Pharmacological and Therapeutic Properties of Punica granatum Phytochemicals: Possible Roles in Breast Cancer

Background: Pomgranate (Punica granatum) represents a high source of polyphenols with great bioavailability. The role of this fruit in the prevention and treatment of various malignant pathologies has been long time cited in both scientific and non-scientific literature, making thus important to identify its involvement in the pathophysiological processes. The treatment for breast cancer had focused on the inhibition of the mechanisms that governs the estrogen activity. These mechanisms are covered either by the antagonism of the estrogen receptor (ER) or by the inhibition of the estrogen synthesis. Our interest in identifying a bioactive compound rich in polyphenols, which induces both the antagonism of the estrogen receptor, and the inhibition of the estrogen synthesis, revealed us the pomegranate fruit and its derivatives: peel and seeds. Pomegranates’ chemical composition include many biological active substances such as flavonols, flavanols, anthocyanins, proanthocyanidins, ellagitannins and gallotannins. Materials and Methods: We performed a review of the scientific literature by using the following keywords: “pomegranate”, “breast cancer”, “Punica granatum”, “pomegranate polyphenols”. Our search was performed in the PubMed and Google Scholar databases, and it included only original research written in English from the last 20 years. None of the articles were excluded due to affiliation. A total number of 28 original papers, which mentioned the beneficial activity of pomegranate against breast cancer, were selected. Both clinical and preclinical studies were considered for this review. Results: Recent discoveries pointed out that polyphenols from Punica granatum possess strong anti-cancer activity, exhibited by a variety of mechanisms, such as anti-estrogenic, anti-proliferative, anti-angiogenetic, anti-inflammatory, and anti-metastatic. Pomegranate extracts induced cell cycle arrest in the G0/G1 phase, and induced cytotoxicity in a dose- and time-dependent manner. Moreover, several polyphenols extracted from pomegranate inhibited the invasion potential, migration and viability of breast cancer cells. The effects of pomegranate juice on serum estrogens and other sexual hormones levels were also investigated on two human cohorts. Conclusions: Punica granatum represents a promising area in oncology. The large availability and low cost, associated with the lack of side effects, made from this natural product a great strategy for the management of breast cancer. There are several mechanistic studies in mouse models and in breast cancer cell lines, suggesting the possible pathways through which polyphenols from pomegranate extracts act, but larger and better-controlled studies are necessary in the future. Only two small clinical trials were conducted on humans until now, but their results are contradictory and should be considered preliminary.     

Identification of novel HDAC8 selective inhibitors through ligand and structure based studies: Exploiting the acetate release channel differences among class I isoforms

Histone deacetylases (HDACs) are key regulators of gene expression and have emerged as crucial therapeutic targets for cancer. Among the HDACs, inhibition of HDAC8 enzyme has been reported to be a novel strategy in the treatment of female-specific cancers. Most of the HDAC inhibitors discovered so far inhibit multiple HDAC isoforms causing toxicities in the clinic thus limiting their potential. Therefore, the discovery of isoform-selective HDAC8 inhibitors is highly desirable. In the present study, a combination of ligand and structure based drug design tools were utilized to build a statistically significant pharmacophore based 3D QSAR model with statistical parameters R²: 0.9964, and Q²: 0.7154, from a series of 31 known HDAC8 inhibitors. Top 1000 hits obtained from Virtual screening of Phase database were subjected to docking studies against HDAC8. Top 100 hits obtained were redocked into HDAC Class I (HDAC 1,2,3) and Class II isoforms (HDAC 4, 6) and rescored with XP Glide Score. Based on fitness score, XP glide score and interacting amino acid residues, five HDAC8 inhibitors (1–5) were selected for in vitro studies. The HDAC8 activity assay followed by enzyme kinetics clearly indicated Compounds 1, 2 and 3 to be potent HDAC8 selective inhibitors with IC₅₀ of 126 pM, 112 nM, and 442 nM respectively. These compounds were cytotoxic to HeLa cells where HDAC8 is overexpressed but not to normal cells, HEK293. Also, they were able to induce apoptosis by modulating Bax/Bcl2, cleavage of PARP and release of Cytochrome C. Molecular Dynamics simulations observed most favorable interaction patterns and presented a rationale for the activities of the identified compounds. Selectivity against HDAC8 was due to exploitation of the architectural difference in the acetate release channel among class I HDAC isoforms.     

