Redox ranking of inducers of a cancer-protective enzyme via the energy of their highest occupied molecular orbital

Induction of phase 2 enzymes is a major strategy in chemoprotection against cancer. Inducers belong to nine different chemical classes. In this study we found that a measure of the tendency of 30 plant phenylpropenoids and synthetic analogs to release electrons correlates linearly with their potency in inducing the activity of NAD(P)H:quinone reductase (NQO1), a prototypic phase 2 cancer-protective enzyme. The tendency to release electrons was determined by the energy of the highest occupied molecular orbital (E(HOMO)), calculated by simple quantum-mechanical methods. The correlations observed establish a clear conclusion: the smaller the absolute E(HOMO) of an agent, A, i.e., the lower its reduction potential, E(A*+/A), the stronger is its electron donor property and the greater its inducer potency. The finding of this redox ranking of the inducers demonstrates the possibility of controlling and predicting the genetic expression of an enzymatic defense against cancer by xenobiotics via one physicochemical parameter, the reduction potential, E(A*+/A).     

Celastrol analogues as inducers of the heat shock response. Design and synthesis of affinity probes for the identification of protein targets

The natural product celastrol (1) possesses numerous beneficial therapeutic properties and affects numerous cellular pathways. The mechanism of action and cellular target(s) of celastrol, however, remain unresolved. While a number of studies have proposed that the activity of celastrol is mediated through reaction with cysteine residues, these observations have been based on studies with specific proteins or by in vitro analysis of a small fraction of the proteome. In this study, we have investigated the spatial and structural requirements of celastrol for the design of suitable affinity probes to identify cellular binding partners of celastrol. Although celastrol has several potential sites for modification, some of these were not synthetically amenable or yielded unstable analogues. Conversion of the carboxylic acid functionality to amides and long-chain analogues, however, yielded bioactive compounds that induced the heat shock response (HSR) and antioxidant response and inhibited Hsp90 activity. This led to the synthesis of biotinylated celastrols (23 and 24) that were used as affinity reagents in extracts of human Panc-1 cells to identify Annexin II, eEF1A, and β-tubulin as potential targets of celastrol.     

Synthesis of hydroxy derivatives of highly potent non-steroidal CYP 17 inhibitors as potential metabolites and evaluation of their activity by a non cellular assay using recombinant human enzyme.

Inhibition of CYP 17 is a promising strategy for the treatment of prostate cancer. Recently two non-steroidal compounds with high in vitro activity were synthesized in our group (BW19 and BW95). However, after a few hours they showed in vivo a strong decrease in their activity. This might be due to a fast biodegradation. Potential hydroxy and epoxy metabolites were synthesized and their inhibitory activities were tested by a new non-cellular assay using recombinant enzyme. As source, membrane fractions of E. coli pJL17/OR coexpressing human CYP 17 and rat NADPH-P450-reductase were, used. Showing a high and constant CYP 17 activity and a fast and easy isolation procedure the new method was advantageous compared with the microsomal assay. Interestingly, all the new synthesized hydroxy and epoxy compounds except one showed a lower inhibition of CYP 17 than the parent compounds. Thus, the loss of in vivo activity may be partly explained.     

Phosphonic and arsonic acids as inhibitors of human red cell acid phosphatase and their use in affinity chromatography.

1. In order to obtain an effective ligand for affinity chromatography of the low molecular weight acid phosphatase (orthophosphoric-monoester phosphohydrolase (acid optimum), EC 3.1.3.2) from human red cells nine phosphonic and two arsonic acid substrate analogues were investigated as potential inhibitors. The two forms of acid phosphatase type B (b1 and b2) were isolated and partially purified using conventional methods and the inhibitory action of the substrate analogs investigated. 2. Four of the phosphonic acids were relatively effective competitive inhibitors. It appears that certain structural and electronic requirements have to be fulfilled by the phosphonic acids in order to exhibit significant affinity for the enzyme. A high affinity appears to require the presence of a bulky, hydrophobic moiety which has to be separated from the phosphorus atom by the distance of one atom. 3. p-Aminobenzylphosphonic acid exerted the highest affinity for acid phosphatase with a pH optimum at 6.5. Ki values of 4 . 10(-4) and 6 . 10(-4) M were found for the b1 and b2 forms, respectively. 4. Coupling of p-aminobenzylphosphonic acid to Agarose yielded an effective and specific affinity medium. By means of affinity chromatography using this medium, acid phosphatase was purified 500-fold in a single step.     

