Design, synthesis, and evaluation of 4'-phosphonomethoxy pyrimidine ribonucleosides as potential anti-influenza agents.

Influenza viruses belong to the Orthomyxoviridae family and cause acute respiratory distress in humans. The developed drug resistance toward existing drugs and the emergence of viral mutants that can escape vaccines mandate the search for novel antiviral drugs. Herein, the synthesis of epimeric 4'-methyl-4'-phosphonomethoxy [4'-C-Me-4'-C-(O-CH2 P═O)] pyrimidine ribonucleosides, their phosphonothioate [4'-C-Me-4'-C-(O-CH2 P═S)] derivatives, and their evaluation against an RNA viral panel are described. Selective formation of the α- l-lyxo epimer, [4'-C-(α)-Me-4'-C-(ß)-(O-CH2 -P(═O)(OEt)2 )] over the ß- d-ribo epimer [4'-C-(ß)-Me-4'-C-(α)-(O-CH2 -P(═O)(OEt)2 )] was explained by DFT equilibrium geometry optimizations studies. Pyrimidine nucleosides having the [4'-C-(α)-Me-4'-C-(ß)-(O-CH2 -P(═O)(OEt)2 )] framework showed specific activity against influenza A virus. Significant anti-influenza virus A (H1N1 California/07/2009 isolate) was observed with the 4'-C-(α)-Me-4'-C-(ß)-O-CH2 -P(═O)(OEt)2 -uridine derivative 1 (EC50 = 4.56 mM, SI50 > 56), 4-ethoxy-2-oxo-1(2H)-pyrimidin-1-yl derivative 3 (EC50 = 5.44 mM, SI50 > 43) and the cytidine derivative 2 (EC50 = 0.81 mM, SI50 > 13), respectively. The corresponding thiophosphonates 4'-C-(α)-Me-4'-C-(ß)-(O-CH2 -P( S)(OEt)2 ) and thionopyrimidine nucleosides were devoid of any antiviral activity. This study shows that the 4'-C-(α)-Me-4'-(ß)-O-CH2 -P(═O)(OEt)2 ribonucleoside can be further optimized to provide potent antiviral agents.

Design, Synthesis, Characterization, and Molluscicidal Activity Screening of New Nicotinonitrile Derivatives against Land Snails, M. cartusiana.

A new series of nicotinonitrile derivatives 2-7 was designed and synthesized from the starting material (E)-3-(4-chlorophenyl)-1-(4-methoxyphenyl)prop-2-en-1-one (1) to assess their molluscicidal activity. The newly synthesized nicotinonitrile compounds 2-7 were characterized based on FTIR, 1H-NMR, and 13C-APT NMR spectra as well as elemental microanalyses. The target compounds 2-7 were screened for their toxicity effect against M. cartusiana land snails and were compared to Acetamiprid as a reference compound. The results demonstrated that the nicotinonitrile-2-thiolate salts 4a and 4b had good mortality compared with that of Acetamiprid. The results of the in vivo effect of the prepared nicotinonitrile molecules 2, 4a, and 4b on biochemical parameters, including AChE, ALT, AST, and TSP, indicated a reduction in the level of AChE and TSP as well as an increase in the concentration of transaminases (ALT and AST). A histopathological study of the digestive gland sections of the M. cartusiana land snails was carried out. The nicotinonitrile-2-thiolate salts 4a,b showed vacuolization, causing the digestive gland to lose its function. It could be concluded that the water-soluble nicotinonitrile-2-thiolate salts 4a,b could be adequate molluscicidal molecules against M. cartusiana land snails.

Quantitative 1HNMR Spectroscopy: Analysis of Zinc Gluconate in Utozinc® Tablets, a Mixture of Zinc Gluconate and Vitamin C.

