Scaffold-Hopping Strategies in Aurone Optimization: A Comprehensive Review of Synthetic Procedures and Biological Activities of Nitrogen and Sulfur Analogues.

Aurones, particular polyphenolic compounds belonging to the class of minor flavonoids and overlooked for a long time, have gained significative attention in medicinal chemistry in recent years. Indeed, considering their unique and outstanding biological properties, they stand out as an intriguing reservoir of new potential lead compounds in the drug discovery context. Nevertheless, several physicochemical, pharmacokinetic, and pharmacodynamic (P3) issues hinder their progression in more advanced phases of the drug discovery pipeline, making lead optimization campaigns necessary. In this context, scaffold hopping has proven to be a valuable approach in the optimization of natural products. This review provides a comprehensive and updated picture of the scaffold-hopping approaches directed at the optimization of natural and synthetic aurones. In the literature analysis, a particular focus is given to nitrogen and sulfur analogues. For each class presented, general synthetic procedures are summarized, highlighting the key advantages and potential issues. Furthermore, the biological activities of the most representative scaffold-hopped compounds are presented, emphasizing the improvements achieved and the potential for further optimization compared to the aurone class.

Correction to "Design and Synthesis of Novel Thieno[3,2-c]quinoline Compounds with Antiproliferative Activity on RET-Dependent Medullary Thyroid Cancer Cells".

[This corrects the article DOI: 10.1021/acsomega.3c03578.].

Design and Synthesis of Novel Thieno[3,2-c]quinoline Compounds with Antiproliferative Activity on RET-Dependent Medullary Thyroid Cancer Cells.

RET kinase gain-of-function mutations represent the main cause of the high aggressiveness and invasiveness of medullary thyroid cancer (MTC). The selective inhibition of the RET kinase is a suitable strategy for the treatment of this endocrine neoplasia. Herein, we performed an innovative ligand-based virtual screening protocol using the DRUDITonline web service, focusing on the RET kinase as a biological target. In this process, thieno[3,2-c]quinolines 6a-e and 7a-e were proposed as new potential RET inhibitors. The selected compounds were synthetized by appropriate synthetic strategies, and in vitro evaluation of antiproliferative properties conducted on the particularly aggressive MTC cell line TT(C634R) identified compounds 6a-d as promising anticancer agents, with IC50 values in the micromolar range. Further structure-based computational studies revealed a significant capability of the most active compounds to the complex RET tyrosine kinase domain. The interesting antiproliferative results supported by in silico predictions suggest that these compounds may represent a starting point for the development of a new series of small heterocyclic molecules for the treatment of MTC.

In Silico Mixed Ligand/Structure-Based Design of New CDK-1/PARP-1 Dual Inhibitors as Anti-Breast Cancer Agents.

CDK-1 and PARP-1 play crucial roles in breast cancer progression. Compounds acting as CDK-1 and/or PARP-1 inhibitors can induct cell death in breast cancer with a selective synthetic lethality mechanism. A mixed treatment by means of CDK-1 and PARP-1 inhibitors resulted in radical breast cancer cell growth reduction. Inhibitors with a dual target mechanism of action could arrest cancer progression by simultaneously blocking the DNA repair mechanism and cell cycle, resulting in advantageous monotherapy. To this aim, in the present work, we identified compound 645656 with a significant affinity for both CDK-1 and PARP-1 by a mixed ligand- and structure-based virtual screening protocol. The Biotarget Predictor Tool was used at first in a Multitarget mode to filter the large National Cancer Institute (NCI) database. Then, hierarchical docking studies were performed to further screen the compounds and evaluate the ligands binding mode, whose putative dual-target mechanism of action was investigated through the correlation between the antiproliferative activity data and the target proteins' (CDK-1 and PARP-1) expression pattern. Finally, a Molecular Dynamics Simulation confirmed the high stability of the most effective selected compound 645656 in complex with both PARP-1 and CDK-1.

Indicaxanthin Induces Autophagy in Intestinal Epithelial Cancer Cells by Epigenetic Mechanisms Involving DNA Methylation.

