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Journal of Pharmaceutical and... Oct 2023Enrichment of pharmaceutically important vinca alkaloids, vinblastine and vincristine, in the leaves of Madagascar periwinkle (Catharanthus roseus) plants through...
UPLC-ESI-QTOF-MS assisted targeted metabolomics to study the enrichment of vinca alkaloids and related metabolites in Catharanthus roseus plants grown under controlled LED environment.
Enrichment of pharmaceutically important vinca alkaloids, vinblastine and vincristine, in the leaves of Madagascar periwinkle (Catharanthus roseus) plants through different pre- or postharvest treatments or cultivation conditions, e.g., exposing the plants to UV-irradiation, has been in focus for decades. Controlled LED environment in the visible light range offers the possibility of monitoring the changes in the concentration of metabolites in the vinca alkaloid-related pathway without involving UV-related abiotic stress. In the frame of our targeted metabolomics approach, 64 vinca alkaloids and metabolites were screened with the help of a UPLC-ESI-QTOF-MS instrumental setup from the leaf extracts of C. roseus plants grown in chambers under control (medium light), low light, and high blue / high red/ high far-red conditions. Out of the 14 metabolites that could be assigned either unambiguously with authentic standards or tentatively with high resolution mass spectrometry-based methods, all three dimer vinca alkaloids, that is, 3',4'-anhydrovinblastine, vinblastine and vincristine showed an at least nine-fold enrichment under high blue irradiation when compared with the control conditions: final concentrations of 961 mg kg dry weight, 33.8 mg kg dry weight, and 11.7 mg kg dry weight could be achieved, respectively. As supported by multivariate statistical analysis, the key metabolites of the vinca alkaloid pathway were highly represented among the metabolites that were specifically stimulated by high blue light application.
Topics: Vinca Alkaloids; Vinblastine; Catharanthus; Vincristine; Antineoplastic Agents; Metabolomics
PubMed: 37542828
DOI: 10.1016/j.jpba.2023.115611 -
Nature Chemical Biology Aug 2023Advances in omics technologies now permit the generation of highly contiguous genome assemblies, detection of transcripts and metabolites at the level of single cells...
Advances in omics technologies now permit the generation of highly contiguous genome assemblies, detection of transcripts and metabolites at the level of single cells and high-resolution determination of gene regulatory features. Here, using a complementary, multi-omics approach, we interrogated the monoterpene indole alkaloid (MIA) biosynthetic pathway in Catharanthus roseus, a source of leading anticancer drugs. We identified clusters of genes involved in MIA biosynthesis on the eight C. roseus chromosomes and extensive gene duplication of MIA pathway genes. Clustering was not limited to the linear genome, and through chromatin interaction data, MIA pathway genes were present within the same topologically associated domain, permitting the identification of a secologanin transporter. Single-cell RNA-sequencing revealed sequential cell-type-specific partitioning of the leaf MIA biosynthetic pathway that, when coupled with a single-cell metabolomics approach, permitted the identification of a reductase that yields the bis-indole alkaloid anhydrovinblastine. We also revealed cell-type-specific expression in the root MIA pathway.
Topics: Catharanthus; Plants, Medicinal; Multiomics; Indole Alkaloids; Antineoplastic Agents; Monoterpenes; Gene Expression Regulation, Plant; Plant Proteins
PubMed: 37188960
DOI: 10.1038/s41589-023-01327-0 -
Metabolic Engineering Communications Jun 2023With over 3000 reported structures, monoterpenoid indole alkaloids (MIAs) constitute one of the largest alkaloid groups in nature, including the clinically important...
