In response to the growing rates of antibiotic resistance in human bacterial pathogens, this study explores the natural products involved in the defensive symbiosis between actinobacteria and fungus-growing ants to uncover new potential antibiotics. This study also seeks to understand the function of natural antibiotics in their ecological contexts, especially those involved in defensive symbioses. Defensive symbiosis can be a beneficial platform for discovering useful antibiotics, because antibiotics in these relationships must be able to selectively inhibit enemies without harming hosts, and are therefore likely more specific and less toxic. Pseudonocardia sp. associated with Trachymyrmex septentrionalis ants demonstrated antibiotic activity against several gram-positive bacteria. Therefore, the natural products from this strain were extracted and purified through activity-guided fractionation. Using mass spectrometry, the structure of the active compound was elucidated as GE37468A, an antibiotic that has been previously identified from Streptomyces sp. ATCC 55365 from Italy. This compound had never before been characterized in a defensive symbiosis, which demonstrates the use of the molecule in a new context. Antibiotic GE37468A is a thiopeptide, which is a group of antibiotics that has previously demonstrated strong activity against many gram-positive bacteria, including bacterial human pathogens. Due to its potency against dangerous bacteria and its likely low toxicity, this antibiotic could therefore hold potential pharmacological uses.

Diss. (sammanfattning) Stockholm : Karol inst., 2003. / Härtill 6 uppsatser.

Thesis (Ph. D.)--West Virginia University, 2010. / Title from document title page. Document formatted into pages; contains xiv, 180 p. : ill. (some col.). Includes abstract. Includes bibliographical references.

No Brasil, poucos estudos investigaram a prevalência de medicamentos potencialmente inapropriados (MPIs) em idosos ambulatoriais. Este estudo visa determinar a prevalência de MPIs prescritos para estes pacientes, identificando os mais comumente envolvidos e verificando se a idade, o sexo e o número de medicamentos estão relacionados à prescrição de tais medicamentos. Foram coletadas prescrições de 3070 pacientes idosos (60 anos) em banco de dados, provenientes dos ambulatórios de Geriatria e Clínica Geral de um hospital universitário de atenção terciária em São Paulo-Brasil entre fevereiro e maio de 2008, que foram divididas de acordo com o sexo e faixa etária (60-69; 70-79 e 80). Os critérios de Beers versão 2003 foram utilizados para a avaliação de MPIs. A maior parte da casuística foi composta por mulheres em ambos os ambulatórios (66,6% na Clínica Geral e 77,7% na Geriatria). Os pacientes da Clínica Geral apresentaram média de idade de 71,3 anos e os da Geriatria, 80,1 anos. Na Clínica Geral a prevalência média de prescrição de MPIs foi de 37,6% e na Geriatria de 26,9%, sendo que em ambos as mulheres de 60-69 foram as que apresentaram a maior prevalência destes medicamentos. Os MPIs mais prescritos nos dois ambulatórios foram o carisoprodol, a fluoxetina e a amitriptilina, sendo que houve diferenças nos perfis de prescrições entre homens e mulheres. A chance de uso de MPI no sexo feminino é maior que no masculino (p<0,001); a chance de uso de MPI na faixa de 70-79 anos é menor que na faixa de 60-69 anos (p=0,030), assim como na faixa de 80 (p=0,024). Estas conclusões não dependem do ambulatório (p=0,164).O efeito de ambulatório depende do número de medicamentos (p=0,009). Se o número de medicamentos é < 9 a chance de uso de MPI na Clínica Geral é maior que na Geriatria (p=0,041). Quando o número de medicamentos é 7 ou 8, a chance de uso de MPI é maior do que quando são prescritos 1-4 medicamentos (p<0,001), nos dois ambulatórios (p=0,098). Quando são usados 9 medicamentos, a chance de uso de MPI depende do ambulatório (p=0,044). Na Geriatria, a chance de uso de um MPI é 8,2 vezes a RC na categoria 1-4 medicamentos; enquanto que na Clínica Geral a razão de chances é 4,6. As prevalências de MPIs encontradas foram semelhantes ao relatado na literatura e estão correlacionadas ao sexo feminino. A chance de prescrição de MPIs foi menor em pacientes com 70 anos; observou-se que se o número de medicamentos for <9, a chance de uso de MPI na Clínica Geral é maior que na Geriatria, porém, se o número de medicamentos for 9 não há diferença entre as chances de uso de MPI nos dois ambulatórios / In Brazil, few studies have investigated the prevalence of potentially inappropriate medications (PIMs) among elderly outpatients. This study aimed to determine the prevalence of PIMs prescribed to such patients, identify the medications most commonly involved and investigate whether age, sex and number of medications are related to the prescription of such medications. Prescriptions issued to 3,070 elderly patients (60 years) were gathered from a database. These patients were attended at the geriatric and general clinical outpatient services of a tertiary-level university hospital in São Paulo, Brazil, between February and May 2008. They were divided according to sex and age group (60- 69; 70-79; and 80 years). The Beers criteria (2003 version) were used to evaluate PIMs. The majority of the sample comprised women, at both outpatient services (66.6% in the general clinic and 77.7% in geriatrics). The mean age of the general clinical patients was 71.3 years and the mean for the geriatric patients was 80.1 years. At the general clinic, the mean prevalence of prescriptions of PIMs was 37.6%, and it was 26.9% at the geriatric clinic. At both outpatient services, women aged 60-69 years presented the highest prevalence of such medications. The PIMs most prescribed at the two outpatient services were carisoprodol, fluoxetine and amitriptyline, and there were differences in the prescription profiles between the men and women. The chances of using PIMs were greater for the women than for the men (p < 0.001). The chances of using PIMs in the 70-79 years group were lower than in the 60-69 years group (p = 0.030), and likewise for the group 80 years (p = 0.024). These conclusions were independent of the outpatient service (p = 0.164). The outpatient effect depended on the number of medications (p = 0.009). If the number of medications was < 9, the chances of using PIMs at the general clinic were greater than the chances at the geriatric clinic (p = 0.041). When the number of medications was 7 or 8, the chances of using PIMs were greater than when prescribed 1-4 medications (p < 0.001), at both outpatient services (p = 0.098). When 9 medications were used, the chances of using PIMs depended on the outpatient service (p = 0.044). At the geriatric clinic, the chances of using PIMs were 8.2 times greater than the chances in the category of 1-4 medications; while at the general clinic, the odds ratio was 4.6. The prevalence of PIMs encountered was similar to what has been reported in the literature, and it correlated with female sex. The chances of being prescribed PIMs were lower among patients 70 years. If the number of medications was < 9, the chances of using PIMs at the general clinic were greater than the chances at the geriatric clinic. However, if the number of medications was 9, there was no difference in the chances of using PIMs between the two outpatient services