Autophagy is an evolutionarily conserved process critical in maintaining cellular homeostasis. Recently, the anticancer potential of autophagy inducers, including phytochemicals, was suggested. Indicaxanthin is a betalain pigment found in prickly pear fruit with antiproliferative and pro-apoptotic activities in colorectal cancer cells associated with epigenetic changes in selected methylation-silenced oncosuppressor genes. Here, we demonstrate that indicaxanthin induces the up-regulation of the autophagic markers LC3-II and Beclin1, and increases autophagolysosome production in Caco-2 cells. Methylomic studies showed that the indicaxanthin-induced pro-autophagic activity was associated with epigenetic changes. In addition to acting as a hypermethylating agent at the genomic level, indicaxanthin also induced significant differential methylation in 39 out of 47 autophagy-related genes, particularly those involved in the late stages of autophagy. Furthermore, in silico molecular modelling studies suggested a direct interaction of indicaxanthin with Bcl-2, which, in turn, influenced the function of Beclin1, a key autophagy regulator. External effectors, including food components, may modulate the epigenetic signature of cancer cells. This study demonstrates, for the first time, the pro-autophagic potential of indicaxanthin in human colorectal cancer cells associated with epigenetic changes and contributes to outlining its potential healthy effect in the pathophysiology of the gastrointestinal tract.

Synthesis of new antiproliferative 1,3,4-substituted-pyrrolo[3,2-c]quinoline derivatives, biological and in silico insights.

A series of biologically unexplored substituted 1,3,4-subtituted-pyrrolo[3,2-c]quinoline derivatives (PQs) was evaluated against a panel of about 60 tumor cells (NCI). Based on the preliminary antiproliferative data, the optimizations efforts permitted us to design and synthesize a new series of derivatives allowing us to individuate a promising hit (4g). The insertion of a 4-benzo[d] [1,3]dioxol-5-yl moiety on increased and extended the activity towards five panel tumor cell lines such as leukemia, CNS, melanoma, renal and breast cancer, reaching IG50 in the low µM range. Replacement of this latter with a 4-(OH-di-Cl-Ph) group (4i) or introduction a Cl-propyl chain in position 1 (5), selectively addressed the activity against the entire leukemia sub-panel (CCRF-CEM, K-562, MOLT-4, RPMI-8226, SR). Preliminary biological assays on MCF-7 such as cell cycle, clonogenic assay, ROS content test alongside a comparison of viability between MCF-7 and non-tumorigenic MCF-10 were investigated. Among the main anticancer targets involved in breast cancer, HSP90 and ER receptors were selected for in silico studies. Docking analysis revealed a valuable affinity for HSP90 providing structural insights on the binding mode, and useful features for optimization.

Antiproliferative Activity Predictor: A New Reliable In Silico Tool for Drug Response Prediction against NCI60 Panel.

In vitro antiproliferative assays still represent one of the most important tools in the anticancer drug discovery field, especially to gain insights into the mechanisms of action of anticancer small molecules. The NCI-DTP (National Cancer Institute Developmental Therapeutics Program) undoubtedly represents the most famous project aimed at rapidly testing thousands of compounds against multiple tumor cell lines (NCI60). The large amount of biological data stored in the National Cancer Institute (NCI) database and many other databases has led researchers in the fields of computational biology and medicinal chemistry to develop tools to predict the anticancer properties of new agents in advance. In this work, based on the available antiproliferative data collected by the NCI and the manipulation of molecular descriptors, we propose the new in silico Antiproliferative Activity Predictor (AAP) tool to calculate the GI50 values of input structures against the NCI60 panel. This ligand-based protocol, validated by both internal and external sets of structures, has proven to be highly reliable and robust. The obtained GI50 values of a test set of 99 structures present an error of less than ±1 unit. The AAP is more powerful for GI50 calculation in the range of 4-6, showing that the results strictly correlate with the experimental data. The encouraging results were further supported by the examination of an in-house database of curcumin analogues that have already been studied as antiproliferative agents. The AAP tool identified several potentially active compounds, and a subsequent evaluation of a set of molecules selected by the NCI for the one-dose/five-dose antiproliferative assays confirmed the great potential of our protocol for the development of new anticancer small molecules. The integration of the AAP tool in the free web service DRUDIT provides an interesting device for the discovery and/or optimization of anticancer drugs to the medicinal chemistry community. The training set will be updated with new NCI-tested compounds to cover more chemical spaces, activities, and cell lines. Currently, the same protocol is being developed for predicting the TGI (total growth inhibition) and LC50 (median lethal concentration) parameters to estimate toxicity profiles of small molecules.

