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Heart and blood vessel disorders, such as coronary heart disease, brain vessel disease, rheumatic heart disease, and others, are together referred to as cardiovascular disease (CVD). In this study, we sought to determine how mitochondrial Leucine Transfer RNA genes and CVDs are related (MT-L1 and MT-L2). From CVD patients in Peshawar, a total of 27 saliva samples were taken. Leu-tRNA genes expressed by mitochondria were amplified using polymerase chain reaction after DNA was removed. Ten samples were sent for sequencing after PCR and gene cleaning. We obtained all of the sequenced results, which were subsequently aligned and evaluated against the mitochondrial revised Cambridge Reference Sequence (rCRS). However, in our sequenced samples, Leu-tRNA MT-L1 and MT-L2 genes were determined to be unaltered. Thus, it is suggested that a large population be taken into account while screening for mutations in the mitochondrial encoded Leu-tRNA MT-L1 and MT-L2 genes of cardiac patients in areas of Pakistan. Additionally, it is recommended that patients with cardiac problems should also have other mitochondrial encoded genes checked for potential mutations. This could result in the identification of genetic markers that could be used for early CVD screening in Pakistan. Resumo Distúrbios do coração e dos vasos sanguíneos, como doença cardíaca coronária, doença dos vasos cerebrais, doença cardíaca reumática entre outros, são referidos juntos como doença cardiovascular (DCV). Neste estudo, procuramos determinar como os genes mitocondriais do RNA de transferência de leucina e as DCVs estão relacionados (MT-L1 e MT-L2). Foi coletado um total de 27 amostras de saliva de pacientes com DCV em Peshawar. Genes de Leu-tRNA expressos por mitocôndrias foram amplificados usando reação em cadeia da polimerase (PCR) após a remoção do DNA. Dez amostras foram enviadas para sequenciamento após PCR e limpeza gênica. Obtivemos todos os resultados sequenciados, que foram posteriormente alinhados e avaliados em comparação com a Sequência de Referência de Cambridge revisada (rCRS). No entanto, em nossas amostras sequenciadas, os genes Leu-tRNA MT-L1 e MT-L2 foram determinados como inalterados. Assim, sugere-se que uma grande população seja levada em consideração durante a triagem de mutações nos genes Leu-tRNA MT-L1 e MT-L2 mitocondriais codificados de pacientes cardíacos em áreas do Paquistão. Além disso, recomenda-se que outros genes mitocondriais codificados de pacientes com problemas cardíacos também sejam verificados quanto a possíveis mutações. Isso pode resultar na identificação de marcadores genéticos que podem ser usados para triagem precoce de DCV no Paquistão.

Life cycle assessment was carried out for a conventional wooden furniture set produced in Mardan division of the Khyber Pakhtunkhwa province of Pakistan during 2018-19. Primary data regarding inputs and outputs were collected through questionnaire surveys from 100 conventional wooden furniture set manufacturers, 50 in district Mardan and 50 in district Swabi. In the present study, cradle-to-gate life cycle assessment approach was applied for a functional unit of one conventional wooden furniture set. Production weighted average data were modelled in the environmental impacts modelling software i.e., SimaPro v.8.5. The results showed that textile used in sofa set, wood preservative for polishing and preventing insects attack and petrol used in generator had the highest contribution to all the environmental impact categories evaluated. Total cumulative energy demand for wooden furniture set manufactured was 30,005 MJ with most of the energy acquired from non-renewable fossil fuel resources. Resumo A abordagem de avaliação do ciclo de vida foi realizada para um conjunto de móveis de madeira convencional produzido na divisão Mardan da província de Khyber Pakhtunkhwa do Paquistão durante 2018-19. Os dados primários sobre entradas e saídas foram coletados por meio de pesquisas por questionário de 100 fabricantes de conjuntos de móveis de madeira convencionais, 50 no distrito de Mardan e 50 no distrito de Swabi. No presente estudo, a abordagem de avaliação do ciclo de vida do berço ao portão foi aplicada para uma unidade funcional de um conjunto de móveis de madeira convencional. Os dados da média ponderada da produção foram modelados no software de modelagem de impactos ambientais, isto é, SimaPro v.8.5. Os resultados mostraram que os têxteis usados no conjunto de sofás, o preservativo de madeira para polir e prevenir o ataque de insetos e a gasolina usada no gerador tiveram a maior contribuição em todas as categorias de impacto ambiental avaliadas. A demanda total acumulada de energia para o conjunto de móveis de madeira fabricado foi de 30.005 MJ, com a maior parte da energia adquirida de recursos de combustíveis fósseis não renováveis.

