Intratumoral IL12 mRNA Therapy Promotes TH1 Transformation of the Tumor Microenvironment.

PURPOSE: While immune checkpoint inhibitors such as anti-PD-L1 are rapidly becoming the standard of care in the treatment of many cancers, only a subset of treated patients have long-term responses. IL12 promotes antitumor immunity in mouse models; however, systemic recombinant IL12 had significant toxicity and limited efficacy in early clinical trials. EXPERIMENTAL DESIGN: We therefore designed a novel intratumoral IL12 mRNA therapy to promote local IL12 tumor production while mitigating systemic effects. RESULTS: A single intratumoral dose of mouse (m)IL12 mRNA induced IFNγ and CD8+ T-cell-dependent tumor regression in multiple syngeneic mouse models, and animals with a complete response demonstrated immunity to rechallenge. Antitumor activity of mIL12 mRNA did not require NK and NKT cells. mIL12 mRNA antitumor activity correlated with TH1 tumor microenvironment (TME) transformation. In a PD-L1 blockade monotherapy-resistant model, antitumor immunity induced by mIL12 mRNA was enhanced by anti-PD-L1. mIL12 mRNA also drove regression of uninjected distal lesions, and anti-PD-L1 potentiated this response. Importantly, intratumoral delivery of mRNA encoding membrane-tethered mIL12 also drove rejection of uninjected lesions with very limited circulating IL12p70, supporting the hypothesis that local IL12 could induce a systemic antitumor immune response against distal lesions. Furthermore, in ex vivo patient tumor slice cultures, human IL12 mRNA (MEDI1191) induced dose-dependent IL12 production, downstream IFNγ expression and TH1 gene expression. CONCLUSIONS: These data demonstrate the potential for intratumorally delivered IL12 mRNA to promote TH1 TME transformation and robust antitumor immunity.See related commentary by Cirella et al., p. 6080.

Automated leukocyte processing by microfluidic deterministic lateral displacement.

We previously developed a Deterministic Lateral Displacement (DLD) microfluidic method in silicon to separate cells of various sizes from blood (Davis et al., Proc Natl Acad Sci 2006;103:14779-14784; Huang et al., Science 2004;304:987-990). Here, we present the reduction-to-practice of this technology with a commercially produced, high precision plastic microfluidic chip-based device designed for automated preparation of human leukocytes (white blood cells; WBCs) for flow cytometry, without centrifugation or manual handling of samples. After a human blood sample was incubated with fluorochrome-conjugated monoclonal antibodies (mAbs), the mixture was input to a DLD microfluidic chip (microchip) where it was driven through a micropost array designed to deflect WBCs via DLD on the basis of cell size from the Input flow stream into a buffer stream, thus separating WBCs and any larger cells from smaller cells and particles and washing them simultaneously. We developed a microfluidic cell processing protocol that recovered 88% (average) of input WBCs and removed 99.985% (average) of Input erythrocytes (red blood cells) and >99% of unbound mAb in 18 min (average). Flow cytometric evaluation of the microchip Product, with no further processing, lysis or centrifugation, revealed excellent forward and side light scattering and fluorescence characteristics of immunolabeled WBCs. These results indicate that cost-effective plastic DLD microchips can speed and automate leukocyte processing for high quality flow cytometry analysis, and suggest their utility for multiple other research and clinical applications involving enrichment or depletion of common or rare cell types from blood or tissue samples. © 2016 International Society for Advancement of Cytometry.

Artemisinin-derived dimer ART-838 potently inhibited human acute leukemias, persisted in vivo, and synergized with antileukemic drugs.

Artemisinins, endoperoxide-containing molecules, best known as antimalarials, have potent antineoplastic activity. The established antimalarial, artesunate (AS), and the novel artemisinin-derived trioxane diphenylphosphate dimer 838 (ART-838) inhibited growth of all 23 tested acute leukemia cell lines, reduced cell proliferation and clonogenicity, induced apoptosis, and increased intracellular levels of reactive oxygen species (ROS). ART-838 was 88-fold more potent that AS in vitro, inhibiting all leukemia cell lines at submicromolar concentrations. Both ART-838 and AS cooperated with several established antileukemic drugs and newer kinase inhibitors to inhibit leukemia cell growth. ART-838 had a longer plasma half-life than AS in immunodeficient NOD-SCID-IL2Rgnull (NSG) mice, remaining at effective antileukemic concentrations for >8h. Intermittent cycles of ART-838 inhibited growth of acute leukemia xenografts and primagrafts in NSG mice, at higher potency than AS. Based on these preclinical data, we propose that AS, with its established low toxicity and low cost, and ART-838, with its higher potency and longer persistence in vivo, should be further developed toward integration into antileukemic regimens.