Phytochemical Profile and Antioxidant, Antiproliferative, and Antimicrobial Properties of Rubus idaeus Seed Powder.

In the context of the contemporary research on sustainable development and circular economy, the quest for effective strategies aimed at revaluation of waste and by-products generated in industrial and agricultural production becomes important. In this work, an ethanolic extract from red raspberry (Rubus idaeus) seed waste (WRSP) was evaluated for its phytochemical composition and functional properties in term of antioxidative, antiproliferative, and antimicrobial activities. Chemical composition of the extract was determined by both HPLC-ESI-MS/MS and spectrophotometric methods. Phytochemical analysis revealed that flavan-3-ols and flavonols were the major phenolic compounds contained in WRSP. The extract demonstrated very high radical-scavenging (4.86 ± 0.06 µmol TE/DW) and antioxidant activity in a cell-based model (0.178 ± 0.03 mg DW/mL cell medium). The WRSP extract also exhibited antiproliferative activity against three different epithelial cancer cell lines (MCF-7, HepG2, and HeLa cells) in a dose-dependent manner. Finally, microbiological assays showed the absence of colonies of bacteria and microscopic fungi (yeasts and molds) and revealed that the WRSP extract has a large inhibition spectrum against spoilage and pathogenic bacteria, without inhibitory activity against pro-technological bacteria. In conclusion, the obtained results show that WRSP is a rich source of phytochemical compounds exerting interesting biological activities. For these reasons WRSP could find applications in the nutritional, nutraceutical, and pharmacological fields.

Targeting SARS-CoV-2 Main Protease for Treatment of COVID-19: Covalent Inhibitors Structure-Activity Relationship Insights and Evolution Perspectives.

The viral main protease is one of the most attractive targets among all key enzymes involved in the SARS-CoV-2 life cycle. Covalent inhibition of the cysteine145 of SARS-CoV-2 MPRO with selective antiviral drugs will arrest the replication process of the virus without affecting human catalytic pathways. In this Perspective, we analyzed the in silico, in vitro, and in vivo data of the most representative examples of covalent SARS-CoV-2 MPRO inhibitors reported in the literature to date. In particular, the studied molecules were classified into eight different categories according to their reactive electrophilic warheads, highlighting the differences between their reversible/irreversible mechanism of inhibition. Furthermore, the analyses of the most recurrent pharmacophoric moieties and stereochemistry of chiral carbons were reported. The analyses of noncovalent and covalent in silico protocols, provided in this Perspective, would be useful for the scientific community to discover new and more efficient covalent SARS-CoV-2 MPRO inhibitors.

Phytochemical profile and antioxidant properties of the edible and non-edible portions of black sapote (Diospyros digyna Jacq.).

This study evaluated the phytochemical profile and antioxidative properties of the edible and non-edible portions of black sapote. The phytochemical analysis highlighted the presence of several bioactive compounds, differently distributed among peel, pulp and seeds. In particular, the peel resulted rich of flavan-3-ols and proanthocyanidins, whereas seeds contained high amount of organic acids, including ferulic, citric and sinapic acids. Concerning functional properties, both edible and non-edible portions showed a significant prevention of lipid peroxidation in a cell-based model. Moreover, the results suggested that the antioxidant protection involved both redox active properties and gene expression modulation. Concerning redox active properties, peel extracts showed an antioxidant activity 7/12-fold higher than the edible portion, while seed extracts were more active in increasing catalase gene expression. The obtained results confirmed that black sapote is a good source of antioxidant phytochemicals and its non-edible portions have a great potential in the production of functional foods and supplements.

Antiproliferative Properties and G-Quadruplex-Binding of Symmetrical Naphtho[1,2-b:8,7-b']dithiophene Derivatives.