With over 3000 reported structures, monoterpenoid indole alkaloids (MIAs) constitute one of the largest alkaloid groups in nature, including the clinically important anticancer drug vinblastine and its semi-synthetic derivatives from (Madagascar's periwinkle). With the elucidation of the complete 28-step biosynthesis for anhydrovinblastine, it is possible to investigate the heterologous production of vinblastine and other medicinal MIAs. In this study, we successfully expressed the flavoenzyme -acetylstemmadenine oxidase in (baker's yeast) by signal peptide modification, which is a vinblastine biosynthetic gene that has not been functionally expressed in this system. We also reported the simultaneous integration of ∼18 kb MIA biosynthetic gene cassettes as single copies into four genomic loci of baker's yeast by CRISPR-Cas9, which enabled the biosynthesis of vinblastine precursors catharanthine and tabersonine from the feedstocks secologanin and tryptamine. We further demonstrated the biosynthesis of fluorinated and hydroxylated catharanthine and tabersonine derivatives using our yeasts, which showed that the MIA biosynthesis accommodates unnatural substrates, and the system can be further explored to produce other complex MIAs.
PubMed: 36569379
DOI: 10.1016/j.mec.2022.e00215 -
Journal of the American Chemical Society Sep 2019A new triarylaminium radical cation promoted coupling of catharanthine with vindoline is disclosed, enlisting tris(4-bromophenyl)aminium hexachlororantimonate (BAHA, 1.1...
A new triarylaminium radical cation promoted coupling of catharanthine with vindoline is disclosed, enlisting tris(4-bromophenyl)aminium hexachlororantimonate (BAHA, 1.1 equiv) in aqueous 0.05 N HCl/trifluoroethanol (1-10:1) at room temperature (25 °C), that provides anhydrovinblastine in superb yield (85%) with complete control of the newly formed quaternary C16' stereochemistry. A definition of the scope of aromatic substrates that participate with catharanthine in the BAHA-mediated diastereoselective coupling reaction and simplified indole substrates other than catharanthine that participate in the reaction are disclosed that identify the key structural features required for participation in the reaction, providing a generalized indole functionalization reaction that bears little structural relationship to catharanthine or vindoline.
Topics: Amines; Cations; Free Radicals; Molecular Structure; Stereoisomerism; Vinblastine; Vinca Alkaloids
PubMed: 31442047
DOI: 10.1021/jacs.9b06968 -
Frontiers in Plant Science 2019contains a variety of monoterpenoid indole alkaloids (MIAs), among which bisindole alkaloids vinblastine and vincristine are well-known to have antitumor effects and...
contains a variety of monoterpenoid indole alkaloids (MIAs), among which bisindole alkaloids vinblastine and vincristine are well-known to have antitumor effects and widely used in clinical treatment. However, their contents in is extremely low and difficult to meet market demands. Therefore, it is of great significance to study the transcriptional regulation mechanism of MIAs biosynthesis for high yielding of bisindole alkaloids in . Studies have shown that MIAs biosynthesis in has complex temporal and spacial specificity and is under tight transcriptional regulation, especially bisindole alkaloids. In this study, an AP2/ERF transcription factor CrERF5 was selected by RNA-seq of organs, and its full-length sequence was cloned and characterized. CrERF5 responds to both ethylene and JA signals and is localized in the nucleus. CrERF5 could activate the transcriptional activity of the TDC promoter. Transient overexpressing CrERF5 in petals caused a significant increase of the expression levels of key genes in both the upstream and downstream pathways of MIAs biosynthesis while silencing resulted in a decrease of them. Accordingly, the contents of bisindole alkaloids anhydrovinblastine and vinblastine, monoindole alkaloids ajmalicine, vindoline, and catharanthine were strongly enhanced in -overexpressing petals while their contents decreased in -silenced plants. These results suggested that CrERF5 is a novel positive ethylene-JA-inducible AP2/ERF transcription factor upregulating the MIAs biosynthetic pathway leading to the bisindole alkaloids accumulation.
PubMed: 31379908
DOI: 10.3389/fpls.2019.00931 -
Frontiers in Plant Science 2018is the sole source of two of the most important anticancer monoterpene indole alkaloids (MIAs), vinblastine and vincristine and their precursors, vindoline and...