BACKGROUND: Zinc is an essential metal for humans and plays key roles in several biological events such as immunity, allergy, growth, and inflammation. The deficiency in zinc causes an increased infection rate with pathogens. Organo-zincates such as zinc gluconate are known for better absorption compared with their inorganic zinc salts. Its role in enhancing the immune system has driven a huge demand for organo-zinc supplements and in the treatment protocol of coronavirus disease, the causative agent of the COVID-19 pandemic. OBJECTIVE: Herein, we report on a quantitative analysis of zinc gluconate in the authentic form in presence of vitamin C, and the method was applied to their dosage form (Utozinc® tablets). The method is simple, accurate, and validated according to ICH guidelines. METHOD: Quantification of zinc gluconate formulated with vitamin C (Utozinc tablets) using Q-1HNMR. Maleic acid and deuterium oxide were used as internal standards and solvents, respectively. RESULTS: The linearity range, the limit of detection and quantification, stability, precision, and accuracy, were validated. The validation of the method within five concentration levels (from 10 to 50 mg/0.5 mL D2O) afforded a limit of detection of 4.58 mg/mL, a quantification limit of 15.27 mg/mL, and excellent linearity. CONCLUSIONS: The method proposed in the present study is simple, fast, nondestructive, and accurate. Zinc gluconate quantification values obtained by the Q-1HNMR method were found to show an acceptable correlation with those obtained by the thin-layer chromatographic technique. HIGHLIGHTS: The method was successfully applied to Utozinc tablets, and the results were compared with the reported reference pharmacopeial method. The salt exchange between maleic acid (IS) and zinc gluconate was tested by noticing the change in the chemical shift of IS and zinc gluconate.

Evaluation of Green Silver Nanoparticles Fabricated by Spirulina platensis Phycocyanin as Anticancer and Antimicrobial Agents.

Green nanotechnology has attracted attention worldwide, especially in treating cancer and drug-resistant section 6 microbes. This work aims to investigate the anticancer activity of green silver nanoparticles synthesized by Spirulina platensis phycocyanin (SPAgNPs) on two cancer cell lines: Lung cancer cell line (A-549) and breast cancer cell line (MCF-7), compared to the normal human lung cell line (A138). We also aimed to investigate the bactericidal activity against Staphylococcus aureus ATCC29737, Bacillus cereus ATCC11778, Escherichia coli ATCC8379, and Klebsiella pneumonia, as well as the fungicidal activity against Candida albicans (ATCC6019) and Aspergillus niger. The obtained SPAgNPs were spherical and crystalline with a size of 30 nm and a net charge of -26.32 mV. Furthermore, they were surrounded by active groups responsible for stability. The SPAgNPs scavenged 85% of the DPPH radical with a relative increase of approximately 30% over the extract. The proliferation of cancer cells using the MTT assay clarified that both cancer cells (A-549 and MCF-7) are regularly inhibited as they grow on different concentrations of SPAgNPs. The maximum inhibitory effect of SPAgNPs (50 ppm) reached 90.99 and 89.51% against A-549 and MCF7, respectively. Regarding antimicrobial activity, no inhibition zones occurred in bacterial or fungal strains at low concentrations of SPAgNPs and the aqueous Spirulina platensis extract. However, at high concentrations, inhibition zones, especially SPAgNPs, were more potent for all tested microorganisms than their positive controls, with particular reference to Staphylococcus aureus, since the inhibition zones were 3.2, 3.8, and 4.3 mm, and Bacillus cereus was 2.37 mm when compared to tetracycline (2.33 mm). SPAgNPs have more potent antifungal activity, especially against Aspergillus niger, compared to their positive controls. We concluded that SPAgNPs are powerful agents against oxidative stress and microbial infection.

Probing the Substrate Requirements of the In†Vitro Geranylation Activity of Selenouridine Synthase (SelU).