Background: G-quadruplex (G4) forming sequences are recurrent in telomeres and promoter regions of several protooncogenes. In normal cells, the transient arrangements of DNA in G-tetrads may regulate replication, transcription, and translation processes. Tumors are characterized by uncontrolled cell growth and tissue invasiveness and some of them are possibly mediated by gene expression involving G-quadruplexes. The stabilization of G-quadruplex sequences with small molecules is considered a promising strategy in anticancer targeted therapy. Methods: Molecular virtual screening allowed us identifying novel symmetric bifunctionalized naphtho[1,2-b:8,7-b']dithiophene ligands as interesting candidates targeting h-Telo and c-MYC G-quadruplexes. A set of unexplored naphtho-dithiophene derivatives has been synthesized and biologically tested through in vitro antiproliferative assays and spectroscopic experiments in solution. Results: The analysis of biological and spectroscopic data highlighted noteworthy cytotoxic effects on HeLa cancer cell line (GI50 in the low µM range), but weak interactions with G-quadruplex c-MYC promoter. Conclusions: The new series of naphtho[1,2-b:8,7-b']dithiophene derivatives, bearing the pharmacophoric assumptions necessary to stabilize G-quadruplexes, have been designed and successfully synthesized. The interesting antiproliferative results supported by computer aided rational approaches suggest that these studies are a significant starting point for a lead optimization process and the isolation of a more efficacious set of G-quadruplexes stabilizers.

Off-Target-Based Design of Selective HIV-1 PROTEASE Inhibitors.

The approval of the first HIV-1 protease inhibitors (HIV-1 PRIs) marked a fundamental step in the control of AIDS, and this class of agents still represents the mainstay therapy for this illness. Despite the undisputed benefits, the necessary lifelong treatment led to numerous severe side-effects (metabolic syndrome, hepatotoxicity, diabetes, etc.). The HIV-1 PRIs are capable of interacting with "secondary" targets (off-targets) characterized by different biological activities from that of HIV-1 protease. In this scenario, the in-silico techniques undoubtedly contributed to the design of new small molecules with well-fitting selectivity against the main target, analyzing possible undesirable interactions that are already in the early stages of the research process. The present work is focused on a new mixed-hierarchical, ligand-structure-based protocol, which is centered on an on/off-target approach, to identify the new selective inhibitors of HIV-1 PR. The use of the well-established, ligand-based tools available in the DRUDIT web platform, in combination with a conventional, structure-based molecular docking process, permitted to fast screen a large database of active molecules and to select a set of structure with optimal on/off-target profiles. Therefore, the method exposed herein, could represent a reliable help in the research of new selective targeted small molecules, permitting to design new agents without undesirable interactions.

Identification of biological targets through the correlation between cell line chemosensitivity and protein expression pattern.

Matching biological data sequences is one of the most interesting ways to discover new bioactive compounds. In particular, matching cell chemosensitivity with a protein expression profile can be a useful approach to predict the activity of compounds against definite biological targets. In this review, we discuss this correlation. First, we analyze case studies in which some known drugs, acting on known targets, show a good correlation between their antiproliferative activities and protein expression when a large panel of tumor cells is considered. Then, we highlight how the application of in silico methods based on the correlation between cell line chemosensitivity and gene/protein expression patterns might be a quick, cheap, and interesting approach to predict the biological activity of investigated molecules.

Quinoline anticancer agents active on DNA and DNA-interacting proteins: From classical to emerging therapeutic targets.

Quinoline is one of the most important and versatile nitrogen heterocycles embodied in several biologically active molecules. Within the numerous quinolines developed as antiproliferative agents, this review is focused on compounds interfering with DNA structure or with proteins/enzymes involved in the regulation of double helix functional processes. In this light, a special focus is given to the quinoline compounds, acting with classical/well-known mechanisms of action (DNA intercalators or Topoisomerase inhibitors). In particular, the quinoline drugs amsacrine and camptothecin (CPT) have been studied as key lead compounds for the development of new agents with improved PK and tolerability properties. Moreover, notable attention has been paid to the quinoline molecules, which are able to interfere with emerging targets involved in cancer progression, as G-quadruplexes or the epigenetic ones (e.g.: histone deacetylase, DNA and histones methyltransferase). The antiproliferative and the enzymatic inhibition data of the reviewed compounds have been analyzed. Furthermore, concerning the SAR (structure-activity relationship) aspects, the most recurrent ligand-protein interactions are summarized, underling the structural requirements for each kind of mechanism of action.

