Development of PDT/PET theranostics: synthesis and biological evaluation of an ¹⁸F-radiolabeled water soluble porphyrin

Synthesis of the first water-soluble porphyrin radiolabeled with fluorine-18 is described: a new molecular theranostic agent which integrates the therapeutic selectivity of photodynamic therapy (PDT) with the imaging efficacy of positron emission tomography (PET). Generation of the theranostic was carried out through the conjugation of a cationic water-soluble porphyrin bearing an azide functionality to a fluorine-18 radiolabeled prosthetic bearing an alkyne functionality through click conjugation, with excellent yields obtained in both cold and hot synthesis. Biological evaluation of the synthesized structures shows the first example of an 18 F-radiolabeled porphyrin retaining photocytotoxicity following radiolabeling and demonstrable conjugate uptake and potential application as a radiotracer in vivo. The promising results gained from biological evaluation demonstrate the potential of this structure as a clinically relevant theranostic agent, offering exciting possibilities for the simultaneous imaging and photodynamic treatment of tumors

In search of a combined brucellosis and tuberculosis vaccine for cattle

Bovine brucellosis is caused by Brucella abortus. The bacterial pathogen causes economic losses because it induces abortion in cattle. Vaccination of calves with live B. abortus strain 19 induces a certain level of protection but induces persistent antibodies against cell envelope lipopolysaccharide that make it difficult to Distinguish Infected from Vaccinated Animals (DIVA). Live vaccine B. abortus strain RB51 was developed to eliminate such interfering antibodies and therefore, facilitate the differentiation of infected from vaccinated animals and help in the eradication of the disease. Vaccination with strain RB51 induces levels of protection similar to strain 19 but neither of the two vaccines give complete protection. We have been working to enhance protection induced by strain RB51 vaccine. Protective Brucella antigens can be over-expressed in strain RB51 by introducing a plasmid containing the leuB gene and the genes encoding such antigens. To avoid the expression of antibiotic resistance genes, we produced a leuB deficient strain RB51 and introduced a plasmid containing the leuB gene and the genes to be over-expressed. This new strain maintains the plasmid and has induced significantly high protection levels in mice. In addition, it allowed the construction of an RB51 vaccine strain able to express Mycobacterium bovis protective antigens so that the vaccine could protect against brucellosis and tuberculosis simultaneously

Phase 1 study of sirolimus in combination with oral cyclophosphamide and topotecan in children and young adults with relapsed and refractory solid tumors.

PurposeTo determine the maximum tolerated dose (MTD), toxicities, and pharmacodynamics effects of sirolimus combined with oral metronomic topotecan and cyclophosphamide in a pediatric population.Materials and methodsPatients who were 1 to 30 years of age with relapsed/refractory solid tumors (including CNS) were eligible. Patients received daily oral sirolimus and cyclophosphamide (25-50 mg/m2/dose) on days 1-21 and oral topotecan (0.8 mg/m2/dose) on days 1-14 in 28-day cycles. Sirolimus steady-state plasma trough concentrations of 3-7.9 ng/mL and 8-12.0 ng/mL were evaluated, with dose escalation based on a 3+3 phase 1 design. Biomarkers of angiogenesis were also evaluated.ResultsTwenty-one patients were treated (median age 18 years; range 9-30). Dose-limiting toxicities included myelosuppression, ALT elevation, stomatitis, and hypertriglyceridemia. The MTD was sirolimus with trough goal of 8-12.0 ng/mL; cyclophosphamide 25 mg/m2/dose; and topotecan 0.8 mg/m2/dose. No objective responses were observed. Four patients had prolonged stable disease > 4 cycles (range 4-12). Correlative biomarker analyses demonstrated reductions in thrombospondin-1 (p=0.043) and soluble vascular endothelial growth factor receptor-2 plasma concentrations at 21 days compared to baseline.ConclusionsThe combination of oral sirolimus, topotecan, and cyclophosphamide was well tolerated and biomarker studies demonstrated modulation of angiogenic pathways with this regimen

Pharmacometabolomics reveals racial differences in response to atenolol treatment.