Due to extensive application of antibiotics as growth promoters in animal feed, antimicrobial resistance has been increased. To overcome this challenge, rumen microbiologists search for new probiotics to improve the rate of livestock production. The present study was aimed to isolate and evaluate breed-specific lactic acid bacteria (LAB) as potential animal probiotics. The current study was conducted during 10 months from July 2020 to April 2021, in which a total of n=12 strains were isolated from different samples including milk, rumen, and feces of Nilli Ravi Buffaloes. These isolates were evaluated for their antimicrobial potential against common animal pathogens (Bacillus spp., E. coli, Staphylococcus aureus, Salmonella spp., Listeria spp.). All the isolates were identified using 16S rRNA gene sequencing and the phylogenetic analyses inferred that these strains showed close relations to the species of various genera; Enterococcus lactis, Pediococcus pentosaceus, Bacillus subtilis Weissella cibaria, Weissella soli, Bacillus tequilensis, Weissella bombi, Bacillus licheniformis, Lactococcus lactis, Bacillus megaterium, Lactobacillus ruminis, and Lactococcus lactis. NMCC-Ru2 has exhibited the enormous potential of antimicrobial activity, 28 mm, for Salmonella typhimurium;23 mm for Listeria monocytogenes 21 mm for E.coil. Highest resistance was seen in NMCC-Ru2 agasint test antbiotic, like 25.5 mm for Tetracycline. Overall results revesl that the probiotic profile of isolates was achieved using standard criteria, particularly with animal probiotic properties Resumo Devido à extensa aplicação de antibióticos como promotores de crescimento na alimentação animal, a resistência aos antimicrobianos aumentou. Para superar esse desafio, os microbiologistas do rúmen buscam novos probióticos para melhorar a produtividade do gado. O presente estudo teve como objetivo isolar e avaliar bactérias lácticas específicas de raças (BAL) como potenciais probióticos animais. 12 cepas foram isoladas de diferentes amostras, incluindo leite, rúmen e fezes de búfalos Nilli Ravi. Esses isolados foram avaliados quanto ao seu potencial antimicrobiano contra patógenos animais comuns (Bacillus spp., E. coli, Staphylococcus aureus, Salmonella spp., Listeria spp.). Todos os isolados foram identificados por meio do sequenciamento do gene 16S rRNA e as análises filogenéticas inferiram que essas cepas apresentaram estreita relação com as espécies de vários gêneros; Enterococcus lactis, Pediococcus pentosaceus, Bacillus subtilis, Weissella cibaria, Weissella soli, Bacillus tequilensis, Weissella bombi, Bacillus licheniformis, Lactococcus lactis, Bacillus megaterium, Lactobacillus ruminis e Lactococcus lactis. O perfil probiótico dos isolados foi obtido usando critérios padrão, particularmente com propriedades probióticas animais.

Background: Sex hormones play an important role in uterus and bone tissue integration. Oral hormonal contraceptive pills (OCPs) were developed over 50 years ago. Aims of the study: to clarify the histological evaluation of different types of hormonal contraceptive effects on uterus, cervix, ovary, endometrium, and bone in the female albino rat. Materials and methods: The study was conducted on thirty albino rats. three groups each group 10 rats one of them considered as a control group and the other two groups considered as a treated group which taking oral hormonal contraceptive once daily for about 8 weeks and another group takes injectable hormonal contraceptive once weekly for about 8 weeks then euthanized for histological examination. Result: After 8 weeks of drug administration histological findings show significant differences in the thickness of articular cartilage, the thickness of metaphyseal plate, the thickness of trabecular bone, and thickness of endometrium between the control group and experimental group. Conclusion: The current study showed that hormonal contraceptives exhibited histological changes on the uterus and bone which can be correlated to different clinical findings.

