Codon-Optimization of Wild-Type Adeno-Associated Virus Capsid Sequences Enhances DNA Family Shuffling while Conserving Functionality

Adeno-associated virus (AAV) vectors have become one of the most widely used gene transfer tools in human gene therapy. Considerable effort is currently being focused on AAV capsid engineering strategies with the aim of developing novel variants with enhanced tropism for specific human cell types, decreased human seroreactivity, and increased manufacturability. Selection strategies based on directed evolution rely on the generation of highly variable AAV capsid libraries using methods such as DNA-family shuffling, a technique reliant on stretches of high DNA sequence identity between input parental capsid sequences. This identity dependence for reassembly of shuffled capsids is inherently limiting and results in decreased shuffling efficiency as the phylogenetic distance between parental AAV capsids increases. To overcome this limitation, we have developed a novel codon-optimization algorithm that exploits evolutionarily defined codon usage at each amino acid residue in the parental sequences. This method increases average sequence identity between capsids, while enhancing the probability of retaining capsid functionality, and facilitates incorporation of phylogenetically distant serotypes into the DNA-shuffled libraries. This technology will help accelerate the discovery of an increasingly powerful repertoire of AAV capsid variants for cell-type and disease-specific applications. Keywords: AAV, library, directed evolution, codon optimization, DNA shuffling, capsi

