An overview on olfaction in the biological, analytical, computational, and machine learning fields.

Recently, the comprehension of odor perception has advanced, unveiling the mysteries of the molecular receptors within the nasal passages and the intricate mechanisms governing signal transmission between these receptors, the olfactory bulb, and the brain. This review provides a comprehensive panorama of odors, encompassing various topics ranging from the structural and molecular underpinnings of odorous substances to the physiological intricacies of olfactory perception. It extends to elucidate the analytical methods used for their identification and explores the frontiers of computational methodologies.

TERRA G-quadruplex stabilization behind the anti-multiple myeloma activity: Novel insights about resveratrol pleiotropic effects.

Resveratrol (RSV) is a nutraceutical compound belonging to the nonflavonoid polyphenol family, whose antioxidants, anti-inflammatory, and antitumoral properties have been widely investigated. The ability of RSV to provide beneficial effects for neurological, cardiovascular, and cancer disorders rekindled the interest to explore the molecular mechanisms behind its pleiotropic effects, which are due to the modulation of coding and noncoding genes involved in many key biological pathways. With a computational approach, including docking studies and thermodynamics calculations followed by 200-ns-long molecular dynamics and a clustering analysis, we hypothesized the stabilizing binding between RSV and G4 structures of telomeric repeat-containing RNA (TERRA), which is a tumor-suppressive long noncoding RNAs (lncRNA) involved in the regulation of telomere maintenance. In vitro studies performed on cellular models of multiple myeloma (MM) strengthened our hypothesis by highlighting that the antiproliferative and apoptotic effect induced by the treatment with RSV is associated with an increase of TERRA transcript and with upregulation of telomeric heterochromatin markers, such as H3K27Me3 and H4K20Me3, and of the hallmark of apoptosis, cleaved-poly(ADP-ribose) polymerase-1. Our results propose innovative insights underlying the multifaceted role of RSV in MM, by pointing out the role of this natural compound in an lncRNA-mediated regulation to counteract cellular immortality.

Targeting SARS-CoV-2 Main Protease: A Successful Story Guided by an In Silico Drug Repurposing Approach.

The SARS-CoV-2 main protease (Mpro) is a crucial enzyme for viral replication and has been considered an attractive drug target for the treatment of COVID-19. In this study, virtual screening techniques and in vitro assays were combined to identify novel Mpro inhibitors starting from around 8000 FDA-approved drugs. The docking analysis highlighted 17 promising best hits, biologically characterized in terms of their Mpro inhibitory activity. Among them, 7 cephalosporins and the oral anticoagulant betrixaban were able to block the enzyme activity in the micromolar range with no cytotoxic effect at the highest concentration tested. After the evaluation of the degree of conservation of Mpro residues involved in the binding with the studied ligands, the ligands' activity on SARS-CoV-2 replication was assessed. The ability of betrixaban to affect SARS-CoV-2 replication associated to its antithrombotic effect could pave the way for its possible use in the treatment of hospitalized COVID-19 patients.

Hit identification of novel small molecules interfering with MALAT1 triplex by a structure-based virtual screening.

Nowadays, RNA is an attractive target for the design of new small molecules with different pharmacological activities. Among several RNA molecules, long noncoding RNAs (lncRNAs) are extensively reported to be involved in cancer pathogenesis. In particular, the overexpression of lncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) plays an important role in the development of multiple myeloma (MM). Starting from the crystallographic structure of the triple-helical stability element at the 3'-end of MALAT1, we performed a structure-based virtual screening of a large commercial database, previously filtered according to the drug-like properties. After a thermodynamic analysis, we selected five compounds for the in vitro assays. Compound M5, characterized by a diazaindene scaffold, emerged as the most promising molecule enabling the destabilization of the MALAT1 triplex structure and antiproliferative activity on in vitro models of MM. M5 is proposed as a lead compound to be further optimized for improving its affinity toward MALAT1.