Natural RNA modifications diversify the structures and functions of existing nucleic acid building blocks. Geranyl is one of the most hydrophobic groups recently identified in bacterial tRNAs. Selenouridine synthase (SelU, also called mnmH) is an enzyme with a dual activity which catalyzes selenation and geranylation in tRNAs containing 2-thiouridine using selenophosphate or geranyl-pyrophosphate as cofactors. In this study, we explored the in vitro geranylation process of tRNA anticodon stem loops (ASL) mediated by SelU and showed that the geranylation activity was abolished when U35 was mutated to A35 (ASL-tRNALys (s2U)UU to ASL-tRNAIle (s2U)AU ). By examining the SelU cofactor geranyl-pyrophosphate (gePP) and its analogues, we found that only the geranyl group, but not dimethylallyl- and farnesyl-pyrophosphate with either shorter or longer terpene chains, could be incorporated into ASL. The degree of tRNA geranylation in the end-point analysis for SelU follows the order of ASLLys (s2UUU) ≃ ASLGln (s2UUG) >ASLGlu (s2UUC) . These findings suggest a putative mechanism for substrate discrimination by SelU and reveal key factors that might influence its enzymatic activity. Given that SelU plays an important role in bacterial translation systems, inhibiting this enzyme and targeting its geranylation and selenation pathways could be exploited as a promising strategy to develop SelU-based antibiotics.

Fluorescence-Based Binding Characterization of Small Molecule Ligands Targeting CUG RNA Repeats.

Pathogenic CUG and CCUG RNA repeats have been associated with myotonic dystrophy type 1 and 2 (DM1 and DM2), respectively. Identifying small molecules that can bind these RNA repeats is of great significance to develop potential therapeutics to treat these neurodegenerative diseases. Some studies have shown that aminoglycosides and their derivatives could work as potential lead compounds targeting these RNA repeats. In this work, sisomicin, previously known to bind HIV-1 TAR, is investigated as a possible ligand for CUG RNA repeats. We designed a novel fluorescence-labeled RNA sequence of r(CUG)10 to mimic cellular RNA repeats and improve the detecting sensitivity. The interaction of sisomicin with CUG RNA repeats is characterized by the change of fluorescent signal, which is initially minimized by covalently incorporating the fluorescein into the RNA bases and later increased upon ligand binding. The results show that sisomicin can bind and stabilize the folded RNA structure. We demonstrate that this new fluorescence-based binding characterization assay is consistent with the classic UV Tm technique, indicating its feasibility for high-throughput screening of ligand-RNA binding interactions and wide applications to measure the thermodynamic parameters in addition to binding constants and kinetics when probing such interactions.

Accuracy of genomic selection for reducing susceptibility to pendulous crop in turkey (Meleagris gallopavo).

Pendulous crop (PC) in the turkey occurs when the crop distends from its normal position, thereby preventing the movement of feed and water from the crop down into the digestive system. This condition negatively impacts the turkey industry at both production and welfare levels. In this study, we estimated the genetic parameters for PC incidence and its genetic correlation with 5 production traits. Additionally, we evaluated the prediction accuracy and bias of breeding values for the selection candidates using pedigree (BLUP) or pedigree-genomic (ssGBLUP) relationships among the animals. A total of 245,783 turkey records were made available by Hybrid Turkeys, Kitchener, Canada. Of these, 6,545 were affected with PC. In addition, the data included 9,634 records for breast meat yield (BMY); 5,592 records for feed conversion ratio (FCR) and residual feed intake (RFI) in males; 170,844 records for body weight (BW) and walking score (WS) between 18 and 20 wk of age for males (71,012) and females (99,832), respectively. Among this population, 36,830 were genotyped using a 65K SNP Illumina Inc. chip. While all animals passed the quality control criteria, only 53,455 SNP markers were retained for subsequent analysis. Heritability for PC was estimated at 0.16 ± 0.00 and 0.17 ± 0.00 using BLUP and ssGBLUP, respectively. The incidence of PC was not genetically correlated with WS or FCR. Low unfavourable genetic correlations with BW (0.12 and 0.14), BMY (0.24 and 0.24) and RFI (-0.33 and -0.28) were obtained using BLUP and ssGBLUP, respectively. Using ssGBLUP showed higher prediction accuracy (0.51) for the breeding values for the selection candidates than the pedigree-based model (0.35). Whereas the bias of the prediction was slightly reduced with ssGBLUP (0.33 ± 0.05) than BLUP (0.30 ± 0.08), both models showed a regression coefficient lower than one, indicating inflation in the predictions. The results of this study suggest that PC is a heritable trait and selection for lower PC incidence rates is feasible. Although further investigation is necessary, selection for BW, BMY, and RFI may increase PC incidence. Incorporating genomic information would lead to higher accuracy in predicting the genetic merit for selection candidates.