Antihypertensive drugs are among the most commonly prescribed drugs for chronic disease worldwide. The response to antihypertensive drugs varies substantially between individuals and important factors such as race that contribute to this heterogeneity are poorly understood. In this study we use metabolomics, a global biochemical approach to investigate biochemical changes induced by the beta-adrenergic receptor blocker atenolol in Caucasians and African Americans. Plasma from individuals treated with atenolol was collected at baseline (untreated) and after a 9 week treatment period and analyzed using a GC-TOF metabolomics platform. The metabolomic signature of atenolol exposure included saturated (palmitic), monounsaturated (oleic, palmitoleic) and polyunsaturated (arachidonic, linoleic) free fatty acids, which decreased in Caucasians after treatment but were not different in African Americans (p<0.0005, q<0.03). Similarly, the ketone body 3-hydroxybutyrate was significantly decreased in Caucasians by 33% (p<0.0001, q<0.0001) but was unchanged in African Americans. The contribution of genetic variation in genes that encode lipases to the racial differences in atenolol-induced changes in fatty acids was examined. SNP rs9652472 in LIPC was found to be associated with the change in oleic acid in Caucasians (p<0.0005) but not African Americans, whereas the PLA2G4C SNP rs7250148 associated with oleic acid change in African Americans (p<0.0001) but not Caucasians. Together, these data indicate that atenolol-induced changes in the metabolome are dependent on race and genotype. This study represents a first step of a pharmacometabolomic approach to phenotype patients with hypertension and gain mechanistic insights into racial variability in changes that occur with atenolol treatment, which may influence response to the drug

Enzymatic, immunological and phylogenetic characterization of Brucella suis urease

<p>Abstract</p> <p>Background</p> <p>The sequenced genomes of the <it>Brucella </it>spp. have two urease operons, <it>ure</it>-1 and <it>ure</it>-2, but there is evidence that only one is responsible for encoding an active urease. The present work describes the purification and the enzymatic and phylogenomic characterization of urease from <it>Brucella suis </it>strain 1330. Additionally, the urease reactivity of sera from patients diagnosed with brucellosis was examined.</p> <p>Results</p> <p>Urease encoded by the <it>ure</it>-1 operon of <it>Brucella suis </it>strain 1330 was purified to homogeneity using ion exchange and hydrophobic interaction chromatographies. The urease was purified 51-fold with a recovery of 12% of the enzyme activity and 0.24% of the total protein. The enzyme had an isoelectric point of 5, and showed optimal activity at pH 7.0 and 28–35°C. The purified enzyme exhibited a Michaelis-Menten saturation kinetics with a <it>K</it>_{<it>m </it>}of 5.60 ± 0.69 mM. Hydroxyurea and thiourea are competitive inhibitors of the enzyme with K_iof 1.04 ± 0.31 mM and 26.12 ± 2.30 mM, respectively. Acetohydroxamic acid also inhibits the enzyme in a competitive way. The molecular weight estimated for the native enzyme was between 130–135 kDa by gel filtration chromatography and 157 ± 7 kDa using 5–10% polyacrylamide gradient non-denaturing gel. Only three subunits in SDS-PAGE were identified: two small subunits of 14,000 Da and 15,500 Da, and a major subunit of 66,000 Da. The amino terminal sequence of the purified large subunit corresponded to the predicted amino acid sequence encoded by <it>ureC1</it>. The UreC1 subunit was recognized by sera from patients with acute and chronic brucellosis. By phylogenetic and cluster structure analyses, <it>ureC1 </it>was related to the <it>ureC </it>typically present in the <it>Rhizobiales</it>; in contrast, the <it>ureC2 </it>encoded in the <it>ure</it>-2 operon is more related to distant species.</p> <p>Conclusion</p> <p>We have for the first time purified and characterized an active urease from <it>B. suis</it>. The enzyme was characterized at the kinetic, immunological and phylogenetic levels. Our results confirm that the active urease of <it>B. suis </it>is a product of <it>ure</it>-1 operon.</p

The s-mote: a versatile heterogeneous multi-radio platform for wireless sensor networks applications

