From bioactive natural products to drug-like small molecules

Natural products have historically been the most productive source of leads for the development of drugs. Thanks to the chemical methodologies of natural products, a vast array of bioactive secondary metabolites from terrestrial and marine sources has been discovered. Many of these natural products became current drug candidates. Therefore, the research of new biologically active compounds through structure elucidation and biological tests is the central issue of these studies. My research is placed in this field and it has been mainly devoted to the discovery and to the chemical and pharmacological investigation of new bioactive natural products as “lead compounds” in the antitumor and antimalarial activities area. My work, described in this PhD thesis, was organized in two different topics: i) isolation and structural characterization of bioactive secondary metabolites from marine invertebrates, ii) synthesis of thiazinoquinones derivatives endowed with cytotoxic and antiplasmodial activities from marine natural metabolites. The achievement of my research project required isolation and extraction procedures. The chemical characterization of the isolated compounds has been performed through an extensive spectroscopic analysis (UV, IR, ECD, 1D and 2D NMR) together with mass spectrometry, computational and electrochemistry methods. I have also used synthetic methods both for the chemical derivatization of the isolated molecules and for the preparation of analogues on the simplified model of natural molecules. During the course of my PhD research, whose results are reported in the following thesis, I have been dealing with the extraction and the chemical re-investigation of a new collection of the Mediterranean ascidian Phallusia fumigata. This analysis led to the isolation of one sulfated sterol, phallusiasterol C, which is a possible modulator of the PXR nuclear receptor. Morover, I have been strongly involved in completing the stereochemistry assignment of phosphoeleganin, a complex acyclic marine natural product, isolated previously from the Mediterranean ascidian Sidnyum elegans. In addition, the electrochemical response of four natural cytotoxic thiazinoquinones, beforehand isolated and characterized from Aplidium species, has been investigated, in order to clarify the mechanism of action which is the basis of their cytotoxicity. The research for new antiplasmodial hits is another main topic of my PhD activity discussed in this thesis. Previously, having identified the thiazinoquinone nucleus as new active chemiotype against P. falciparum, my research started from the development of two new series of methoxy and amide derivatives inspired by two marine metabolites isolated from the Mediterranean ascidian Aplidium conicum. Recently, in order to refine this pharmacophore model and improve the pharmacokinetic and pharmacodynamic properties, I have performed a rational design and synthesis of new modified analogues with simplified side chains and different substituents. In collaboration with the Department of Biomedical, Surgical and Dental sciences (University of Milan), the synthetic analogues of natural quinones have been tested for their in vitro antiplasmodial activity against both chloroquine (CQ)-sensitive (D10) and -resistant (W2) strains of P. falciparum, although some of them were strongly cytotoxic. Some of the synthetic derivatives showed significant antiplasmodial activity together with some important structural requirements. Additionally, in order to rationalize the structure-activity relationships (SARs), an integrated approach based on computational and electrochemistry studies was performed. These studies were carried out by a further collaborating external research group. The above results clearly evidence that quinone natural products represent an excellent source of novel “drug-like” small molecules for drug discovery in antimalarial research

Development of nature inspired antiplasmodial hits possessing the thiazinoquinone pharmacophore

Malaria accounts globally for more than 200 million new cases and 438,000 deaths per year. Since malaria is a disease of worldwide implications, combating it is one of the highest priority programs of the WHO. A worrisome increase in the number of fatal cases has been registered in recent years and it is principally due to the diffusion of multi-drug resistant strains of Plasmodium, making less effective the limited armamentarium of available drugs. Therefore, there is an urgent need of new antimalarial drugs with high efficacy against resistant strains and broad stage mode of action. To reach these challenging aims, the identification and selection of new lead compounds constitutes a crucial point. In this regard, nature remains an ever evolving resource. Recently, the antiplasmodial activity of marine secondary metabolites characterized by a quinone scaffold has been reported. In particular, it is worth to point out that a number of quinones have been shown to be effective antimalarials. The observed effects are most likely related to the most prominent chemical feature of these kind of molecules, that is their ability to undergo redox reaction i) shuttling electrons from reduced flavoproteins to acceptors such as hemoglobin-associated or free Fe(III)-protoporphyrin IX or ii) inhibiting the mitochondrial electron transport chain. In this context, recently, we were inspired by two marine metabolites Aplidinone A and B isolated from the Mediterranean ascidian Aplidium conicum, and we developed a series of synthetic analogues featuring the thiazinoquinone chemotype present in the natural metabolites with simplified side chains and different substituents. Manipulation of this chemical scaffold afforded additional analogues with improved pharmacological proprieties compared to the starting hits identified in the previous series

Assignment of the Absolute Configuration of Phosphoeleganin via Synthesis of Model Compounds