Synthesis of 6-Aza-2-Hydroxyimino-5-Methylpyrimidine Nucleosides for Antiviral Evaluation.

The syntheses of a series of novel 6-aza-2-hydroxyimino-5-methylpyrimidine and related nucleosides are described. A suitably protected 2-methylthiopyrimidine nucleoside was selected as the precursor for installing a hydroxyimino moiety at the C-2 position. The starting nucleobase 6-aza-5-methyl-2-thiouracil is prepared in two steps from thiosemicarbazone and ethyl pyruvate. This is subjected to coupling with 1-O-acetyl-2,3,5-tri-O-benzoyl-ß-D-ribofuranose under Vorbrüggen glycosylation conditions to provide the corresponding nucleoside in high yield. Activation of the nucleoside to the corresponding 2-methylthio derivative followed by treatment with hydroxylamine hydrochloride in pyridine provides the corresponding 2-hydroxyimino derivative in high yield. Finally, the synthesis of five free modified nucleoside analogs is described. The newly synthesized nucleosides have been evaluated against an RNA viral panel and moderate activity was observed against hepatitis C virus, Zika virus, and human respiratory syncytial virus. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Preparation of 6-aza-5-methyl-2-thiouracil Basic Protocol 2: Preparation of 6-aza-5-methyl-2-thiouridine and 6-aza-5-methyluridine Basic Protocol 3: Preparation of 6-aza-2-hydroxyimino-5-methyluridine Basic Protocol 4: Preparation of 6-aza-2-hydroxyimino-5-methyl-4-thiouridine and 6-aza-2-hydroxyimino-5-methylcytosine.

Exosomes in Alzheimer's Disease: From Being Pathological Players to Potential Diagnostics and Therapeutics.

Exosomes (EXOs) were given attention as an extracellular vesicle (EV) with a pivotal pathophysiological role in the development of certain neurodegenerative disorders (NDD), such as Parkinson's and Alzheimer's disease (AD). EXOs have shown the potential to carry pathological and therapeutic cargo; thus, researchers have harnessed EXOs in drug delivery applications. EXOs have shown low immunogenicity as natural drug delivery vehicles, thus ensuring efficient drug delivery without causing significant adverse reactions. Recently, EXOs provided potential drug delivery opportunities in AD and promising future clinical applications with the diagnosis of NDD and were studied for their usefulness in disease detection and prediction prior to the emergence of symptoms. In the future, the microfluidics technique will play an essential role in isolating and detecting EXOs to diagnose AD before the development of advanced symptoms. This review is not reiterative literature but will discuss why EXOs have strong potential in treating AD and how they can be used as a tool to predict and diagnose this disorder.

Discovering lethal alleles across the turkey genome using a transmission ratio distortion approach.