Targeting SARS-CoV-2 nsp13 Helicase and Assessment of Druggability Pockets: Identification of Two Potent Inhibitors by a Multi-Site In Silico Drug Repurposing Approach.

The SARS-CoV-2 non-structural protein 13 (nsp13) helicase is an essential enzyme for viral replication and has been identified as an attractive target for the development of new antiviral drugs. In detail, the helicase catalyzes the unwinding of double-stranded DNA or RNA in a 5' to 3' direction and acts in concert with the replication-transcription complex (nsp7/nsp8/nsp12). In this work, bioinformatics and computational tools allowed us to perform a detailed conservation analysis of the SARS-CoV-2 helicase genome and to further predict the druggable enzyme's binding pockets. Thus, a structure-based virtual screening was used to identify valuable compounds that are capable of recognizing multiple nsp13 pockets. Starting from a database of around 4000 drugs already approved by the Food and Drug Administration (FDA), we chose 14 shared compounds capable of recognizing three out of four sites. Finally, by means of visual inspection analysis and based on their commercial availability, five promising compounds were submitted to in vitro assays. Among them, PF-03715455 was able to block both the unwinding and NTPase activities of nsp13 in a micromolar range.

Key genetic elements, single and in clusters, underlying geographically dependent SARS-CoV-2 genetic adaptation and their impact on binding affinity for drugs and immune control.

OBJECTIVES: To define key genetic elements, single or in clusters, underlying SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2) evolutionary diversification across continents, and their impact on drug-binding affinity and viral antigenicity. METHODS: A total of 12 150 SARS-CoV-2 sequences (publicly available) from 69 countries were analysed. Mutational clusters were assessed by hierarchical clustering. Structure-based virtual screening (SBVS) was used to select the best inhibitors of 3-chymotrypsin-like protease (3CL-Pr) and RNA-dependent RNA polymerase (RdRp) among the FDA-approved drugs and to evaluate the impact of mutations on binding affinity of these drugs. The impact of mutations on epitope recognition was predicted following Grifoni et al. (Cell Host Microbe 2020. 27: 671-80.). RESULTS: Thirty-five key mutations were identified (prevalence: ≥0.5%), residing in different viral proteins. Sixteen out of 35 formed tight clusters involving multiple SARS-CoV-2 proteins, highlighting intergenic co-evolution. Some clusters (including D614GSpike + P323LRdRp + R203KN + G204RN) occurred in all continents, while others showed a geographically restricted circulation (T1198KPL-Pr + P13LN + A97VRdRp in Asia, L84SORF-8 + S197LN in Europe, Y541CHel + H504CHel + L84SORF-8 in America and Oceania). SBVS identified 20 best RdRp inhibitors and 21 best 3CL-Pr inhibitors belonging to different drug classes. Notably, mutations in RdRp or 3CL-Pr modulate, positively or negatively, the binding affinity of these drugs. Among them, P323LRdRp (prevalence: 61.9%) reduced the binding affinity of specific compounds including remdesivir while it increased the binding affinity of the purine analogues penciclovir and tenofovir, suggesting potential hypersusceptibility. Finally, specific mutations (including Y541CHel + H504CHel) strongly hampered recognition of Class I/II epitopes, while D614GSpike profoundly altered the structural stability of a recently identified B cell epitope target of neutralizing antibodies (amino acids 592-620). CONCLUSIONS: Key genetic elements reflect geographically dependent SARS-CoV-2 genetic adaptation, and may play a potential role in modulating drug susceptibility and hampering viral antigenicity. Thus, a close monitoring of SARS-CoV-2 mutational patterns is crucial to ensure the effectiveness of treatments and vaccines worldwide.

Current status of antivirals and druggable targets of SARS CoV-2 and other human pathogenic coronaviruses.