Resistance to glucocorticoid (GC) therapy results in poor prognosis for acute lymphoblastic leukemia (ALL) patients. Utilizing a whole genome shRNA screen our lab identified several novel mechanisms of GC resistance. My thesis work established that an orphan nuclear receptor, the Estrogen Related Receptor Beta (ESRRB), is critical for induction of apoptotic genes following treatment with the GC dexamethasone. ESRRB has mostly been implicated in maintenance of pluripotency in mouse embryonic stem cells. We find that repression of ESRRB results in GC resistance in ALL and define ESRRB as a novel cooperating transcription factor in GC-induced gene expression. We also show that agonists to ESRRB synergize with dexamethasone and increase dexamethasone induced apoptosis in relapse ALL patient samples. Interestingly, our shRNA screen identified another factor important in stem cell maintenance: SOX2. While we originally hypothesized that ESRRB and SOX2 may cooperate in ALL, RNA-sequencing studies revealed that these factors mediate GC resistance by independent mechanisms. Our data define SOX2 as a repressor of key signaling pathways in ALL. Upon SOX2 knockdown, we observe activation of pro-survival gene expression including activation of the MAPK pathway, which has previously been implicated in GC resistance. MAPK activation may be explained by an increase in EGFR expression observed in Sox2 knockdown cells and GC resistant patients, suggesting EGFR inhibitors may re-sensitize patients to GCs. Overall my thesis work identifies mechanisms of GC resistance in ALL and utilizes these findings to define novel therapeutic strategies for GC resistant ALL patients.