Binding Free Energy (BFE) Calculations and Quantitative Structure–Activity Relationship (QSAR) Analysis of Schistosoma mansoni Histone Deacetylase 8 (smHDAC8) Inhibitors

Histone-modifying proteins have been identified as promising targets to treat several diseases including cancer and parasitic ailments. In silico methods have been incorporated within a variety of drug discovery programs to facilitate the identification and development of novel lead compounds. In this study, we explore the binding modes of a series of benzhydroxamates derivatives developed as histone deacetylase inhibitors of Schistosoma mansoni histone deacetylase (smHDAC) using molecular docking and binding free energy (BFE) calculations. The developed docking protocol was able to correctly reproduce the experimentally established binding modes of resolved smHDAC8–inhibitor complexes. However, as has been reported in former studies, the obtained docking scores weakly correlate with the experimentally determined activity of the studied inhibitors. Thus, the obtained docking poses were refined and rescored using the Amber software. From the computed protein–inhibitor BFE, different quantitative structure–activity relationship (QSAR) models could be developed and validated using several cross-validation techniques. Some of the generated QSAR models with good correlation could explain up to ~73% variance in activity within the studied training set molecules. The best performing models were subsequently tested on an external test set of newly designed and synthesized analogs. In vitro testing showed a good correlation between the predicted and experimentally observed IC₅₀ values. Thus, the generated models can be considered as interesting tools for the identification of novel smHDAC8 inhibitors.     

Target Design of Novel Histone Deacetylase 6 Selective Inhibitors with 2-Mercaptoquinazolinone as the Cap Moiety

Epigenetic alterations found in all human cancers are promising targets for anticancer therapy. In this sense, histone deacetylase inhibitors (HDACIs) are interesting anticancer agents that play an important role in the epigenetic regulation of cancer cells. Here, we report 15 novel hydroxamic acid-based histone deacetylase inhibitors with quinazolinone core structures. Five compounds exhibited antiproliferative activity with IC₅₀ values of 3.4–37.8 µM. Compound 8 with a 2-mercaptoquinazolinone cap moiety displayed the highest antiproliferative efficacy against MCF-7 cells. For the HDAC6 target selectivity study, compound 8 displayed an IC₅₀ value of 2.3 µM, which is 29.3 times higher than those of HDAC3, HDAC4, HDAC8, and HDAC11. Western blot assay proved that compound 8 strongly inhibited tubulin acetylation, a substrate of HDAC6. Compound 8 also displayed stronger inhibition activity against HDAC11 than the control drug Belinostat. The inhibitory mechanism of action of compound 8 on HDAC enzymes was then explored using molecular docking study. The data revealed a high binding affinity (−7.92 kcal/mol) of compound 8 toward HDAC6. In addition, dock pose analysis also proved that compound 8 might serve as a potent inhibitor of HDAC11.     

Design and Synthesis of Non-Covalent Imidazo[1,2-a]quinoxaline-Based Inhibitors of EGFR and Their Anti-Cancer Assessment

A series of 30 non-covalent imidazo[1,2-a]quinoxaline-based inhibitors of epidermal growth factor receptor (EGFR) were designed and synthesized. EGFR inhibitory assessment (against wild type) data of compounds revealed 6b, 7h, 7j, 9a and 9c as potent EGFR^WT inhibitors with IC₅₀ values of 211.22, 222.21, 193.18, 223.32 and 221.53 nM, respectively, which were comparable to erlotinib (221.03 nM), a positive control. Furthermore, compounds exhibited excellent antiproliferative activity when tested against cancer cell lines harboring EGFR^WT; A549, a non-small cell lung cancer (NSCLC), HCT-116 (colon), MDA-MB-231 (breast) and gefitinib-resistant NSCLC cell line H1975 harboring EGFR^L858R/T790M. In particular, compound 6b demonstrated significant inhibitory potential against gefitinib-resistant H1975 cells (IC₅₀ = 3.65 μM) as compared to gefitinib (IC₅₀ > 20 μM). Moreover, molecular docking disclosed the binding mode of the 6b to the domain of EGFR (wild type and mutant type), indicating the basis of inhibition. Furthermore, its effects on redox modulation, mitochondrial membrane potential, cell cycle analysis and cell death mode in A549 lung cancer cells were also reported.     

