Antiplasmodial activity of p-substituted benzyl thiazinoquinone derivatives and their potential against parasitic infections

Malaria is a life-threatening disease and, what is more, the resistance to available antimalarial drugs is a recurring problem. The resistance of Plasmodium falciparum malaria parasites to previous generations of medicines has undermined malaria control efforts and reversed gains in child survival. This paper describes a continuation of our ongoing efforts to investigate the effects against Plasmodium falciparum strains and human microvascular endothelial cells (HMEC-1) of a series of methoxy p-benzyl-substituted thiazinoquinones designed starting from a pointed antimalarial lead candidate. The data obtained from the newly tested compounds expanded the structure-activity relationships (SARs) of the thiazinoquinone scaffold, indicating that antiplasmodial activity is not affected by the inductive effect but rather by the resonance effect of the introduced group at the para position of the benzyl substituent. Indeed, the current survey was based on the evaluation of antiparasitic usefulness as well as the selectivity on mammalian cells of the tested p-benzyl-substituted thiazinoquinones, upgrading the knowledge about the active thiazinoquinone scaffold

Dysregulation of haematopoietic stem cell regulatory programs in acute myeloid leukaemia.

Haematopoietic stem cells (HSC) are situated at the apex of the haematopoietic differentiation hierarchy, ensuring the life-long supply of mature haematopoietic cells and forming a reservoir to replenish the haematopoietic system in case of emergency such as acute blood loss. To maintain a balanced production of all mature lineages and at the same time secure a stem cell reservoir, intricate regulatory programs have evolved to control multi-lineage differentiation and self-renewal in haematopoietic stem and progenitor cells (HSPCs). Leukaemogenic mutations commonly disrupt these regulatory programs causing a block in differentiation with simultaneous enhancement of proliferation. Here, we briefly summarize key aspects of HSPC regulatory programs, and then focus on their disruption by leukaemogenic fusion genes containing the mixed lineage leukaemia (MLL) gene. Using MLL as an example, we explore important questions of wider significance that are still under debate, including the importance of cell of origin, to what extent leukaemia oncogenes impose specific regulatory programs and the relevance of leukaemia stem cells for disease development and prognosis. Finally, we suggest that disruption of stem cell regulatory programs is likely to play an important role in many other pathologies including ageing-associated regenerative failure

Time-lapse Whole-field fluorescence imaging of microglia processes motility in acute mouse hippocampal slices and analysis

Microglia are the resident immune cells of the central nervous system (CNS). In the last year, the improvements in the transgenic mouse technologies and imaging techniques have shed light on microglia functions under physiological conditions. Microglia continuously scan the brain parenchyma with their highly motile processes, maintaining tissue homeostasis and participating in neuronal circuits refinement. Here, we describe a protocol that enables us to perform time-lapse imaging of microglial cells in acute hippocampal slices, making image acquisition possible on an electrophysiology rig equipped with a standard imaging system. Using this ex vivo approach, we investigated microglial processes scanning abilities under physiological condition in hippocampus

Synthesis and antiplasmodial activity of novel bioinspired imidazolidinedione derivatives

Malaria is an enormous threat to public health, due to the emergence of Plasmodium falciparum resistance to widely-used antimalarials, such as chloroquine (CQ). Current antimalarial drugs are aromatic heterocyclic derivatives, most often containing a basic component with an added alkyl chain in their chemical structure. While these drugs are effective, they have many side effects. This paper presents the synthesis and preliminary physicochemical characterisation of novel bioinspired imidazolidinedione derivatives, where the imidazolidinedione core was linked via the alkylene chain and the basic piperazine component to the bicyclic system. These compounds were tested against the asexual stages of two strains of P. falciparum—the chloroquine-sensitive (D10) and chloroquine-resistant (W2) strains. In parallel, in vitro cytotoxicity was investigated on a human keratinocyte cell line, as well as their hemolytic activity. The results demonstrated that the antiplasmodial effects were stronger against the W2 strain (IC50 between 2424.15–5648.07 ng/mL (4.98–11.95 µM)), compared to the D10 strain (6202.00–9659.70 ng/mL (12.75–19.85 µM)). These molecules were also non-hemolytic to human erythrocytes at a concentration active towards the parasite, but with low toxicity to mammalian cell line. The synthetized derivatives, possessing enhanced antimalarial activity against the CQ-resistant strain of P. falciparum, appear to be interesting antimalarial drug candidates

Investigating the antiparasitic potential of the marine sesquiterpene avarone, its reduced form avarol, and the novel semisynthetic thiazinoquinone analogue thiazoavarone

The chemical analysis of the sponge Dysidea avara afforded the known sesquiterpene quinone avarone, along with its reduced form avarol. To further explore the role of the thiazinoquinone scaffold as an antiplasmodial, antileishmanial and antischistosomal agent, we converted the quinone avarone into the thiazinoquinone derivative thiazoavarone. The semisynthetic compound, as well as the natural metabolites avarone and avarol, were pharmacologically investigated in order to assess their antiparasitic properties against sexual and asexual stages of Plasmodium falciparum, larval and adult developmental stages of Schistosomamansoni (eggs included), and also against promastigotes and amastigotes of Leishmania infantum and Leishmania tropica. Furthermore, in depth computational studies including density functional theory (DFT) calculations were performed. A toxic semiquinone radical species which can be produced starting both from quinone- and hydroquinone-based compounds could mediate the anti-parasitic effects of the tested compounds