Fatores de personalidade e evolu??o cl?nica em pacientes transplantados de rim

Submitted by Setor de Tratamento da Informa??o - BC/PUCRS ([email protected]) on 2015-10-28T22:00:26Z No. of bitstreams: 1 475925 - Texto Completo.pdf: 4376108 bytes, checksum: 6bf0771ea7aea9aa52f43e4e367e7bab (MD5)Made available in DSpace on 2015-10-28T22:00:26Z (GMT). No. of bitstreams: 1 475925 - Texto Completo.pdf: 4376108 bytes, checksum: 6bf0771ea7aea9aa52f43e4e367e7bab (MD5) Previous issue date: 2015-08-31Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPESConselho Nacional de Pesquisa e Desenvolvimento Cient?fico e Tecnol?gico - CNPqThe relationship between personality and health is frequently studied in scientific research. This study aimed to investigate the clinical/biochemical course of kidney transplant patients and its relationship with personality traits in the 3rd, 6th and 9th month after transplantation. Participants were 114 kidney transplant patients, 68 men and 46 women, with an average age of 47.72 years (SD=11.4). Personality was assessed using the Brazilian Factorial Personality Inventory (BFP). Patient charts were used to record clinical/biochemical variables over nine months following transplantation (hypertension, acute rejection, graft loss, death, creatinine and estimated glomerular filtration rate/eGFR). In addition to sociodemographic variables, information was also collected on transfusions prior to transplantation and panel reactive antibodies (HLA I and II).Two groups with personality types were differentiated by psychological characteristics (hierarchical cluster analysis): Cluster 1- average Neuroticism, high Surgency, Agreeableness and Conscientiousness, and low Openness; Cluster 2- high Neuroticism, average Surgency and Agreeableness, average Conscientiousness and low Openness. There was no statistically significant difference between the two clusters in terms of hypertension, acute infection, graft loss, death and HLA I and II panel reactive antibodies. Creatinine levels, eGFR and transfusions were associated with personality types. Cluster 1 contained significantly higher creatinine levels than Cluster 2 and these remained high on all three assessment occasions, with transfusion prior to transplantation less frequent in this group. Cluster 1 exhibited a slight decrease in average eGFR over time, with an increase observed in cluster 2. In individual analyses, Neuroticism was higher in patients with lower schooling levels (p=0.002) and exhibited a significant positive correlation with average eGFR (r=0.250; p=0.008). Agreeableness was significantly higher in men patients (p<0.001) and those without infection (p=0.050). Agreeableness also showed a significant positive correlation with average creatinine levels (r= 0.250; p= 0.007) and a negative association with eGFR (r=-0.208; p=0.027). Higher levels of Conscientiousness were observed in participants with children (p=0.026) not taking medication for depression (p=0.033), as well as a positive correlation with HLA I panel reactive antibodies (r=0.223; p=0.018). In relation to Openness, the only statistically significant difference identified was in relation to schooling, with higher average values found among participants who had completed higher education (p=0.037). The results suggest that personality traits may be associated with transplant results. Monitoring these patients over a longer period may provide a better understanding of the relationship between personality traits and clinical course during the posttransplant period.A rela??o entre personalidade e sa?de tem sido objeto de estudo frequente em pesquisas cient?ficas. Este estudo teve como objetivo principal investigar a evolu??o cl?nica/laboratorial de pacientes transplantados de rim e sua rela??o com tra?os de personalidade no 30, 60 e 90 m?s ap?s o transplante. Participaram do estudo 114 pacientes transplantados renais, 68 homens e 46 mulheres, com idade m?dia de 47,72 anos (DP=11,4). Para a avalia??o da personalidade foi utilizada a Bateria Fatorial de Personalidade (BFP). A partir dos prontu?rios, foram registradas as vari?veis cl?nicas/laboratoriais ao longo de nove meses ap?s o transplante (hipertens?o arterial (HA), rejei??o aguda, infec??o, perda do enxerto, ?bito, creatinina e taxa de filtra??o glomerular estimada/ TFGe). Al?m das vari?veis sociodemogr?ficas, coletaram-se ainda informa??es sobre as transfus?es pr?vias ao transplante e Painel de ant?genos HLA (classe I e II). Dois grupos com perfis de personalidade foram diferenciados pelas caracter?sticas psicol?gicas (an?lise de cluster hierarquizado): Cluster 1- Neuroticismo m?dio, Extrovers?o, Socializa??o e Realiza??o altos e Abertura baixo; Cluster 2- Neuroticismo alto, Extrovers?o, Socializa??o, Realiza??o m?dio e Abertura baixo. Para HA, rejei??o aguda, infec??o, perda do enxerto, ?bito e Painel de ant?genos HLA, classe I e II n?o houve diferen?a estatisticamente significativa entre os dois Clusters. Os n?veis de creatinina, TFGe e transfus?es tiveram associa??o com os perfis de personalidade. Verificou-se que o Cluster 1 concentrou n?veis de creatinina significativamente mais altos que o Cluster 2 e que se mantiveram mais elevados nos tr?s momentos de avalia??o, sendo a transfus?o pr?via ao transplante menos frequente neste grupo. No Cluster 1 ocorreu leve redu??o da m?dia de TFGe ao longo do tempo, enquanto que no Cluster 2 houve aumento. Nas an?lises individuais, Neuroticismo apresentou m?dias mais elevadas nos pacientes com menor escolaridade (p=0,002) e correla??o significativa e positiva com a m?dia de TFGe (r=0,250; p=0,008). O fator Socializa??o apresentou m?dias significativamente mais elevadas nos pacientes homens (p<0,001) e em pacientes sem infe??o (p=0,050). Al?m disto, o fator Socializa??o indicou correla??o significativa positiva com a m?dia de creatinina (r= 0,250; p= 0,007) e negativa com TFGe (r=-0,208; p=0,027). O fator Realiza??o apontou n?veis mais altos nos participantes com filhos (p=0,026) e sem uso de medica??es para depress?o (p=0,033), al?m de correla??o positiva com Reatividade contra painel de ant?genos HLA I (r=0,223; p=0,018). No fator Abertura a ?nica diferen?a estat?stica significativa identificada foi em rela??o ? escolaridade que apontou maiores m?dias nos investigados com Ensino Superior (p=0,037). Os resultados sugerem que as caracter?sticas de personalidade podem estar associadas aos resultados do transplante O acompanhamento destes pacientes durante um per?odo maior poder? levar a um melhor entendimento da rela??o entre fatores de personalidade e evolu??o cl?nica no per?odo p?s-transplante

Long-range angular correlations on the near and away side in p-Pb collisions at root S-NN=5.02 TeV

Angular correlations between charged trigger and associated particles are measured by the ALICE detector in p-Pb collisions at a nucleon-nucleon centre-of-mass energy of 5.02 TeV for transverse momentum ranges within 0.5 < P-T,P-assoc < P-T,P-trig < 4 GeV/c. The correlations are measured over two units of pseudorapidity and full azimuthal angle in different intervals of event multiplicity, and expressed as associated yield per trigger particle. Two long-range ridge-like structures, one on the near side and one on the away side, are observed when the per-trigger yield obtained in low-multiplicity events is subtracted from the one in high-multiplicity events. The excess on the near-side is qualitatively similar to that recently reported by the CMS Collaboration, while the excess on the away-side is reported for the first time. The two-ridge structure projected onto azimuthal angle is quantified with the second and third Fourier coefficients as well as by near-side and away-side yields and widths. The yields on the near side and on the away side are equal within the uncertainties for all studied event multiplicity and p(T) bins, and the widths show no significant evolution with event multiplicity or p(T). These findings suggest that the near-side ridge is accompanied by an essentially identical away-side ridge. (c) 2013 CERN. Published by Elsevier B.V. All rights reserved