A logically derived diet to stop MS relapses.

This brief document outlines reasons for expecting that diets which cause gout should help prevent multiple sclerosis relapses.

Endocrine and phototransduction effects in the prevention of multiple sclerosis.

In addressing the question of what seems to prevent multiple sclerosis (MS) in the tropics this paper reviews work done by various researchers and suggests that MS incidence may be affected in the ensemble by the endocrine system's response to environment temperature, the skin's response to sunlight, and by the retina's response to brightness. It shows how the hypouricemia which is a reliable indicator in MS patients can leave the blood-brain barrier unsealed in general but allow retinoids to block their access to the central nervous system. It presents published studies as evidence and suggests a number of straightforward tests of these theories which could allow clinicians to advise their MS patients to take appropriate actions to help slow down or prevent disease progression.

Endocrine system dynamics and MS epidemiology.

In the kidney there is a co-transport relationship in the nephron between the reabsorption of positive Na(+) ions and the reabsorption of negative ions such as uric acid anions. Uric acid acts as an anti-oxidant and it has been shown to have a sealing effect on the blood-brain barrier. The theory developed here is that chronic neurological vasoconstriction in cool environmental conditions injects an offset into the rennin-angiotensin-aldosterone system (RAAS) blood pressure control loop and reduces demand for angiotensin and aldosterone. (Aldosterone is produced in the adrenal gland and has a direct effect on renal reabsorption of Na(+) ions.) Via co-transport these conditions will reduce the body's ability to reabsorb uric acid and this in turn will weaken the integrity of the blood-brain barrier. Also, in cool environments, where levels of vasopressin (ADH) and aldosterone are lower, the gain of the hypothalamus-pituitary-adrenal gland (HPA) axis is reduced so that the production average levels of ACTH, cortisone and aldosterone will be biased at a lower level and the kidney-local levels of aldosterone in particular will remain lower. This paper develops these ideas and suggests that they can help explain the traditionally-recognized latitudinal gradient in MS epidemiology. Also, acclimatization to heat encourages sweating, which should create a greater demand for the renal reabsorption of Na(+) ions which enables greater reabsorption of uric acid. Therefore people living at low latitudes should have a lower chance of hypouricemia and a lower chance of developing MS. In fact people who spend their first fifteen years in the tropics almost never go onto develop MS. And MS patients in relapse are consistently hypouricemic. This hypothesis can explain both of these facts. The paper goes onto show how the MS condition will tend to progress because of a number of self-sustaining effects: over time the immune system becomes more targeted to myelin, MS patients are unlikely to become acclimatized to heat because they tend to avoid heat since demyelinated nerve function is worsened by elevated temperature, and the normal circadian excitation of the HPA axis gets weaker under the benign environmental conditions typically adopted by MS patients as the disease develops.

Linking technology with processes for payment collection.

To improve patient payment collections, providers should: Assess and improve communications with patients regarding personal out-of-pocket healthcare expenses at appropriate times within the revenue cycle. Use the optimal technology to access payment networks and the banking system. Develop and implement a payment training program for patient access associates and other appropriate staff.

Adjunctive atropine is unnecessary during ketamine sedation in children.

BACKGROUND: The prophylactic coadministration of atropine or other anticholinergics during dissociative sedation has historically been considered mandatory to mitigate ketamine-associated hypersalivation. Emergency physicians (EPs) are known to omit this adjunct, so a prospective study to describe the safety profile of this practice was initiated. OBJECTIVES: To quantify the magnitude of excessive salivation, describe interventions for hypersalivation, and describe any associated airway complications. METHODS: In this prospective observational study of emergency department (ED) pediatric patients receiving dissociative sedation, treating physicians rated excessive salivation on a 100-mm visual analog scale and recorded the frequency and nature of airway complications and interventions for hypersalivation. RESULTS: Of 1,090 ketamine sedations during the 3-year study period, 947 (86.9%) were performed without adjunctive atropine. Treating physicians assigned the majority (92%) of these subjects salivation visual analog scale ratings of 0 mm, i.e., "none," and only 1.3% of ratings were >or= 50 mm. Transient airway complications occurred in 3.2%, with just one (brief desaturation) felt related to hypersalivation (incidence 0.11%, 95% confidence interval = 0.003% to 0.59%). Interventions for hypersalivation (most commonly suctioning) occurred in 4.2%, with no occurrences of assisted ventilation or intubation. CONCLUSIONS: When adjunctive atropine is omitted during ketamine sedation in children, excessive salivation is uncommon, and associated airway complications are rare. Anticholinergic prophylaxis is not routinely necessary in this setting.

