Estudio de las regiones implicadas en la funcionalidad y resistencia a fármacos de la ADN topoisomerasa IB de leishmania

278 p.Las leishmaniosis son un complejo de enfermedades causadas por parásitos del género Leishmania spp. que presentan una elevada prevalencia a nivel mundial. Afectan sobretodo a países en vías de desarrollo, aunque recientemente, debido a diversos factores, se ha producido una propagación hacia regiones aparentemente exentas de riesgo. Por todo ello, la OMS considera la leishmaniosis una enfermedad priorizada dentro de su ¿Special Programme for Research and Training in Tropical Diseases¿. Dado que no existen vacunas eficaces contra estas enfermedades y que los tratamientos médicos actuales están basados en fármacos obsoletos con una elevada toxicidad, es vital abrir nuevas vías de lucha frente a este parásito mediante la identificación de nuevas dianas terapéuticas y el consiguiente diseño de inhibidores específicos. Recientemente, se han identificado las ADN topoisomerasas como prometedoras dianas de intervención terapéutica en tripanosomátidos. Estas enzimas están involucradas en resolver los problemas topológicos generados durante la replicación, trascripción y recombinación del ADN. La ADN topoisomerasa IB de Leishmania spp. es un complejo heterodimérico con escasa homología con respecto a la proteína del hospedador mamífero, por lo que las regiones no conservadas de la enzima de Leishmania constituyen una diana terapéutica altamente selectiva. En el presente trabajo se ha realizado un estudio exhaustivo de las regiones no conservadas de la ADN topoisomerasa IB de Leishmania, identificando las secuencias específicas responsables de las interacciones entre las dos subunidades del heterodímero así como de la conformación del dominio linker de la proteína, probando además la importancia de este dominio tanto en la capacidad de relajación de la enzima como en la sensibilidad a diferentes tipos de inhibidores. En segundo lugar se han identificado y caracterizado las tres secuencias responsables de la translocación de la proteína al interior del núcleo; una en el extremo C-terminal de la subunidad grande y dos en la subunidad pequeña. La primera de ellas correspondió con una secuencia de 10 aminoácidos localizados en el extremo N-terminal, mientras la que la segunda fue delimitada en las inmediaciones del residuo Tyr catalítico del extremo C-terminal. En este estudio no se encontró ninguna señal específica de translocación al kinetoplasto. Finalmente se evaluó la enzima de Leishmania como diana molecular de intervención terapéutica con tres familias de compuestos: derivados de la camptotecina, indenoisoquinolinas y ácidos grasos tanto acetilénicos como alquinoicos. Con ello se demostró que varios derivados camptotecínicos e indenoisoquinolínicos, especialmente el gimatecan y el LMP400, son inhibidores preferenciales de la enzima de Leishmania frente a la humana, encontrándose una buena correlación entre los ensayos in vitro e in vivo realizados. De igual manera se demostró el uso potencial de los ácidos grasos insaturados de cadena larga como inhibidores de la ADN topoisomerasa, por un mecanismo de acción diferente al de la camptotecina, en el que dichos ácidos grasos impiden el corte del ADN mediado por la enzima pero no la unión entre ambos

Trypanosomatids topoisomerase re-visited. New structural findings and role in drug discovery

AbstractThe Trypanosomatidae family, composed of unicellular parasites, causes severe vector-borne diseases that afflict human populations worldwide. Chagas disease, sleeping sickness, as well as different sorts of leishmaniases are amongst the most important infectious diseases produced by Trypanosoma cruzi, Trypanosoma brucei and Leishmania spp., respectively. All these infections are closely related to weak health care services in low-income populations of less developed and least economically developed countries. Search for new therapeutic targets in order to hit these pathogens is of paramount priority, as no effective vaccine is currently in use against any of these parasites. Furthermore, present-day chemotherapy comprises old-fashioned drugs full of important side effects. Besides, they are prone to produce tolerance and resistance as a consequence of their continuous use for decades. DNA topoisomerases (Top) are ubiquitous enzymes responsible for solving the torsional tensions caused during replication and transcription processes, as well as in maintaining genomic stability during DNA recombination. As the inhibition of these enzymes produces cell arrest and triggers cell death, Top inhibitors are among the most effective and most widely used drugs in both cancer and antibacterial therapies. Top relaxation and decatenation activities, which are based on a common nicking–closing cycle involving one or both DNA strands, have been pointed as a promising drug target. Specific inhibitors that bind to the interface of DNA-Top complexes can stabilize Top-mediated transient DNA breaks. In addition, important structural differences have been found between Tops from the Trypanosomatidae family members and Tops from the host. Such dissimilarities make these proteins very interesting for drug design and molecular intervention. The present review is a critical update of the last findings regarding trypanosomatid’s Tops, their new structural features, their involvement both in the physiology and virulence of these parasites, as well as their use as promising targets for drug discovery

