Alzheimer's (AD) and Parkinson's (PD) diseases are the main neurodegenerative disorders of the central nervous system, affecting millions of people worldwide. The absence of effective and curative therapies to slow down the progression of neurodegenerative processes constitutes a serious medical concern. In this sense, the development of new neuroprotective therapies becomes imperative. Glypromate is an endogenous neuropeptide with the sequence of Gly-Pro-Glu which displays evidence of neuroprotective activity in many in vitro models of AD and PD. Despite its neuroprotective potential use in the clinical, this neuropeptide exhibits low intestinal absorption, liability towards enzymatic proteolysis, and reduced blood-brain barrier permeability. In fact, clinical trials led by Neuren Pharmaceuticals with Glypromate failed in phase III. The use of constrained proline mimetics and capping strategies have been employed in the assembly of bioactive Glypromate analogues to improve lipophilicity and enhance enzymatic stability. Considering this rationale, the NeuroPro project aims at the design, synthesis, and biological evaluation of novel constrained Glypromate analogues. NeuroPro also explores the chemical conjugation of these constrained peptidomimetics with relevant active pharmaceutical ingredients (APIs) used in AD and PD therapy. This approach is expected to deliver new neuroprotective hits with higher metabolic resistance while exploring synergism between Glypromate analogues and APIs. In this work, the synthesis of 52 new Glypromate conjugates with Amantadine, Memantine, and Aminoindane is disclosed. These peptide-conjugates are currently undergoing biological evaluation to assess their cytotoxicity in human differentiated SH-SY5Y cells. The conjugates with the lowest cytotoxicity will be selected to proceed with neuroprotection studies.

Calcium channel Orai1 is currently considered as an emerging and relevant target in cancer due to its indirect contribution, particularly in cell migration/invasion and metastatic spread. Therefore, the calcium channel Orai1 represents a promising therapeutic lead and accessibility to a selective inhibitor is a challenge for preventive treatment of metastases. During a Structure Activity Relationship study around SKF-96365 (a molecule used as reference for measuring calcium influx since 1991), delikine molecules were identified as a new family of selective inhibitors targeting SOCE calcium influx controlled by the membrane protein Orai1. Thus, the delikine DAD3.4733 was identified and patented in 2020 as an active "hit" on pancreatic carcinoma cells PANC1 and on breast carcinoma cells MDA-H321 during the RSA study. To date, the mechanism of action of industrial SOCE/Orai1 inhibitors (GSK-96365 and CM4620) in clinical studies, is not or is poorly controlled. In this context and to try to build proof of concept around this new SOCE/Orai1 inhibitor, we decided to develop fluorescence probes derived from the delikine DAD3.473. To achieve this objective, we opted for the use of a short linker (3 or 4 carbon atoms) so as not to disturb the intrinsic character of this SOCE inhibitor, and to graft it onto the West, East or North of this inhibitor. The terminal part of the linker comprises the fluorophore NBD (7-nitro-1,2,3-benzoxadiazole). For this 8^th ECMC presentation, the results of the multi-step syntheses of these fluorescence probes will be presented as well as the cell bio-imaging works.

Black currants have antioxidant, anti-inflammatory and chemoprotective properties. Anthocyanins, which are components of black currants, are powerful antioxidants, being able to inhibit the growth of tumor cells and to induce apoptosis. In this context, our study was designed to investigate the antitumoral effects following exposure to the total ethanolic extract obtained from blackcurrant powder, rich in anthocyanins, on cervical cancer. The content of total phenolic compounds was analyzed using Folin-Ciocalteu reagent, and the concentration of anthocyanins was determined by HPLC. The in vitro characterization of the extracts included common tests to measure antioxidant capacity, cell viability and inflammation tests on HeLa cervix cells, and measurement of reduced glutathione level and catalase and glutathione S transferase activities, as well as flow cytometry analysis to evaluate the cell cycle phases. Our study demonstrated that the extract highest concentration in anthocyanins (delphinidin and malvidin), with an antiproliferative capacity was the one obtained after 48 hours of extraction with ethanol, which induced a time- and dose-dependent decrease in cancer cell viability. An increase in enzyme activity of catalase and glutathione S-transferase was noted after the first 24 hours of incubation, suggesting a tendency of the cells to counteract the oxidative stress induced by anthocyanins. Incubation of cells with blackcurrant extract resulted in cell cycle arrest in the G1 and sub-G1 phases after 24 and 72 hours, respectively. These data support the antioxidant and anti-proliferative efficiency of anthocyanins from black currants, being valuable for further in vivo studies.

