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Small molecule screening is commonly used to discover lead compounds for drug development, but it can also be a powerful way to identify chemical probes for studying biological mechanisms. Our lab uses small molecules to study the mechanisms by which the protozoan parasite Toxoplasma gondii infects and replicates within its hosts. In this work, we employed a fluorescence-based assay to screen the Medicines for Malaria Venture (MMV) Open Access Malaria box for compounds that affect T. gondii growth. The box contains 400 previously identified small-molecule inhibitors of the related parasite, Plasmodium falciparum. We identified 79 hits, including a 2,4-diaminoquinazoline (MMV006169; IC50=1.15µM) that strongly inhibits T. gondii intracellular replication and invasion with no evidence of toxicity to mammalian cells. Extensive structure-activity relationship analyses with T. gondii identified a number of analogs with changed potency and altered effects on replication and invasion. These structure-activity analyses provided the information necessary to synthesize a bivalent chemical inducer of dimerization (CID) containing MMV006169 for use in yeast three-hybrid experiments. Yeast growth competition assays showed that this CID is capable of entering the yeast nucleus, as required for yeast three-hybrid screening. Yeast three-hybrid was used in a targeted format to test the hypothesis that MMV006169 works by inhibiting parasite CDC48, an ATPase involved in trafficking and the degradation of misfolded proteins. Large-scale cDNA library screening by yeast three-hybrid suggests that the compound may instead be working through inhibition of a host cell target. This work has provided insight into how MMV006169 affects the parasite's lytic cycle and generated a testable hypothesis for the biologically relevant target of the compound.

Submitted by JÚLIO HEBER SILVA (julioheber@yahoo.com.br) on 2017-03-15T17:36:59Z No. of bitstreams: 2 Dissertação - Sidney Tadeu Santiago Costa - 2017.pdf: 1406618 bytes, checksum: 291719e6f7eaaff496ec405e241ce518 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-03-20T12:39:15Z (GMT) No. of bitstreams: 2 Dissertação - Sidney Tadeu Santiago Costa - 2017.pdf: 1406618 bytes, checksum: 291719e6f7eaaff496ec405e241ce518 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-03-20T12:39:15Z (GMT). No. of bitstreams: 2 Dissertação - Sidney Tadeu Santiago Costa - 2017.pdf: 1406618 bytes, checksum: 291719e6f7eaaff496ec405e241ce518 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-03-03 / With the note gotten in the Exame Nacional do Ensino Médio - ENEM the students can applay the vacant in diverse public institutions of superior education and programs of the government, for example, the program Universidade para Todos(Prouni) and the Fundo de Financiamento Estudantil (Fies). The ENEM uses a methodology of correction of the objective questions called Theory of Reply to the Item - TRI, that has some aspects that are different of the Classic Theory of the Tests - TCT. The main factor that determines the result of a citizen in a avaliativo process where if uses the TCT, is the number of correct answers, while in the TRI, beyond the amount of rightnesss is basic if to analyze which answers they are correct. The objective of this work is to explain what it is the TRI and as if it applies this methodology in evaluations of wide scale. A historical boarding of the logistic models used by the TRI and the justification of the existence of each parameter will be made that composes the main equation of the modeling. To determine each parameter that composes the model of the TRI and to calculate the final note of each candidate, a procedure of called optimization will be used Method of Maximum Probability - MMV. The computational tools in the work had been software R, with packages developed for application of the TRI and the Visual programming language beginner’s all-purpose symbolic instruction code to program functions, called as macros, in electronic spread sheets. / Com a nota obtida no Exame Nacional do Ensino Médio - ENEM os estudantes podem se candidatar a vagas em diversas instituições públicas de ensino superior e programas do governo, por exemplo, o programa Universidade para Todos (Prouni) e o Fundo de Financiamento Estudantil (Fies). O ENEM utiliza uma metodologia de correção das questões objetivas denominada Teoria de Resposta ao Item - TRI, que possui vários aspectos que são diferentes da Teoria Clássica dos Testes - TCT. O principal fator que determina o resultado de um sujeito em um processo avaliativo onde se utiliza a TCT, é o número de respostas corretas, enquanto na TRI, além da quantidade de acertos é fundamental se analisar quais respostas estão corretas. O objetivo deste trabalho é explicar o que é a TRI e como se aplica essa metodologia em avaliações de larga escala. Será feita uma abordagem histórica dos modelos logísticos utilizados pela TRI e a justificativa da existência de cada parâmetro que compõe a equação principal da modelagem. Para determinar cada parâmetro que compõe o modelo da TRI e calcular a nota final de cada candidato, será utilizado um procedimento de otimização denominado Método da Máxima Verossimilhança - MMV. As ferramentas computacionais no trabalho foram o software R, com pacotes desenvolvidos para aplicação da TRI e a linguagem de programação Visual Basic para programar funções, denominadas como macros, em planilhas eletrônicas.

The ophthalmic trigeminal placode (opV) is the birth place of one cell type of sensory neurons contributing to the trigeminal ganglion. Signals from the neural tube induce placodal identity within the surface ectoderm. Specified opV placode cells then up-regulate neuron differentiation markers and migrate to the ganglion. Several molecular pathways have been shown to act in opV placode cell development. Despite this, signals that specify individual neurons from within the opV placode remain unknown. However, it is known that components of the Notch signaling pathway are expressed in the opV placode. I tested the role of Notch signaling in opV placode development by separately inhibiting and over-activating the pathway. Using DAPT, an inhibitor of gamma-secretase, I inhibited Notch signaling in 13-15 somite stage chick embryo heads. Attenuated Notch signaling caused increased neuronal differentiation of opV cells at 13-15 somites. I also observed an increase in migratory opV placode (Pax3+) cells in the mesenchyme and expression of neuronal marker Islet1 in the ectoderm. Further, I activated Notch signaling by misexpressing the Notch intracellular domain (NICD) by in ovo electroporation of 10-12 somite stage chick embryos. This resulted in Pax3+ targeted cells failing to differentiate and remain instead in the ectoderm. Thus, Notch/Delta signaling plays an important role in selecting ophthalmic trigeminal cells to differentiate and migrate to the trigeminal ganglion.

To overcome the curse of dimensionality, dimension reduction is important and necessary for understanding the underlying phenomena in a variety of fields. Dimension reduction is the transformation of high-dimensional data into a meaningful representation in the low-dimensional space. It can be further classified into feature selection and feature extraction. In this thesis, which is composed of four projects, the first two focus on feature selection, and the last two concentrate on feature extraction. The content of the thesis is as follows. The first project presents several efficient methods for the sparse representation of a multiple measurement vector (MMV); some theoretical properties of the algorithms are also discussed. The second project introduces the NP-hardness problem for penalized likelihood estimators, including penalized least squares estimators, penalized least absolute deviation regression and penalized support vector machines. The third project focuses on the application of manifold learning in the analysis and prediction of 24-hour electricity price curves. The last project proposes a new hessian regularized nonlinear time-series model for prediction in time series.