Cardiac troponin I as a predictor of mortality for pediatric submersion injuries requiring out-of-hospital cardiopulmonary resuscitation.

BACKGROUND: It is difficult to predict ultimate survivors to hospital discharge in children who are successfully resuscitated after a cardiorespiratory arrest associated with a submersion injury. Serum measurements of organ injury or dysfunction may serve as a surrogate marker of the degree of hypoxic injury. We designed a prospective study whose purpose was to assess the predictive value for outcome of serum cardiac troponin I measurements after submersion injury and cardiorespiratory arrest. METHODS: This is a prospective, observational study of children admitted to a postintensive care unit after experiencing an out-of-hospital cardiorespiratory arrest associated with a submersion event. Cardiac troponin I measurements were examined upon admission to the postoperative intensive care unit after successful emergency department resuscitation. RESULTS: Nine patients were admitted, and 2 patients (22%) survived to hospital discharge. The area under the receiver operating characteristic curve is 0.786 (95% confidence interval, 0.481-1.0). This suggests that cardiac troponin I has a moderate degree of discriminatory power in selecting children who did not survive to hospital discharge.

Signs and symptoms of cerebrospinal fluid shunt malfunction in the pediatric emergency department.

OBJECTIVES: Pediatric patients with cerebrospinal fluid shunts frequently present to the emergency department for evaluation of possible shunt malfunction. Most shunt studies appear in the neurosurgical literature. To our knowledge, none have reviewed presenting signs and symptoms of shunt malfunction in patients who present to the pediatric emergency department. The study objective was to evaluate the medical record of children with cerebrospinal fluid shunts who presented to a pediatric emergency department to determine if any signs and/or symptoms were predictive of shunt malfunction. METHODS: A retrospective chart review was conducted on 352 pediatric patients aged 0 to 18 years, who presented to the pediatric emergency department between January 1, 1998, and December 31, 2002, with signs and/or symptoms that prompted an evaluation for possible shunt malfunction. RESULTS: Univariate analysis of all signs and symptoms revealed lethargy (odds ratio, 1.99; 95% confidence interval, 1.15-3.42; P = 0.02) and shunt site swelling (odds ratio, 2.56, 95% confidence interval, 1.08-6.07, P = 0.03) to be significantly predictive of shunt malfunction. Logistic regression analysis continued to show significance for lethargy (odds ratio, 2.20; bias-corrected 95% confidence interval, 1.11-3.63) and shunt site swelling (odds ratio, 3.10; bias-corrected 95% confidence interval, 1.38-9.05), but found no other study variable to be significant. Bootstrap resampling validated the importance of the significant variables identified in the regression analysis. CONCLUSIONS: In this study, lethargy and shunt site swelling were predictive of shunt malfunction. Other signs and symptoms studied did not reach statistical significance; however, one must maintain a high index of suspicion when evaluating children with an intracranial shunt because the presentation of malfunction is widely varied. A missed diagnosis can result in permanent neurological sequelae or even death.

Rate of palivizumab administration in accordance with current recommendations among hospitalized children.

INTRODUCTION: The American Academy of Pediatrics recommends palivizumab prophylaxis for children born premature or with chronic lung disease to reduce the severity of respiratory syncytial virus (RSV) infection. The purpose of the current study is to examine palivizumab administration among children hospitalized with RSV infection. METHOD: This is a retrospective medical record review at a tertiary care children's hospital. The study included children <2 years of age hospitalized between October 1, 2000, and April 1, 2001, with the diagnosis of RSV infection. RESULTS: The records of 264 children meeting inclusion criteria were reviewed. Forty children qualified for administration of palivizumab using American Academy of Pediatrics recommendations. Of these, 14 (35%) received palivizumab prior to admission. Palivizumab administration rate was not affected by age, race, or insurance coverage. DISCUSSION: Exact barriers to the administration of palivizumab remain unclear. The identification of high-risk children, prevention of RSV by use of palivizumab, and collaboration between hospital and community health care providers will help increase the use of palivizumab and decrease the incidence of RSV.

The removal of coins from the upper esophageal tract of children by emergency physicians: a pilot study.

