PPARγ transcription effect on naturally occurring O-prenyl cinnamaldehydes and cinnamyl alcohol derivatives.

Background: PPARγ is known to be a key regulator of metabolism and storage of lipids and glucose and to be implicated in the pathology of severe syndromes like obesity, diabetes, atherosclerosis and cancer. Methods: As a continuation of the authors' studies on oxyprenylated secondary metabolites as effective PPARγ agonists, the authors describe herein the chemical synthesis of natural O-prenyl cinnamaldehydes and cinnamyl alcohols and preliminary data on their in vitro effects on PPARγ transcription. Results: Among the panel of eight compounds tested, three - namely, (2E)-3-(4-((E)3,7-dimethylocta-2,6-dienyloxy)-3-methoxyphenyl)acrylaldehyde, (2E)-3-(4-((E)3,7-dimethylocta-2,6-dienyloxy)-3-methoxyphenyl)prop-2-en-1-ol and boropinal A - exerted activity in a dose-dependent manner. Conclusion:O-prenyl cinnamaldehydes and cinnamyl alcohols have the potential to effectively interact with PPARγ receptor.

Syntheses of Complex Terpenes from Simple Polyprenyl Precursors.

From structure elucidation and biogenesis to synthetic methodology and total synthesis, terpene natural products have profoundly influenced the development of organic chemistry. Moreover, their myriad functional attributes range from fragrance to pharmaceuticals and have had great societal impact. Ruzicka's formulation of the "biogenetic isoprene rule," a Nobel Prize winning discovery now over 80 years old, allowed for identification of higher order terpene (aka "isoprenoid") structures from simple five-carbon isoprene fragments. Notably, the isoprene rule still holds pedagogical value to students of organic chemistry today. Our laboratory has completed syntheses of over two dozen terpene and meroterpene structures to date, and the isoprene rule has served as a key pattern recognition tool for our synthetic planning purposes. At the strategic level, great opportunity exists in finding unique and synthetically simplifying ways to connect the formal C5 isoprene fragments embedded in terpenes. Biomimetic cationic polyene cyclizations represent the earliest incarnation of this idea, which has facilitated expedient routes to certain terpene polycycle classes. Nonetheless, a large swath of terpene chemical space remains inaccessible using this approach.In this Account, we describe strategic insight into our endeavors in terpene synthesis published over the last five years. We show how biosynthetic understanding, combined with a desire to utilize abundant and inexpensive [C5]n building blocks, has led to efficient, abiotic syntheses of multiple complex terpenes with disparate ring systems. Informed by nature, but unconstrained by its processes, our synthetic assembly exploits chemical reactivity across diverse reaction types-including radical, anionic, pericyclic, and metal-mediated transformations.First, we detail an eight-step synthesis of the cembrane diterpene chatancin from dihydrofarnesal using a bioinspired-but not -mimetic-cycloaddition. Next, we describe the assembly of the antimalarial cardamom peroxide using a polyoxygenation cascade to fuse multiple units of molecular oxygen onto a dimeric skeleton. This three-to-four-step synthesis arises from (-)-myrtenal, an inexpensive pinene oxidation product. We then show how a radical cyclization cascade can forge the hallmark cyclooctane ring system of the complex sesterterpene 6-epi-ophiobolin N from two simple polyprenyl precursors, (-)-linalool and farnesol. To access the related, more complex metabolite 6-epi-ophiobolin A, we exploited the plasticity of our synthetic route and found that use of geraniol (C10) rather than farnesol (C15) gave us the flexibility needed to address the additional oxidation found in this congener. Following this work, we describe two strategies to access several guaianolide sesquiterpenes. Retrosynthetic disconnection to monoterpenes, carvone or (-)-linalool, coupled with a powerful allylation strategy allowed us to address guaianolides with disparate stereochemical motifs. Finally, we examine a semisynthetic approach to the illicium sesquiterpenes from the abundant 15-carbon feedstock terpene (+)-cedrol using an abiotic ring shift and multiple C-H oxidation reactions inspired by a postulated biosynthesis of this natural product class.

A novel prenyl-polybasic domain code determines lipid-binding specificity of the K-Ras membrane anchor.