The present study conducted to study epipelic diatoms in the Tigris River within Wasit province for one year from June 2015 to May 2016, five sites have been chosen along the river between Al-Aziziyah and Kut .The aim of the study was to identify and main distributed to them in the Tigris River data were collected by the qualitative and quantitative study of epipelic diatoms. A total number of 161 species of epipelic diatoms was recorded belongs to 36 genera, where the number of central diatoms species recorded were 14 diatoms belonged to 7 genera by (8.69%) and The pinnate diatoms species were 147 species belonging to 27 genera by (91.30%). Seasonal and spatial variations were observed in the total numbers of benthic diatom during the study period, the results recorded the lowest rate of both central and pinnate diatoms were (745.29× 104/cm3) in site1 and the highest rate (1122.92 × 104 /cm3) in site4.Some genera have recorded higher number species during the study period; these genera were Nitzschia, Navicula, Cymbella, Gomphonema, Amphora, Cyclotella, Melosira, Cocconies, Caloneis, Diatoma, Fragiaria, Surirella and Synedra. The study revealed that Bacillariophyceae were abundant, reflecting the quality of water and mud and determine the extent of pollution and polluted type

Context Total pancreatectomy with islet autotransplantation (TPIAT) is a definitive management for intractable pain in patients with chronic pancreatitis (CP). Islet autotransplantation (IAT) allows for the preservation of beta cells to prevent complications of long-term diabetes. Objective Our study follows TPIAT recipients for up to 12 years to determine the efficacy of the procedure completed with an off-site islet isolation facility. Methods Patient demographics, mixed meal tolerance test measures, glycosylated hemoglobin, insulin requirements, and homeostatic model assessment for insulin resistance values were collected prior to surgery and at the most recent follow-up assessment. Results Forty-four patients (median age, 46.0 years; range, 20-78 years) underwent TPIAT for CP. At an overall median follow-up time of 845.5 days (range, 195-4470 days) 8 patients were insulin independent and 36 patients were insulin dependent. At the most recent follow-up time point, islet yield per kilogram was the strongest indicator of insulin independence. Homeostatic model assessment for insulin resistance values were comparable between insulin independent and dependent cohorts. Conclusions Our long-term follow-up data suggest that IAT can effectively reduce insulin requirements and improve postoperative glycemic control.

Bile duct injury (BDI) is a severe and sometimes life-threatening complication of cholecystectomy. Several series have described a 0.5% to 0.6% incidence of BDI during laparoscopic cholecystectomy. We received an emergency call from the operating theater by the surgery team to assess an iatrogenic BDI in a 58-year-old man with cirrhosis who presented for laparoscopic cholecystectomy. After many trials by endoscopic retrograde cholangiopancreatography (ERCP) the guide wire passed to the peritoneal cavity and failed to pass proximally. Laparoscopy resumed, and the surgeon tried to pass the flexible guide wire proximally unsuccessfully. Then, a decision to hold the sphincterotome by laparoscopy and passing it proximally in harmony with ERCP was taken, which was successful. A regular ERCP with 10F plastic stent insertion was carried out, and the perforation was secured by the inserted stent without any further surgical intervention. Laparoscopy-assisted ERCP may give new insights into the immediate repair of iatrogenic bile duct injuries.

Enteropathogenic Escherichia coli (EPEC) were considered prime reason of diarrhea among neonatal livestock animals in developing countries and could be of public health importance via contaminated milk and meat. Continual attempts were conducted to combat this illness using various antigenic determinants. This study was performed on four E. coli serotypes O26, O45, O103 and O111, which were previously recovered from diarrheic camel calves in Giza, Egypt. Extraction of fimbrial proteins was carried out through dialysis then evaluation of their immunogenic activity was preceded. SDS-PAGE electrophoresis was performed on crude extracted fimbrial proteins and revealed single band for each isolate ranged from 22 to 33kDa. Immunobloting was implemented on the extracted crude fimbrial proteins against E.coli O26 antisera formerly prepared in rabbit.These findings suggested that the fimbrial proteins are of immunogenicity importance and can serve as a protective passive vaccine antigen in prevention of diarrhea caused by EPEC and ETEC infection in camel calves.