OBJECTIVE: There are few reports in the medical literature describing removal of a coin from the upper esophageal tract of a child by an emergency physician. However, given the nature of their training and practice, emergency physicians are well suited to perform this common procedure. We describe our experience with this procedure. METHODS: This was a retrospective review of a continuous quality improvement data set from a university-based tertiary care pediatric emergency department between Nov. 1, 2003, and Mar. 31, 2004. RESULTS: Thirteen children, with a median age of 20 months, underwent rapid sequence intubation and had coins successfully removed from their upper esophageal tract by emergency physicians. In 10 cases, the coin was visible at laryngoscopy and removed with Magill forceps. In 3 cases this approach failed and a Foley catheter was used to remove the coin. One child suffered a tonsillar abrasion and two sustained minor lip trauma, but all were extubated and discharged home from the emergency department with no significant complications. Eleven of the 13 patients were successfully followed up, and the parents reported no problems. CONCLUSIONS: This pilot study suggests that the removal of a coin from the upper esophageal tract by an emergency physician can be both safe and effective. A larger study is needed before this procedure can be generally recommended.

Investigation of afebrile neonates with a history of fever.

OBJECTIVE: Our objective was to describe clinically significant infections in a cohort of afebrile neonates who underwent an emergency department (ED) septic workup because of the history of a measured fever at home. METHODS: Retrospective medical record review of all infants (3/4)28 days of age who presented to our tertiary care pediatric ED between Jan. 1, 1999, and Aug. 22, 2002, underwent lumbar puncture in the ED, had a reported temperature at home of >or=38 degrees C, and an ED triage temperature of <38 degrees C. Laboratory and radiographic results were tabulated. RESULTS: During the study period, 206 neonates underwent lumbar puncture in our ED. Of these, 108 were excluded because their home temperature was not documented, and 71 were excluded because they were still febrile on presentation to the ED. The study group consisted of the remaining 27 subjects, 4 of whom had received acetaminophen prior to ED arrival. Infections were confirmed in 10 (37%) subjects (3 urinary tract infections, 2 aseptic meningitis, 1 enterovirus meningitis, 1 respiratory syncytial virus bronchiolitis, 1 rotavirus enteritis and 2 pneumonias). CONCLUSIONS: Clinically important infections are not uncommon among afebrile neonates undergoing ED septic workup because of a measured fever at home. Some diagnostic testing is warranted in this group, although the clinical utility and indications for specific test modalities remain unclear.

Blunt pediatric head trauma requiring neurosurgical intervention: how subtle can it be?

Recent literature on pediatric head injuries has suggested that important intracranial injuries might present to the ED without typical signs or symptoms. The objective of our study was to review our institutional experience with head-injured infants and young children to assess the subtlety of the ED presentation. We performed a retrospective medical record review of head-injured children

Cardiac troponin I as a predictor of respiratory failure in children hospitalized with respiratory syncytial virus (RSV) infections: a pilot study.

This pilot study was performed to assess the use of cardiac troponin I to predict respiratory failure in children admitted to the hospital with respiratory syncytial virus (RSV) infections. We enrolled a prospective convenience sample of children under 5 years of age who were admitted to our university-based, tertiary care children's hospital from December 1, 2000, to February 1, 2002, with RSV infections. A cardiac troponin I was drawn at admission. We assessed the test characteristics for positive cardiac troponin I (defined as >0.3 ng/mL, the manufacturer's upper limit of normal) in correctly identifying children who had respiratory failure as evidenced by the need for endotracheal intubation. Twenty-five children from 9 days to 38 months of age were included. Ten children had a positive cardiac troponin I and 3 of these children had respiratory failure. A positive cardiac troponin I demonstrated a sensitivity of 100%, specificity 68%, positive predictive value of 30%, negative predictive value of 100%, and accuracy of 72%. The area under the receiver operating characteristic curve was 0.939 (95% confidence interval, 0.820-1.0), suggesting a high degree of discriminatory power in selecting children with respiratory failure. A sample size calculation revealed that a follow-up study of 359 patients is needed before the clinical use of cardiac troponin I for this purpose.

Myocardial injury in children following resuscitation after cardiac arrest.

