IL16 and factor V gene variations are associated with asparaginase-related thrombosis in childhood acute lymphoblastic leukemia patients.

Aim: We previously conducted exome-wide association study in acute lymphoblastic leukemia patients and identified association of five SNPs with asparaginase-related thrombosis. Here we aimed to replicate these findings in an independent patient cohort and through analyses in vitro. Patients & methods: SNPs located in IL16, MYBBP1A, PKD2L1, RIN3 and MPEG1 genes were analyzed in patients receiving Dana-Farber Cancer Institute acute lymphoblastic leukemia treatment protocols 05-001 and 11-001. Thrombophilia-related variations were also analysed. Results: IL16 rs11556218 conferred higher risk of thrombosis and higher in vitro sensitivity to asparaginase. The association was modulated by the treatment protocol, risk group and immunophenotype. A crosstalk between factor V Leiden, non-O blood groups and higher risk of thrombosis was also seen. Conclusion: IL16 and factor V Leiden variations are implicated in asparaginase-related thrombosis.

This study looked at how certain genetic variations are related to a higher risk of blood clots in children with a type of cancer called acute lymphoblastic leukemia who are receiving a certain treatment (asparaginase). The study found that one specific genetic variation (IL16 rs11556218) was linked to a higher risk of blood clots (thrombosis), and that this risk was influenced by disease and treatment features. The study also found that a certain genetic variation (factor V Leiden), which makes blood more likely to clot, and blood type (non-O) were linked to a higher risk of thrombosis. The conclusion of this study is that genetic variations may play a role in blood clots in children with acute lymphoblastic leukemia receiving asparaginase, and if further confirmed, these variations can serve to advance personalized treatment strategies.

Pharmacodynamic characterization of rytvela, a novel allosteric anti-inflammatory therapeutic, to prevent preterm birth and improve fetal and neonatal outcomes.

BACKGROUND: Preterm birth is the leading cause of neonatal morbidity and mortality. Studies have shown that interleukin 1 plays a major role in the pathophysiology of preterm birth by inducing the production of proinflammatory mediators and uterine activation proteins leading to labor. More importantly, uteroplacental inflammation, associated with preterm birth parturition pathways, is detrimental to fetal tissues and leads to long-term sequelae. Our group has developed an allosteric antagonist of the interleukin 1 receptor, rytvela, found to be potent and safe in preventing preterm birth by suppressing inflammation via the inhibition of the mitogen-activated protein kinase pathway while preserving the Nuclear factor kappa B pathway (important in immune vigilance). Rytvela has been shown to inhibit inflammatory up-regulation and uterine activation while preserving fetal development. OBJECTIVE: This study aimed to further the preclinical development of rytvela by evaluating its optimal dose and minimal duration of treatment to inhibit the inflammatory cascade, prolong gestation, and promote neonatal outcomes. STUDY DESIGN: Pregnant CD-1 mice were administered with lipopolysaccharide (10 µg, intraperitoneal administration) or interleukin 1 (1 µg/kg, intrauterine administration) on gestational day 16 to induce preterm labor. Rytvela was administered at different doses (0.1, 0.5, 1.0, 2.0, 4.0 mg/kg/d subcutaneously) from gestational days 16 to 18.5. To evaluate the minimal duration of treatment, the mice were administered with rytvela (2 mg/kg/d subcutaneously) for 24, 36, or 48 hours. The rate of prematurity (gestational day <18.5) and neonate survival and weight were evaluated. Gestational tissues were collected at gestational day 17.5 to quantify cytokines, proinflammatory mediators, and uterine activating proteins by real-time quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. The neonatal lungs and intestines were collected from postnatal days 5 to 7 and analyzed by histology. RESULTS: Rytvela exhibited a dose-response profile and achieved maximum efficacy at a dose of 2 mg/kg/d by reducing 70% of lipopolysaccharide-induced preterm births and 60% of interleukin 1ß-induced preterm births. In addition, rytvela attained maximum efficacy at a dose of 1 mg/kg/d by increasing neonate survival by up to 65% in both models of preterm birth. Rytvela protected fetuses from inflammatory insult as of 24 hours, preserving lung and intestinal integrity, and prevented preterm birth and fetal mortality by 60% and 50%, respectively, as of 36 hours of treatment. CONCLUSION: The maximum efficacy of rytvela was achieved at 2 mg/kg/d with improved birth outcomes and prevented inflammatory up-regulation upon 36 hours (only) of treatment. Rytvela exhibited desirable properties for the safe prevention of preterm birth and fetal protection.