Coronaviridae is a peculiar viral family, with a very large RNA genome and characteristic appearance, endowed with remarkable tendency to transfer from animals to humans. Since the beginning of the 21st century, three highly transmissible and pathogenic coronaviruses have crossed the species barrier and caused deadly pneumonia, inflicting severe outbreaks and causing human health emergencies of inconceivable magnitude. Indeed, in the past two decades, two human coronaviruses emerged causing serious respiratory illness: severe acute respiratory syndrome coronavirus (SARS-CoV-1) and Middle Eastern respiratory syndrome coronavirus (MERS-CoV), causing more than 10,000 cumulative cases, with mortality rates of 10 % for SARS-CoV-1 and 34.4 % for MERS-CoV. More recently, the severe acute respiratory syndrome coronavirus virus 2 (SARS-CoV-2) has emerged in China and has been identified as the etiological agent of the recent COVID-19 pandemic outbreak. It has rapidly spread throughout the world, causing nearly 22 million cases and ∼ 770,000 deaths worldwide, with an estimated mortality rate of ∼3.6 %, hence posing serious challenges for adequate and effective prevention and treatment. Currently, with the exception of the nucleotide analogue prodrug remdesivir, and despite several efforts, there is no known specific, proven, pharmacological treatment capable of efficiently and rapidly inducing viral containment and clearance of SARS-CoV-2 infection as well as no broad-spectrum drug for other human pathogenic coronaviruses. Another confounding factor is the paucity of molecular information regarding the tendency of coronaviruses to acquire drug resistance, a gap that should be filled in order to optimize the efficacy of antiviral drugs. In this light, the present review provides a systematic update on the current knowledge of the marked global efforts towards the development of antiviral strategies aimed at coping with the infection sustained by SARS-CoV-2 and other human pathogenic coronaviruses, displaying drug resistance profiles. The attention has been focused on antiviral drugs mainly targeting viral protease, RNA polymerase and spike glycoprotein, that have been tested in vitro and/or in clinical trials as well as on promising compounds proven to be active against coronaviruses by an in silico drug repurposing approach. In this respect, novel insights on compounds, identified by structure-based virtual screening on the DrugBank database endowed by multi-targeting profile, are also reported. We specifically identified 14 promising compounds characterized by a good in silico binding affinity towards, at least, two of the four studied targets (viral and host proteins). Among which, ceftolozane and NADH showed the best multi-targeting profile, thus potentially reducing the emergence of resistant virus strains. We also focused on potentially novel pharmacological targets for the development of compounds with anti-pan coronavirus activity. Through the analysis of a large set of viral genomic sequences, the current review provides a comprehensive and specific map of conserved regions across human coronavirus proteins which are essential for virus replication and thus with no or very limited tendency to mutate. Hence, these represent key druggable targets for novel compounds against this virus family. In this respect, the identification of highly effective and innovative pharmacological strategies is of paramount importance for the treatment and/or prophylaxis of the current pandemic but potentially also for future and unavoidable outbreaks of human pathogenic coronaviruses.

Ligand binding to telomeric G-quadruplex DNA investigated by funnel-metadynamics simulations.

G-quadruplexes (G4s) are higher-order DNA structures typically present at promoter regions of genes and telomeres. Here, the G4 formation decreases the replicative DNA at each cell cycle, finally leading to apoptosis. The ability to control this mitotic clock, particularly in cancer cells, is fascinating and passes through a rational understanding of the ligand/G4 interaction. We demonstrate that an accurate description of the ligand/G4 binding mechanism is possible using an innovative free-energy method called funnel-metadynamics (FM), which we have recently developed to investigate ligand/protein interaction. Using FM simulations, we have elucidated the binding mechanism of the anticancer alkaloid berberine to the human telomeric G4 (d[AG3(T2AG3)3]), computing also the binding free-energy landscape. Two ligand binding modes have been identified as the lowest energy states. Furthermore, we have found prebinding sites, which are preparatory to reach the final binding mode. In our simulations, the ions and the water molecules have been explicitly represented and the energetic contribution of the solvent during ligand binding evaluated. Our theoretical results provide an accurate estimate of the absolute ligand/DNA binding free energy ([Formula: see text] = -10.3 ± 0.5 kcal/mol) that we validated through steady-state fluorescence binding assays. The good agreement between the theoretical and experimental value demonstrates that FM is a most powerful method to investigate ligand/DNA interaction and can be a useful tool for the rational design also of G4 ligands.

