Ernesto G. Occhiato

A new class of conjugated strigolactone analogues with fluorescent properties: synthesis and biological activity

A new class of strigolactone analogues has been synthesized. They differ from known molecules, both of natural and synthetic origin, in two main features. The conjugated system extends from the enol ether bridge to the A ring, the B ring is a heterocycle while the C ring is a cyclic ketone instead of a gamma-lactone. The key step of the synthesis is a Nazarov cyclization on activated substrates. Bioassays using Orobanche seeds have revealed that all the molecules strongly stimulate germination; in particular the oxygen containing analogues are the most active. Interestingly, some of the new molecules show fluorescent properties.

Selective non-steroidal inhibitors of 5α-reductase type 1

The enzyme 5α-reductase (5αR) catalyses the reduction of testosterone (T) into the more potent androgen dihydrotestosterone (DHT). The abnormal production of DHT is associated to pathologies of the main target organs of this hormone: the prostate and the skin. Benign prostatic hyperplasia (BPH), prostate cancer, acne, androgenetic alopecia in men, and hirsutism in women appear related to the DHT production. Two isozymes of 5α-reductase have been cloned, expressed and characterized (5αR-1 and 5αR-2). They share a poor homology, have different chromosomal localization, enzyme kinetic parameters, and tissue expression patterns. Since 5αR-1 and 5αR-2 are differently distributed in the androgen target organs, a different involvement of the two isozymes in the pathogenesis of prostate and skin disorders can be hypothesized. High interest has been paid to the synthesis of inhibitors of 5α-reductase for the treatment of DHT related pathologies, and the selective inhibition of any single isozyme represents a great challenge for medical and pharmaceutical research in order to have more specific drugs. At present, no 5αR-1 inhibitor is marketed for the treatment of 5αR-1 related pathologies but pharmaceutical research is very active in this field. This paper will review the major classes of 5αR inhibitors focusing in particular on non-steroidal inhibitors and on structural features that enhance the selectivity versus the type 1 isozyme. Biological tests to assess the inhibitory activity towards the two 5αR isozymes will be also discussed.

Low molecular weight, non-peptidic agonists of TrkA receptor with NGF-mimetic activity

Exploitation of the biologic activity of neurotrophins is desirable for medical purposes, but their protein nature intrinsically bears adverse pharmacokinetic properties. Here, we report synthesis and biologic characterization of a novel class of low molecular weight, non-peptidic compounds with NGF (nerve growth factor)-mimetic properties. MT2, a representative compound, bound to Trk (tropomyosin kinase receptor)A chain on NGF-sensitive cells, as well as in cell-free assays, at nanomolar concentrations and induced TrkA autophosphorylation and receptor-mediated internalization. MT2 binding involved at least two amino-acid residues within TrkA molecule. Like NGF, MT2 increased phosphorylation of extracellular signal-regulated kinase1/2 and Akt proteins and production of MKP-1 phosphatase (dual specificity phosphatase 1), modulated p38 mitogen-activated protein kinase activation, sustained survival of serum-starved PC12 or RDG cells, and promoted their differentiation. However, the intensity of such responses was heterogenous, as the ability of maintaining survival was equally possessed by NGF and MT2, whereas the induction of differentiation was expressed at definitely lower levels by the mimetic. Analysis of TrkA autophosphorylation patterns induced by MT2 revealed a strong tyrosine (Tyr)490 and a limited Tyr785 and Tyr674/675 activation, findings coherent with the observed functional divarication. Consistently, in an NGF-deprived rat hippocampal neuronal model of Alzheimer Disease, MT2 could correct the biochemical abnormalities and sustain cell survival. Thus, NGF mimetics may reveal interesting investigational tools in neurobiology, as well as promising drug candidates.

Structure–Function Relations of Strigolactone Analogs: Activity as Plant Hormones and Plant Interactions

Strigolactones (SLs) have several functions as signaling molecules in their interactions with symbiotic arbuscular mycorrhizal (AM) fungi and the parasitic weeds Orobanche and Striga. SLs are also a new class of plant hormone regulating plant development. In all three organisms, a specific and sensitive receptor-mediated perception system is suggested. By comparing the activity of synthetic SL analogs on Arabidopsis root-hair elongation, Orobanche aegyptiaca seed germination, and hyphal branching of the AM fungus Glomus intraradices, we found that each of the tested organisms differs in its response to the various examined synthetic SL analogs. Structure-function relations of the SL analogs suggest substitutions on the A-ring as the cause of this variation. Moreover, the description of competitive antagonistic analogs suggests that the A-ring of SL can affect not only affinity to the receptor, but also the molecules ability to activate it. The results support the conclusion that Arabidopsis, Orobanche, and AM fungi possess variations in receptor sensitivity to SL analogs, probably due to variation in SL receptors among the different species.