The Succinate Receptor SUCNR1 Resides at the Endoplasmic Reticulum and Relocates to the Plasma Membrane in Hypoxic Conditions.

The GPCR SUCNR1/GPR91 exerts proangiogenesis upon stimulation with the Krebs cycle metabolite succinate. GPCR signaling depends on the surrounding environment and intracellular localization through location bias. Here, we show by microscopy and by cell fractionation that in neurons, SUCNR1 resides at the endoplasmic reticulum (ER), while being fully functional, as shown by calcium release and the induction of the expression of the proangiogenic gene for VEGFA. ER localization was found to depend upon N-glycosylation, particularly at position N8; the nonglycosylated mutant receptor localizes at the plasma membrane shuttled by RAB11. This SUCNR1 glycosylation is physiologically regulated, so that during hypoxic conditions, SUCNR1 is deglycosylated and relocates to the plasma membrane. Downstream signal transduction of SUCNR1 was found to activate the prostaglandin synthesis pathway through direct interaction with COX-2 at the ER; pharmacologic antagonism of the PGE2 EP4 receptor (localized at the nucleus) was found to prevent VEGFA expression. Concordantly, restoring the expression of SUCNR1 in the retina of SUCNR1-null mice renormalized vascularization; this effect is markedly diminished after transfection of the plasma membrane-localized SUCNR1 N8A mutant, emphasizing that ER localization of the succinate receptor is necessary for proper vascularization. These findings uncover an unprecedented physiologic process where GPCR resides at the ER for signaling function.

Performance of New Hypothermic Corneal Storage Media With an Antimycotic Tablet in Comparison to Traditional Hypothermic Media During Simulated Eye Bank Processing.

PURPOSE: To compare the performance of Kerasave (AL.CHI.MI.A. S.R.L., Ponte San Nicolò, Italy) containing 2.5 µg/mL of amphotericin B and Optisol-GS (Bausch & Lomb, Bridgewater, NJ) cold corneal storage media on donor corneas during routine eye bank procedures. METHODS: Forty-four paired donor corneas were preserved after swab sample collection and povidone-iodine decontamination. Right and left corneas were immersed in Kerasave and Optisol-GS, respectively, and stored at 4°C before the initial evaluation. Paired corneas were assigned to processing subgroups for penetrating keratoplasty (n = 20), Descemet stripping automated endothelial keratoplasty (n = 14), or Descemet membrane endothelial keratoplasty (n = 10). Endothelial cell density, central corneal thickness, slit-lamp examination, and endothelial cell damage were assessed at different intervals. Sterility testing was performed on media samples. RESULTS: At the initial evaluation, after 25.6 ± 3.2 hours of storage, the mean central corneal thickness of all corneas in Kerasave (n = 22) was greater than those in Optisol-GS (n = 22) (571 ± 12 µm vs. 526 ± 10 µm, respectively; P = 0.006). All other metrics were comparable between Kerasave and Optisol-GS in processing subgroups at all time intervals. Corneal swabs were positive in 90% of corneas before decontamination with povidone-iodine. At the initial evaluation, fungal contamination was detected in 24% and 19% of Kerasave and Optisol-GS, respectively. At the final evaluation, no fungi was detected in Kerasave and 1 Optisol-GS sample was positive (P = 0.999). CONCLUSIONS: Metrics of corneas stored in Kerasave and Optisol-GS were comparable. Kerasave might be considered an antifungal-possessing alternative to Optisol-GS.

The survival of patients enrolled in a global direct-to-patient cancer medicine donation program: The Glivec International Patient Assistance Program (GIPAP).

BACKGROUND: The Glivec International Patient Assistance Program (GIPAP) is a unique direct-to-patient program that provides imatinib (Glivec) at no cost to eligible patients in low- and middle-income countries (LMICs) with chronic myelogenous leukemia (CML) or gastrointestinal stromal tumor (GIST). This paper analyses the output, outcome and impact of the program between 2001 and 2014 using the data collected by the Max Foundation. METHOD: We extracted data on GIPAP patients' country of residence, sex, diagnosis, date of enrollment in GIPAP, age at enrollment, case closure date, and reason for closure from The Max Foundation database covering the period 2001 to 2014. We used Kaplan-Meier method to assess the survival rate of patients in GIPAP and used the proportional hazard regression model to estimate the effect of different variables on patients' survival. FINDINGS: About 63,000 GIPAP patients in 93 countries received over 71 million defined daily doses (DDD) of imatinib between 2001 and 2014. Our analysis showed that GIPAP patients had a 5-year survival rate of 89% which compares favorably to survival in high income countries despite the challenges of delivering cancer care in LMICs. Age at enrollment into the program, sex, duration between diagnosis and enrollment into program, year of enrollment, and patients' diagnosis (CML vs non-CML) were factors that influenced survival. INTERPRETATION: The GIPAP program has improved the survival of CML and GIST patients in LMICs, most of whom would not have had access to imatinib in the absence of the donation and therapeutic support of the program. FUNDING: This work was funded as part of Access Accelerated case studies. Access Accelerated is an initiative of more than 20 global biopharmaceutical companies in partnership with the World Bank and Union of International Cancer Control that seeks to reduce barriers to prevention, treatment and care for non-communicable diseases in LMICs.