The aim of the thesis was to analyze the historical pharmaceutical preparations, including the determination of the active substance and identify theirs possible degradation products. A historical pharmaceutical preparation of naloxone was analyzed by mass spectrometry. Historical pharmaceutical preparations of adrenaline and ephedrine were analyzed by UHPLC-MS and were quantified using a calibration curve. In the historical injection solution of naloxone, "NARCAN", dated around 1980, there were no significant degradation products and the measured mass and UV spectrum was consistent with the spectrum of naloxone. The analyzed sample of naloxone was stable even after 35 years of storage. In the analyzed historical injection solution of adrenaline, "Adrenalin Hydrochlor., Dr. Heisler" (dated between 1917 and 1938) was determined 5.26 ± 0.11 % of the declared amount of adrenaline. In the measured spectras were noticeable degradation products, which have not been described in the literature yet and their identification was beyond the scope of this paper. The analyzed sample of adrenaline was almost completely degraded during about ninety years. The stability test carried out with four standard solutions of adrenaline proved influence of oxygen, light, temperature and time on the degradation of adrenaline. In...

HIV-1 protease is a very important drug target for AIDS therapy. Nine protease inhibitors have been proved by FDA and used in AIDS treatment. Due to the high replication rate and the lack of fidelity of the HIV-1 reverse transcriptase, HIV-1 virus developed various drug-resistant variants. Although experimental methods such as crystallography and isothermal titration calorimetry provide structural and thermodynamic data on drug-resistant variants, they are unable to discern the mechanism by which the mutations confer resistance to inhibitors. Understanding the drug-resistance mechanism is crucial for developing new inhibitors more tolerant to the drug-resistant mutations. Computational methods such as free energy calculations and molecular dynamic simulations can provide insights to the drug resistance mechanism at an atomic level. In this thesis, I have focused on the elucidation of the energetic and dynamics of key drug-resistant variants of HIV-1 protease. Two multi-drug resistant variants, in comparison with wild-type HIV-1 protease were used for the comparisons: Flap+ (L10I, G48V, I54V, and V82A) which contains a combination of flap and active site mutations and ACT (V82T, I84V) that only contains active site mutations. In Chapter II, I applied free energy simulations and decomposition methods to study the differential mechanism of resistance to the two variants, Flap+ and ACT, to the recently FDA-approved protease inhibitor darunavir (DRV). In this study, the absolute and relative binding free energies of DRV with wild-type protease and the two protease variants were calculated with MM-PB/GBSA and thermodynamic integration methods, respectively. And the predicted results are in good agreement with the ITC experimental results. Free energy decomposition elucidates the mutations alter not only its own interaction with DRV but also other residues by changing the geometry of binding pocket. And the VdW interactions between the bis-THF group of DRV is predominant even in the drug-resistant variants. At the end of this chapter, I offer suggestions on developing new inhibitors that are based on DRV but might be less susceptible to drug-resistant mutations. In Chapter III, 20-ns MD simulations of the apo wildtype protease and the apo drug-resistant protease variant Flap+ are analyzed and compared. In these studies, these mutations have been found to decrease the protease flexibility in the apo form but increase the mobility when the protease is binding with inhibitor. In Chapter IV, more details of the free energy simulation and decomposition are discussed. NMR relaxation experiments were set up as a control for the MD simulation study of the dynamics of the Flap+ variant. The difficulty of finishing the NMR experiment is discussed and the solution and some preliminary results are shown. In summary, the scope of this thesis was to use computational methods to study drug-resistant protease variants’ thermodynamic and dynamic properties to illuminate the mechanism of protease drug resistance. This knowledge will contribute to rational design of new protease inhibitors which bind more tightly to the protease and hinder the development of drug-resistant mutations.

Background To investigate plasma microRNA (miRNA) profiles indicative of hepatotoxicity in the setting of lethal acetaminophen (APAP) toxicity in mice. Methods Using plasma from APAP poisoned mice, either lethally (500 mg/kg) or sublethally (150 mg/kg) dosed, we screened commercially available murine microRNA libraries (SABiosciences, Qiagen Sciences, MD) to evaluate for unique miRNA profiles between these two dosing parameters. Results We distinguished numerous, unique plasma miRNAs both up- and down-regulated in lethally compared to sublethally dosed mice. Of note, many of the greatest up- and down-regulated miRNAs, included, but were not limited to, 574-5p, 466g, 466f-3p, 375, 29c, and 148a. There was a statistically significant increase in alanine aminotransferase levels in the lethal compared to sublethal APAP dosing groups at the 12 h time point ( P < 0.001). There was 90% mortality in the lethally compared to sublethally dosed mice at the 48 h time point ( P = 0.011). Conclusion We identified unique plasma miRNAs both up- and down-regulated in lethally dosed APAP poisoned mice.