Deviation from Mendelian inheritance expectations (transmission ratio distortion, TRD) has been observed in several species, including the mouse and humans. In this study, TRD was characterized in the turkey genome using both allelic (specific- and unspecific-parent TRD) and genotypic (additive- and dominance-TRD) parameterizations within a Bayesian framework. In this study, we evaluated TRD for 23 243 genotyped Turkeys across 56 393 autosomal SNPs. The analyses included 500 sires, 2013 dams and 11 047 offspring (trios). Three different haplotype sliding windows of 4, 10 and 20 SNPs were used across the autosomal chromosomes. Based on the genotypic parameterizations, 14 haplotypes showed additive and dominance TRD effects highlighting regions with a recessive TRD pattern. In contrast, the allelic model uncovered 12 haplotype alleles with the allelic TRD pattern which showed an underrepresentation of heterozygous offspring in addition to the absence of homozygous animals. For regions with the allelic pattern, only one particular region showed a parent-specific TRD where the penetrance was high via the dam, but low via the sire. The gene set analysis uncovered several gene ontology functional terms, Reactome pathways and several Medical Subject Headings that showed significant enrichment of genes associated with TRD. Many of these gene ontology functional terms (e.g. mitotic spindle assembly checkpoint, DRM complex and Aneuploidy), Reactome pathways (e.g. Mismatch repair) and Medical Subject Headings (e.g. Adenosine monophosphate) are known to be related to fertility, embryo development and lethality. The results of this study revealed potential novel candidate lethal haplotypes, functional terms and pathways that may enhance breeding programs in Turkeys through reducing mortality and improving reproduction rate.

Synthesis of 9-(6-Deoxy-α-L-Talofuranosyl)-6-Methylpurine and 9-(6-Deoxy-ß-D-Allofuranosyl)-6-Methylpurine Nucleosides.

6-Methylpurine (MeP) is a cytotoxic adenine analog that does not exhibit selectivity when administered systemically and could be very useful in a gene therapy approach to cancer treatment involving Escherichia coli purine nucleoside phosphorylase (PNP). 9-(6-Deoxy-ß-D-allofuranosyl)-6-methylpurine [methyl(allo)-MePR, 18] and 9-(6-deoxy-α-L-talofuranosyl)-6-methylpurine [methyl(talo)-MePR, 21] were synthesized as potential prodrugs for MeP in the E. coli PNP/prodrug cancer gene therapy approach. The detailed syntheses of [methyl(allo)-MePR] and [methyl(talo)-MePR] are described. The glycosyl donors, 1,2-di-O-acetyl-3,5-di-O-benzyl-α-D-allofuranose (12) and 1-O-acetyl-3-O-benzyl-2,5-di-O-benzoyl-α-L-talofuranose (16) were prepared from 1,2:5,6-di-O-isopropylidene-α-D-glucofuranose (4) in nine and eleven steps, respectively. Vorbrüggen coupling of the latter glycosyl donors with 6-methylpurine (3), followed by deprotection of the sugar hydroxyl groups, gave the title compounds in good overall yields. © 2020 by John Wiley & Sons, Inc. Basic Protocol 1: Preparation of 6-methylpurine Basic Protocol 2: Preparation of the D-allofuranose derivative (12) Basic Protocol 3: Preparation of 6-deoxy-α-L-talofuranoside Basic Protocol 4: Preparation of methyl(allo)-MePR (18) Basic Protocol 5: Preparation of methyl(talo)-MePR (21).

Genetic parameters for clutch and broodiness traits in turkeys (Meleagris Gallopavo) and their relationship with body weight and egg production.

The objective of this study was to estimate phenotypic and genetic parameters for clutch and broodiness (BR) traits in turkeys and their relationship with body weight and egg production. Data on dam line hens was available and included: body weight at 18 wk of age (BW18), body weight at lighting (BWL, 29 to 33 wk), age at first egg (AFE), egg number (EN), rate of lay (RL), clutch length (CL), maximum clutch length (MCL), pause length (PL), maximum PL (MPL) and BR. BR was defined as the average number of consecutive pause days between clutches that was higher than the average PL per hen. Heritability estimates for BW18 and BWL were 0.50 and 0.53, respectively. The heritability for egg production, clutch, and pause traits varied from low (MPL = 0.15; BR = 0.15) to moderate (AFE = 0.22; EN = 0.28; RL = 0.29; CL = 0.21; MCL = 0.27; PL = 0.25). Genetic correlations were negative between body weight traits and EN (rg (BW18, EN) = -0.27; rg(BWL, EN) = -0.33) and CL (rg(BW18, CL) = -0.40; rg(BWL, CL) = -0.33). BR was negatively genetically correlated with EN (rg(BR, EN) = -0.85) and CL (rg(BR, CL) = -0.30), and positively genetically correlated with PL (rg(BR, PL) = 0.93) and AFE (rg(BR, AFE) = 0.21). EN had a positive (0.73) and a negative (-0.84) genetic correlation with CL and PL, respectively. Overall, the results of this study confirmed the negative (unfavorable) correlations between egg production and body weight. Despite unfavorable genetic and phenotypic correlations between egg production traits and those relating to BR, the inclusion of BR in a selection program through incorporation of clutch length traits and pause length traits is feasible. Integration of either clutch length traits or pause length traits in a selection index is likely to increase egg number while decreasing broodiness.