In Silico Identification of Small Molecules as New Cdc25 Inhibitors through the Correlation between Chemosensitivity and Protein Expression Pattern.

The cell division cycle 25 (Cdc25) protein family plays a crucial role in controlling cell proliferation, making it an excellent target for cancer therapy. In this work, a set of small molecules were identified as Cdc25 modulators by applying a mixed ligand-structure-based approach and taking advantage of the correlation between the chemosensitivity of selected structures and the protein expression pattern of the proposed target. In the first step of the in silico protocol, a set of molecules acting as Cdc25 inhibitors were identified through a new ligand-based protocol and the evaluation of a large database of molecular structures. Subsequently, induced-fit docking (IFD) studies allowed us to further reduce the number of compounds biologically screened. In vitro antiproliferative and enzymatic inhibition assays on the selected compounds led to the identification of new structurally heterogeneous inhibitors of Cdc25 proteins. Among them, J3955, the most active inhibitor, showed concentration-dependent antiproliferative activity against HepG2 cells, with GI50 in the low micromolar range. When J3955 was tested in cell-cycle perturbation experiments, it caused mitotic failure by G2/M-phase cell-cycle arrest. Finally, Western blotting analysis showed an increment of phosphorylated Cdk1 levels in cells exposed to J3955, indicating its specific influence in cellular pathways involving Cdc25 proteins.

The dimer-monomer equilibrium of SARS-CoV-2 main protease is affected by small molecule inhibitors.

The maturation of coronavirus SARS-CoV-2, which is the etiological agent at the origin of the COVID-19 pandemic, requires a main protease Mpro to cleave the virus-encoded polyproteins. Despite a wealth of experimental information already available, there is wide disagreement about the Mpro monomer-dimer equilibrium dissociation constant. Since the functional unit of Mpro is a homodimer, the detailed knowledge of the thermodynamics of this equilibrium is a key piece of information for possible therapeutic intervention, with small molecules interfering with dimerization being potential broad-spectrum antiviral drug leads. In the present study, we exploit Small Angle X-ray Scattering (SAXS) to investigate the structural features of SARS-CoV-2 Mpro in solution as a function of protein concentration and temperature. A detailed thermodynamic picture of the monomer-dimer equilibrium is derived, together with the temperature-dependent value of the dissociation constant. SAXS is also used to study how the Mpro dissociation process is affected by small inhibitors selected by virtual screening. We find that these inhibitors affect dimerization and enzymatic activity to a different extent and sometimes in an opposite way, likely due to the different molecular mechanisms underlying the two processes. The Mpro residues that emerge as key to optimize both dissociation and enzymatic activity inhibition are discussed.

Quinoline-Based Molecules Targeting c-Met, EGF, and VEGF Receptors and the Proteins Involved in Related Carcinogenic Pathways.

The quinoline ring system has long been known as a versatile nucleus in the design and synthesis of biologically active compounds. Currently, more than one hundred quinoline compounds have been approved in therapy as antimicrobial, local anaesthetic, antipsychotic, and anticancer drugs. In drug discovery, indeed, over the last few years, an increase in the publication of papers and patents about quinoline derivatives possessing antiproliferative properties has been observed. This trend can be justified by the versatility and accessibility of the quinoline scaffold, from which new derivatives can be easily designed and synthesized. Within the numerous quinoline small molecules developed as antiproliferative drugs, this review is focused on compounds effective on c-Met, VEGF (vascular endothelial growth factor), and EGF (epidermal growth factor) receptors, pivotal targets for the activation of important carcinogenic pathways (Ras/Raf/MEK and PI3K/AkT/mTOR). These signalling cascades are closely connected and regulate the survival processes in the cell, such as proliferation, apoptosis, differentiation, and angiogenesis. The antiproliferative biological data of remarkable quinoline compounds have been analysed, confirming the pivotal importance of this ring system in the efficacy of several approved drugs. Furthermore, in view of an SAR (structure-activity relationship) study, the most recurrent ligand-protein interactions of the reviewed molecules are summarized.