This analysis examines the flow of Sutterby nanofluid having bioconvection due to a stretching sheet and heated convective boundary condition. The heat sink/source was imposed in terms of the energy equation. The entropy minimization is also considered. Through the appropriate transformation of the system of nonlinear PDE’s setting, the existing problem has altered into nonlinear ODEs and then numerically utilized via the bvp4c method. The numerical values of skin friction, Nusselt number, Sherwood number, and motile density profiles are revealed in a tabular form. In comparison, other governing variables on velocity, temperature, concentration, and bioconvection are seen through various plots and discussed. For example, increment in the Deborah number of heat flux depreciated in temperature distribution while opposite observation was carried out for heat source and sink parameter. Moreover, it has also been investigated that the shear-thinning fluid is entirely reverse to that of the shear thickening fluid. Further, the increase in the magnetic number accelerates in the Bejan number and irreversibility ratio. Finally, the comparison has been made with previous literature and found an excellent agreement.

Fifteen alfalfa populations were tested for resistance to the seedling damping-off disease sourced by

Knowledge of soil weed seed bank is important for population dynamics studied, establishment of appropriate weed management programs, a little effort in understanding weed seed bank can give valuable information about what weeds to expect in growing season, weed density, and when most weed germination will take place. In this study, a two - year's, two sites were carried out with the aim of assessing weed seed bank status of the soil throughout 2018 and 2019. A site was worked out in Sakha Agriculture Research farm act as a clay soil, Kafr El-Sheikh Governorate, Agriculture Research Center (ARC). Another site was worked out in El-Ismailia Agr; Res; farm act as sandy soil, El-Ismailia Governorate, ARC. At each site, soil samples were selected from nine different places as like three Zigzag shapes divided into three, six and nine sites, "W" to act the whole soil area (30 faddan in Sakha farm, and 15 faddan in El-Ismailia farm). The soil samples were taken from topsoil 0-10 cm depth with an auger (core) 10 cm diameter the soils without tillage and before sowing the summer crop. The result of present the study in two different stations and soils, revealed that the number of soil samples to estimate weed seed banks should be either six or nine sites; each sample weighted 0.50 Kg soil with zigzag shape act a direct seed extraction technique to able recognize the abundance of weed species into the soil and their seed density. The aim is to improve integrated weed control.

Pyrethroid pesticides are commonly used for pest control in agriculture setup, veterinary and home garden. They are now posing increased risks to non-targeted organisms associated to human beings due to their considerable use. The present work deals with the isolation of bacteria with tolerance to high concentrations of bifenthrin and cypermethrin from contaminated soil. Enrichment culture technique (bifenthrin concentration = 50-800 mg/L) was used for bacterial isolation. Bacteria that showed growth on minimal media with bifenthrin were also sub-cultured on minimal media with cypermethrin. Bacteria showing luxurious growth on both the pyrethroid, were screened out based on their morphological, biochemical parameters and by API 20NE Kit. Phylogenetic studies revealed that, one bacterial isolate (MG04) belonging to Acinetobacter lwoffii and other five bacterial isolates (MG06, MG05, MG01, MG03 and MG02) cluster with Pseudomonas aeruginosa, Pseudomonas putida respectively. Isolated members of genera Pseudomonas and Acinetobacter could be used for further detailed degradation studies by using FTIR, HPLC-MS or GC-MS analysis. Resumo Os pesticidas piretróides são comumente usados para controle de pragas na agricultura, veterinária e hortas domésticas. Atualmente eles apresentam riscos aumentados para organismos não-alvo associados a seres humanos devido ao seu uso considerável. O presente trabalho analisou o isolamento de bactérias com tolerância a altas concentrações de bifentrina e cipermetrina de solo contaminado. A técnica de cultura de enriquecimento (concentração de bifentrina = 50-800 mg/L) foi utilizada para o isolamento bacteriano. Bactérias que apresentaram crescimento em meio mínimo com bifentrina também foram subcultivadas em meio mínimo com cipermetrina. Bactérias apresentando crescimento luxuoso em ambos os piretróides foram triadas com base em seus parâmetros morfológicos, bioquímicos e pelo Kit API 20NE. Estudos filogenéticos revelaram que, um isolado bacteriano (MG04) pertencente a Acinetobacter lwoffii e outros cinco isolados bacterianos (MG06, MG05, MG01, MG03 e MG02) agrupam-se com Pseudomonas aeruginosa, Pseudomonas putida respectivamente. Membros isolados dos gêneros Pseudomonas e Acinetobacter podem ser usados para estudos de degradação mais detalhados usando análises de FTIR, HPLC-MS ou GC-MS.