3-Trifluoromethylpyrazolones derived nucleosides: Synthesis and antiviral evaluation.

Dengue (DENV) viral infection is a global public health problem that infrequently develops life threatening diseases such as dengue hemorrhagic fever (DFS) and dengue shock syndrome (DSS). Middle East respiratory syndrome coronavirus (MERS-CoV) is a highly pathogenic human corona virus with 38% fatality rate of infected patients. A series of 4-arylhydrazono-5-trifluoromethyl-pyrazolones, their ribofuranosyl, and 5'-deoxyribofuranosyl nucleosides were synthesized, geometry optimized using Density functional theory (DFT), and evaluated for their antiviral activity. 2-Nitrophenylhydrazonopyra-zolone derivative 5 showed significant activity against MERS-CoV (EC50 = 4.6 µM). The nucleoside analog 8 showed moderate activity against DENV-2 (EC50 = 10 µM), while the activity was abolished with the corresponding 5'-deoxyribonucleoside analogs. The identified hits in this study set this category of compounds for further future optimizations.

Construction and structure studies of DNA-bipyridine complexes as versatile scaffolds for site-specific incorporation of metal ions into DNA.

The facile construction of metal-DNA complexes using 'Click' reactions is reported here. A series of 2'-propargyl-modified DNA oligonucleotides were initially synthesized as structure scaffolds and were then modified through 'Click' reaction to incorporate a bipyridine ligand equipped with an azido group. These metal chelating ligands can be placed in the DNA context in site-specific fashion to provide versatile templates for binding various metal ions, which are exchangeable using a simple EDTA washing-and-filtration step. The constructed metal-DNA complexes were found to be thermally stable. Their structures were explored by solving a crystal structure of a propargyl-modified DNA duplex and installing the bipyridine ligands by molecular modeling and simulation. These metal-DNA complexes could have wide applications as novel organometallic catalysts, artificial ribonucleases, and potential metal delivery systems.

Cyano Modification on Uridine Decreases Base-Pairing Stability and Specificity through Neighboring Disruption in RNA Duplex.

5-Cyanomethyluridine (cnm5 U) and 5-cyanouridine (cn5 U), the two uridine analogues, were synthesized and incorporated into RNA oligonucleotides. Base-pairing stability and specificity studies in RNA duplexes indicated that cnm5 U slightly decreased the stability of the duplex but retained the base-pairing preference. In contrast, cn5 U dramatically decreased both base-pairing stability and specificity between U:A and other noncanonical U:G, U:U, and U:C pairs. In addition, the cn5 U:G pair was found to be stronger than the cn5 U:A pair and the other mismatched pairs in the context of a RNA duplex; this implied that cn5 U might slightly prefer to recognize G over A. Our mechanistic studies by molecular simulations showed that the cn5 U modification did not directly affect the base pairing of the parent nucleotide; instead, it weakened the neighboring base pair in the 5' side of the modification in the RNA duplexes. Consistent with the simulation data, replacing the Watson-Crick A:U pair to a mismatched C:U pair in the 5'-neighboring site did not affect the overall stability of the duplex. Our work reveals the significance of the electron-withdrawing cyano group in natural tRNA systems and provides two novel building blocks for constructing RNA-based therapeutics.