Ras proteins must localize to the plasma membrane (PM) for biological function. The membrane anchor of the K-Ras4B isoform comprises a farnesylated and methylated C-terminal cysteine together with an adjacent hexa-lysine polybasic domain (PBD). Traditionally, polybasic sequences have been thought to interact electrostatically with negatively charged membranes showing no specificity for anionic lipid head groups. By contrast we recently showed that the K-Ras membrane anchor actually exhibits a very high degree of specificity for phosphatidylserine (PtdSer). The selectivity for PtdSer is determined by a combinatorial code comprising the PBD sequence plus the prenyl anchor. Lipid binding specificity is therefore altered by PBD point mutations that in turn modulate signaling output. For example, mutating Lys177 or Lys178 to glutamine switches K-Ras4B lipid affinity from PtdSer to phosphoinositol 4,5-bisphosphate (PIP2). Changing the lipid anchor from farnesyl to geranylgeranyl or the PBD lysines to arginines also changes lipid binding specificity. All-atom molecular dynamics simulations reveal the structural basis for these K-Ras anchor lipid-binding preferences. Here we examine the PM interactions of a series of geranylgeranylated PBD mutants and provide further evidence that the precise PBD sequence and prenyl lipid determines lipid sorting specificity of the K-Ras anchor and hence biological function.

Lipid-Sorting Specificity Encoded in K-Ras Membrane Anchor Regulates Signal Output.

K-Ras is targeted to the plasma membrane by a C-terminal membrane anchor that comprises a farnesyl-cysteine-methyl-ester and a polybasic domain. We used quantitative spatial imaging and atomistic molecular dynamics simulations to examine molecular details of K-Ras plasma membrane binding. We found that the K-Ras anchor binds selected plasma membrane anionic lipids with defined head groups and lipid side chains. The precise amino acid sequence and prenyl group define a combinatorial code for lipid binding that extends beyond simple electrostatics; within this code lysine and arginine residues are non-equivalent and prenyl chain length modifies nascent polybasic domain lipid preferences. The code is realized by distinct dynamic tertiary structures of the anchor on the plasma membrane that govern amino acid side-chain-lipid interactions. An important consequence of this specificity is the ability of such anchors when aggregated to sort subsets of phospholipids into nanoclusters with defined lipid compositions that determine K-Ras signaling output.

Estimated exposure of hands inside the protective gloves used by non-occupational handlers of agricultural pesticides.

Exposure of handlers'/operators' hands is a main route of agricultural pesticides entry into their body. Non-occupational handlers still lack information about appropriate selection of protective gloves to minimize exposure and reduce adverse effects of these chemicals. According to the results of our previous survey, six commercially available, water-resistant gloves commonly used by non-professional gardeners were evaluated for permeation of Acetamiprid, Pirimicarb, and Chlorpyrifos-methyl (Chlorp-m) pesticides by means of in vitro testing. In-use conditions were mimicked as close as possible. Chlorp-m through latex was observed inside the glove from >10 to ⩽15 min; however, Acetamiprid and Pirimicarb through neoprene/latex and all the three pesticides through butyl were not observed inside gloves for the duration of the experiments (the Breakthrough time (BT)>8 h). The 1-h exposure proved the interior glove contamination with Chlorp-m through disposable latex, vinyl, and nitrile gloves (51, 33, and 41% of applied dose (AD), respectively) just as with Acetamiprid and Pirimicarb through latex glove (11 and 14%AD, respectively). However, when storing the used gloves for 4 days after the exposure, no release of the three pesticides from the butyl and Acetamiprid from neoprene/latex gloves was detected. In all other cases, pesticides were found in the interior glove (36-79, 31-63, and 51-81%AD for Acetamiprid, Pirimicarb, and Chlorp-m, respectively). If used repeatedly, gloves contaminated in this way lose their protective function but give the user a false sense of security. The results suggest that (i) water-resistant gloves are not necessarily pesticide resistant; (ii) disposable latex gloves commonly worn by non-professional gardeners provide inadequate protection even for a short-time contact with pesticides; (iii) to assess the efficiency of reusable gloves, not only BT value but also the reservoir/release effect of parent pesticide and its degradation products should be evaluated; and (iv) awareness-raising activities for non-occupational handlers of pesticides should be enhanced.

