![Yeast-based p53 dual-luciferase transactivation assay using cells (A) expressing human wt/mutant p53 or (B) co-expressing human wt p53 and a cofactor (e.g. MDM2 or 53BP1). The assay exploits the variable expression of p53 proteins and utilizes the Firefly and Renilla luminescent reporters integrated as single copies at different chromosomal loci in haploid strains or at the same chromosomal location in diploid strains (heteroalleles). While a common minimal promoter controls low-level constitutive expression of both reporters, p53-dependent expression of the Firefly reporter is attained through a specific p53 response element (RE) placed upstream of the minimal promoter (pCYC1) [20]. The p53 transcriptional activity is evaluated by quantification of luciferase activity used as reporter gene, which is directly proportional to the light units measured in a plate reader.](https://www.researchgate.net/profile/Lucilia-Saraiva/publication/225084949/figure/fig1/AS:393880815063049@1470919912231/Fig-1-Yeast-based-p53-dual-luciferase-transactivation-assay-using-cells-A_Q320.jpg)
![Principle of DNA microarray assays. Typically, by this methodology DNA, cDNA or mRNA from yeast controls and samples (e.g. expression of a foreign gene, mutation, drug treatment) are isolated and differentially labeled with two fluorophores. Both labeled populations are mixed and hybridized to the DNA microarray. DNA microarrays are arrays of thousands of DNA sequences (e.g. PCR products or oligonucleotides), representing in the case of yeast all 6,200 genes, printed at high density onto a glass slide. Hybridization to spotted cDNA is detected by laser scanning, and the ratio of the dual-color fluorescent signal at any cDNA spot reveals for each target gene the difference in expression between control and sample. The large amount of results obtained is analyzed by clustering algorithms and data visualization tools [122,123].](https://www.researchgate.net/profile/Lucilia-Saraiva/publication/225084949/figure/fig4/AS:393880815063062@1470919912502/Fig-4-Principle-of-DNA-microarray-assays-Typically-by-this-methodology-DNA-cDNA-or_Q320.jpg)
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New Therapeutic Strategies for Cancer and Neurodegeneration Emerging from Yeast Cell-based Systems
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Despite great advances in understanding the molecular etiology of cancer and neurodegeneration, therapeutic strategies against these diseases are still largely lacking. Hence, acceleration of the discovery of new therapeutic agents against these pathologies is of enormous interest. This review is focused on the role of multi-faceted and expanding yeast cell-based systems in the search for new drugs and therapeutic targets in cancer and neurodegeneration. Though the obvious limitations of using a microorganism to address human diseases, when used in the early phase and with complementary mammalian systems, it can have a tremendous impact in the discovery of new therapeutic opportunities. In this review, many evidence are provided demonstrating the valuable contribution of yeast in this area. Additionally, several yeast target-based drug screening approaches based on a readily screenable phenotype on genomic technologies increasingly oriented towards genetic and chemical high-throughput analysis are addressed. Altogether, with this review, we intend not only to recognize previous successes and ongoing work in this area, but also to point out new opportunities that may be of interest for yeast as a model organism and as a powerful system in the discovery of new lead compounds that have the potential to become novel drugs in cancer and neurodegeneration.
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... The impairment of antioxidant defense systems is among the main effects provoked by exposures to mercury compounds in experimental studies [4]. Yeast antioxidant machinery presents a wide range of enzymes that in response to oxidative insults can be accurately managed [49][50][51]. In S. cerevisiae, the expression of antioxidant enzyme genes is tightly controlled by the transcription factor Yap1, a functional homologous of mammalian AP-1 factor (Activator protein 1) [32]. ...
... IC50 values for MeHg and EtHg on cell growth in WT and mutant strains. ...
... This group is thought to be responsible for 1.5 billion cancer cases and 1.2 billion deaths worldwide [31]. In addition, with an estimated 8.5 billion people, total fatalities are predicted to reach 2.14 billion by 2030 [32,33]. Lung and breast cancers are the most frequent causes of mortality in women and men, respectively, with 522,000 deaths in 2012. ...