Lyophilised yeasts: easy-to-handle biocatalysts for stereoselective reduction of ketones

Abstract The use of lyophilised yeasts as biocatalysts for the reduction of carbonyl compounds has been studied. First, a comparison of the performances of fresh and lyophilised cells of seven yeasts was performed using ethyl acetoacetate and acetophenone as typical substrates. Lyophilised cells gave from low to high conversions but, with a few exceptions, always good enantioselectivity, and they were then employed for the reduction of structurally different carbonyl compounds. Highly enantioselective reduction of carbonyls was often achieved, even with aromatic ketones. Both enantiomers of the alcohols were, in most cases, obtained with high enantiomeric excess by simply choosing a suitable yeast.

Synthesis of Weinreb Amides via Pd-Catalyzed Aminocarbonylation of Heterocyclic-Derived Triflates

The direct transformation of lactam-, lactone-, and thiolactone-derived triflates into N-methoxy-N-methyl or morpholine Weinreb amides has been realized using Pd-catalyzed aminocarbonylation under CO atmospheric pressure and at room temperature. The carbonylative coupling can be efficiently carried out with 2% of catalyst in the presence of Xantphos as a ligand. The amides smoothly react with nucleophiles to afford acylated aza-, oxa-, and thio-heterocycles. The proposed methodology could be advantageously exploited for the synthesis of dienones in which one of the double bonds is embedded in a heterocyclic moiety, as useful substrates for Nazarov cyclization.

Microbial biotransformations in water/organic solvent system. Enantioselective reduction of aromatic β- and γ-nitroketones

Abstract The production of single enantiomers of γ- and β-nitroalcohols by microbial bioreduction has been studied. A restricted screening among 14 yeasts was performed using 1-phenyl-4-nitro-1-butanone 1 as substrate. Pichia minuta (CBS 1708) and Pichia etchellsii (CBS 2011) gave the highest enantiopreference for (S)-alcohol 3 formation (e.e. 85% and 80%, respectively), while Kluyveromyces marxianus (CBS 397) was the only strain able to preferentially furnish the (R) enantiomer (e.e. 70%). These three microorganisms, along with bakers yeast, were then employed in reactions performed in water/organic solvent systems using different solvents (hexane, benzene and dibutyl ether) affording alcohol 3 in high enantiomeric excesses (>95–97%). These strains were also employed to reduce 1-phenyl-3-nitro-1-propanone 2, maintaning the same stereobias observed with γ-nitroketone 1 and showing high enantioselectivity in both simple aqueous (>85–97%) and biphasic media (>97%).

Synthesis of a new enantiopure bicyclic γ/δ-amino acid (BTKa) derived from tartaric acid and α-amino acetophenone

Abstract The synthesis of a novel enantiopure γ/δ-amino acid having the 3-aza-6,8-dioxabicyclo[3.2.1]octane structure was realized by the combination of tartaric acid derivatives and α-amino acetophenone followed by a trans-acetalisation process. This amino acid, which has a rigid skeleton and carries substituents at the 3, 5 and 7 positions of the scaffold, could find different applications in organic and peptidomimetic synthesis. Two different synthetic strategies were studied, one of them allowing the multigram scale preparation of the amino acid.

Baker's yeast reduction of prochiral γ-nitroketones. II.1 straightforward enantioselective synthesis of 2,7-dimethyl-1,6-dioxaspiro[4.4]nonanes

Abstract The bakers yeast reduction of 5-nitro-2,8-nonanedione 2 afforded the corresponding (2 S ,8 S )-5-nitro-2,8-nonanediol 3 with complete diastereoselectivity and high enantioselectivity. The conversion of 3 into the thermodynamic (E,E) (Z,Z) (3:1) mixture of optically active (95% e.e.) 2,7-dimethyl-1,6-dioxaspiro[4.4]nonanes 5 was then achieved by spontaneous cyclization under the acidic conditions of the Nef reaction.

Strigolactone Analogs as Molecular Probes in Chasing the (SLs) Receptor/s: Design and Synthesis of Fluorescent Labeled Molecules

Originally identified as allelochemicals involved in plant-parasite interactions, more recently, Strigolactones (SLs) have been shown to play multiple key roles in the rhizosphere communication between plants and mycorrhizal fungi. Even more recent is the hormonal role ascribed to SLs which broadens the biological impact of these relatively simple molecules. In spite of the crucial and multifaceted biological role of SLs, there are no data on the receptor(s) which bind(s) such active molecules, neither in the producing plants nor in parasitic weeds or AM fungi. Information about the putative receptor of SLs can be gathered by means of structural, molecular, and genetic approaches. Our contribution on this topic is the design and synthesis of fluorescent labeled SL analogs to be used as probes for the detection in vivo of the receptor(s). Knowledge of the putative receptor structure will boost the research on analogs of the natural substrates as required for agricultural applications.