Editorial: Signaling in stress sensing and resistance in parasitic protozoa

Protozoan parasites are a large and highly diverse group of unicellular eukaryotes infecting humans and animals, posing enormous health and socio-economic impacts. More than 1 million deaths annually are caused globally by protozoan parasites (Andrews et al., 2014). A significant proportion of the world population is at risk of being afflicted by parasitic diseases like malaria, African trypanosomiasis, Chagas disease, and leishmaniases, widely considered “Neglected global infectious diseases” for which the dearth of clinically approved vaccines and safer and efficacious chemotherapeutic options (https://www.who.int/health-topics/neglected-tropical-diseases#tab=tab_1) still persists. Protozoan pathogens lead complex life cycles, from one host to another, surviving in a variety of morphotypic forms through this complex continuum. The developmental stages can be free-living, extracellular parasitic or intracellular parasitic. The key trigger identified for such transitions has often been stresses such as the alteration of temperature or pH, hypoxia, oxidative burst, or nutrient depletion (Vonlaufen et al., 2008). Of note, stress-associated modulations confer endurance to drug exposure to augment resistance or develop quiescent or ‘persister-like’ forms. Hence, the ability to sense and respond to stress and thrive in less than optimal conditions, hence, are crucial for pathogenicity of the parasites, manifestation of diseases and for combating chemotherapeutic challenge (Bhattacharya et al., 2020). Some of the major stress-response pathways, characterized to date, are elicited by posttranslational modifications (PTM), redox systems, chaperon proteins that mediate protein folding and secondary messengers like cyclic adenosine monophosphate or calcium (cAMP or Ca2+) which modulate chemotaxis, antioxidant defence of the parasite (Kelly et al., 2021; Quintana et al., 2021). Against this backdrop, the primary goal of the Research Topic has been to envision the link between adaptive stress response with pathogenicity and drug response. With three original articles, the Research Topic offers a glance hinting towards the significance of PTM in oxidative stress response in T. cruzi, alteration of membrane dynamics in drug resistance in L. donovani, and development of a potential nanoformulation inducting hallmarks of stress response in L. donovani. Alongside this, an illustrative review highlights a putative trypanosmatid G-protein coupled receptor signaling in a sensing host environment.Fil: Bhattacharya, Arijit. Adamas University; IndiaFil: Fernandez Prada, Christopher. University of Montreal; CanadáFil: Alonso, Guillermo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Biswas, Arunima. University Of Kalyani; Indi

Different mutations in a P-type ATPase transporter in Leishmania parasites are associated with cross-resistance to two leading drugs by distinct mechanisms

Work in TKS’s lab is supported by the Wellcome Trust grant 093228 and European Community’s Seventh Framework Programme under grant agreement No. 602773 (Project KINDRED).Leishmania infantum is an etiological agent of the life-threatening visceral form of leishmaniasis. Liposomal amphotericin B (AmB) followed by a short administration of miltefosine (MF) is a drug combination effective for treating visceral leishmaniasis in endemic regions of India. Resistance to MF can be due to point mutations in the miltefosine transporter (MT). Here we show that mutations in MT are also observed in Leishmania AmB-resistant mutants. The MF-induced MT mutations, but not the AmB induced mutations in MT, alter the translocation/uptake of MF. Moreover, mutations in the MT selected by AmB or MF have a major impact on lipid species that is linked to cross-resistance between both drugs. These alterations include changes of specific phospholipids, some of which are enriched with cyclopropanated fatty acids, as well as an increase in inositolphosphoceramide species. Collectively these results provide evidence of the risk of cross-resistance emergence derived from current AmB-MF sequential or co-treatments for visceral leishmaniasis.Publisher PDFPeer reviewe

Evolution in the Halo Masses of Isolated Galaxies between z~1 and z~0: From DEEP2 to SDSS