... Imidazoles have anticancer drug properties, according to several study articles. The development of imidazoles as antineoplastic drugs began with dacarbazine, Fig. 11 Imidazole-based anticancer drugs' chemical structures [23][24][25][26][27][28][29][30][31][32][33][34] Fig. 12 Literature survey on recent reports about imidazole derivatives [45][46][47][48] which piqued interest in imidazole compounds. Various types of structured heterocyclic compounds, including imidazole, have been created to treat cancer by focusing on distinct targets. ...
Imidazole and its derivatives are one of the most important and widely used heterocycles in medicinal chemistry, natural products, and synthetic chemicals. Because of the imidazole ring's unique structural features, imidazole derivatives may easily attach to a wide range of enzymes and receptors via a number of weak interactions, demonstrating a wide range of biological and pharmacological effects. Several imidazole derivatives have been widely used in the clinic to treat a wide range of illnesses, suggesting their enormous potential for further research. In the realm of medicinal chemistry, this chapter presents a comprehensive summary of current findings in imidazole derivatives as anti-inflammatory, anticancer, antiviral, antibacterial, and other therapeutic agents.
... The estimate is that 1.5 billion cancer cases will bediagnosed, and that 1.2 billion fatalities will occur (Bray and Møller, 2006). In addition, with an anticipated 8.5 billion people on the planet, global mortality is expected to hit 2.14 billion by 2030 (Ali et al., 2011;Pereira et al., 2012). Cancers of the breast and lung are major causes of death in both women and men (5,22,000 deaths in 2012). ...
In this work, we synthesize the sulfonated Schiff bases of the chitosan derivatives 2a-2j without the use of a catalyst in two moderately straightforward steps with good yield within a short reaction time. The morphology and chemical structure of chitosan derivatives were investigated using FT-IR, NMR (1 H-13 C), XRD, and SEM. Furthermore, our chitosan derivatives were tested for their anticancer activity against the MCF-7 cancer cell line, and doxorubicin was used as a standard. In addition, the normal cell lines of the breast cancer cell MCF-10A, and of the lung cell MRC-5 were tested. Compound 2 h, with a GI 50 value of 0.02 µM for MCF-7, is highly active compared with the standard doxorubicin and other compounds. The synthesized compounds 2a-2j exhibit low cytotoxicity, with IC 50 > 100 μg/ml, against normal cell lines MCF-10A, MRC-5. We also provide the results of an in-silico study involving the Methoxsalen protein (1Z11). Compound 2h exhibits a higher binding affinity for 1Z11 protein (−5.9 kcal/mol) and a lower binding affinity for Doxorubicin (−5.3 kcal/mol) than certain other compounds. As a result of the aforementioned findings, the use of compound 2h has an anticancer drug will be researched in the future.
... The estimate is that 1.5 billion cancer cases will bediagnosed, and that 1.2 billion fatalities will occur (Bray and Møller, 2006). In addition, with an anticipated 8.5 billion people on the planet, global mortality is expected to hit 2.14 billion by 2030 (Ali et al., 2011;Pereira et al., 2012). Cancers of the breast and lung are major causes of death in both women and men (5,22,000 deaths in 2012). ...
In this work, we synthesize the sulfonated Schiff bases of the chitosan derivatives 2a-2j without the use of a catalyst in two moderately straightforward steps with good yield within a short reaction time. The morphology and chemical structure of chitosan derivatives were investigated using FT-IR, NMR (¹H—¹³C), XRD, and SEM. Furthermore, our chitosan derivatives were tested for their anticancer activity against the MCF-7 cancer cell line, and doxorubicin was used as a standard. In addition, the normal cell lines of the breast cancer cell MCF-10A, and of the lung cell MRC-5 were tested. Compound 2 h, with a GI50 value of 0.02 µM for MCF-7, is highly active compared with the standard doxorubicin and other compounds. The synthesized compounds 2a-2j exhibit low cytotoxicity, with IC50 > 100 μg/ml, against normal cell lines MCF-10A, MRC-5. We also provide the results of an in-silico study involving the Methoxsalen protein (1Z11). Compound 2h exhibits a higher binding affinity for 1Z11 protein (−5.9 kcal/mol) and a lower binding affinity for Doxorubicin (−5.3 kcal/mol) than certain other compounds. As a result of the aforementioned findings, the use of compound 2h has an anticancer drug will be researched in the future.