To improve the phosphorus (P) recovery efficiency from livestock wastewater, a novel MgO doped mildewed corn biochar with thermal pre-puffing treatment (Mg-PBC) and without pre-puffing (Mg-BC) was synthesized and tested. The thermal-puffing pretreatment improved the effectiveness of metal soaking and MgO dispersion. P recovery time with Mg-PBC (7 h) was significantly shorter than that with Mg-BC (12 h). Moreover, Mg-PBC showed significantly higher P recovery capacity (241 mg g−1) than Mg-BC (96.6 mg g−1). P recovery capacity of the Mg-PBC fitted to the Thomas model was 90.7 mg g−1, which was 4 times higher than that of Mg-BC (22.9 mg g−1) under column test conditions. The mechanisms involved in P recovery included precipitation, surface complexation, and electrostatic interaction. After adsorption, both Mg-BC and Mg-PBC showed relatively low regeneration abilities. The P loaded Mg-BC (Mg-BC-P) and Mg-PBC (Mg-PBC-P), the later particularly, obviously increased the available P content and promoted plant growth. The release of P increased with time in the Mg-PBC-P treated soil, while it decreased with time in the P fertilizer treated soil. A cost–benefit analysis revealed that thermal-puffing pretreatment greatly increased the profit of MgO doped biochar from −0.66 to 5.90 US$ kg−1. These findings highlight that biomass pre-puffing is a feasible treatment to produce MgO modified biochar and to recover P from livestock wastewater, and that the Mg-PBC-P can be used as a slow-release P fertilizer. Graphical Abstract

Crystal morphology of metal oxides in engineered metal-biochar composites governs the removal of phosphorus (P) from aqueous solutions. Up to our best knowledge, preparation of bio-assembled MgO-coated biochar and its application for the removal of P from solutions and kitchen waste fermentation liquids have not yet been studied. Therefore, in this study, a needle-like MgO particle coated tea waste biochar composite (MTC) was prepared through a novel biological assembly and template elimination process. The produced MTC was used as an adsorbent for removing P from a synthetic solution and real kitchen waste fermentation liquid. The maximum P sorption capacities of the MTC, deduced from the Langmuir model, were 58.80 mg g−1 from the solution at pH 7 and 192.8 mg g−1 from the fermentation liquid at pH 9. The increase of ionic strength (0–0.1 mol L−1 NaNO3) reduced P removal efficiency from 98.53% to 93.01% in the synthetic solution but had no significant impact on P removal from the fermentation liquid. Precipitation of MgHPO4 and Mg(H2PO4)2 (76.5%), ligand exchange (18.0%), and electrostatic attraction (5.5%) were the potential mechanisms for P sorption from the synthetic solution, while struvite formation (57.6%) and ligand exchange (42.2%) governed the sorption of P from the kitchen waste fermentation liquid. Compared to previously reported MgO-biochar composites, MTC had a lower P sorption capacity in phosphate solution but a higher P sorption capacity in fermentation liquid. Therefore, the studied MTC could be used as an effective candidate for the removal of P from aqueous environments, and especially from the fermentation liquids. In the future, it will be necessary to systematically compare the performance of metal-biochar composites with different metal oxide crystal morphology for P removal from different types of wastewater. Graphical Abstract