New 1,3,4-Thiadiazole Derivatives with Anticancer Activity

We designed and synthesized the 1,3,4-thiadiazole derivatives differing in the structure of the substituents in C2 and C5 positions. The cytotoxic activity of the obtained compounds was then determined in biological studies using MCF-7 and MDA-MB-231 breast cancer cells and normal cell line (fibroblasts). The results showed that in both breast cancer cell lines, the strongest anti-proliferative activity was exerted by 2-(2-trifluorometylophenylamino)-5-(3-methoxyphenyl)-1,3,4-thiadiazole. The IC₅₀ values of this compound against MCF-7 and MDA-MB-231 breast cancer cells were 49.6 µM and 53.4 µM, respectively. Importantly, all new compounds had weaker cytotoxic activity on normal cell line than on breast cancer cell lines. In silico studies demonstrated a possible multitarget mode of action for the synthesized compounds. The most likely mechanism of action for the new compounds is connected with the activities of Caspase 3 and Caspase 8 and activation of BAX proteins.     

Syntheses of amamistatin fragments and determination of their HDAC and antitumor activity

[reaction: see text] Amamistatins A and B are natural products found to have anti-proliferative effects against MCF-7, A549, and MKN45 human tumor cell lines (IC50 0.24-0.56 microM). It was proposed that their activity was due to histone deacetylase (HDAC) inhibition mediated by the N-formyl-N-hydroxy lysine moiety. Amamistatin B fragment analogs were synthesized and screened for biological activity. These compounds were modest HDAC inhibitors and showed antitumor activity against MCF-7 and PC-3 human tumor cells.     

Evaluation of Adding Natural Gum to Pectin Extracted from Ecuadorian Citrus Peels as an Eco-Friendly Corrosion Inhibitor for Carbon Steel

The production and use of eco-friendly corrosion inhibitors allows valuable compounds contained in plant waste to be identified and repurposed while reducing the use of polluting synthetic substances. Pectin extracted from Tahiti limes (Citrus latifolia) and King mandarin (Citrus nobilis L.) in addition to natural gums—xanthan gum and latex from the “lechero” plant (Euphorbia laurifolia)—were used to create an eco-friendly corrosion inhibitor. The optimal extraction conditions for pectin were determined from different combinations of pH, temperature, and time in a 2³ factorial design and evaluated according to the obtained pectin yield. The highest pectin extraction yields (38.10% and 41.20% from King mandarin and lime, respectively) were reached at pH = 1, 85 °C, and 2 h. Extraction of pectic compounds was confirmed using Fourier-transform infrared spectroscopy, differential scanning calorimetry, and thermogravimetry analyses. Subsequently, a simplex-centroid mixture design was applied to determine the formulation of extracted pectin and natural gums that achieved the highest corrosion inhibitor effect (linear polarization and weight loss methods in NACE 1D-196 saline media using API-5LX52 carbon steel). Impedance spectroscopy analysis showed that the addition of xanthan gum to pectin (formulation 50% pectin–50% xanthan gum) improved the corrosion inhibitor effect from 29.20 to 78.21% at 400 ppm due to higher adsorption of inhibitory molecules on the metal surface.     