Lithium Hexamethyldisilazane Transformation of Transiently Protected 4-Aza/Benzimidazole Nitriles to Amidines and their Dimethyl Sulfoxide Mediated Imidazole Ring Formation.

Trimethylsilyl-transient protection successfully allowed the use of lithium hexamethyldisilazane to prepare benzimidazole (BI) and 4-azabenzimidazole (azaBI) amidines from nitriles in 58-88% yields. This strategy offers a much better choice to prepare BI/azaBI amidines than the lengthy, low-yielding Pinner reaction. Synthesis of aza/benzimidazole rings from aromatic diamines and aldehydes was affected in dimethyl sulfoxide in 10-15 min, while known procedures require long time and purification. These methods are important for the BI/azaBI-based drug industry and for developing specific DNA binders for expanded therapeutic applications.

Genome-wide association mapping and pathway analysis of leukosis incidence in a US Holstein cattle population.

Bovine leukosis virus is an oncogenic virus that infects B cells, causing bovine leukosis disease. This disease is known to have a negative impact on dairy cattle production and, because no treatment or vaccine is available, finding a possible genetic solution is important. Our objective was to perform a comprehensive genetic analysis of leukosis incidence in dairy cattle. Data on leukosis occurrence, pedigree and molecular information were combined into multitrait GBLUP models with milk yield (MY) and somatic cell score (SCS) to estimate genetic parameters and to perform whole-genome scans and pathway analysis. Leukosis data were available for 11 554 Holsteins daughters of 3002 sires from 112 herds in 16 US states. Genotypes from a 60K SNP panel were available for 961 of those bulls as well as for 2039 additional bulls. Heritability for leukosis incidence was estimated at about 8%, and the genetic correlations of leukosis disease incidence with MY and SCS were moderate at 0.18 and 0.20 respectively. The genome-wide scan indicated that leukosis is a complex trait, possibly modulated by many genes. The gene set analysis identified many functional terms that showed significant enrichment of genes associated with leukosis. Many of these terms, such as G-Protein Coupled Receptor Signaling Pathway, Regulation of Nucleotide Metabolic Process and different calcium-related processes, are known to be related to retrovirus infection. Overall, our findings contribute to a better understanding of the genetic architecture of this complex disease. The functional categories associated with leukosis may be useful in future studies on fine mapping of genes and development of dairy cattle breeding strategies.

Short communication: Genetic correlation of bovine leukosis incidence with somatic cell score and milk yield in a US Holstein population.

Bovine leukosis (BL) is a retroviral disease caused by the bovine leukosis virus (BLV), which affects only cattle. Dairy cows positive for BL produce less milk and have more days open than cows negative for BL. In addition, the virus also affects the immune system and causes weaker response to vaccines. Heritability estimates of BL incidence have been reported for Jersey and Holstein populations at about 0.08, indicating an important genetic component that can potentially be exploited to reduce the prevalence of the disease. However, before BL is used in selection programs, it is important to study its genetic associations with other economically important traits such that correlated responses to selection can be predicted. Hence, this study aimed to estimate the genetic correlations of BL with milk yield (MY) and with somatic cell score (SCS). Data of a commercial assay (ELISA) used to detect BLV antibodies in milk samples were obtained from Antel BioSystems (Lansing, MI). The data included continuous milk ELISA scores and binary milk ELISA results for 11,554 cows from 112 dairy herds across 16 US states. Continuous and binary milk ELISA were analyzed with linear and threshold models, respectively, together with MY and SCS using multitrait animal models. Genetic correlations (posterior means ± standard deviations) between BL incidence and MY were 0.17 ± 0.077 and 0.14 ± 0.076 using ELISA scores and results, respectively; with SCS, such estimates were 0.20 ± 0.081 and 0.17 ± 0.079, respectively. In summary, the results indicate that selection for higher MY may lead to increased BLV prevalence in dairy herds, but that the inclusion of BL (or SCS as an indicator trait) in selection indexes may help attenuate this problem.