Hereditary spinocerebellar degenerations (SCDs) is an umbrella term that covers a group of monogenic conditions that share common pathogenic mechanisms and include hereditary spastic paraplegia (HSP), cerebellar ataxia, and spinocerebellar ataxia. They are often complicated with axonal neuropathy and/or intellectual impairment and overlap with many neurological conditions, including neurodevelopmental disorders. More than 200 genes and loci inherited through all modes of Mendelian inheritance are known. Autosomal recessive inheritance predominates in consanguineous communities; however, autosomal dominant and X-linked inheritance can also occur. Sudan is inhabited by genetically diverse populations, yet it has high consanguinity rates. We used next-generation sequencing, genotyping, bioinformatics analysis, and candidate gene approaches to study 90 affected patients from 38 unrelated Sudanese families segregating multiple forms of SCDs. The age-at-onset in our cohort ranged from birth to 35 years; however, most patients manifested childhood-onset diseases (the mean and median ages at onset were 7.5 and 3 years, respectively). We reached the genetic diagnosis in 63% and possibly up to 73% of the studied families when considering variants of unknown significance. Combining the present data with our previous analysis of 25 Sudanese HSP families, the success rate reached 52–59% (31–35/59 families). In this article we report candidate variants in genes previously known to be associated with SCDs or other phenotypically related monogenic disorders. We also highlight the genetic and clinical heterogeneity of SCDs in Sudan, as we did not identify a major causative gene in our cohort, and the potential for discovering novel SCD genes in this population.

Background and Aim: Environmental contaminants such as pesticides have shown immunomodulatory effects that can make animals highly susceptible to pathogenic invasion. The current work aims to study the incidence of udder infections in a single dairy herd of 160 cows in Qalyoubia Governorate, in relation to the potential intoxication of dairy cattle with organochlorine (OCs) pesticides. The study also aims to investigate the impact of udder infections on milk composition. Materials and Methods: The dairy herd was screened for udder infections using the California mastitis test and measurement of somatic cell count (SCC), followed by bacteriological and molecular analysis. In parallel, the milk samples were also tested for residues of 15 OCs compounds using gas chromatographic analysis. Results: The examined herd showed a high prevalence of mastitis (37.5%) and Mycoplasma was identified as the main bacterial pathogen. OCs residues were detected in milk of 45 cows out of 160 with a higher incidence in mastitic (43.3%) than in healthy cows (19%). Further, the biochemical analysis of milk showed a significant drop in major electrolytes combined with a significant rise in blood-borne electrolytes (Na and Cl) and total protein. This was more extreme in the case of Mycoplasam mastitis compared to non-Mycoplasma mastitis. In addition, Mycoplasma mastitic milk revealed a high level of malondialdehyde associated with reduced antioxidant enzymes (glutathione peroxidase, superoxide dismutase and catalase), compared to non-Mycoplasma mastitis. Conclusion: Mycoplasma mastitis was shown to be associated with increased SCC and, in turn, appeared significantly correlated with increased biochemical changes in milk, indicating the serious impact of Mycoplasma mastitis on the dairy industry. Our data also show a strong correlation between increased SCC and biochemical changes in milk, suggesting that tested biochemical parameters might serve as potential biomarkers for the early detection of mastitis. The study also suggested a potential relationship between poisoning with OCs and susceptibility to bacterial udder infections. However, further studies are required to examine the immune status of a dairy herd in relation to the level of OCs in cow's blood, as well as the water sources used, grass forage and soil.

This study was performed on the Gymnura altavela (Linnaeus, 1758), during the period between July 2011- June 2012, in the Syrian coast. A total of 58 examined samples (26 males and 32 females) showed that the size which female start maturation at 701 mm of disk width (DW), while the males start maturation at 682 mm. Females were always larger than males, the largest female and male were 1342 mm, 893 mm (DW), and the total weigh were (20170 g, 5071 g) respectively. G. altavela is a non-placental ovoviviparous species. The female has two ovaries, the left one is active, and the right one is atrophic. Female gestate the embryos in two uteri. Gestation could continue 9-12 months. Ovulation occurs at end of summer and at beginning of autumn, parturition occurs at end of spring and beginning of summer. One reproductive cycle occurs per year. Average of disk width and weight of fully-developed fetuses were 290 mm and 246 g. respectively. Fecundity ranges one to four newborns per litter, embryos and fetuses are symmetrically distributed in two uteruses, with increasing to left one, the ratio was 1:1.2, average of uterine fecundity was (3), average of ovarian fecundity was (6.3).