... The use of a simple yeast genetic model to perform large-scale screenings has provided insights into the molecular mechanisms of several human diseases [105]. For example, several yeast models have been developed to study α-synuclein toxicity and better understand disease-related defects in vesicular trafficking, oxidative stress, and mitochondrial dysfunction [106][107][108][109]. ...
Mitophagy, the selective degradation of mitochondria by autophagy, is one of the most important mechanisms of mitochondrial quality control, and its proper functioning is essential for cellular homeostasis. In this review, we describe the most important milestones achieved during almost 2 decades of research on yeasts, which shed light on the molecular mechanisms, regulation, and role of the Atg32 receptor in this process. We analyze the role of ROS in mitophagy and discuss the physiological roles of mitophagy in unicellular organisms, such as yeast; these roles are very different from those in mammals. Additionally, we discuss some of the different tools available for studying mitophagy.
Topoisomerases I and II are the functionally two forms of DNA topoisomerase. In anticancer research, novel anticancer chemotherapeutical capable of blocking topoisomerase enzymes have been discovered. Most commonly, topoisomerase causes replication fork arrest and doublestrand breaks, and this is how a clinically successful topoisomerase-targeting anticancer medicines work. Unfortunately, this novel mechanism of action has been linked to the development of secondary malignancies as well as cardiotoxicity. The specific binding locations and mechanisms of topoisomerase poisons have been identified by studying the structures of topoisomerase-drug-DNA ternary complexes. Recent breakthroughs in science have revealed that isoform-specific human topoisomerase II poison could be created as safer anticancer drug molecules. It may also be able to develop catalytic inhibitors of topoisomerases by focusing on their inactive conformations. In addition to this, the discovery of new bacterial topoisomerase inhibitor molecules and regulatory proteins could lead to the discovery of new human topoisomerase inhibitors. As a result, biologists, organic chemists, and medicinal chemists worldwide have been identifying, designing, synthesizing, and testing a variety of novel topoisomerase-targeting bioactive compounds. This review focused on topoisomerase inhibitors, their mechanisms of action, and different types of topoisomerase inhibitors that have been developed during the last ten years.
Bu çalışmada N-benziletilendiamin ile 2-Hidroksi-5-metilbenzaldehit, 2-Hidroksi-5-metilasetofenon, 2-Hidroksi-5-kloroasetofenon bileşiklerinin kondenzasyon reaksiyonu sonucu üç yeni Schiff bazı sentezlendi (L1, L2 ve L3). Sentezlenen yeni bileşiklerin yapıları, 1H NMR, 13C NMR, FT-IR ve elemental analiz yöntemleri ile karakterize edildi. Daha sonra, elde edilen bileşiklerin sitotoksik aktiviteleri, insan kolon kanser hücre dizisi (Caco-2, kolon ve HT-29 kolon karsinoma hücre dizisi) kullanılarak araştırıldı. Elde edilen sonuçlar standart kemoterapi ilacı 5-Fluorourasil (5-FU) ile karşılaştırıldı. Sitotoksisite sonuçlarına göre L2 bileşiğinin Caco-2 (kolon) ve HT-29 (kolon) hücre hatlarında en yüksek antikanser aktiviteye sahip olduğu bulundu.
A novel Schiff base namely 3,5‐di‐tert‐butyl‐6‐((2‐(perfluorophenyl) hydrazono) methyl) phenol was successfully synthesized and characterized using FTIR and 1H‐NMR, 13C‐NMR, and 19F‐NMR. The crystal structure analysis of the Schiff base compound was also characterized with single crystal X‐ray diffraction studies and supported the spectroscopic results. The cytotoxicity, anti‐bacterial properties, and enzyme inhibition of the compound were also investigated. The molecular docking studies were performed in order to explain the interactions of the synthesized compound with target enzymes. The newly synthesized hydrazone derivative Schiff base compound showed high cellular toxicity on MCF‐7 and PC‐3 cells. Also, this compound caused low antibacterial effect on E. coli and S. aureus. Besides, the compound exhibited the inhibitory effect against pancreatic cholesterol esterase and carbonic anhydrase isoenzyme I, II with IC50 values 63, 99, and 188 µM, respectively. Consequently, it has been determined that the prepared Schiff base is an active compound in terms of cytotoxicity, enzyme inhibition, and anti‐bacterial properties. These results provide preliminary information for some biological features of the compound and can play a major role in drug applications of the Schiff base compound.