The full absolute configuration assignment of phosphoeleganin (1), a recently discovered marine-derived phosphorylated polyketide with protein tyrosine phosphatase 1B inhibitory activity, was achieved. It was based on the synthesis of model diasteroisomeric compounds of the C-8-C-12 segment portion of phosphoeleganin, chiral derivatization methods, and application of the universal NMR database concept

Phallusiasterol C, A New Disulfated Steroid from the Mediterranean Tunicate Phallusia fumigata

A new sulfated sterol, phallusiasterol C (1), has been isolated from the Mediterranean ascidian Phallusia fumigata and its structure has been determined on the basis of extensive spectroscopic (mainly 2D NMR) analysis. The possible role in regulating the pregnane X receptor (PXR) activity of phallusiasterol C has been investigated; although the new sterol resulted inactive, this study adds more items to the knowledge of the structure-PXR regulating activity relationships in the case of sulfated steroids

High-Resolution Monthly Precipitation Fields (1913–2015) over a Complex Mountain Area Centred on the Forni Valley (Central Italian Alps)

Mountain environments are extremely influenced by climate change but are also often affected by the lack of long and high-quality meteorological data, especially in glaciated areas, which limits the ability to investigate the acting processes at local scale. For this reason, we checked a method to reconstruct high-resolution spatial distribution and temporal evolution of precipitation. The study area is centred on the Forni Glacier area (Central Italian Alps), where an automatic weather station is present since 2005. We set up a model based on monthly homogenised precipitation series and we spatialised climatologies and anomalies on a 30-arc-second-resolution DEM, using Local Weighted Linear Regression (LWLR) and Regression Kriging (RK) of precipitation versus elevation, in order to test the most suitable approach for this complex terrain area. The comparison shows that LWLR has a better reconstruction ability for winter while RK slightly prevails during summer. The results of precipitation spatialisation were compared with station observations and with data collected at the weather station on Forni Glacier, which were not used to calibrate the model. A very good agreement between observed and modelled precipitation records was pointed out for most station sites. The agreement is lower, but encouraging, for Forni Glacier station data

Differential Effects of 3,5-Diiodo-L-Thyronine and 3,5,3'-Triiodo-L-Thyronine On Mitochondrial Respiratory Pathways in Liver from Hypothyroid Rats.

Both 3,5-diiodo-L-thyronine (3,5-T2) and 3,5,3'-triiodo-L-tyronine (T3) affect energy metabolism having mitochondria as a major target. However, the underlying mechanisms are poorly understood. Here, using a model of chemically induced hypothyroidism in male Wistar rats, we investigated the effect of administration of either 3,5-T2 or T3 on liver oxidative capacity through their influence on mitochondrial processes including: proton-leak across the mitochondrial inner membrane; complex I-, complex II- and glycerol-3-phosphate-linked respiratory pathways; respiratory complex abundance and activities as well as individual complex aggregation into supercomplexes. Background/Aims: Both 3,5-diiodo-L-thyronine (3,5-T2) and 3,5,3'-triiodo-L-tyronine (T3) affect energy metabolism having mitochondria as a major target. However, the underlying mechanisms are poorly understood. Here, using a model of chemically induced hypothyroidism in male Wistar rats, we investigated the effect of administration of either 3,5-T2 or T3 on liver oxidative capacity through their influence on mitochondrial processes including: proton-leak across the mitochondrial inner membrane; complex I-, complex II- and glycerol-3-phosphate-linked respiratory pathways; respiratory complex abundance and activities as well as individual complex aggregation into supercomplexes. Methods: Hypothyroidism was induced by propylthiouracil and iopanoic acid; 3,5-T2 and T3 were intraperitoneally administered at 25 and 15 μg/100 g BW for 1 week, respectively. Resulting alterations in mitochondrial function were studied by combining respirometry, Blue Native-PAGE followed by in-gel activity, and Western blot analyses. Results: Administration of 3,5-T2 and T3 to hypothyroid (hypo) rats enhanced mitochondrial respiration rate with only T3 effectively stimulating proton-leak (450% vs. Hypo). T3 significantly enhanced complex I (+145% vs. Hypo), complex II (+66% vs. Hypo), and glycerol-3 phosphate dehydrogenase (G3PDH)-linked oxygen consumptions (about 6- fold those obtained in Hypo), while 3,5-T2 administration selectively restored Euthyroid values of complex II- and increased G3PDH- linked respiratory pathways (+165% vs. Hypo). The mitochondrial abundance of all respiratory complexes and of G3PDH was increased by T3 administration whereas 3,5-T2 only increased complex V and G3PDH abundance. 3,5-T2 enhanced complex I and complex II in gel activities with less intensity than did T3, and T3 also enhanced the activity of all other respiratory complexes tested. In addition, only T3 enhanced individual respiratory component complex assembly into supercomplexes. Conclusions: The reported data highlight novel molecular mechanisms underlying the effect elicited by iodothyronine administration to hypothyroid rats on mitochondrial processes related to alteration in oxidative capacity in the liver. The differential effects elicited by the two iodothyronines indicate that 3,5-T2, by influencing the kinetic properties of specific mitochondrial respiratory pathways, would promote a rapid response of the organelle, while T3, by enhancing the abundance of respiratory chain component and favoring the organization of respiratory chain complex in supercomplexes, would induce a slower and prolonged response of the organelle