Gas-Phase Fragmentation Behavior of Oxidized Prenyl Peptides by CID and ETD Tandem Mass Spectrometry.

Farnesylation and geranylgeranylation are the two types of prenyl modification of proteins. Prenylated peptides are highly hydrophobic and their abundances in biological samples are low. In this report, we studied the oxidized prenylated peptides by electrospray ionization mass spectrometry and identified them by collision-induced dissociation (CID) and electron-transfer dissociation (ETD) tandem mass spectrometry. Modified prenyl peptides were generated utilizing strong and low strength oxidizing agents to selectively oxidize and epoxidize cysteine sulfur and prenyl side chain. We selected three peptides with prenyl motifs and synthesized their prenylated versions. The detailed characteristic fragmentations of oxidized and epoxidized farnesylated and geranylgeranylated peptides were studied side by side with two popular fragmentation techniques. CID and ETD mass spectrometry clearly distinguished the modified version of these peptides. ETD mass spectrometry provided sequence information of the highly labile modified prenyl peptides and showed different characteristic fragmentations compared with CID. A detailed fragmentation of modified geranylgeranylated peptides was compared by CID and ETD mass spectrometry for the first time. Graphical Abstract ᅟ.

A cis-prenyltransferase from Methanosarcina acetivorans catalyzes both head-to-tail and nonhead-to-tail prenyl condensation.

Cis-prenyltransferase usually consecutively catalyzes the head-to-tail condensation reactions of isopentenyl diphosphate to allylic prenyl diphosphate in the production of (E,Z-mixed) polyprenyl diphosphate, which is the precursor of glycosyl carrier lipids. Some recently discovered homologs of the enzyme, however, catalyze the nonhead-to-tail condensation reactions between allylic prenyl diphosphates. In this study, we characterize a cis-prenyltransferase homolog from a methanogenic archaeon, Methanosarcina acetivorans, to obtain information on the biosynthesis of the glycosyl carrier lipids within it. This enzyme catalyzes both head-to-tail and nonhead-to-tail condensation reactions. The kinetic analysis shows that the main reaction of the enzyme is consecutive head-to-tail prenyl condensation reactions yielding polyprenyl diphosphates, while the chain lengths of the major products seem shorter than expected for the precursor of glycosyl carrier lipids. On the other hand, a subsidiary reaction of the enzyme, i.e., nonhead-to-tail condensation between dimethylallyl diphosphate and farnesyl diphosphate, gives a novel diterpenoid compound, geranyllavandulyl diphosphate.

Prenyl Ammonium Salts--New Carriers for Gene Delivery: A B16-F10 Mouse Melanoma Model.

PURPOSE: Prenyl ammonium iodides (Amino-Prenols, APs), semi-synthetic polyprenol derivatives were studied as prospective novel gene transfer agents. METHODS: AP-7, -8, -11 and -15 (aminoprenols composed of 7, 8, 11 or 15 isoprene units, respectively) were examined for their capacity to form complexes with pDNA, for cytotoxicity and ability to transfect genes to cells. RESULTS: All the carriers were able to complex DNA. The highest, comparable to commercial reagents, transfection efficiency was observed for AP-15. Simultaneously, AP-15 exhibited the lowest negative impact on cell viability and proliferation--considerably lower than that of commercial agents. AP-15/DOPE complexes were also efficient to introduce pDNA to cells, without much effect on cell viability. Transfection with AP-15/DOPE complexes influenced the expression of a very few among 44 tested genes involved in cellular lipid metabolism. Furthermore, complexes containing AP-15 and therapeutic plasmid, encoding the TIMP metallopeptidase inhibitor 2 (TIMP2), introduced the TIMP2 gene with high efficiency to B16-F10 melanoma cells but not to B16-F10 melanoma tumors in C57BL/6 mice, as confirmed by TIMP2 protein level determination. CONCLUSION: Obtained results indicate that APs have a potential as non-viral vectors for cell transfection.

Cytotoxic tetraprenylated alkaloids from the South China Sea gorgonian Euplexaura robusta.