Natural Products Extracted from Fungal Species as New Potential Anti-Cancer Drugs: A Structure-Based Drug Repurposing Approach Targeting HDAC7.

Mushrooms can be considered a valuable source of natural bioactive compounds with potential polypharmacological effects due to their proven antimicrobial, antiviral, antitumor, and antioxidant activities. In order to identify new potential anticancer compounds, an in-house chemical database of molecules extracted from both edible and non-edible fungal species was employed in a virtual screening against the isoform 7 of the Histone deacetylase (HDAC). This target is known to be implicated in different cancer processes, and in particular in both breast and ovarian tumors. In this work, we proposed the ibotenic acid as lead compound for the development of novel HDAC7 inhibitors, due to its antiproliferative activity in human breast cancer cells (MCF-7). These promising results represent the starting point for the discovery and the optimization of new HDAC7 inhibitors and highlight the interesting opportunity to apply the "drug repositioning" paradigm also to natural compounds deriving from mushrooms.

Multi-Targeting Bioactive Compounds Extracted from Essential Oils as Kinase Inhibitors.

Essential oils (EOs) are popular in aromatherapy, a branch of alternative medicine that claims their curative effects. Moreover, several studies reported EOs as potential anti-cancer agents by inducing apoptosis in different cancer cell models. In this study, we have considered EOs as a potential resource of new kinase inhibitors with a polypharmacological profile. On the other hand, computational methods offer the possibility to predict the theoretical activity profile of ligands, discovering dangerous off-targets and/or synergistic effects due to the potential multi-target action. With this aim, we performed a Structure-Based Virtual Screening (SBVS) against X-ray models of several protein kinases selected from the Protein Data Bank (PDB) by using a chemoinformatics database of EOs. By evaluating theoretical binding affinity, 13 molecules were detected among EOs as new potential kinase inhibitors with a multi-target profile. The two compounds with higher percentages in the EOs were studied more in depth by means Induced Fit Docking (IFD) protocol, in order to better predict their binding modes taking into account also structural changes in the receptor. Finally, given its good binding affinity towards five different kinases, cinnamyl cinnamate was biologically tested on different cell lines with the aim to verify the antiproliferative activity. Thus, this work represents a starting point for the optimization of the most promising EOs structure as kinase inhibitors with multi-target features.

Pathway involving the N155H mutation in HIV-1 integrase leads to dolutegravir resistance.

Background: Dolutegravir, an integrase strand-transfer inhibitor (STI), shows a high genetic barrier to resistance. Dolutegravir is reported to be effective against viruses resistant to raltegravir and elvitegravir. In this study, we report the case of a patient treated with dolutegravir monotherapy. Failure of dolutegravir treatment was observed concomitant with the appearance of N155H-K211R-E212T mutations in the integrase (IN) gene in addition to the polymorphic K156N mutation that was present at baseline in this patient. Methods: The impact of N155H-K156N-K211R-E212T mutations was studied in cell-free, culture-based assays and by molecular modelling. Results: Cell-free and culture-based assays confirm that selected mutations in the patient, in the context of the polymorphic mutation K156N present at the baseline, lead to high resistance to dolutegravir requiring that the analysis be done at timepoints longer than usual to properly reveal the results. Interestingly, the association of only N155H and K156N is sufficient for significant resistance to dolutegravir. Modelling studies showed that dolutegravir is less stable in IN/DNA complexes with respect to the WT sequence. Conclusions: Our results indicate that the stability of STI IN/DNA complexes is an important parameter that must be taken into account when evaluating dolutegravir resistance. This study confirms that a pathway including N155H can be selected in patients treated with dolutegravir with the help of the polymorphic K156N that acts as a secondary mutation that enhances the resistance to dolutegravir.