MS Dereplication for Rapid Discovery of Structurally New or Novel Natural Products

In order to accelerate the isolation and characterisation of structurally new or novel natural products, it is crucial to develop efficient strategies that prioritise samples with greatest promise early in the workflow so that resources can be utilised in a more efficient and cost-effective manner. Two complementary approaches have been developed: One is based on targeted identification of known compounds held in a database based on high resolution MS and predicted LC retention time data [1]. The second is an MS metrics-based approach where the software algorithm calculates metrics for sample novelty, complexity, and diversity after interrogating databases of known compounds, and contaminants. These metrics are then used to prioritise samples for isolation and structure elucidation work [2]. Both dereplication approaches have been validated using natural product extracts resulting in the isolation and characterization of new or novel natural products

RNA Nanotherapeutics for the Amelioration of Astroglial Reactivity.

In response to injuries to the CNS, astrocytes enter a reactive state known as astrogliosis, which is believed to be deleterious in some contexts. Activated astrocytes overexpress intermediate filaments including glial fibrillary acidic protein (GFAP) and vimentin (Vim), resulting in entangled cells that inhibit neurite growth and functional recovery. Reactive astrocytes also secrete inflammatory molecules such as Lipocalin 2 (Lcn2), which perpetuate reactivity and adversely affect other cells of the CNS. Herein, we report proof-of-concept use of the packaging RNA (pRNA)-derived three-way junction (3WJ) motif as a platform for the delivery of siRNAs to downregulate such reactivity-associated genes. In vitro, siRNA-3WJs induced a significant knockdown of Gfap, Vim, and Lcn2 in a model of astroglial activation, with a concomitant reduction in protein expression. Knockdown of Lcn2 also led to reduced protein secretion from reactive astroglial cells, significantly impeding the perpetuation of inflammation in otherwise quiescent astrocytes. Intralesional injection of anti-Lcn2-3WJs in mice with contusion spinal cord injury led to knockdown of Lcn2 at mRNA and protein levels in vivo. Our results provide evidence for siRNA-3WJs as a promising platform for ameliorating astroglial reactivity, with significant potential for further functionalization and adaptation for therapeutic applications in the CNS.The authors wish to acknowledge J. Bernstock and G. Pluchino for their critical insights throughout the execution of the study. This work was funded by the European Research Council (ERC) under the ERC-2010-StG grant agreement n° 260511-SEM_SEM, the Bascule Charitable Trust (RG 75149 to SP), the International Foundation for Research in Paraplegia (RG 69318 to S.P.), Wings for Life (RG 82921 to S.P.) and a core support grant from the Wellcome Trust and Medical Research Council to the Wellcome Trust – MRC Cambridge Stem Cell Institute. LPJ was supported by a research training fellowship from the Wellcome Trust (RRZA/057 RG79423)

Azacarbazole n-3 and n-6 polyunsaturated fatty acids ethyl esters nanoemulsion with enhanced efficacy against Plasmodium falciparum

Alternative therapies are necessary for the treatment of malaria due to emerging drug resistance. However, many promising antimalarial compounds have poor water solubility and suffer from the lack of suitable delivery systems, which seriously limits their activity. To address this problem, we synthesized a series of azacarbazoles that were evaluated for antimalarial activity against D10 (chloroquine-sensitive) and W2 (chloroquine-resistant) strains of P. falciparum. The most active compound, 9H-3-azacarbazole (3), was encapsulated in a novel o/w nanoemulsion consisting of ethyl esters of polyunsaturated fatty acids n-3 and n-6 obtained from flax oil as the oil phase, Smix (Tween 80 and Transcutol HP) and water. This formulation was further analyzed using transmission electron microscopy, dynamic light scattering and in vitro and in vivo studies. It was shown that droplets of the 3-loaded nanosystem were spherical, with satisfactory stability, without cytotoxicity towards fibroblasts and intestinal cell lines at concentrations corresponding to twice the IC50 for P. falciparum. Moreover, the nanoemulsion with this type of oil phase was internalized by Caco-2 cells. Additionally, pharmacokinetics demonstrated rapid absorption of compound 3 (tmax = 5.0 min) after intragastric administration of 3-encapsulated nanoemulsion at a dose of 0.02 mg/kg in mice, with penetration of compound 3 to deep compartments. The 3-encapsulated nanoemulsion was found to be 2.8 and 4.2 times more effective in inhibiting the D10 and W2 strains of the parasite, respectively, compared to non-encapsulated 3. Our findings support a role for novel o/w nanoemulsions as delivery vehicles for antimalarial drugs

Microglia complement signaling promotes neuronal elimination and normal brain functional connectivity

Complement signaling is thought to serve as an opsonization signal to promote the phagocytosis of synapses by microglia. However, while its role in synaptic remodeling has been demonstrated in the retino-thalamic system, it remains unclear whether complement signaling mediates synaptic pruning in the brain more generally. Here we found that mice lacking the Complement receptor 3, the major microglia complement receptor, failed to show a deficit in either synaptic pruning or axon elimination in the developing mouse cortex. Instead, mice lacking Complement receptor 3 exhibited a deficit in the perinatal elimination of neurons in the cortex, a deficit that is associated with increased cortical thickness and enhanced functional connectivity in these regions in adulthood. These data demonstrate a role for complement in promoting neuronal elimination in the developing cortex