Synthesis of a terphenyl substituted 4-aza-2,3-didehydropodophyllotoxin analogues as inhibitors of tubulin polymerization and apoptosis inducers

A series of terphenyl based 4-aza-2,3-didehydropodophyllotoxin conjugates (8a–r) were synthesized by a straightforward one-step multicomponent synthesis that demonstrated anticancer activity against five human cancer cell lines (lung, colon, renal, prostate and cervical). All the tested compounds showed potent anticancer activity with IC₅₀ values ranging from 0.87 to 16.59 μM. Among them compounds 8n and 8p showed significant anticancer activity in lung cancer cells with IC₅₀ values 0.91 and 0.87 μM, respectively. Flow cytometric analysis revealed that these compounds induced cell cycle arrest in G₂/M phase in A549 cell line leading to caspase-3 dependent apoptotic cell death. The tubulin polymerization assay and immunofluorescence analysis showed that these compounds effectively inhibit microtubule assembly at both molecular and cellular levels in A549 cells. Further, Hoechst staining, DNA fragmentation analysis also suggested that these compounds induced cell death by apoptosis. Overall, the current study demonstrated that the synthesis of terphenyl based 4-aza-2,3-didehydropodophyllotoxin conjugates as promising anticancer agents with G₂/M cell cycle arrest and apoptotic-inducing activities via targeting tubulin.     

Aminoacyl-tRNA synthetases and their inhibitors as a novel family of antibiotics

The emergence of multidrug-resistant strains of pathogenic microorganisms and the slow progress in new antibiotic development has led in recent years to a resurgence of infectious diseases that threaten the well-being of humans. The result of many microorganisms becoming immune to major antibiotics means that fighting off infection by these pathogens is more difficult. The best strategy to get around drug resistance is to discover new drug targets, taking advantage of the abundant information that was recently obtained from genomic and proteomic research, and explore them for drug development. In this regard, aminoacyl-tRNA synthetases (ARSs) provide a promising platform to develop novel antibiotics that show no cross-resistance to other classical antibiotics. During the last few years there has been a comprehensive attempt to find the compounds that can specifically target ARSs and inhibit bacterial growth. In this review, the current status in the development of ARS inhibitors will be briefly summarized, based on their chemical structures and working mechanisms.     

Design,synthesis, molecular modeling and biological evaluation of novel Benzoxazole-Benzamide conjugates via a 2-Thioacetamido linker as potential anti-proliferative agents,VEGFR-2 inhibitors and apoptotic inducers

A novel series of 2-thioacetamide linked benzoxazole-benzamide conjugates 1–15 was designed as potential inhibitors of the vascular endothelial growth factor receptor-2 (VEGFR-2). The prepared compounds were evaluated for their potential antitumor activity and their corresponding selective cytotoxicity was estimated using normal human fibroblast (WI-38) cells. Compounds 1, 9–12 and 15 showed good selectivity and displayed excellent cytotoxic activity against both HCT-116 and MCF-7 cancer cell lines compared to sorafenib, used as a reference compound. Furthermore, compounds 1 and 11 showed potent VEGFR-2 inhibitory activity. The cell cycle progression assay showed that 1 and 11 induced cell cycle arrest at G2/M phase, with a concomitant increase in the pre-G1 cell population. Further pharmacological studies showed that 1 and 11 induced apoptosis and inhibited the expression of the anti-apoptotic Bcl-2 and Bcl-xL proteins in both cell lines. Therefore, compounds 1 and 11 might serve as promising candidates for future anticancer therapy development.     

Perindopril and ramipril phosphonate analogues as a new class of angiotensin converting enzyme inhibitors

A series of phosphonate analogues related to perindopril and ramipril were prepared and tested to estimate their ability to inhibit angiotensin converting enzyme. These new synthesized compounds were active ACE inhibitors with a promising activity.     

Oxathiolene oxide synthesis via chelation-controlled addition of organometallic reagents to alkynols followed by addition of sulfur electrophiles and evaluation of oxathiolene oxides as anticarcinogenic enzyme inducers

A number of alkynols have been prepared by Sonogashira coupling of propargyl alcohol to aromatic halides. Chelation-controlled addition of organometallic nucleophiles to these alkynols was then effected followed by the addition of the sulfur electrophiles, sulfur dioxide or thionyl chloride. This methodology was used to prepare a number of oxathiolene oxides, which have been screened as NQO1 (quinone oxidoreductase) inducers.     

Synthesis of angiotensin converting enzyme inhibitors containing residues of substituted quinolines and a study of their biological activity]

Inhibitors of the angiotensin-converting enzyme were synthesized by substituting N-and C-terminal amino acid residues of tripeptide Bz-Phe-Ala-Pro by the residues of 8-methoxy-5-sulphoquinoline and carboxy-1,2,3,4-tetrahydroquinoline, respectively, and their in vivo and in vitro biological activity was determined. The enzyme's S2' site proved to be non specific to the position of the carboxylic group in the C-terminal heterocyclic part of the inhibitor molecule. Introducing a modified quinoline residue into the N-terminal part of the inhibitor does not increase its specific interaction with the hydrophobic pocket of the angiotensin-converting enzyme.     

