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Background: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a pos-itive-sense, single stranded RNA virus, responsible for the pandemic outbreak called COVID-19. The pandemic, still ongoing, had presented unprecedented challenges in terms of finding appropriate pharmacological treatments. Methods: Starting from the recent literature that demonstrates how ALKBH5 inhibitors could be used as a new strategy to reduce SARS-CoV-2 replication, we decided to repurpose our newly dis-covered ALKBH5 inhibitor MV1035, previously tested and proved effective against glioblastoma, for its putative antiviral activity against SARS-CoV-2. We demonstrated a reduction in SARS-CoV-2-induced CPE after 72 h incubation using MV1035 (50 µM), for SARS-CoV-2 wild type (Wuhan strain) and South African variant. Results: The results show how MV1035 seems to be able to reduce SARS-CoV-2 replication through an indirect mechanism of action, which might involve an interaction with the host cell rather than with a virus protein. Conclusion: This may be particularly interesting as it lays the foundation for the rational design of molecules in principle not subject to drug resistance, as host cell proteins are not affected by virus mutations.

The insurgence of drug resistance in treating Multiple Myeloma (MM) still represents a major hamper in finding effective treatments, although over the past decades new classes of drugs, such as proteasome inhibitors and immunomodulatory drugs, have been discovered. Recently, our research team, within a Nature-Aided Drug Discovery project, isolated from Hibiscus Sabdariffa L. calyces the secondary metabolite called Hib-ester which possesses antiproliferative properties against human multiple myeloma RPMI 8226 cells, reduces migration and cell invasion and inhibits proteasome without neurotoxic effects. In the present study, we explored the chemical spaces of the hit compound Hib-ester. We explored the structure-activity relationships (SAR), and we optimized the scaffold through sequentially modifying Hib-ester subunits. Compound screening was performed based on cytotoxicity against the RPMI 8226 cells to assess the potential efficacy toward human MM. The ability of the most effective molecules to inhibit the proteasome was evaluated and the binding mode of the most promising compounds in the proteasome chymotrypsin binding pocket was deciphered through molecular modeling simulations. Compounds 13 and 14 are more potent than Hib-ester, demonstrating that our strategy was suitable for the identification of a novel chemotype for developing possible drug candidates and hopefully widening the drug armamentarium against MM.

In this manuscript we report on the design, synthesis and evaluation of dual Sigma 1 Receptor (S1R) modulators/Acetylcholinesterase (AChE) inhibitors endowed with antioxidant and neurotrophic properties, potentially able to counteract neurodegeneration. The compounds based on arylalkylaminoketone scaffold integrate the pharmacophoric elements of RRC-33, a S1R modulator developed by us, donepezil, a well-known AChE inhibitor, and curcumin, a natural antioxidant compound with neuroprotective properties. A small library of compounds was synthesized and preliminary in vitro screening performed. Some compounds showed good S1R binding affinity, selectivity towards S2R and N-Methyl-D-Aspartate (NMDA) receptor, AChE relevant inhibiting activity and are potentially able to bypass the BBB, as predicted by the in silico study. For the hits 10 and 20, the antioxidant profile was assessed in SH-SY5Y human neuroblastoma cell lines by evaluating their protective effect against H2O2 cytotoxicity and reactive oxygen species (ROS) production. Tested compounds resulted effective in decreasing ROS production, thus ameliorating the cellular survival. Moreover, compounds 10 and 20 showed to be effective in promoting the neurite elongation of Dorsal Root Ganglia (DRG), thus demonstrating a promising neurotrophic activity. Of note, the tested compounds did not show any cytotoxic effect at the concentration assayed. Relying on these encouraging results, both compounds will undergo a structure optimization program for the development of therapeutic candidates for neurodegenerative diseases treatment.

