA5 can influence bone formation via regulation of osteoprogenitor proliferation, differentiation, and responsiveness to cytokines in addition to their well-studied function in matrix vesicles.

ERK oscillation-dependent gene expression patterns and deregulation by stress response.

Studies were undertaken to determine whether extracellular signal regulated kinase (ERK) oscillations regulate a unique subset of genes in human keratinocytes and subsequently whether the p38 stress response inhibits ERK oscillations. A DNA microarray identified many genes that were unique to ERK oscillations, and network reconstruction predicted an important role for the mediator complex subunit 1 (MED1) node in mediating ERK oscillation-dependent gene expression. Increased ERK-dependent phosphorylation of MED1 was observed in oscillating cells compared to nonoscillating counterparts as validation. Treatment of keratinocytes with a p38 inhibitor (SB203580) increased ERK oscillation amplitudes and MED1 and phospho-MED1 protein levels. Bromate is a probable human carcinogen that activates p38. Bromate inhibited ERK oscillations in human keratinocytes and JB6 cells and induced an increase in phospho-p38 and a decrease in phospho-MED1 protein levels. Treatment of normal rat kidney cells and primary salivary gland epithelial cells with bromate decreased phospho-MED1 levels in a reversible fashion upon treatment with p38 inhibitors (SB202190; SB203580). Our results indicate that oscillatory behavior in the ERK pathway alters homeostatic gene regulation patterns and that the cellular response to perturbation may manifest differently in oscillating vs nonoscillating cells.

Randomized trial of the anti-FGF23 antibody KRN23 in X-linked hypophosphatemia.

BACKGROUND: X-linked hypophosphatemia (XLH) is the most common heritable form of rickets and osteomalacia. XLH-associated mutations in phosphate-regulating endopeptidase (PHEX) result in elevated serum FGF23, decreased renal phosphate reabsorption, and low serum concentrations of phosphate (inorganic phosphorus, Pi) and 1,25-dihydroxyvitamin D [1,25(OH)2D]. KRN23 is a human anti-FGF23 antibody developed as a potential treatment for XLH. Here, we have assessed the safety, tolerability, pharmacokinetics (PK), pharmacodynamics (PD), and immunogenicity of KRN23 following a single i.v. or s.c. dose of KRN23 in adults with XLH. METHODS: Thirty-eight XLH patients were randomized to receive a single dose of KRN23 (0.003-0.3 mg/kg i.v. or 0.1-1 mg/kg s.c.) or placebo. PK, PD, immunogenicity, safety, and tolerability were assessed for up to 50 days. RESULTS: KRN23 significantly increased the maximum renal tubular threshold for phosphate reabsorption (TmP/GFR), serum Pi, and 1,25(OH)2D compared with that of placebo (P<0.01). The maximum serum Pi concentration occurred later following s.c. dosing (8-15 days) compared with that seen with i.v. dosing (0.5-4 days). The effect duration was dose related and persisted longer in patients who received s.c. administration. Changes from baseline in TmP/GFR, serum Pi, and serum 1,25(OH)2D correlated with serum KRN23 concentrations. The mean t1/2 of KRN23 was 8-12 days after i.v. administration and 13-19 days after s.c. administration. Patients did not exhibit increased nephrocalcinosis or develop hypercalciuria, hypercalcemia, anti-KRN23 antibodies, or elevated serum parathyroid hormone (PTH) or creatinine. CONCLUSION: KRN23 increased TmP/GFR, serum Pi, and serum 1,25(OH)2D. The positive effect of KR23 on serum Pi and its favorable safety profile suggest utility for KRN23 in XLH patients. Trial registration. Clinicaltrials.gov NCT00830674. Funding. Kyowa Hakko Kirin Pharma, Inc.

ALDH1A1 Deficiency in Gorlin Syndrome Suggests a Central Role for Retinoic Acid and ATM Deficits in Radiation Carcinogenesis.

We hypothesize that aldehyde dehydrogenase 1A1 (ALDH1A1) deficiency will result in impaired ataxia-telangiectasia mutated (ATM) activation in a retinoic acid-sensitive fashion. Data supporting this hypothesis include (1) reduced ATM activation in irradiated primary dermal fibroblasts from ALDH1A1-deficient Gorlin syndrome patients (GDFs), relative to ALDH1A1-positive normal human dermal fibroblasts (NHDFs) and (2) increased ATM activation by X-radiation in GDFs pretreated with retinoic acid, however, the impact of donor variability on ATM activation in fibroblasts was not assessed and is a prudent consideration in future studies. Clonogenic survival of irradiated cells showed differential responses to retinoic acid as a function of treatment time. Long-term (5 Day) retinoic acid treatment functioned as a radiosensitizer and was associated with downregulation of ATM protein levels. Short-term (7 h) retinoic acid treatment showed a trend toward increased survival of irradiated cells and did not downregulate ATM protein levels. Using a newly developed IncubATR technology, which defines changes in bulk chemical bond patterns in live cells, we can discriminate between the NHDF and GDF phenotypes, but treatment of GDFs with retinoic acid does not induce reversion of bulk chemical bond patterns associated with GDFs toward the NHDF phenotype. Collectively, our preliminary investigation of the Gorlin phenotype has identified deficient ALDH1A1 expression associated with deficient ATM activation as a possible susceptibility factor that is consistent with the high incidence of spontaneous and radiation-induced carcinogenesis in these patients. The IncubATR technology exhibits sufficient sensitivity to detect phenotypic differences in live cells that may be relevant to radiation health effects.

In situ live cell sensing of multiple nucleotides exploiting DNA/RNA aptamers and graphene oxide nanosheets.

Nucleotides, for example, adenosine-5'-triphosphate (ATP) and guanosine-5'-triphosphate (GTP), are primary energy resources for numerous reactions in organisms including microtubule assembly, insulin secretion, ion channel regulation, and so on. In order to advance our understanding of the production and consumption of nucleoside triphosphates, a versatile sensing platform for simultaneous visualization of ATP, GTP, adenosine derivates, and guanosine derivates in living cells has been built up in the present work based on graphene oxide nanosheets (GO-nS) and DNA/RNA aptamers. Taking advantage of the robust fluorescence quenching ability, unique adsorption for single-strand DNA/RNA probes, and efficient intracellular transport capacity of GO-nS, selective and sensitive visualization of multiple nucleoside triphosphates in living cells is successfully realized with the designed aptamer/GO-nS sensing platform. Moreover, GO-nS displays good biocompatibility to living cells and high protecting ability for DNA/RNA probes from enzymatic cleavage. These results demonstrate that the aptamers/GO-nS-based sensing platform is capable of selective, simultaneous, and in situ detection of multiple nucleotides, which hold a great potential for analyzing other biomolecules in living cells.