Elemental sulfur, an industrial by-product from petroleum industries worldwide, has drawn sufficient attention to researchers. The limited scope of application has caused a colossal surplus amount of elemental sulfur stacked in the open places. Several polysulfide synthesis processes, including condensation, free-radical process, and ionic copolymerization technique, were used but resulted in unstable products. A new polymerization technique, termed inverse vulcanization, has been introduced, which enabled different types of crosslinkers for polysulfide production and their scopes to explore numerous applications. The current paper concisely reviews the evolution and advances of using vegetable oils and plant extracts in inverse vulcanization to produce polysulfides. The alluring applications and properties have also been discussed briefly.

The pathophysiology underlying the loss of dopaminergic neurons in Parkinson's disease (PD) is unclear. A gap of knowledge in the molecular and cellular events leading to degeneration of the nigrostriatal DA system is a major barrier to the development of effective therapies for PD. 1-methyl-4-phenylpyridinium (MPP

Saudi journal of biological sciences 29(1), 190-196 (2022). doi:10.1016/j.sjbs.2021.08.072, Published by Elsevier, Amsterdam [u.a.]

In vitro hydrolysis of NKTR-214 under physiological conditions generating NKTR-214 released IL-2 conjugates

Binding curves from surface plasmon resonance studies of purified active 1-PEG-IL-2 released from NKTR-214 binding to IL-2Rαβ and IL-2Rβ

NKTR-214 produces potent immune activation in cynomolgus monkeys and is well tolerated

Antitumor immunity stimulated by NKTR-214 and anti-CTLA-4 in mouse EMT6 tumors requires innate and adaptive immune responses

Purpose: Aldesleukin, recombinant human IL2, is an effective immunotherapy for metastatic melanoma and renal cancer, with durable responses in approximately 10% of patients; however, severe side effects limit maximal dosing and thus the number of patients able to receive treatment and potential cure. NKTR-214 is a prodrug of conjugated IL2, retaining the same amino acid sequence as aldesleukin. The IL2 core is conjugated to 6 releasable polyethylene glycol (PEG) chains. In vivo, the PEG chains slowly release to generate active IL2 conjugates.Experimental Design: We evaluated the bioactivity and receptor binding of NKTR-214 and its active IL2 conjugates in vitro; the tumor immunology, tumor pharmacokinetics, and efficacy of NKTR-214 as a single agent and in combination with anti–CTLA-4 antibody in murine tumor models. Tolerability was evaluated in non-human primates.Results: In a murine melanoma tumor model, the ratio of tumor-killing CD8+ T cells to Foxp3+ regulatory T cells was greater than 400 for NKTR-214 compared with 18 for aldesleukin, supporting preferential activation of the IL2 receptor beta over IL2 receptor alpha, due to the location of PEG molecules. NKTR-214 provides a 500-fold greater exposure of the tumor to conjugated IL2 compared with aldesleukin. NKTR-214 showed efficacy as a single agent and provided durable immunity that was resistant to tumor rechallenge in combination with anti–CTLA-4 antibody. NKTR-214 was well tolerated in non-human primates.Conclusions: These data support further evaluation of NKTR-214 in humans for a variety of tumor types, adding to the repertoire of potent and potentially curative cancer immunotherapies. Clin Cancer Res; 22(3); 680–90. ©2016 AACR.

Supplemental Materials and Methods. ELISA study materials; NKTR-214 chemistry; Mapping of PEGylation sites; Identification of active released IL-2 conjugates; Binding affinity to IL-2 receptors; In vivo immune cell phenotyping, In vivo immune cell phenotyping; In vivo T cell antigen staining; In vivo depletion of immune cells; Hematology parameters for evaluation of immune markers in non-human primates. Supplemental Figure Legends S1-S4. Supplemental Tables S1 and S2. Table S1: Fold change of affinity to IL-2αβ and IL-2β for NKTR-214 and its released active IL-2 conjugates compared to aldesleukin; Table S2: Activation of Primary Cynomulgus Monkey and Human T Cells in Response to IL-2 and the Active Metabolite of NKTR-214