Nine achiral tetraprenylated alkaloids, including three new compounds, named malonganenones I-K (1-3, resp.), together with six known analogs, 4-9, were isolated from the gorgonian Euplexaura robusta collected from Weizhou Island of Guangxi Province, China. The structures of compounds 1-3 were elucidated by extensive spectral analyses, especially of their 1D- and 2D-NMR data. Compounds 1, 4, 6, and 7 showed moderate cytotoxicities against K562 and HeLa tumor cell lines with IC(50) values ranging from 0.35 to 10.82 µM. Compound 6 also showed moderate inhibitory activity against c-Met kinase at a concentration of 10 µM.

New cationic polyprenyl derivative proposed as a lipofecting agent.

Cationic linear poly-cis-isoprenoid prepared from natural plant polyprenol in a mixture with dioleyl phosphatidylethanolamine was found to be an effective lipofection agent for eukaryotic cells. The transfecting activity is related to the poly-cis structure of the polyprenyl chain.

Structure-activity relationship of prenyl-substituted polyphenols from Artocarpus heterophyllus as inhibitors of melanin biosynthesis in cultured melanoma cells.

A series of prenylated, flavone-based polyphenols, compounds 1-8, were isolated from the wood of Artocarpus heterophyllus. These compounds, which have previously been shown not to inhibit tyrosinase activity, were found to be active inhibitors of the in vivo melanin biosynthesis in B16 melanoma cells, with little or no cytotoxicity. To clarify the structural requirement for inhibition, some structure-activity relationships were studied, in comparison with related compounds lacking prenyl side chains. Our experiments indicate that both prenyl and OH groups, as well as the type of substitution pattern, are crucial for the inhibition of melanin production in B16 melanoma cells.

Synthesis of polyprenylated acylphloroglucinols using bridgehead lithiation: the total synthesis of racemic clusianone and a formal synthesis of racemic garsubellin A.

The synthesis of polyprenylated phloroglucinol natural products, including clusianone, nemorosone, and garsubellin A, was pursued by a strategy involving construction of a core bicyclo[3.3.1]nonanetrione structure and subsequent elaboration via organolithium intermediates. Appropriate bridged core structures were obtained through the cyclization of a suitably substituted cyclohexanone enol ether or enol silane with malonyl dichloride. Additional substituents were then introduced by means of regioselective lithiation reactions, including the generation of bridgehead enolates, thus enabling the total synthesis of clusianone and also of an advanced intermediate toward nemorosone. In the case of garsubellin A, an additional THF-like ring was elaborated by a biomimetic 5-exo-tet cyclization of an enol ether (or enol) with a side-chain epoxide. This enabled a formal synthesis of racemic garsubellin A by accessing one of the late intermediates in the Danishefsky synthesis.

Chemical constituents of malagasy liverworts, part V: prenyl bibenzyls and clerodane diterpenoids with nitric oxide inhibitory activity from Radula appressa and Thysananthus spathulistipus.

3Beta,4beta:15,16-diepoxy-13(16),14-clerodadiene (1) and a new clerodane diterpenoid designated thysaspathone (2) were isolated from the liverwort Thysananthus spathulistipus, while Radula appressa produced radulannin A (3), radulannin L (4), 2-geranyl-3,5-dihydroxybibenzyl (5), 2(S)-2-methyl-2-(4-methyl-3-pentenyl)-7-hydroxy-5-(2-phenylethyl) chromene (o-cannabichromene) (6), 6-hydroxy-4-(2-phenylethyl) benzofuran (7), and o-cannabicyclol (8). All of the isolated compounds inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells and the greatest inhibition was attributed to compound 5, with an IC50 value of 4.5 microM.

Permeability of 13 different gloves to 13 cytotoxic agents under controlled dynamic conditions.