Discovery of inhibitors of Pseudomonas aeruginosa virulence through the search for natural-like compounds with a dual role as inducers and substrates of efflux pumps

Multidrug efflux pumps are ancient elements encoded in every genome, from bacteria to humans. In bacteria, in addition to antibiotics, efflux pumps extrude a wide range of substrates, including quorum sensing signals, bacterial metabolites, or plant-produced compounds. This indicates that their original functions may differ from their recently acquired role in the extrusion of antibiotics during human infection. Concerning plant-produced compounds, some of them are substrates and inducers of the same efflux pump, suggesting a coordinated plant/bacteria coevolution. Herein we analyse the ability of 1243 compounds from a Natural Product-Like library to induce the expression of P. aeruginosa mexCD-oprJ or mexAB-oprM efflux pumps' encoding genes. We further characterized natural-like compounds that do not trigger antibiotic resistance in P. aeruginosa and that act as virulence inhibitors, choosing those that were not only inducers but substrates of the same efflux pump. Four compounds impair swarming motility, exotoxin secretion through the Type 3 Secretion System (T3SS) and the ability to kill Caenorhabditis elegans, which might be explained by the downregulation of genes encoding flagellum and T3SS. Our results emphasize the possibility of discovering new anti-virulence drugs by screening natural or natural-like libraries for compounds that behave as both, inducers and substrates of efflux pumps.     

Interaction between lymphocytes and inflammatory exudate cells. II. A proteolytic enzyme released by PMN as a possible mediator for enhancement of thymocyte response.

Mouse polymorphonuclear leukocytes (PMN) were harvested from the peritoneal cavity stimulated with sodium caseinate. The cells were cultivated in vitro and the supernatant of these cultures (SUP) was tested for enhancing potency on DNA synthesis by syngeneic thymocytes stimulated with phytohemagglutinin (PHA). The enhancing potency of the SUP was markedly influenced by duration of the donor cultures of PMN, population density of the cultures, and protein concentrations in the medium, respectively. The enhancing factor in the SUP was found to be non-dialysable, heat-labile, and stable in the pH ranging between 3 and 9 but labile in the pH below 2 or above 10; its m.w. was approximately 19.000 when measured by gel-filtration on Sephadex G-75. The factor had a proteolytic activity on 3H-acetyl hemoglobin (3HHb) at neutral pH (7.2). Both the proteolytic activity and thymocyte-helping potency of the SUP were similarly abolished by adding protease inhibitor (Trasylol) in soluble form, or by passing through a column of the inhibitor insolubilized. It was thus assumed that the enhancing effect of PMN on thymocyte response was associated with a neutral protease released from the cells: it may be termed a lymphocyte-helping protease (LHP).     

Synthesis and evaluation of 1H-pyrrole-2,5-dione derivatives as cholesterol absorption inhibitors for suppressing the formation of foam cells and inflammatory response

Excess lipid accumulation in the arterial intima and formation of macrophage-derived foam cells in the plaque could cause atherosclerotic lesion. Cholesterol absorption inhibitors could suppress the lipid accumulation of human macrophage, inflammatory response and the development of atherosclerosis. In this research, a series of 1H-pyrrole-2,5-dione derivatives were synthesized as cholesterol absorption inhibitor and tested in in vitro experiments. One of the most active inhibitors, compound 20 exhibited stronger in vitro cholesterol absorption activity than ezetimibe, no cytotoxicity in HEK293 and RAW264.7 cell lines and satisfied lipophilicity. The further study indicated that 20 could inhibit lipid accumulation of macrophage and reduce the secretion of LDH, MDA, TNF-α and ROS in a concentration-dependent manner. In conclusion, as a novel and potent cholesterol absorption inhibitor, compound 20 could suppress the formation of foam cells and inflammatory response.     

Synthetic staurosporines via a ring closing metathesis strategy as potent JAK3 inhibitors and modulators of allergic responses

The synthesis and biological evaluation of JAK3 based staurosporine compounds is described. The compounds are constructed completely de novo, and a ring closing metathesis strategy is used to assemble the sugar mimetic portion. These analogs show potent JAK3 activity against isolated enzyme and in T-cells. One analog (32) showed unique biological effects during in vitro and in vivo tests including inhibition of STAT5 phosphorylation, blockade of mast cell responses, and reduction of JAK3 based effects in mice models of allergic disease.     

Synthesis of 6- or 4-functionalized indoles via a reductive cyclization approach and evaluation as aromatase inhibitors

Two new series of benzonitrile derivatives on position 6 or 4 of indole ring were successfully synthesized via a Leimgruber-Batcho reaction. All the compounds were evaluated in vitro on the inhibition of aromatase (CYP19) and 17alpha-hydroxylase-C17,20-lyase (CYP17). The racemate, 4-[(1H-imidazol-1-yl)(1H-indol-4-yl)methyl]benzonitrile 9, showed high level of inhibitory activity towards CYP19 (IC(50)=11.5 nM).