Marie Togashi

O conceito de pessoa variou historicamente, com controvérsias de natureza hegemônica. Portanto, apresenta-se à sociedade moderna o desafio de revalorizar e ressignificar o ser humano como pessoa, independente de qualquer outra característica adjetiva ² . Na perspectiva bioética, trata-se de unidade de corpo e espírito; e uma totalidade porque deve ser considerada em todas as suas dimensões (física, psíquica, espiritual, social e moral) ³ . Refere-se não só ao sujeito de direito, mas ao ser concreto, com aptidões, necessidades, possibilidades e, principalmente, singularidades ⁴ .

Peroxisome proliferator-activated receptors (PPARs) are involved in the control of carbohydrate and lipid metabolism and are considered important targets to treat diabetes mellitus and metabolic syndrome. The available PPAR ligands have several side effects leading to health risks justifying the search for new bioactive ligands to activate the PPAR subtypes, in special PPARδ, the less studied PPAR isoform. Here, we used a structure-based virtual screening protocol in order to find out new PPAR ligands. From a lead-like subset of purchasable compounds, we identified 5 compounds with potential PPAR affinity and, from preliminary in vitro assays, 4 of them showed promising biological activity. Therefore, from our in silico and in vitro protocols, new PPAR ligands are potential candidates to treat metabolic diseases.

Thiazolidinediones (TZDs) act through peroxisome proliferator activated receptor (PPAR) γ to increase insulin sensitivity in type 2 diabetes (T2DM), but deleterious effects of these ligands mean that selective modulators with improved clinical profiles are needed. We obtained a crystal structure of PPARγ ligand binding domain (LBD) and found that the ligand binding pocket (LBP) is occupied by bacterial medium chain fatty acids (MCFAs). We verified that MCFAs (C8–C10) bind the PPARγ LBD in vitro and showed that they are low-potency partial agonists that display assay-specific actions relative to TZDs; they act as very weak partial agonists in transfections with PPARγ LBD, stronger partial agonists with full length PPARγ and exhibit full blockade of PPARγ phosphorylation by cyclin-dependent kinase 5 (cdk5), linked to reversal of adipose tissue insulin resistance. MCFAs that bind PPARγ also antagonize TZD-dependent adipogenesis in vitro. X-ray structure B-factor analysis and molecular dynamics (MD) simulations suggest that MCFAs weakly stabilize C-terminal activation helix (H) 12 relative to TZDs and this effect is highly dependent on chain length. By contrast, MCFAs preferentially stabilize the H2-H3/β-sheet region and the helix (H) 11-H12 loop relative to TZDs and we propose that MCFA assay-specific actions are linked to their unique binding mode and suggest that it may be possible to identify selective PPARγ modulators with useful clinical profiles among natural products.

Protein quality and stability are critical during protein purification for X‐ray crystallography. A target protein that is easy to manipulate and crystallize becomes a valuable product useful for high‐throughput crystallography for drug design and discovery. In this work, a single surface mutation, D355R, was shown to be crucial for converting the modestly stable monomeric ligand binding domain of the human thyroid hormone receptor (TR LBD) into a stable dimer. The structure of D335R TR LBD mutant was solved using X‐ray crystallography and refined to 2.2 Å resolution with R_free/R values of 24.5/21.7. The crystal asymmetric unit reveals the TR dimer with two molecules of the hormone‐bound LBD related by twofold symmetry. The ionic interface between the two LBDs comprises residues within loop H10‐H11 and loop H6‐H7 as well as the C‐terminal halves of helices 8 of both protomers. Direct intermolecular contacts formed between the introduced residue Arg 355 of one TR molecule and Glu 324 of the second molecule become a part of the extended dimerization interface of 1330 Ų characteristic for a strong complex assembly that is additionally strengthened by buffer solutes. Proteins 2009.

The development of nuclear hormone receptor antagonists that directly inhibit the association of the receptor with its essential coactivators would allow useful manipulation of nuclear hormone receptor signaling. We previously identified 3-(dibutylamino)-1-(4-hexylphenyl)-propan-1-one (DHPPA), an aromatic β-amino ketone that inhibits coactivator recruitment to thyroid hormone receptor β (TRβ), in a high-throughput screen. Initial evidence suggested that the aromatic β-enone 1-(4-hexylphenyl)-prop-2-en-1-one (HPPE), which alkylates a specific cysteine residue on the TRβ surface, is liberated from DHPPA. Nevertheless, aspects of the mechanism and specificity of action of DHPPA remained unclear. Here, we report an x-ray structure of TRβ with the inhibitor HPPE at 2.3-Å resolution. Unreacted HPPE is located at the interface that normally mediates binding between TRβ and its coactivator. Several lines of evidence, including experiments with TRβ mutants and mass spectroscopic analysis, showed that HPPE specifically alkylates cysteine residue 298 of TRβ, which is located near the activation function-2 pocket. We propose that this covalent adduct formation proceeds through a two-step mechanism: 1) β-elimination to form HPPE; and 2) a covalent bond slowly forms between HPPE and TRβ. DHPPA represents a novel class of potent TRβ antagonist, and its crystal structure suggests new ways to design antagonists that target the assembly of nuclear hormone receptor gene-regulatory complexes and block transcription.

The role of thyroid hormone [l-3,5,3′-triiodothyronine (T₃)] and the thyroid hormone receptor (TR) in regulating growth, development, and metabolic homeostasis is well established. It is also emerging that T₃ is associated with oxidative stress through the regulation of the activity of superoxide dismutase-1 (SOD-1), a key enzyme in the metabolism of oxygen free radicals. We found that T₃ reverses the activation of the SOD-1 promoter caused by the free radical generators paraquat and phorbol 12-myristate 13-acetate through the direct repression of the SOD-1 promoter by liganded TR. Conversely, the SOD-1 promoter is significantly stimulated by unliganded TRs. This regulation requires the DNA-binding domain of the TR, which is recruited to an inhibitory element between -157 and +17 of the SOD-1 promoter. TR mutations, which abolish recruitment of coactivator proteins, block repression of the SOD-1 promoter. Conversely, a mutation that inhibits corepressor binding to the TR prevents activation. Together, our findings suggest a mechanism of negative regulation in which TR binds to the SOD-1 promoter but coactivator and corepressor binding surfaces have an inverted function. This effect may be important in T₃ induction of oxidative stress in thyroid hormone excess.