Title: Treatment of CMV Vitritis in a Preterm Newborn Author’s Section: Remil Simon1, Darshan Shah1, Peter Blosser1, Demetrio Macariola1, Jeffrey Carlsen2 1.Department of Pediatrics, Quillen College of Medicine, East Tennessee State University, Johnson City, TN 2.Johnson City Eye Clinic, Johnson City, TN Body: Cytomegalovirus (CMV) infection in the neonate is an infrequent occurrence in the developing world, and observing the symptoms of ocular CMV infection such as vitritis is rare. Treating CMV infection promptly is necessary to prevent mortality and potential neurological deficits including blindness and hearing loss. We encountered a preterm infant presenting with CMV sepsis immediately after birth. Our question was: will the current standard of treatment for CMV sepsis prevent CMV ocular infection? With our method of treatment, we followed the current standard of treatment for CMV infection by administering intravenous Gancyclovir for 6 weeks and oral Valgancyclovir for 6 months. Despite using the standard treatment to prevent neurological sequelae, the patient developed CMV vitritis and retinitis bilaterally. Although the treatment did not prevent CMV ocular infection, the severity of CMV retinitis and vitritis improved with treatment, and full resolution of vitritis was noted by day of life 61.

Neste trabalho, é proposto o desenvolvimento de instrumentação e procedimentos analíticos automáticos empregando fotometria em fase sólida para determinação de zinco em produtos farmacêuticos e água. Para implementação do procedimento analítico, todo o sistema foi acoplado a um computador através de uma interface eletrônica. Um software escrito em linguagem QuickBASIC 4.5 permite que o computador efetue o controle da adição das soluções da amostra e do eluente e faça aquisição de dados. O sistema de detecção é constituído de uma cela de fluxo contendo, um LED e um fotodiodo. A geometria da cela de fluxo possibilitava variar o comprimento do caminho óptico. O procedimento para determinação do zinco foi baseado na retenção do analito na fase sólida (TAN-C18), previamente inserida na cela de fluxo, seguido de uma etapa de eluição. Com os parâmetros analíticos otimizados obteve-se resposta linear na faixa de 0,05 a 0,85 mg L-1 (R=0,995), limite detecção de 9,3 \'mü\'g L-1, coeficiente de variação de 1,4% (n=10) e frequência de amostragem de 36 det h-1. O módulo de análise foi aplicado em amostras de produtos farmacêuticos e a exatidão dos resultados foi averiguada comparando com os resultados obtidos por espectrometria de emissão óptica com plasma acoplado indutivamente ICP-OES. Aplicando-se tratamento estatístico apropriado, observou-se que não havia diferença significativa ao nível de confiança de 95 %. Estes resultados comprovam a viabilidade do emprego de fotômetro de LED em fotometria em fase sólida / In this work, it is propose the instrumentation and automatic analytic procedures development using solid phase spectrophotometry for determination of zinc in pharmaceutical preparations and water. For analytic procedure implementation, the whole system was coupled to a computer through an electronic interface. A written software in language QuickBASIC 4.5 allows the computer makes the sample solutions addition control and of eluent and do data acquisition. The detection system is constituted of a flow cell contend, a LED and a photodiode. The flow cell geometry enabled vary the length of the optical path. The procedure for zinc determination was going based in analyte retention in the solid phase (TAN-C18), previously inserted in the flow cell, followed by an elution stage. With the optimized analytic parameters it btained lineal answer in the band of 0,05 to 0,85 mg L-1 (R=0,995), limit detection of 9,3 \'mü\'g L-1, variation coefficient of 1,4% (n=10) and sampling throughput of 36 det h-1. The analysis module was going applied in pharmaceutical preparations samples and the exactness of the results was going ascertained comparing with the results obtained for inductively coupled plasma optic emission spectrometry of with ICP- its. Applying appropriated statistical treatment, it observed that there wasn\'t significant difference to the reliable level of 95 %. These results prove photometer job viability of LED in photometry in solid phase