We measure the evolution in the virial mass-to-light ratio (M_{200}/L_B) and virial-to-stellar mass ratio (M_{200}/M_\ast) for isolated ~ L* galaxies between z~1 and z~0 by combining data from the DEEP2 Galaxy Redshift Survey and the Sloan Digital Sky Survey. Utilizing the motions of satellite galaxies around isolated galaxies, we measure line-of-sight velocity dispersions and derive dark matter halo virial masses for these host galaxies. At both epochs the velocity dispersion of satellites correlates with host galaxy stellar mass, \sigma\propto M_\ast^{0.4+/-0.1}, while the relation between satellite velocity dispersion and host galaxy B-band luminosity may grow somewhat shallower from \sigma\propto L_B^{0.6+/-0.1} at z~1 to \sigma\propto L_B^{0.4+/-0.1} at z~0. The evolution in M_200/M_\ast from z~1 to z~0 displays a bimodality insofar as host galaxies with stellar mass below M_\ast ~10^{11} M_Sun/h maintain a constant ratio (the intrinsic increase is constrained to a factor of 1.1+/-0.7) while host galaxies above M_\ast ~10^{11} M_Sun/h experience a factor of 4+/-3 increase in their virial-to-stellar mass ratio. This result can be easily understood if galaxies below this stellar mass scale continue to form stars while star formation in galaxies above this scale is quenched and the dark matter halos of galaxies both above and below this scale grow in accordance with LCDM cosmological simulations. Host galaxies that are red in U-B color have larger satellite dispersions and hence reside on average in more massive halos than blue galaxies at both z~1 and z~0. The redshift and host galaxy stellar mass dependence of M_200/M_\ast agrees qualitatively with the Millennium Run semi-analytic model of galaxy formation. (ABRIDGED)Comment: 20 pages, 8 figures, submitted to Ap

Influence of N- methylation and conformation on almiramide anti-leishmanial activity

The almiramide N-methylated lipopeptides exhibit promising activity against trypanosomatid parasites. A structure–activity relationship study has been performed to examine the influences of N-methylation and conformation on activity against various strains of leishmaniasis protozoan and on cytotoxicity. The synthesis and biological analysis of twenty-five analogs demonstrated that derivatives with a single methyl group on either the first or fifth residue amide nitrogen exhibited greater activity than the permethylated peptides and relatively high potency against resistant strains. Replacement of amino amide residues in the peptide, by turn inducing α‑amino γ‑lactam (Agl) and N-aminoimidazalone (Nai) counterparts, reduced typically anti-parasitic activity; however, peptide amides possessing Agl residues at the second residue retained significant potency in the unmethylated and permethylated series. Systematic study of the effects of methylation and turn geometry on anti-parasitic activity indicated the relevance of an extended conformer about the central residues, and conformational mobility by tertiary amide isomerization and turn geometry at the extremities of the active peptides

Productivity links morphology, symbiont specificity, and bleaching in the evolution of Caribbean octocoral symbioses

Many cnidarians host endosymbiotic dinoflagellates from the genus Symbiodinium. It is generally assumed that the symbiosis is mutualistic, where the host benefits from symbiont photosynthesis while providing protection and photosynthetic substrates. Diverse assemblages of symbiotic gorgonian octocorals can be found in hard bottom communities throughout the Caribbean. While current research has focused on the phylo- and population genetics of gorgonian symbiont types and their photo-physiology, relatively less work has focused on biogeochemical benefits conferred to the host and how these benefits vary across host species. Here, we examine this symbiosis among 11 gorgonian species collected in Bocas del Toro, Panama. By coupling light and dark bottle incubations (P/R) with 13C-bicarbonate tracers, we quantified the link between holobiont oxygen metabolism with carbon assimilation and translocation from symbiont to host. Our data show that P/R varied among species, and was correlated with colony morphology and polyp size. Sea fans and sea plumes were net autotrophs (P/R > 1.5) while nine species of sea rods were net heterotrophs with most below compensation (P/R < 1.0). 13C assimilation corroborated the P/R results, and maximum δ13Chost values were strongly correlated with polyp size, indicating higher productivity by colonies with high polyp SA:V. A survey of gorgonian-Symbiodinium associations revealed that productive species maintain specialized, obligate symbioses and are more resistant to coral bleaching, whereas generalist and facultative associations are common among sea rods that have higher bleaching sensitivities. Overall, productivity and polyp size had strong phylogenetic signals with carbon fixation and polyp size showing evidence of trait covariance.published_or_final_versio