Natural Compounds Isolated from Stachybotrys chartarum Are Potent Inhibitors of Human Protein Kinase CK2

A large number of secondary metabolites have been isolated from the filamentous fungus Stachybotrys chartarum and have been described before. Fourteen of these natural compounds were evaluated in vitro in the present study for their inhibitory activity towards the cancer target CK2. Among these compounds, stachybotrychromene C, stachybotrydial acetate and acetoxystachybotrydial acetate turned out to be potent inhibitors with IC₅₀ values of 0.32 µM, 0.69 µM and 1.86 µM, respectively. The effects of these three compounds on cell proliferation, growth and viability of MCF7 cells, representing human breast adenocarcinoma as well as A427 (human lung carcinoma) and A431 (human epidermoid carcinoma) cells, were tested using EdU assay, IncuCyte^® live-cell imaging and MTT assay. The most active compound in inhibiting MCF7 cell proliferation was acetoxystachybotrydial acetate with an EC₅₀ value of 0.39 µM. In addition, acetoxystachybotrydial acetate turned out to inhibit the growth of all three cell lines completely at a concentration of 1 µM. In contrast, cell viability was impaired only moderately, to 37%, 14% and 23% in MCF7, A427 and A431 cells, respectively.     

Antioxidant and Antitumor Activities of Newly Synthesized Hesperetin Derivatives

Hesperetin is a class of natural products with a wide range of sources and remarkable biological activities. In this study, we described the synthesis of a series of novel hesperetin derivatives and evaluated the in vitro antioxidant and antitumor activity of these compounds. Eleven novel compounds were synthesized in moderate yields. The compounds synthesized in this work exhibited antioxidant activities against DPPH and ABTS free radicals in a dose-dependent manner. Among them, compound 3f had the best antioxidant activity, with IC₅₀ of 1.2 μM and 24 μM for DPPH and ABTS, respectively. The antitumor activity of the compounds against human cancer cell lines, such as breast MCF-7, liver HepG2, and cervical Hela, was determined by a standard 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Three compounds had moderate IC₅₀ values. Interestingly, compound 3f had better biological activity than hesperetin, which matches the prediction by Maestro from Schrödinger. Therefore, the new hesperidin derivative is a promising drug for the treatment of cancer due to its effective antitumor activity. The results also suggested that the antitumor activities of hesperetin derivatives may be related to their antioxidant activities.     

The Development of BTK Inhibitors: A Five-Year Update

Bruton’s tyrosine kinase (BTK) represented, in the past ten years, an important target for the development of new therapeutic agents that could be useful for cancer and autoimmune disorders. To date, five compounds, able to block BTK in an irreversible manner, have been launched in the market, whereas many reversible BTK inhibitors (BTKIs), with reduced side effects that are more useful for long-term administration in autoimmune disorders, are under clinical investigation. Despite the presence in the literature of many articles and reviews, studies on BTK function and BTKIs are of great interest for pharmaceutical companies as well as academia. This review is focused on compounds that have appeared in the literature from 2017 that are able to block BTK in an irreversible or reversible manner; also, new promising tunable irreversible inhibitors, as well as PROTAC molecules, have been reported. This summary could improve the knowledge of the chemical diversity of BTKIs and provide information for future studies, particularly from the medicinal chemistry point of view. Data reported here are collected from different databases (Scifinder, Web of Science, Scopus, Google Scholar, and Pubmed) using “BTK” and “BTK inhibitors” as keywords.     

Comparative Study of Volatile Compounds and Expression of Related Genes in Fruit from Two Apple Cultivars during Different Developmental Stages

Aromatic volatile compounds are important contributors to fruit quality that vary among different cultivars. Herein, headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry was used to determine changes in volatile compounds and related gene expression patterns in “Ruixue” and “Fuji” apples (Malus domestica Borkh.) during fruit development and maturation. Volatile compounds detected in the fruit of both cultivars exhibited similar trends across different developmental stages. In the early stages of “Ruixue” fruit development (60 days after full bloom), there were fewer volatile compounds, mainly aldehydes (87.0%). During fruit maturation (180 days after full bloom), the types and amounts of volatile compounds increased, mainly including esters (37.6%), and alkenes (23.2%). The total volatile concentration, the types of major volatile compounds, and their relative content in both cultivars varied across different stages. Gene expression analysis indicated that the upregulation of MdLOX, MdAAT2, and MdADH3 was associated with increased aroma compound content, especially esters, during fruit development in both cultivars. Changes in the expression of MdArAT, MdACPD, MdADH3, MdAAT2, and MdLOX may lead to differences in volatile compounds between apple cultivars.     