In the present study, new Schiff bases derived from pentafluorophenyl-hydrazine (L1, L2, L3, L4) were synthesized and their structures were characterized by 1 H nuclear magnetic resonance spectroscopy (NMR), 13 C NMR, 19 F NMR, fourier transform infrared spectroscopy, and elemental analysis methods. Then, the anticancer activities of the obtained compounds were investigated using three human cancer cell lines (A2780, over; Caco-2, colon; and HT-29 colon carcinoma cell lines). According to the obtained cytotoxicity results, compound number L4 was found to have the highest anticancer activity in A2780 (over) and Caco-2 (colon) cell lines. Furthermore, in silico, ADMET properties, where studies play an important role in the development and prediction of drug compounds, were calculated using web-based platforms. In addition, molecular docking studies were performed to evaluate the binding interactions between the synthesized pentafluorophenyl-hydrazone compounds and the MDM2 protein (4JSC). Both in vitro and in silico results showed that the synthesized compounds could act as potent anticancer agents.
Starches from cowpea (Vigna Unguiculata), Bambara groundnut (Vigna Subterranea), and six variety of common beans (Phaseolus Vulgaris) cultivated in the Far North, Adamaoua and Western regions of Cameroon, were study. We determined the physical properties and the water absorption kinetics of the seeds, isolated the starches and determine their physicochemical properties and finally evaluate their functional properties. The starches were isolated according to the alkaline method. The physical properties (granule shape and size), chemical composition (moisture content, starch and amylose content) and functional properties (solubility index, swelling power, viscosity, gelation and syneresis) of the isolated starches were determined. Important results have been obtained, highlighting interesting physicochemical properties of these starches. Indeed, there are significant differences (p < 0,05) between the physical properties of seeds both between varieties of the same species and between the different seed species used. The proposed Peleg model provided water absorption capacity values approximately equal to those obtained experimentally. The soaking temperature significantly influenced the Peleg kinetic water absorption constants. The purity of the obtained starches was highly satisfactory. However, the extraction yields were relatively low. In addition, the average size of the native granules varied with multiple and irregular shapes. Their values were closed to those of legume starches from other regions in the world. Theirs sizes were approximately twice as larger as that of maize and cassava starches. The legume starches analyzed were higher amylose contain. Temperatures of gelatinization were high. Stacking behavior analysis showed a low peak viscosity demonstrating the resistance of these starches to shear and swelling. The evaluation of the conservation of starch gels studied showed that these starches have a low retrogradation and an average syneresis. The study demonstrated the potential of local legume starches for outstanding successfully bioprocessing. These legume starches have approximately the same characteristics than modified starches.
Keywords: Common bean, Cowpea, Bambara groundnut, starch, physicochemical properties, pasting properties.
Selective protein kinase inhibitors were developed on the basis of the unexpected binding mode of 2,6,9-trisubstituted purines
to the adenosine triphosphate–binding site of the human cyclin-dependent kinase 2 (CDK2). By iterating chemical library synthesis
and biological screening, potent inhibitors of the human CDK2–cyclin A kinase complex and of Saccharomyces cerevisiae Cdc28p were identified. The structural basis for the binding affinity and selectivity was determined by analysis of a three-dimensional
crystal structure of a CDK2-inhibitor complex. The cellular effects of these compounds were characterized in mammalian cells
and yeast. In the latter case the effects were characterized on a genome-wide scale by monitoring changes in messenger RNA
levels in treated cells with high-density oligonucleotide probe arrays. Purine libraries could provide useful tools for analyzing
a variety of signaling and regulatory pathways and may lead to the development of new therapeutics.