An ulcer that develops on the mucosal surface of the oral cavity is known as a mouth ulcer, also known as an oral ulcer or a mucosal ulcer. A mucus membrane ulcer is an open sore that is distinguished by the removal of inflammatory dead tissue. The most typical type of oral ulcer is aphthous stomatitis. This investigation focuses on temperature-sensitive in-situ gel formulations, which change their phase on response to body heat from liquid to semisolid gel. These are easily administered into the buccal cavity at the ulcer site and are a free-flowing liquid at room temperature. Utilizing various polymers, a temperature-sensitive in-situ gel comprising phytoconstituents was developed utilising the cold technique. To optimise various types and concentrations of polymers, including carbopol, Poloxamer 188 (P 188), Poloxamer 407 (P 407), and others, preliminary research was conducted. For the formulation, 20% P 188 and 15% P 407 were employed because there is a correlation between the amount of poloxamers and thermogelling transition temperatures (Tsol-gel). A blend of phytoconstituents found in the extracts of Glycyrrhizin glabra and Psidium guava are used in the formulation of mouth ulcers because, as we know, they have less negative effects than synthetic chemicals. The outcomes demonstrated improved homogeneity, stability, gelation temperature, and spreadability for the developed product, which was regarded satisfactory. The created formulation can also lessen dose variation and treat oral ulcers in the most effective way with improved patient compliance.

The current therapeutic agents for Type 2 diabetes (like Insulin, Sulphonylureas, Biguanides, α-Glucosidase inhibitors, PPAR agonist and GLP-1 agonist), although effective in increasing insulin secretion, are associated with some safety issue and undesirable side effects, including hypoglycemia, abnormalities in cardiovascular responses and β-cell apoptosis. DPP-4 inhibitors offer several potential advantages over existing therapies including decreased risk of hypoglycemia, potential for weight loss, and the potential for regeneration and differentiation of pancreatic β-cells. Moreover, DPP-4 inhibitors can also be administered orally. Overall, DPP 4 inhibitors are promising new class of antidiabetics and intense research in this area has resulted in the launch of sitagliptin, vildagliptin, saxagliptin, alogliptin and linagliptin as new weapon in the arsenal of oral antihyperglycemic agents. Among all DPP-4 inhibitor derivatives, 2-Cyano pyrrolidine-based inhibitors have been studied most extensively. Apart from behaving as a proline mimic, the presence of the nitrile on the five-membered ring was shown to provide (i) nanomolar inhibition of DPP-4 and (ii) chemical stability adequate for oral administration. These intermediate was fused with 2-amino-5-aryl-1, 3, 4-thiadiazole derivative to get series of novel 1-(2-(5-aryl-1,3,4-thiadiazol-2-ylamino)acetyl)pyrrolidine-2-carbonitrile derivatives. The synthesized DPP-4 inhibitor derivatives were evaluated by fluorescence assay using Gly-Pro-AMC as a DPP-4-specific fluorescent substrate.

With antimicrobial resistance reaching critical levels worldwide, the development of new compounds that are effective against resistant bacterial pathogens or that can potentiate the effect of known antibiotics is an urgent need. The formation of biofilms and the overexpression of efflux pumps are some of the most common causes of drug resistance. Previous work from our group has shown that alkaloids related to the fumiquinazolines have antibacterial potential. Herein we aimed to synthesize a small library of fumiquinazoline related alkaloids and to study their antibacterial and anti-biofilm activities as well as their capacity to inhibit bacterial efflux pumps. To achieve these goals, two naturally occurring alkaloids, as well as several new derivatives, were synthesized through a multi-step synthetic pathway. The screening of their antibacterial activities was achieved by determination of the minimum inhibitory concentration of each compound against a panel of clinically relevant bacterial species. Several compounds exhibited promising activities against Gram-positive bacteria. Then, using the ethidium bromide accumulation assay it was possible to identify some compounds with capacity to inhibit efflux pumps. Some of the synthesized alkaloids also showed anti-biofilm potential, reinforcing the idea that fumiquinazoline related alkaloids can constitute a key strategy to fighting antimicrobial resistance.

Breast cancer is the most common cancer worldwide. Santolina chamaecyparissus L. has successfully inhibited the MCF-7 cancer cell line. This study aims to evaluate the chemopreventive effects of S. chamaecyparissus aqueous extract (SCE) on female rats' physiological parameters with mammary cancer induced by N-methyl-N-nitrosourea (MNU).

The institutional ethics committee approved this study. Twenty-eight four-week-old female Wistar rats were divided into Control, MNU-induced (IND), SCE and SCE+IND. SCE was supplemented in drinking water (120µg/mL). At 50 days of age, MNU was intraperitoneally administered. Humane endpoints were evaluated weekly. After twenty-one weeks, animals were sacrificed by ketamine/xylazine overdose, and blood was collected. A complete blood count was performed using an automated haematology analyser. An autoanalyser was used to measure serum markers (albumin, cholesterol, glucose, and triglycerides). SCE’s chemical characterisation was performed by LC-MS, being found nineteen phenolic compounds, being the main molecules myricetin-O-glucuronide and 1,3-O-dicaffeoylquinic acid.