Hibiscus Sabdariffa (HS) is a plant of the Malvacee family commonly cultured in tropical and subtropical countries. It is mainly known as the main ingredient for the preparation of cold drink called Karkadè. Calices and leaves of HS plant are also used in folk medicine thanks to their antioxidant and anti-inflammatory properties. In recent years, HS has also gained great interest as a possible antitumoral agent. In the present PhD project, we evaluated the antitumoral effects of HS against multiple mye-loma cells in vitro. Multiple myeloma is the most frequent hematological malignancy world-wide. In recent years, new drugs have increased the survival expectancy of patients. Despite this, new therapeutic approaches are necessary, especially for high multiple myeloma hetero-geneity and for relapsed or refractory multiple myeloma. The project was organized in three distinct phases: 1- Evaluation of antitumoral effects of HS against RPMI 8226 human multiple myeloma cells. We demonstrated by MTT and Trypan blue assays that a total HS extract (HSE) and one of its fraction obtained by liquid-liquid extraction (HSEC) were able to impair cell viability of human multiple myeloma RPMI 8226 in a dose and time dependent manner. HSE cell viability reduction was due to a cytostatic action, while HSEC was more cytotoxic and induced a caspase dependent apoptosis. Moreover, both HSE and HSEC impaired cell migration and invasion of RPMI 8226 cells in a Boyden chamber as-say. We also demonstrated in in vitro model of neurotoxicity (dorsal root ganglia model) that HSE and HSEC concentrations used in our experiments were not neurotoxic. In RPMI 8226 cells autophagy and proteasome activity were impaired by both HSE and HSEC. MAPK p38 activation was observed in the first 6h of treatment, while ERK 1 and ERK 2 activation occurred between 16 and 48h. 2- Evaluation of combinations between Bortezomib (BTZ) and HSE or HSEC against RPMI 8226 multiple myeloma cells. We evaluated several combinat, Hibiscus Sabdariffa (HS) is a plant of the Malvacee family commonly cultured in tropical and subtropical countries. It is mainly known as the main ingredient for the preparation of cold drink called Karkadè. Calices and leaves of HS plant are also used in folk medicine thanks to their antioxidant and anti-inflammatory properties. In recent years, HS has also gained great interest as a possible antitumoral agent. In the present PhD project, we evaluated the antitumoral effects of HS against multiple mye-loma cells in vitro. Multiple myeloma is the most frequent hematological malignancy world-wide. In recent years, new drugs have increased the survival expectancy of patients. Despite this, new therapeutic approaches are necessary, especially for high multiple myeloma hetero-geneity and for relapsed or refractory multiple myeloma. The project was organized in three distinct phases: 1- Evaluation of antitumoral effects of HS against RPMI 8226 human multiple myeloma cells. We demonstrated by MTT and Trypan blue assays that a total HS extract (HSE) and one of its fraction obtained by liquid-liquid extraction (HSEC) were able to impair cell viability of human multiple myeloma RPMI 8226 in a dose and time dependent manner. HSE cell viability reduction was due to a cytostatic action, while HSEC was more cytotoxic and induced a caspase dependent apoptosis. Moreover, both HSE and HSEC impaired cell migration and invasion of RPMI 8226 cells in a Boyden chamber as-say. We also demonstrated in in vitro model of neurotoxicity (dorsal root ganglia model) that HSE and HSEC concentrations used in our experiments were not neurotoxic. In RPMI 8226 cells autophagy and proteasome activity were impaired by both HSE and HSEC. MAPK p38 activation was observed in the first 6h of treatment, while ERK 1 and ERK 2 activation occurred between 16 and 48h. 2- Evaluation of combinations between Bortezomib (BTZ) and HSE or HSEC against RPMI 8226 multiple myeloma cells. We evaluated several combinat

Cancer is a leading cause of death worldwide. Despite therapeutic improvements, some cancers are still untreatable. Recently there has been an increasing interest in the use of natural substances for cancer prevention and treatment. Hibiscus sabdariffa (HS) is a plant, belonging to Malvaceae family, widespread in South Asia and Central Africa. HS extract (HSE) used in folk medicine, gained researchers' interest thanks to its antioxidant, anti-inflammatory, and chemopreventive properties. In the present study, we initially assessed HSE effect on a panel of human tumor cell lines. Then we focused our study on the following that are most sensitive to HSE action cell lines: Multiple Myeloma (MM) cells (RPMI 8226) and Oral Squamous Cell Carcinoma (OSCC) cells (SCC-25). In both RPMI 8226 and SCC-25 cells, HSE impaired cell growth, exerted a reversible cytostatic effect, and reduced cell motility and invasiveness. We evaluated the involvement of MAPKs ERK1/2 and p38 in HSE effects by using specific inhibitors, U0126 and SB203580, respectively. For both SCC-25 and RPMI 8226, HSE cytostatic effect depends on p38 activation, whereas ERK1/2 modulation is crucial for cell motility and invasiveness. Our results suggest that HSE may be a potential therapeutic agent against MM and OSCC.