Real-life 3D therapy failure: Analysis of NS5A 93H RAS plus 108 K polymorphism in complex with ombitasvir by molecular modeling.

We report a real-life 3D therapy failure in a patient treated with ombitasvir (OMV)/paritaprevir/ritonavir and dasabuvir without ribavirin (3D-R). He had therapy failure at week 12 after the end of treatment. We detected resistance-associated substitutions (RASs) plus polymorphisms on NS3, NS5A, and NS5B target regions by population sequencing (15% cut-off) at baseline, at relapse and during follow-up. About this, NS5A RASs generally persist longer than resistances in the other target genes and may impact treatment outcome. Therefore, to evaluate OMV drug-resistance mechanism, we studied the acquired RAS plus polymorphisms on NS5A phosphoprotein by computational studies. OMV showed a higher affinity towards baseline and 93H/108 K mutant structure (follow-up) with respect to 93H/R108 mutant structure (relapse) on phosphoprotein. By Molecular Dynamics simulations (MDs), structural information about the protein stability in presence of OMV were observed. According to our data, molecular modeling approach has proved to be a powerful method to evaluate the impact of these RASs plus specific amino acid (AA) changes on phosphoprotein.

Identification of G-quadruplex DNA/RNA binders: Structure-based virtual screening and biophysical characterization.

BACKGROUND: Recent findings demonstrated that, in mammalian cells, telomere DNA (Tel) is transcribed into telomeric repeat-containing RNA (TERRA), which is involved in fundamental biological processes, thus representing a promising anticancer target. For this reason, the discovery of dual (as well as selective) Tel/TERRA G-quadruplex (G4) binders could represent an innovative strategy to enhance telomerase inhibition. METHODS: Initially, docking simulations of known Tel and TERRA active ligands were performed on the 3D coordinates of bimolecular G4 Tel DNA (Tel2) and TERRA (TERRA2). Structure-based pharmacophore models were generated on the best complexes and employed for the virtual screening of ~257,000 natural compounds. The 20 best candidates were submitted to biophysical assays, which included circular dichroism and mass spectrometry at different K+ concentrations. RESULTS: Three hits were here identified and characterized by biophysical assays. Compound 7 acts as dual Tel2/TERRA2 G4-ligand at physiological KCl concentration, while hits 15 and 17 show preferential thermal stabilization for Tel2 DNA. The different molecular recognition against the two targets was also discussed. CONCLUSIONS: Our successful results pave the way to further lead optimization to achieve both increased selectivity and stabilizing effect against TERRA and Tel DNA G4s. GENERAL SIGNIFICANCE: The current study combines for the first time molecular modelling and biophysical assays applied to bimolecular DNA and RNA G4s, leading to the identification of innovative ligand chemical scaffolds with a promising anticancer profile. This article is part of a Special Issue entitled "G-quadruplex" Guest Editor: Dr. Concetta Giancola and Dr. Daniela Montesarchio.

Incomplete APOBEC3G/F Neutralization by HIV-1 Vif Mutants Facilitates the Genetic Evolution from CCR5 to CXCR4 Usage.