PURPOSE: The permeability of 13 different gloves to 13 cytotoxic agents under controlled dynamic conditions is described. METHODS: Thirteen cytotoxic agents were prepared at the highest concentrations normally encountered by pharmacy personnel. Four glove types--neoprene, natural rubber latex, nitrile, and vinyl--were exposed to the cytotoxic agents for 15, 30, and 60 minutes. Tests were conducted using the middle finger of each glove. Linearity, reproducibility, and sensitivity were evaluated for each drug tested. Assays were run using liquid chromatographic tandem mass spectrometry (LC/MS/MS) and high-performance liquid chromatography with ultraviolet light (HPLC-UV). Permeability testing was conducted using an original system designed to evaluate dynamic constraints, such as rubbing, stretching, and tension. RESULTS: Linearity by LC/MS/MS and HPLC-UV was confirmed at concentrations up to 1000 ng/mL for all drugs. Most glove materials were permeable at rates below ASTM recommendations over the one-hour testing period. Vinyl was the most permeable material. Carmustine permeated the widest variety of materials. Due to the high sensitivity of the analytic methods, all materials displayed low but significant permeability for at least one drug after one hour. Higher resistance to permeation was recorded for all neoprene, some natural rubber latex, and one nitrile glove. CONCLUSION: Neoprene, natural rubber latex, and nitrile gloves displayed the highest resistance to permeation of the 13 cytotoxic agents studied. Additional factors, such as duration of exposure, glove thickness, and drug liposolubility and molecular weight, also affected permeability.

Artelastocarpin and carpelastofuran, two new flavones, and cytotoxicities of prenyl flavonoids from Artocarpus elasticus against three cancer cell lines.

Further study of one of the fractions from the wood of Artocarpus elasticus furnished two new prenylated flavonoids artelastocarpin and carpelastofuran as well as ethyl 2,4-dihydroxybenzoate. The two flavonoids and the prenylated flavonoids artelastin, artelastochromene, artelasticin, artocarpesin, and cyclocommunin isolated earlier from this species were tested for cytotoxicity in vitro against three human cell lines. All seven flavonoids were active, the cytotoxic effect varying from strong to moderate and with artelastin showing the most potent activity.

Characterization of hydrophobic prenyl groups of isoprenylated proteins in human cancer cells.

Extensive protease digestion of delipidated [3H]mevalonate (MVA)-labeled proteins, followed by HPLC separation of the products, is one approach to identify and study prenyl cysteines. Using this methodology three major [3H]MVA-labeled peaks appeared. Two of them represent farnesyl cysteine (FC) and geranylgeranyl cysteine (GGC). The third peak represents unknown products that are considerably more hydrophobic than FC and GGC, here designated HPC. Previously, we provided evidence that cysteine residues may also be modified by dolichyl groups. Dolichyl cysteines (DolC) belong to HPC. However, as shown in the present study, DolC only represents a minor portion of HPC. Data obtained from different sets of experiments, including [3H]GGOH-labeling and use of prenyl transferase inhibitors, suggest that HPC mainly involves CXC or CC residues with double-linked GG groups. In turn this points to the possibility that proteins modified by double GG groups are quite common, and may probably involve other proteins than the rab family of GTPases.

Exposures to asbestos arising from bandsawing gasket material.

A simulation of bandsawing sheet asbestos gasket material was performed as part of a retrospective exposure evaluation undertaken to assist in determining causation of a case of mesothelioma. The work was performed by bandsawing a chrysotile asbestos (80%)/neoprene gasket sheet with a conventional 16-inch woodworking bandsaw inside a chamber. Measurements of airborne asbestos were made using conventional area and personal sampling methods, with analysis of collected samples by transmission electron microscopy (TEM) and phase contrast microscopy (PCM). These were supplemented by qualitative scanning electron microscopy (SEM) examinations of some of the airborne particles collected on the filters. In contrast with findings from studies examining manual handling (installation and removal) of gaskets, airborne asbestos concentrations from this operation were found to be well above current Occupational Safety and Health Administration (OSHA) permissible exposure limit (PEL) (eight-hour time-weighted average [TWA]) and excursion limit (30-minute) standards. Although some "encapsulation" effect of the neoprene matrix was seen on the particles in the airborne dust, unencapsulated individual fiber bundles were also seen. Suggestions for the implications of the work are given. In summary, the airborne asbestos concentrations arising from this work were quite high, and point to the need for careful observation of common sense precautions when manipulation of asbestos-containing materials (even those believed to have limited emissions potential) may involved machining operations.