The two greatest medical fears of the aging population are cancer and Alzheimer’s disease. Despite dramatic advances in understanding the molecular etiology of these disorders, therapeutic options for many patients with advanced disease have changed little and outcomes remain dismal. Paradoxically, recent findings suggest that some of the same molecules and biochemical processes underlying cancer may also participate in neurodegeneration. Therefore, it would be very useful to bring together experts from the fields of cancer research and neurodegeneration for discussions of the latest advances and ideas, with a particular emphasis on areas of overlap, to stimulate transdisciplinary interactions with the hope of accelerating progress. Cancer arises as a consequence of a breakdown in the genetic and epigenetic processes governing cell proliferation and cell death. Alterations in several classes of signaling molecules, both oncogenes and tumor suppressor genes, lead to uncontrolled cell growth. Over the past two decades, details of the intricate signaling pathways, from cell surface receptors through protein kinase cascades, transcription factors and modulators of chromatin, as well as the DNA damage response pathways linked to cell cycle control that guard the genome, have been uncovered. In some instances, key regulatory proteins have provided novel targets for development of small molecule inhibitors that are currently being tested in the clinic. The development of the nervous system relies on many of the signaling pathways and growth control processes that go awry in cancer. However, in mature neurons, the very same signaling proteins participate in transduction cascades linking short-term stimuli, elicited by synaptic stimulation, to long-term alterations in neuronal circuits through the regulation of gene expression and chromatin structure. These long-term adaptive modifications lead to changes in synaptic structure and function that contribute to learning and memory. The persistence of growth regulatory molecules in postmitotic neurons provides an opportunity for their contribution to pathophysiological processes resulting in neuronal loss. Recently, evidence has accumulated suggesting an association of cell cycle proteins and signal transduction proteins with neurodegeneration. Indeed, inhibitors of histone deacetylation have shown promise both as anti-cancer agents and in the prevention of neuronal loss
Site-specific proteases play critical roles in regulating many cellular processes. To identify novel site-specific proteases, their regulators, and substrates, we have designed a general reporter system in Saccharomyces cerevisiae in which a transcription factor is linked to the intracellular domain of a transmembrane protein by protease cleavage sites. Here, we explore the efficacy of this approach by using caspases, a family of aspartate-specific cysteine proteases, as a model. Introduction of an active caspase into cells that express a caspase-cleavable reporter results in the release of the transcription factor from the membrane and subsequent activation of a nuclear reporter. We show that known caspases activate the reporter, that an activator of caspase activity stimulates reporter activation in the presence of an otherwise inactive caspase, and that caspase inhibitors suppress caspase-dependent reporter activity. We also find that, although low or moderate levels of active caspase expression do not compromise yeast cell growth, higher level expression leads to lethality. We have exploited this observation to isolate clones from a Drosophila embryo cDNA library that block DCP-1 caspase-dependent yeast cell death. Among these clones, we identified the known cell death inhibitor DIAP1. We showed, by using bacterially synthesized proteins, that glutathione S-transferase–DIAP1 directly inhibits DCP-1 caspase activity but that it had minimal effect on the activity of a predomainless version of a second Drosophila caspase, drICE.
6,11,12,14-tetrahydroxy-abieta-5,8,11,13-tetraene-7-one (coleon U) is a diterpene compound isolated from Plectranthus grandidentatus with an antiproliferative effect on several human cancer cell lines. Herein, we studied the modulatory activity of coleon U on individual isoforms of the three protein kinase C (PKC) subfamilies, classical (cPKC-α and -βI), novel (nPKC-δ and -ɛ) and atypical (aPKC-ζ), using a yeast PKC assay. The results showed that, whereas the PKC activator phorbol-12-myristate-13-acetate (PMA) activated every PKC tested except aPKC, coleon U had no effect on aPKC and cPKCs. Besides, the effect of coleon U on nPKCs was higher than that of PMA. This revealed that coleon U was a potent and selective activator of nPKCs. The isoform-selectivity of coleon U for nPKC-δ and -ɛ was confirmed using an in vitro PKC assay. Most importantly, while PMA activated nPKCs inducing an isoform translocation from the cytosol to the plasma membrane and a G2/M cell cycle arrest, coleon U induced nPKCs translocation to the nucleus and a metacaspase- and mitochondrial-dependent apoptosis. This work therefore reconstitutes in yeast distinct subcellular translocations of a PKC isoform and the subsequent distinct cellular responses reported for mammalian cells. Together, our study identifies a new isoform-selective PKC activator with promising pharmacological applications. Indeed, since coleon U has no effect on cPKCs and aPKC, recognised as anti-apoptotic proteins, and selectively induces an apoptotic pathway dependent on nPKC-δ and -ɛ activation, it represents a promising compound for evaluation as an anti-cancer drug.