Incomplete APOBEC3G/F neutralization by a defective HIV-1Vif protein can promote genetic diversification by inducing G-to-A mutations in the HIV-1 genome. The HIV-1 Env V3 loop, critical for coreceptor usage, contains several putative APOBEC3G/F target sites. Here, we determined if APOBEC3G/F, in the presence of Vif-defective HIV-1 virus, can induce G-to-A mutations at V3 positions critical to modulation of CXCR4 usage. Peripheral blood mononuclear cells (PBMC) and monocyte-derived macrophages (MDM) from 2 HIV-1-negative donors were infected with CCR5-using 81.A-VifWT virus (i.e., with wild-type [WT] Vif protein), 81.A-VifE45G, or 81.A-VifK22E (known to incompletely/partially neutralize APOBEC3G/F). The rate of G-toA mutations was zero or extremely low in 81.A-VifWT- and 81.A-VifE45G-infected PBMC from both donors. Conversely, G-to-A enrichment was detected in 81.A-VifK22E-infected PBMC (prevalence ranging from 2.18% at 7 days postinfection [dpi] to 3.07% at 21 dpi in donor 1 and from 10.49% at 7 dpi to 8.69% at 21 dpi in donor 2). A similar scenario was found in MDM. G-to-A mutations occurred at 8 V3 positions, resulting in nonsynonymous amino acid substitutions. Of them, G24E and E25K strongly correlated with phenotypically/genotypically defined CXCR4-using viruses (P = 0.04 and 5.5e-7, respectively) and increased the CXCR4 N-terminal binding affinity for V3 (WT, -40.1 kcal/mol; G24E, -510 kcal/mol; E25K, -522 kcal/mol). The analysis of paired V3 and Vif DNA sequences from 84 HIV-1-infected patients showed that the presence of a Vif-defective virus correlated with CXCR4 usage in proviral DNA (P = 0.04). In conclusion, incomplete APOBEC3G/F neutralization by a single Vif amino acid substitution seeds a CXCR4-using proviral reservoir. This can have implications for the success of CCR5 antagonist-based therapy, as well as for the risk of disease progression.

SI113, a specific inhibitor of the Sgk1 kinase activity that counteracts cancer cell proliferation.

BACKGROUND/AIMS: Published observations on serum and glucocorticoid regulated kinase 1 (Sgk1) knockout murine models and Sgk1-specific RNA silencing in the RKO human colon carcinoma cell line point to this kinase as a central player in colon carcinogenesis and in resistance to taxanes. METHODS: By in vitro kinase activity inhibition assays, cell cycle and viability analysis in human cancer model systems, we describe the biologic effects of a recently identified kinase inhibitor, SI113, characterized by a substituted pyrazolo[3,4-d]pyrimidine scaffold, that shows specificity for Sgk1. RESULTS: SI113 was able to inhibit in vitro cell growth in cancer cells derived from tumors with different origins. In RKO cells, this kinase inhibitor blocked insulin-dependent phosphorylation of the Sgk1 substrate Mdm2, the main regulator of p53 protein stability, and induced necrosis and apoptosis when used as a single agent. Finally, SI113 potentiated the effects of paclitaxel on cell viability. CONCLUSION: Since SI113 appears to be effective in inducing cell death in RKO cells, potentiating paclitaxel sensitivity, we believe that this new molecule could be efficiently employed, alone or in combination with paclitaxel, in colon cancer chemotherapy.

New raltegravir resistance pathways induce broad cross-resistance to all currently used integrase inhibitors.

OBJECTIVES: The possibility of replacing raltegravir or elvitegravir with dolutegravir in heavily treatment-experienced patients failing on raltegravir/elvitegravir has been evaluated in VIKING trials. All studied patients failed by the most common pathways, Y143, Q148 and N155, and dolutegravir demonstrated efficacy except for Q148 viruses. The aim of this study was to explore, in the same way, the behaviour of dolutegravir in comparison with raltegravir and elvitegravir against the atypical resistance integrase profiles, G118R and F121Y, described in HIV-1 patients failing on raltegravir therapy. METHODS: The behaviour of integrases with mutations G118R and F121Y towards raltegravir, elvitegravir and dolutegravir was analysed by evaluating phenotypic susceptibility and by means of in silico techniques (investigating binding affinities and the stabilization of the inhibitors in terms of their hydrogen bond network). RESULTS: The phenotypic analysis of G118R and F121Y showed high resistance to raltegravir, elvitegravir and dolutegravir with a fold change >100 when the clinically derived integrase was used, and resistance was also seen when mutations were tested alone in an NL43 backbone, but more often with a lower fold change. In silico, results showed that G118R and F121Y enzymes were associated with reduced binding affinities to each of the inhibitors and with a decreased number of hydrogen bonds compared with the wild-type complexes. CONCLUSIONS: This study showed that G118R and F121Y mutations, rarely described in patients failing on raltegravir, induced broad cross-resistance to all currently used integrase inhibitors. These results are in accordance with our thermodynamic and geometric analysis indicating decreased stability compared with the wild-type complexes.