Development of Real-Time Isothermal Amplification Assays for On-Site Detection of Phytophthora infestans in Potato Leaves.

Real-time loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) assays were developed targeting the internal transcribed spacer 2 region of the ribosomal DNA of Phytophthora infestans, the potato late blight causal agent. A rapid crude plant extract (CPE) preparation method from infected potato leaves was developed for on-site testing. The assay's specificity was tested using several species of Phytophthora and other potato fungal and oomycete pathogens. Both LAMP and RPA assays showed specificity to P. infestans but also to the closely related species P. andina, P. mirabilis, P. phaseoli, and P. ipomoeae, although the latter are not reported as potato pathogen species. No cross-reaction occurred with P. capsici or with the potato pathogens tested, including P. nicotianae and P. erythroseptica. The sensitivity was determined using P. infestans pure genomic DNA added into healthy CPE samples. Both LAMP and RPA assays detected DNA at 50 fg/µl and were insensitive to CPE inhibition. The isothermal assays were tested with artificially inoculated and naturally infected potato plants using a Smart-DART platform. The LAMP assay effectively detected P. infestans in symptomless potato leaves as soon as 24 h postinoculation. A rapid and accurate on-site detection of P. infestans in plant material using the LAMP assay will contribute to improved late blight diagnosis and early detection of infections and facilitate prompt management decisions.

Double-Pass Retina Point Imaging for the Evaluation of Optical Light Scatter, Retinal Image Quality, and Staging of Keratoconus.

PURPOSE: To measure retinal image quality using point spread function (PSF) analysis by double-pass retina point imaging in patients with keratoconus and to correlate visual quality with disease severity. METHODS: Patients diagnosed as having keratoconus by clinical examination, topography, and tomography and normal eyes were included in this study. A commercially available double-pass retina point imaging instrument (OQAS 108 II AcuTarget HD; Visiometrics S.L., Terrassa, Spain) was used to collect Objective Scatter Index (OSI) values in 21 keratoconic and 22 normal eyes. Eyes were also subjected to corneal topography and tomography, and staged using the Keratoconus Severity Score (KSS) and Amsler-Krumeich (AK) scales. RESULTS: The OSI was increased in keratoconic eyes (5.85 ± 0.98) versus control eyes (0.83 ± 0.12; mean ± SEM), in AK stages 1 to 4, and KSS stages 3 and 4. Receiver-operator characteristic analysis obtained an area under the curve (AUC) of 0.859 when evaluating the OSI as a unimodal diagnostic indicator for any KSS stage and 0.993 for KSS stages 3 and higher. An AUC of 0.949 was obtained in comparing eyes with lower severity topographic aberrations (KSS 1 and 2) versus mild to moderate keratoconus (KSS 3 and 4). Increasing corneal steepening patterns on tomography and topography were associated with PSF broadening and increased OSI. CONCLUSIONS: Double-pass retina point imaging is useful in correlating retinal image quality with keratoconus severity. The OSI may represent a clinically significant parameter for staging keratoconus with a unique ability to directly evaluate quality of vision in this population. [J Refract Surg. 2016;32(11):760-765.].

Modern laser in situ keratomileusis outcomes.