Aim: Nowadays, there is a great interest in the therapeutic potential of sigma1 receptor ligands for treating different CNS pathologies. Our previous investigations led to identify (R)-RC-33 as a potent and selective S1R agonist. Results: Herein, we report the gram-scale synthesis, pharmacokinetic profile and CNS distribution of (R)-RC-33 in the mouse to determine the most suitable dosage schedule for in vivo administration. For comparative purposes, the same experiments were also performed with PRE-084, the most widely used S1R agonist commonly in pharmacological experiments. Discussion: (R)-RC-33 shows a similar pharmacokinetic profile and a better CNS distribution when compared with PRE-084. Conclusion: (R)-RC-33 may be a promising candidate for in vivo studies in animal models of neurodegenerative diseases.

Cholangiocarcinoma (CCA) is the second most frequent primary liver neoplasia. It mainly arises from the malignant transformation of biliary epithelial cells, although it might originate from either hepatic progenitor cells at the Hering canals or transformed hepatocytes. CCA is a highly aggressive tumor with extremely poor prognosis and limited therapeutic approaches. Baicalin (BA) is one of the main bioactive flavonoids identified in the Scutellaria Baicalensis Georgi root dried extract which is extensively used in the Chinese tra-ditional medicine. Together with the anti-inflammatory effect, the anti-neoplastic action is the most relevant BA property demonstrated on cancer cells of different origin. Being aware of the need of new therapeutic weapons for CCA treatment, we in-vestigated whether Baicalin could exert anti-proliferative and anti-migratory effect on EGI-1 cells, a highly metastatic CCA cell line derived from bile duct carcinoma. We first tested different BA concentrations (from 5 to 200µM) in limiting EGI-1 via-bility using MTT assay. After 24h and 48h treatment, 5 and 10µM BA had no effect while rising from 25µM to 200µM (i.e. 25,50,100 and 200µM) BA exerted a significant cell viability reduction already at 24h and increased after 48h BA expo-sure. This reduction well correlated with the adherent absolute cell number de-crease and it cannot be due to BA induced cell cycle impairment after neither 24 nor 48h treatment. We also evaluated the anti-migratory BA potential by a wound healing assay adding different BA concentrations (5, 25, 50,100 and 200µM) to the culture medium immediately after performing a wound on confluent cell cultures. All BA concen-trations but 5µM induced a significant reduction in the EGI-1 migration rate after 24h treatment. Moreover 25, 50 and 10µM BA showed similar migration inhibition extent at 24 and 48h whilst 200µM BA exerted a stronger inhibitory effect already after 24h exposure which increased with time in a significant w

Epidemiological data consistently demonstrate a reduced cancer risk associated with a polyphenols rich diet. Hibiscus sabdarifa (HS), a polyphenols rich plant widely consumed worldwide as beverage and used in folk medicine, has recently gained interest thanks to its antioxidant, anti-inflammatory and chemopreventive properties. In the present study we investigated the antitumoral potential of HS extract in two different human tumor cell lines: Multiple Myeloma cells (RPMI 8226) and Oral Squamous Cell Carcinoma cells (SCC-25). MTT assays showed that HS extract induced a dose-dependent viability re-duction in both the cells lines. For the subsequent experiments we used HS at the concentration of 5 mg/ml that was the most effective in inducing cell viability reduction after 48h of treatment. Viable cell count using trypan blue staining demonstrated that the HS extract induced decrease in cell growth of both the cell lines and this was due to a reversible cytostatic rather than a cytotoxic effect. Wound-healing and cell invasion assays, respectively performed by a scratch of cell monolayer and Boyden Chamber transwell test, demonstrated that HS ex-tract was able to reduce motility and invasiveness in both RPMI 8226 and SCC-25 cells. The chemical inhibition of ERK1/ERK2 and PI3K, with U0126 and wortmannin re-spectively, reduces proliferation and migration of both SSC-25 and RPMI cells and HB extract treatment played an additive action with the inhibitors. In conclusion, our results suggest that HS extract have antitumoral properties, since it proved to inhibit tumoral cell growth and cell migration and invasiveness. It is interesting to note that HS extract is effective against two very different tumor cell lines. In fact, RPMI 8226 cells are of hematopoietic origin and grow in suspension, whereas SCC-25 cells derive from epithelium and are characterized by adherent cell growth. Therefore, although further studies are needed to clarify the molecular mechanisms involved i