In Silico Insights into the SARS CoV-2 Main Protease Suggest NADH Endogenous Defences in the Control of the Pandemic Coronavirus Infection.

COVID-19 is a pandemic health emergency faced by the entire world. The clinical treatment of the severe acute respiratory syndrome (SARS) CoV-2 is currently based on the experimental administration of HIV antiviral drugs, such as lopinavir, ritonavir, and remdesivir (a nucleotide analogue used for Ebola infection). This work proposes a repurposing process using a database containing approximately 8000 known drugs in synergy structure- and ligand-based studies by means of the molecular docking and descriptor-based protocol. The proposed in silico findings identified new potential SARS CoV-2 main protease (MPRO) inhibitors that fit in the catalytic binding site of SARS CoV-2 MPRO. Several selected structures are NAD-like derivatives, suggesting a relevant role of these molecules in the modulation of SARS CoV-2 infection in conditions of cell chronic oxidative stress. Increased catabolism of NAD(H) during protein ribosylation in the DNA damage repair process may explain the greater susceptibility of the elderly population to the acute respiratory symptoms of COVID-19. The molecular modelling studies proposed herein agree with this hypothesis.

Design of antitumor drugs targeting c-kit receptor by a new mixed ligand-structure based method.

An important challenge, in the medicinal chemistry field, is the research of novel forceful drugs to overcome tumor-acquired resistance. The c-Kit tyrosine kinase receptor (TKR) represents a suitable target for the carcinogenesis control of gastro-intestinal stromal (GIST), leukemia, and mastocytosis tumors; nevertheless, several hotspot mutations of the protein limit the efficacy of a few clinical administered TKRs inhibitors. In this study, a new in silico protocol based on ligand and structure-based combined method is proposed, with the aim to identify a set of new c-Kit inhibitors able to complex c-Kit mutated proteins. A recent and freely available web-server DRUDIT is used for the ligand-based method. The protocol application allows for identifying a new generation of potential TKR inhibitors, which, in silico, complex the V654A and T670I mutated proteins and potentially overcome resistant mutations (D816H). The structure-based analysis is performed by Induced Fit Docking (IFD) studies. The comparison between the explored ligands and well-known drugs highlights the possibility to overcome tumor-acquired resistance. The best-selected structures (630705 and SML1348) provide valuable binding affinities with the mutated c-Kit forms (respectively T670I and V654A).

Identification of an LPS-Induced Chemo-Attractive Peptide from Ciona robusta.

BACKGROUND: Previously published work has demonstrated that the LPS injection of Ciona robusta leads to the overexpression of a truncated form of an immune-related mRNA (C8short) by means of Ciona robusta (CR) alternative polyadenylation (APA) (CR-APA). METHODS: The 3D structure of the C8short-derived Ciona robusta chemo-attractive peptide (CrCP) was evaluated by homology modeling. The biological activity of the CrCP was studied in vitro using a primary human dermal cell line (HuDe). Real-Time PCR was used to investigate the expression levels of genes involved in cell motility. NF-κB signaling was studied by western blotting. RESULTS: In silico modeling showed that CrCP displayed structural characteristics already reported for a short domain of the vertebrate CRK gene, suggesting its possible involvement in cell migration mechanisms. In vitro assays demonstrated that CrCP was capable of inducing the motility of HuDe cells in both wound healing and chemo-attractive experiments. qPCR demonstrated the capability of CrCP to modulate the expression of the matrix metalloproteinase-7 (MMP-7) and E-cadherin genes. Finally, western blot analysis demonstrated that treatment with CrCP induced activation of the NF-κB signaling pathway. CONCLUSION: Our results describe the characterization of the 3D structure and chemo-attractive activity of an LPS-induced CrCP peptide from Ciona robusta.