A One-Pot Six-Component Reaction for the Synthesis of 1,5-Disubstituted Tetrazol-1,2,3-Triazole Hybrids and Their Cytotoxic Activity against the MCF-7 Cell Line

A high-order multicomponent reaction involving a six-component reaction to obtain the novel linked 1,5-disubstituted tetrazole-1,2,3-triazole hybrids in low to moderate yield is described. This one-pot reaction is carried out under a cascade process consisting of three sequential reactions: Ugi-azide, bimolecular nucleophilic substitution (S_N2), and copper-catalyzed alkyne–azide reaction (CuAAC), with high atom and step-economy due the formation of six new bonds (one C-C, four C-N, and one N-N). Thus, the protocol developed offers operational simplicity, mild reaction conditions, and structural diversity. Finally, to evaluate the antitumoral potential of the synthetized molecules, a proliferation study was performed in the breast cancer (BC) derived cell line MCF-7. The hybrid compounds showed several degrees of cell proliferation inhibition with a remarkable effect in those compounds with cyclohexane and halogens in their structures. These compounds represent potential drug candidates for breast cancer treatment. However, additionally assays are needed to elucidate their complete effect over the cellular hallmarks of cancer.     

The Antiproliferative and Apoptotic Effect of a Novel Synthesized S-Triazine Dipeptide Series,and Toxicity Screening in Zebrafish Embryos

Several derivatives containing morpholine/piperidine, anilines, and dipeptides as pending moieties were prepared using s-triazine as a scaffold. These compounds were evaluated for their anticancer activity against two human breast cancer cell lines (MCF-7 and MDA-MB-231), a colon cancer cell line (HCT-116), and a non-tumorigenic cell line (HEK 293). Tamoxifen was used as a reference. Animal toxicity tests were carried out in zebrafish embryos. Most of these compounds showed a higher activity against breast cancer than colon cancer. Compound 3a—which contains morpholine, aniline, and glycylglycinate methyl ester—showed a high level of cytotoxicity against MCF-7 cells with IC₅₀ values of less than 1 µM. This compound showed a much lower level of toxicity against the non-tumorigenic HEK-293 cell line, and in the in vivo studies using zebrafish embryos. Furthermore, it induced cell cycle arrest at the G2/M phase, and apoptosis in MCF-7 cells. On the basis of our results, 3a emerges as a potential candidate for further development as a therapeutic drug to treat hormone receptor-positive breast cancer.     

Bioactive Metabolites from Marine Algae as Potent Pharmacophores against Oxidative Stress-Associated Human Diseases: A Comprehensive Review

In addition to cancer and diabetes, inflammatory and ROS-related diseases represent one of the major health problems worldwide. Currently, several synthetic drugs are used to reduce oxidative stress; nevertheless, these approaches often have side effects. Therefore, to overcome these issues, the search for alternative therapies has gained importance in recent times. Natural bioactive compounds have represented, and they still do, an important source of drugs with high therapeutic efficacy. In the “synthetic” era, terrestrial and aquatic photosynthetic organisms have been shown to be an essential source of natural compounds, some of which might play a leading role in pharmaceutical drug development. Marine organisms constitute nearly half of the worldwide biodiversity. In the marine environment, algae, seaweeds, and seagrasses are the first reported sources of marine natural products for discovering novel pharmacophores. The algal bioactive compounds are a potential source of novel antioxidant and anticancer (through modulation of the cell cycle, metastasis, and apoptosis) compounds. Secondary metabolites in marine Algae, such as phenolic acids, flavonoids, and tannins, could have great therapeutic implications against several diseases. In this context, this review focuses on the diversity of functional compounds extracted from algae and their potential beneficial effects in fighting cancer, diabetes, and inflammatory diseases.     