Regarding haemoglobin concentration, there was a difference (p=0.050) between SCE and Control (16.38±0.41g/dL and 15.18±0.29g/dL, respectively). Mean Platelet Volume differed between SCE+IND (8.29±0.15fL) and IND (9.03±0.26fL) (p=0.014). Platelet Distribution Width differed between 9.06±0.14fL (SCE+IND) and 10.58±0.42fL (IND) (p<0.001), but also between SCE (8.78±0.16fL) and SCE+IND versus control (9.86±0.17fL) (p=0.007 and p=0.034, respectively). SCE had no effect on the humane endpoints or serum markers.

Platelet size appears to be significantly affected by SCE. SCE supplementation had no effect on liver or kidney function or well-being in animals, implying it could be a viable treatment option for breast cancer. Histological analysis will help confirm SCE’s toxicological profile.

Ionic liquids (ILs) containing active pharmaceutical ingredients (APIs) have been reported as a successful approach to improve drug delivery and to overcome other drawbacks of pharmaceutical industry. Moreover, ILs-API derived from antibiotics revealed antimicrobial activity against sensitive bacteria and, particularly, increased for resistant species. The higher antimicrobial activity can be attributed to the improved drug delivery and solubility, but also to some specific interactions. On the other hand, the search for alternative and effective therapies to fight cancer pointed out ionic liquids as potential therapeutic agents with antitumor properties. In this context, several ILs with valproate (VPA) as API were synthesized and studied in terms of their bioactivity against neuroblastoma. The toxicity of the prepared ionic liquids was evaluated by MTT cell metabolic assay in human neuroblastoma SH-SY5Y and human primary Gingival Fibroblast (GF) cell lines, in which they showed inhibitory effects. Low cytotoxicity against GF cell lines was also observed, suggesting that these compounds are not toxic to human cell lines. 1-(2-hydroxyethyl)-2,3-dimethylimidazolium 2-propylpentanoate, [C₂OHDMiM][VPA], demonstrated an outstanding antitumor activity against SH-SY5Y and lower activity against the non-neoplastic GF line. The herein assessed compounds played an important role in the modulation of the signaling pathways involved in the cellular behavior. This work also highlights the potential of these ILs-API as possible antitumor agents.

Iron oxide nanoparticles (IONPs) have high relativity for Magnetic Resonance Imaging (MRI) and have shown a long half-life which can covalently bind with drugs and antibodies that can be used for increased contrast while imaging with MRI. IONPs are approved by the U.S Food and Drug Administration to be used as a contrast agent for MRI in vivo. The Cuban Neuroscience Center has developed a new family of naphthalene derivatives compounds called Amylovis^®, to be used to diagnose and treatment of Alzheimer's disease (AD). The goal of this work is to synthesize IONPs with different coatings, to be conjugated to Amylovis^® as contrast agents for MRI. Coprecipitation method was employed to obtain the magnetic nuclei and two synthesis methodologies were performed to coating the IONPs (post-synthesis or in situ). The different coatings used were: (3-aminopropyl)triethoxysilane (APTES), dicarboxylic polyethyleneglycol, galic acid, polyacrylic acid and citrate. Carbodiimide method was used as synthetic protocol to conjugate the Amylovis^® moiety to different coatings. The nanoparticles were characterized by DRX, FT-IR, Electrophoretic Light Scattering (ELS) and Dynamic Light Scattering (DLS). Moreover, the stability of nanoparticles in water was evaluated using a sedimentation curve. In order to determine the potential interaction of nanosystems with the beta amyloid peptides a molecular docking was carried out. The theoretical study showed that coatings do not affect the interaction of Amylovis^® with the receptor and the new polar groups incorporated increased the affinity to receptor.

Abstract

Annona atemoya is a commercially important fruiting plant belonging to the Annonaceae family. It is widely cultivated in tropical and subtropical continents. It is also known as the custard apple which is a hybrid between two Annonaceae species: (Annona cherimola) and (Annona squamosa). This study aimed to investigate the phytochemical constituents and pharmacological activity of various parts of A. atemoya including leaves and fruit’s pulp. The leaves and fruits of A. atemoya were collected in July 2020 from a local farm in Queensland, air dried at room temperature (3 days – 1 week) and then ground to a powder. The leaves and pulp were separately extracted with hexane, ethyl acetate and finally ethanol for three days each. Preliminary results of TLC and NMR experiments indicated the presence of annonaceous acetogenins for the first time in the leaves and pulp as pink bands after reacting with Kedde reagent. For the leaves, ten compounds were identified, two of them were isolated and the other confirmed via NMR and MS analysis. The ethyl acetate extract was the richest in an abundance of acetogenins in comparison to the hexane extract. With regards to the fruit’s pulp, the concentration of acetogenins was very low and compared to the leaves. Future studies will focus on testing either isolated compounds or crude extracts using various cancer cell lines.