6-Methylpurine derived sugar modified nucleosides: Synthesis and evaluation of their substrate activity with purine nucleoside phosphorylases.

6-Methylpurine (MeP) is cytotoxic adenine analog that does not exhibit selectivity when administered systemically, and could be very useful in a gene therapy approach to cancer treatment involving Escherichia coli PNP. The prototype MeP releasing prodrug, 9-(ß-d-ribofuranosyl)-6-methylpurine, MeP-dR has demonstrated good activity against tumors expressing E. coli PNP, but its antitumor activity is limited due to toxicity resulting from the generation of MeP from gut bacteria. Therefore, we have embarked on a medicinal chemistry program to identify non-toxic MeP prodrugs that could be used in conjunction with E. coli PNP. In this work, we report on the synthesis of 9-(6-deoxy-ß-d-allofuranosyl)-6-methylpurine (3) and 9-(6-deoxy-5-C-methyl-ß-d-ribo-hexofuranosyl)-6-methylpurine (4), and the evaluation of their substrate activity with several phosphorylases. The glycosyl donors; 1,2-di-O-acetyl-3,5-di-O-benzyl-α-d-allofuranose (10) and 1-O-acetyl-3-O-benzyl-2,5-di-O-benzoyl-6-deoxy-5-C-methyl-ß-d-ribohexofuran-ose (15) were prepared from 1,2:5,6-di-O-isopropylidine-α-d-glucofuranose in 9 and 11 steps, respectively. Coupling of 10 and 15 with silylated 6-methylpurine under Vorbrüggen glycosylation conditions followed conventional deprotection of the hydroxyl groups furnished 5'-C-methylated-6-methylpurine nucleosides 3 and 4, respectively. Unlike 9-(6-deoxy-α-l-talo-furanosyl)-6-methylpurine, which showed good substrate activity with E. coli PNP mutant (M64V), the ß-d-allo-furanosyl derivative 3 and the 5'-di-C-methyl derivative 4 were poor substrates for all tested glycosidic bond cleavage enzymes.

6-Methylpurine derived sugar modified nucleosides: Synthesis and in vivo antitumor activity in D54 tumor expressing M64V-Escherichia coli purine nucleoside phosphorylase.

Impressive antitumor activity has been observed with fludarabine phosphate against tumors that express Escherichia coli purine nucleoside phosphorylase (PNP) due to the liberation of 2-fluoroadenine in the tumor tissue. 6-Methylpurine (MeP) is another cytotoxic adenine analog that does not exhibit selectivity when administered systemically, and could be very useful in a gene therapy approach to cancer treatment involving E. coli PNP. The prototype MeP releasing prodrug 9-(2-deoxy-ß-d-ribofuranosyl)-6-methylpurine (1) [MeP-dR] has demonstrated good activity against tumors expressing E. coli PNP, but its antitumor activity is limited due to toxicity resulting from the generation of MeP from gut bacteria. Therefore, we have embarked on a medicinal chemistry program to identify a combination of non-toxic MeP prodrugs and non-human adenosine glycosidic bond cleaving enzymes. The two best MeP-based substrates with M64V-E coli PNP, a mutant which was engineered to tolerate modification at the 5'-position of adenosine and its analogs, were 9-(6-deoxy-α-l-talofuranosyl)-6-methylpurine (3) [methyl(talo)-MeP-R] and 9-(α-l-lyxofuranosyl)6-methylpurine (4) [lyxo-MeP-R]. The detailed synthesis methyl(talo)-MeP-R and lyxo-MeP-R, and the evaluation of their substrate activity with 4 enzymes not normally associated with cancer patients is described. In addition, we have determined the intraperitoneal pharmacokinetic (ip-PK) properties of methyl(talo)-MeP-R and have determined its in vivo bystander activity in mice bearing D54 tumors that express M64V PNP. The observed good in vivo bystander activity of [methyl(talo)-MeP-R/M64V-E coli PNP combination suggests that these agents could be useful for the treatment of cancer.