Expression of certain mammalian protein kinase C (PKC) isoforms inhibits the proliferation of Schizosaccharomyces pombe (Goode et al., Mol. Biol. Cell 5 (1994) 907–920). We have taken advantage of this fact to determine the in vivo isoform preference of a number of PKC inhibitors, using a microtitre plate assay which allows rapid screening. This in vivo model has revealed previously unreported preferences; calphostin C is a more efficient inhibitor of the novel PKCδ than chelerythrine chloride whereas the efficiencies are reversed for inhibition of the classical PKCγ. We have also shown that the anti-leukaemic agent bryostatin 1 inhibits or activates in vivo in an isoform-specific manner.
The effect of the xanthonolignoids trans-(±)-kielcorin C, cis-(±)-kielcorin C, trans-(±)-kielcorin D, trans-(±)-isokielcorin D and trans-(±)-kielcorin E on isoforms α, βI, δ, η and ζ of protein kinase C (PKC) was studied using the yeast phenotypic assay. All the compounds tested revealed an effect compatible with PKC inhibition, similar to that exhibited by the well established PKC inhibitor chelerythrine, and with differences in their potency towards the distinct isoforms tested, being, in general, potent inhibitors of the atypical PKC isoform (PKC-ζ). PKC inhibition caused by these kielcorins was confirmed using an in vitro kinase assay. The present study constitutes the first attempt to unravel the molecular mechanism of kielcorins activity, and shows that xanthonolignoids are a promising group of compounds to investigate for isoform selective PKC inhibitors.
The effect of the xanthonolignoids trans-(+/-)-kielcorin C, cis-(+/-)-kielcorin C, trans-(+/-)-kielcorin D, trans-(+/-)-isokielcorin D and trans-(+/-)-kielcorin E on isoforms alpha, betaI, delta, eta and zeta of protein kinase C (PKC) was studied using the yeast phenotypic assay. All the compounds tested revealed an effect compatible with PKC inhibition, similar to that exhibited by the well established PKC inhibitor chelerythrine, and with differences in their potency towards the distinct isoforms tested, being, in general, potent inhibitors of the atypical PKC isoform (PKC-zeta). PKC inhibition caused by these kielcorins was confirmed using an in vitro kinase assay. The present study constitutes the first attempt to unravel the molecular mechanism of kielcorins activity, and shows that xanthonolignoids are a promising group of compounds to investigate for isoform selective PKC inhibitors.
The cytoplasmic tail of the amyloid precursor protein (APP) contains two putatively cytotoxic peptides, Jcasp and C31, derived by caspase cleavage of APP. Jcasp is a fragment starting from the ε-secretase site to position 664, while C31 is a fragment from position 665 to the C-terminus. Our studies now showed that compared to C31, Jcasp appeared to play a minor role in cytotoxicity. In particular, inhibition of Jcasp generation by treatment of γ-secretase inhibitor did not lead to any attenuation of C31-induced toxicity. Secondly, because C31 toxicity is largely absent in cells lacking endogenous APP, we determined, using a split β-galactosidase complementary assay to monitor protein–protein interactions, the presence of APP associated complexes. Our results demonstrated that both APP homomeric and C31/APP heteromeric complexes were correlated with cell death, indicating that C31 complexes with APP to recruit the interacting partners that initiate the signals related to cellular toxicity.