Measuring costs of community mental health care in Italy: A prevalence-based study

AbstractBackground:Information on individual mental healthcare costs and utilization patterns in Italy is scant. We analysed the use and the annual costs of community mental health services (MHS) in an Italian local health authority (LHA). Our aims are to compare the characteristics of patients in the top decile of costs with those of the remaining 90%, and to investigate the demographic and clinical determinants of costs.Methods:This retrospective study is based on administrative data of adult patients with at least one contact with MHS in 2013. Costs of services were estimated using a microcosting method. We defined as high cost (HC) those patients whose community mental health services costs place them in the top decile of the cost distribution. The predictors of costs were investigated using multiple linear regression.Results:The overall costs borne for 7601 patients were 17 million €, with HC accounting for 87% of costs and 73% of services. Compared with the rest of the patients, HC were younger, more likely to be male, to have a diagnosis of psychosis, and longer and more intensive MHS utilization. In multiple linear regression, younger age, longer duration of contact with MHS, psychosis, bipolar disorder, personality disorder, depression, dementia and Italian citizenship accounted for 20.7% of cost variance.Conclusion:Direct mental health costs are concentrated among a small fraction of patients who receive intensive socio-rehabilitation in community services. One limitation includes the unavailability of hospital costs. Our methodology is replicable and useful for national and cross-national benchmarking

Studies of Complex Biological Systems with Applications to Molecular Medicine: The Need to Integrate Transcriptomic and Proteomic Approaches

Omics approaches to the study of complex biological systems with potential applications to molecular medicine are attracting great interest in clinical as well as in basic biological research. Genomics, transcriptomics and proteomics are characterized by the lack of an a priori definition of scope, and this gives sufficient leeway for investigators (a) to discern all at once a globally altered pattern of gene/protein expression and (b) to examine the complex interactions that regulate entire biological processes. Two popular platforms in “omics” are DNA microarrays, which measure messenger RNA transcript levels, and proteomic analyses, which identify and quantify proteins. Because of their intrinsic strengths and weaknesses, no single approach can fully unravel the complexities of fundamental biological events. However, an appropriate combination of different tools could lead to integrative analyses that would furnish new insights not accessible through one-dimensional datasets. In this review, we will outline some of the challenges associated with integrative analyses relating to the changes in metabolic pathways that occur in complex pathophysiological conditions (viz. ageing and altered thyroid state) in relevant metabolically active tissues. In addition, we discuss several new applications of proteomic analysis to the investigation of mitochondrial activity

Exercise with Energy Restriction as a Means of Losing Body Mass while Preserving Muscle Quality and Ameliorating Co-morbidities: Towards a Therapy for Obesity?

Exercise with Energy Restriction as a Means of Losing Body Mass while Preserving Muscle Quality and Ameliorating Co-morbidities: Towards a Therapy for Obesity? Antonia Giacco1*, Elena Silvestri1*, Rosalba Senese2, Federica Cioffi1, Arianna Cuomo2, Assunta Lombardi3, Maria Moreno1, Antonia Lanni2 and Pieter de Lange()2 1Dipartimento di Science e Tecnologie, Università degli Studi del Sannio, Benevento, Italy 2Dipartimento di Scienze e Tecnologie Ambientali, Biologiche e Farmaceutiche, Università degli Studi della Campania "Luigi Vanvitelli," Caserta, Italy 3Dipartimento di Biologia, Università degli Studi di Napoli "Federico II," Napoli, Italy © The Authors Abstract Obesity and related co-morbidities are a major public health threat worldwide, and efforts to counteract obesity by means of physiological interventions are currently being explored and applied. Here we present an overview of the literature on the effect of dietary/exercise-based programs on loss of different components of body mass. We also discuss gain or lack of loss of lean mass in view of muscle quality maintenance, which is an important aspect to consider when employing weight-loss strategies to tackle obesity. By comparing results obtained in participants with mild to severe obesity with those obtained in lean participants, we highlight variations in the success of these interventions. Furthermore, we briefly address the observation that although certain interventions may not always affect body composition they can nevertheless ameliorate co-morbidities (insulin resistance, non-alcoholic fatty liver disease). Based on what is currently known, in this narrative review we include data from human and animal studies related to the process of unravelling the mechanisms underlying conservation of functional muscle mass