This paper presents a novel architecture and its implementation for a versatile, miniaturised mote which can communicate concurrently using a variety of combinations of ISM bands, has increased processing capability, and interoperability with mainstream GSM technology. All these features are integrated in a small form factor platform. The platform can have many configurations which could satisfy a variety of applications’ constraints. To the best of our knowledge, it is the first integrated platform of this type reported in the literature. The proposed platform opens the way for enhanced levels of Quality of Service (QoS), with respect to reliability, availability and latency, in addition to facilitating interoperability and power reduction compared to existing platforms. The small form factor also allows potential of integration with other mobile platforms including smart phones

Molecular targets for rapid identification of Brucella spp

BACKGROUND: Brucella is an intracellular pathogen capable of infecting animals and humans. There are six recognized species of Brucella that differ in their host preference. The genomes of the three Brucella species have been recently sequenced. Comparison of the three revealed over 98% sequence similarity at the protein level and enabled computational identification of common and differentiating genes. We validated these computational predictions and examined the expression patterns of the putative unique and differentiating genes, using genomic and reverse transcription PCR. We then screened a set of differentiating genes against classical Brucella biovars and showed the applicability of these regions in the design of diagnostic tests. RESULTS: We have identified and tested set of molecular targets that are associated in unique patterns with each of the sequenced Brucella spp. A comprehensive comparison was made among the published genome sequences of B. abortus, B. melitensis and B. suis. The comparison confirmed published differences between the three Brucella genomes, and identified subsets of features that were predicted to be of interest in a functional comparison of B. melitensis and B. suis to B. abortus. Differentiating sequence regions from B. abortus, B. melitensis and B. suis were used to develop PCR primers to test for the existence and in vitro transcription of these genes in these species. Only B. suis is found to have a significant number of unique genes, but combinations of genes and regions that exist in only two out of three genomes and are therefore useful for diagnostics were identified and confirmed. CONCLUSION: Although not all of the differentiating genes identified were transcribed under steady state conditions, a group of genes sufficient to discriminate unambiguously between B. suis, B. melitensis, and B. abortus was identified. We present an overview of these genomic differences and the use of these features to discriminate among a number of Brucella biovars

A Versatile Computational Pipeline for Bacterial Genome Annotation Improvement and Comparative Analysis, with \u3cem\u3eBrucella\u3c/em\u3e as a Use Case

We present a bacterial genome computational analysis pipeline, called GenVar. The pipeline, based on the program GeneWise, is designed to analyze an annotated genome and automatically identify missed gene calls and sequence variants such as genes with disrupted reading frames (split genes) and those with insertions and deletions (indels). For a given genome to be analyzed, GenVar relies on a database containing closely related genomes (such as other species or strains) as well as a few additional reference genomes. GenVar also helps identify gene disruptions probably caused by sequencing errors. We exemplify GenVar’s capabilities by presenting results from the analysis of four Brucella genomes. Brucella is an important human pathogen and zoonotic agent. The analysis revealed hundreds of missed gene calls, new split genes and indels, several of which are species specific and hence provide valuable clues to the understanding of the genome basis of Brucella pathogenicity and host specificity

Spectroscopic Target Selection in the Sloan Digital Sky Survey: The Quasar Sample

We describe the algorithm for selecting quasar candidates for optical spectroscopy in the Sloan Digital Sky Survey. Quasar candidates are selected via their non-stellar colors in "ugriz" broad-band photometry, and by matching unresolved sources to the FIRST radio catalogs. The automated algorithm is sensitive to quasars at all redshifts lower than z=5.8. Extended sources are also targeted as low-redshift quasar candidates in order to investigate the evolution of Active Galactic Nuclei (AGN) at the faint end of the luminosity function. Nearly 95% of previously known quasars are recovered (based on 1540 quasars in 446 square degrees). The overall completeness, estimated from simulated quasars, is expected to be over 90%, whereas the overall efficiency (quasars:quasar candidates) is better than 65%. The selection algorithm targets ultraviolet excess quasars to i^*=19.1 and higher-redshift (z>3) quasars to i^*=20.2, yielding approximately 18 candidates per square degree. In addition to selecting ``normal'' quasars, the design of the algorithm makes it sensitive to atypical AGN such as Broad Absorption Line quasars and heavily reddened quasars.Comment: 62 pages, 15 figures (8 color), 8 tables. Accepted by AJ. For a version with higher quality color figures, see http://archive.stsci.edu/sdss/quasartarget/RichardsGT_qsotarget.preprint.p