BACKGROUND: Myocardial dysfunction occurs immediately after successful cardiac resuscitation. Our purpose was to determine whether measurement of cardiac troponin I in children with acute out-of-hospital cardiac arrest predicts the severity of myocardial injury. METHODS AND RESULTS: This prospective, observational study was performed in the Pediatric Intensive Care Unit (PICU) on 24 patients following arrest, ranging in age from 8 months to 17 years. Troponin measurements were obtained on admission, and at 12, 24, and 48 h. Transthoracic echocardiograms were performed within 24 h after admission. Survival to hospital discharge was 29% (7/24). The mean age was 5.9+/-4.6 years for survivors and 4.2+/-5.3 years for non-survivors. The median (range) duration of cardiac arrest times for survivors was 6 min (3 to 63 min) versus 34 min (4 to 70 min) for nonsurvivors (P=0.02). Survivors received 1.3+/-2.2 doses of epinephrine (adrenaline) compared with 2.9+/-1.6 doses for non-survivors (P=0.02). Only one patient had ventricular fibrillation and defibrillation was unsuccessful. The ejection fraction for survivors averaged 73.2+/-11.2%, but for nonsurvivors only 55.4+/-19.8% (P=0.04). Ejection fraction correlated inversely with troponin at 12 h (r=-0.54, P=0.01) and at 24 h (r=-0.59, P=0.02). Circumferential fiber shortening for survivors was 37.5+/-7.8 and 25.5+/-10.7% for nonsurvivors (P=0.02). It also correlated inversely with troponin (r=-0.46, P=0.03 for survivors and r=-0.65, P=0.01, for nonsurvivors). CONCLUSION: After cardiac arrest and resuscitation in pediatric patients, the severity of myocardial dysfunction was reflected in troponin I levels.

Crotaline Fab antivenom for the treatment of children with rattlesnake envenomation.

OBJECTIVE: There is little data regarding safety or efficacy of Crotalinae polyvalent immune Fab (ovine) antivenom (crotaline Fab) in pediatric patients. Our objective was to provide the first information regarding safety and effectiveness of this new drug in children. METHODS: Data were collected prospectively and retrospectively for all consecutive pediatric patients bitten by rattlesnakes and treated at 2 urban hospitals during 2001. Cases were included if there were signs of envenomation at presentation, patient age was 13 years or less, and there was administration of crotaline Fab. Cases were excluded if antivenin (Crotalidae) polyvalent (equine origin, the conventional antivenom) was given. Primary outcome variables were snakebite severity scores throughout the course of therapy, number of vials of crotaline Fab administered, occurrence of allergic reactions, adjunct surgical therapy, and the presence of permanent sequelae or serum sickness identified at follow-up. RESULTS: In the 12 cases studied, ages ranged from 14 months to 13 years (mean: 6.9; standard deviation: 4.2). Presentation snakebite severity scores ranged from 2 to 9 (mean: 5.3; standard deviation: 2.3). Total crotaline Fab doses ranged from 4 to 22 vials (mean: 12.7; standard deviation: 5.4). Initial control of symptoms was achieved with 4 to 16 vials (mean: 7.7; standard deviation: 3.7), and severity scores stabilized or improved within 24 hours in all patients. Recurrence of local swelling occurred in 1 case despite scheduled repeat doses of antivenom. No cases required surgical intervention, and no permanent sequelae were identified. No immediate or delayed hypersensitivity reactions occurred. CONCLUSION: In this group of pediatric patients treated for rattlesnake envenomation, crotaline Fab antivenom was safe and seemed to be effective.

Crotalidae polyvalent immune Fab (ovine) antivenom is efficacious for envenomations by Southern Pacific rattlesnakes (Crotalus helleri).

STUDY OBJECTIVE: Southern Pacific rattlesnake (Crotalus helleri ) venom is not 1 of the 4 venoms used to produce Crotalidae polyvalent immune Fab (ovine) (CroFab; FabAV). There is currently no published clinical experience regarding the efficacy of this new antivenom for confirmed C helleri envenomation, and animal data suggest greatly diminished efficacy. We assessed the efficacy of FabAV for patients with confirmed C helleri envenomation. METHODS: We conducted a prospective observational study of 23 consecutive rattlesnake envenomations that were treated with FabAV at our center. Patients were excluded if the species of snake could not be confirmed, if FabAV antivenom was not given, or if Antivenin (Crotalidae) polyvalent (equine) was given. We collected serial physical examination and laboratory data over a 24-hour period to serially evaluate the severity score and performed follow-up to evaluate delayed reactions. RESULTS: There were 15 patients who received FabAV and had the species of rattlesnake confirmed (9 C helleri, 4 C scutulatus scutulatus, 1 C mitchellii pyrrhus, 1 C ruber ruber ). C helleri envenomations demonstrated similar improvement in serial snakebite severity scores to those of other species. Three patients treated with scheduled dosing had recurrence of progressive swelling (2 C helleri and 1 C mitchellii pyrrhus ) during the 24-hour study period. CONCLUSION: We observed similar improvement in FabAV-treated patients with C helleri envenomation compared with those of other species and conclude that this treatment in standard doses appears efficacious for bites by this species. Progressive swelling may recur despite scheduled dosing.