UNLABELLED: Laser in situ keratomileusis (LASIK) articles published between 2008 and 2015 that contain clinical outcomes data were reviewed and graded for quality, impression, and potential bias. All 97 relevant articles (representing 67 893 eyes) provided a positive or neutral impression of LASIK. Industry bias was not evident. The aggregate loss of 2 or more lines of corrected distance visual acuity was 0.61% (359/58 653). The overall percentage of eyes with uncorrected distance visual acuity better than 20/40 was 99.5% (59 503/59 825). The spherical equivalent refraction was within ±1.0 diopter (D) of the target refraction in 98.6% (59 476/60 329) of eyes, with 90.9% (59 954/65 974) within ±0.5 D. In studies reporting patient satisfaction, 1.2% (129/9726) of patients were dissatisfied with LASIK. Aggregate outcomes appear better than those reported in summaries of the safety and effectiveness of earlier laser refractive surgery systems approved by the U.S. Food and Drug Administration. Modern results support the safety, efficacy, and patient satisfaction of the procedure. FINANCIAL DISCLOSURE: Proprietary or commercial disclosures are listed after the references.

A pyrosequencing-based method to quantify genetic substitutions associated with resistance to succinate dehydrogenase inhibitor fungicides in Botrytis spp. populations.

BACKGROUND: The genetic underlying resistance mechanisms in the population of the phytopathogenic fungus Botrytis cinerea are well documented. Specifically, several genetic substitutions associated with succinate dehydrogenase inhibitor (SDHI)-based fungicide resistance have been identified in the succinate dehydrogenase gene. The objective of the present work was to develop a molecular tool for accurate quantification of these genetic substitutions within Botrytis populations. A test using the PyroMark Q24 instrument was designed to detect and quantify five genetic substitutions associated with SDHI resistance. RESULTS: The technique is based on sequencing by synthesis, and it generated quantitative and accurate data with a limit of quantification of a minimum of 500 spores. There was a linear relationship between the known and estimated percentages of spores with the targeted genetic substitutions and wild-type strains at ratios of 0-100%, with a 20% increment. CONCLUSION: With the pyrosequencing assay developed in this study, a large number of Botrytis spp. individuals can be characterised in a timely fashion with greater accuracy than by commonly used methods. Hence, pyrosequencing-based methods will be useful for improving our understanding of fungicide resistance, detecting the arrival of new genetic substitutions, monitoring shifts in fungal populations and assessing the effectiveness of antiresistance strategies, and for routine monitoring of fungicide resistance.

Infection Efficiency of Four Phytophthora infestans Clonal Lineages and DNA-Based Quantification of Sporangia.

The presence and abundance of pathogen inoculum is with host resistance and environmental conditions a key factor in epidemic development. Therefore, several spore-sampling devices have been proposed to monitor pathogen inoculum above fields. However, to make spore sampling more reliable as a management tool and to facilitate its adoption, information on infection efficiency and molecular tools for estimating airborne sporangia concentration are needed. Experiments were thus undertaken in a growth chamber to study the infection efficiency of four clonal lineages of P. infestans (US-8, US-11, US-23, and US-24) by measuring the airborne sporangia concentration and resulting disease intensity. The relationship between the airborne sporangia concentration and the number of lesions per leaf was exponential. For the same concentration, the sporangia of US-23 caused significantly more lesions than the sporangia of the other clonal lineages did. Under optimal conditions, an airborne sporangia concentration of 10 sporangia m-3 for US-23 was sufficient to cause one lesion per leaf, whereas for the other clonal lineages, it took 15 to 25 sporangia m-3 to reach the same disease intensity. However, in terms of diseased leaf area, there was no difference between clonal lineages US-8, US-23 and US-24. Also, a sensitive quantitative real-time polymerase chain reaction (qPCR) tool was developed to quantify P. infestans airborne sporangia with detection sensitivity of one sporangium. The specificity of the qPCR assay was rigorously tested for airborne inoculum and was either similar to, or an improvement on, other published PCR assays. This assay allows rapid and reliable detection and quantification of P. infestans airborne sporangia and thereby, facilitates the implementation of spores-sampling network.

Management of Botrytis Leaf Blight of Onion: The Québec Experience of 20 Years of Continual Improvement.

Botrytis leaf blight (BLB) of onion (Allium cepa) is caused by Botrytis squamosa. The disease has been reported on onion crops in several of the onion production areas of the world including North and South America, Europe, Asia, and Australia, although it is not a problem in arid production regions such as the western United States. In eastern Canada, the disease is generally present every year and is especially severe on cultivars of yellow globe onion. The pathogen biology and disease epidemiology have been intensively researched. Over the last few decades, in the organic soil area of Quebec, extensive research effort has been devoted to the development and evaluation of predictive models and disease management strategies. There has been an active integrated pest management program for onions since the early 1980s, and scouting for disease has played a major role in disease management. In this article, the story of BLB management in eastern Canada over a period of two decades is summarized.