Cytotoxic 13,28 Epoxy Bridged Oleanane-Type Triterpenoid Saponins from the Roots of Ardisia crispa

Ardisiacrispin D–F (1–3), three new 13,28 epoxy bridged oleanane-type triterpenoid saponins, together with four known analogues (4–7) were isolated from the roots of Ardisia crispa. The structures of 1–7 were elucidated based on 1D and 2D-NMR experiments and by comparing their spectroscopic data with values from the published literatures. Ardisiacrispin D–F (1–3) are first examples that the monosaccharide directly linked to aglycone C-3 of triterpenoid saponins in genus Ardisia are non-arabinopyranose. In the present paper, all compounds are evaluated for the cytotoxicity against three cancer cell lines (HeLa, HepG2 and U87 MG) in vitro. The results show that compounds 1, 4 and 6 exhibited significant cytotoxicity against Hela and U87 MG cells with IC₅₀ values in the range of 2.2 ± 0.6 to 9.5 ± 1.8 µM. The present investigation suggests that roots of A. crispa could be a potential source of natural anti-tumor agents and their triterpenoid saponins might be responsible for cytotoxicity.     

Cheminformatic Profiling and Hit Prioritization of Natural Products with Activities against Methicillin-Resistant Staphylococcus aureus (MRSA)

Several natural products (NPs) have displayed varying in vitro activities against methicillin-resistant Staphylococcus aureus (MRSA). However, few of these compounds have not been developed into potential antimicrobial drug candidates. This may be due to the high cost and tedious and time-consuming process of conducting the necessary preclinical tests on these compounds. In this study, cheminformatic profiling was performed on 111 anti-MRSA NPs (AMNPs), using a few orally administered conventional drugs for MRSA (CDs) as reference, to identify compounds with prospects to become drug candidates. This was followed by prioritizing these hits and identifying the liabilities among the AMNPs for possible optimization. Cheminformatic profiling revealed that most of the AMNPs were within the required drug-like region of the investigated properties. For example, more than 76% of the AMNPs showed compliance with the Lipinski, Veber, and Egan predictive rules for oral absorption and permeability. About 34% of the AMNPs showed the prospect to penetrate the blood–brain barrier (BBB), an advantage over the CDs, which are generally non-permeant of BBB. The analysis of toxicity revealed that 59% of the AMNPs might have negligible or no toxicity risks. Structure–activity relationship (SAR) analysis revealed chemical groups that may be determinants of the reported bioactivity of the compounds. A hit prioritization strategy using a novel “desirability scoring function” was able to identify AMNPs with the desired drug-likeness. Hit optimization strategies implemented on AMNPs with poor desirability scores led to the design of two compounds with improved desirability scores.     

A “Double-Locked” and Enzyme/pH-Activated Theranostic Agent for Accurate Tumor Imaging and Therapy

Theranostic agents for concurrent cancer therapy and diagnosis have begun attracting attention as a promising modality. However, accurate imaging and identification remains a great challenge for theranostic agents. Here, we designed and synthesized a novel theranostic agent H6M based on the “double-locked” strategy by introducing an electron-withdrawing nitro group into 1-position of a pH-responsive 3-amino-β-carboline and further covalently linking the hydroxamic acid group, a zinc-binding group (ZBG), to the 3-position of β-carboline to obtain histone deacetylase (HDAC) inhibitory effect for combined HDAC-targeted therapy. We found that H6M can be specifically reduced under overexpressed nitroreductase (NTR) to produce H6AQ, which emits bright fluorescence at low pH. Notably, H6M demonstrated a selective fluorescence imaging via successive reactions with NTR (first “key”) and pH (second “key”), and precisely identified tumor margins with a high S/N ratio to guide tumor resection. Finally, H6M exerted robust HDAC1/cancer cell inhibitory activities compared with a known HDAC inhibitor SAHA. Therefore, the NTR/pH-activated theranostic agent provided a novel tool for precise diagnosis and efficient tumor therapy.