Detecting and understanding genetic and structural features in HIV-1 B subtype V3 underlying HIV-1 co-receptor usage.

MOTIVATION: To define V3 genetic elements and structural features underlying different HIV-1 co-receptor usage in vivo. RESULTS: By probabilistically modeling mutations in the viruses isolated from HIV-1 B subtype patients, we present a unique statistical procedure that would first identify V3 determinants associated with the usage of different co-receptors cooperatively or independently, and then delineate the complicated interactions among mutations functioning cooperatively. We built a model based on dual usage of CXCR4 and CCR5 co-receptors. The molecular basis of our statistical predictions is further confirmed by phenotypic and molecular modeling analyses. Our results provide new insights on molecular basis of different HIV-1 co-receptor usage. This is critical to optimize the use of genotypic tropism testing in clinical practice and to obtain molecular-implication for design of vaccine and new entry-inhibitors.

Aryl ethynyl anthraquinones: a useful platform for targeting telomeric G-quadruplex structures.

Aryl ethynyl anthraquinones have been synthesized by Sonogashira cross-coupling and evaluated as telomeric G-quadruplex ligands, by the FRET melting assay, circular dichroism, the DNA synthesis arrest assay and molecular docking. Both the binding properties and G-quadruplex vs. duplex selectivity are controlled by the structures of the aryl ethynyl moieties.

In silico identification and biological evaluation of novel selective serum/glucocorticoid-inducible kinase 1 inhibitors based on the pyrazolo-pyrimidine scaffold.

The serum/glucocorticoid-inducible kinase 1 (Sgk1) has demonstrated antiapoptotic function and the capability to regulate cell survival, proliferation, and differentiation. A pivotal role of Sgk1 in carcinogenesis and in resistance to anticancer therapy has been suggested. With the aim of identifying new Sgk1 modulators, 322 pyrazolo-pyrimidine derivatives have been virtually screened with respect to a crystallographic model of Sgk1. The top five ranked compounds have been evaluated demonstrating Sgk1 inhibition in vitro and selectivity compared to RAC-alpha serine/threonine-protein kinase (Akt1).

Structure-based virtual screening of novel natural alkaloid derivatives as potential binders of h-telo and c-myc DNA G-quadruplex conformations.

Several ligands can bind to the non-canonical G-quadruplex DNA structures thereby stabilizing them. These molecules can act as effective anticancer agents by stabilizing the telomeric regions of DNA or by regulating oncogene expression. In order to better interact with the quartets of G-quadruplex structures, G-binders are generally characterized by a large aromatic core involved in π-π stacking. Some natural flexible cyclic molecules from Traditional Chinese Medicine have shown high binding affinity with G-quadruplex, such as berbamine and many other alkaloids. Using the structural information available on G-quadruplex structures, we performed a high throughput in silico screening of commercially available alkaloid derivative databases by means of a structure-based approach based on docking and molecular dynamics simulations against the human telomeric sequence d[AG3(T2AG3)3] and the c-myc promoter structure. We identified 69 best hits reporting an improved theoretical binding affinity with respect to the active set. Among them, a berberine derivative, already known to remarkably inhibit telomerase activity, was related to a better theoretical affinity versus c-myc.