Terpene Moiety Enhancement by Overexpression of Geranyl(geranyl) Diphosphate Synthase and Geraniol Synthase Elevates Monomeric and Dimeric Monoterpene Indole Alkaloids in Transgenic .
is the sole source of two of the most important anticancer monoterpene indole alkaloids (MIAs), vinblastine and vincristine and their precursors, vindoline and catharanthine. The MIAs are produced from the condensation of precursors derived from indole and terpene secoiridoid pathways. It has been previously reported that the terpene moiety limits MIA biosynthesis in . Here, to overcome this limitation and enhance MIAs levels in , bifunctional geranyl(geranyl) diphosphate synthase [G(G)PPS] and geraniol synthase (GES) that provide precursors for early steps of terpene moiety (secologanin) formation, were overexpressed transiently by agroinfiltration and stably by -mediated transformation. Both transient and stable overexpression of and co-expression of + significantly enhanced the accumulation of secologanin, which in turn elevated the levels of monomeric MIAs. In addition, transgenic plants exhibited increased levels of root alkaloid ajmalicine. The dimeric alkaloid vinblastine was enhanced only in but not in + transgenic lines that correlated with transcript levels of peroxidase-1 () involved in coupling of vindoline and catharanthine into 3',4'-anhydrovinblastine, the immediate precursor of vinblastine. Moreover, first generation (T) lines exhibited comparable transcript and metabolite levels to that of T lines. In addition, transgenic lines displayed normal growth similar to wild-type plants indicating that the bifunctional G(G)PPS enhanced flux toward both primary and secondary metabolism. These results revealed that improved availability of early precursors for terpene moiety biosynthesis enhanced production of MIAs in at the whole plant level. This is the first report demonstrating enhanced accumulation of monomeric and dimeric MIAs including root MIA ajmalicine in through transgenic approaches.
PubMed: 30034406
DOI: 10.3389/fpls.2018.00942 -
Molecules (Basel, Switzerland) Feb 2015Catharanthus roseus is a medicinal plant belonging to the family Apocynaceae which produces terpenoid indole alkaloids (TIAs) of high medicinal importance. Indeed, a... (Review)
Review
Catharanthus roseus is a medicinal plant belonging to the family Apocynaceae which produces terpenoid indole alkaloids (TIAs) of high medicinal importance. Indeed, a number of activities like antidiabetic, bactericide and antihypertensive are linked to C. roseus. Nevertheless, the high added value of this plant is based on its enormous pharmaceutical interest, producing more than 130 TIAs, some of which exhibit strong pharmacological activities. The most striking biological activity investigated has been the antitumour effect of dimeric alkaloids such as anhydrovinblastine, vinblastine and vincristine which are already in pre-, clinical or in use. The great pharmacological importance of these indole alkaloids, contrasts with the small amounts of them found in this plant, making their extraction a very expensive process. To overcome this problem, researches have looked for alternative sources and strategies to produce them in higher amounts. In this sense, intensive research on the biosynthesis of TIAs and the regulation of their pathways has been developed with the aim to increase by biotechnological approaches, the production of these high added value compounds. This review is focused on the different strategies which improve TIA production, and in the analysis of the beneficial effects that these compounds exert on human health.
Topics: Anti-Bacterial Agents; Antineoplastic Agents; Antioxidants; Antiviral Agents; Catharanthus; China; Humans; Hypoglycemic Agents; Plant Extracts; Plant Shoots; Plants, Medicinal; Secologanin Tryptamine Alkaloids
PubMed: 25685907
DOI: 10.3390/molecules20022973 -
ACS Medicinal Chemistry Letters Sep 2013A series of disubstituted C20'-urea derivatives of vinblastine were prepared from 20'-aminovinblastine that was made accessible through a unique Fe(III)/NaBH- mediated...
A series of disubstituted C20'-urea derivatives of vinblastine were prepared from 20'-aminovinblastine that was made accessible through a unique Fe(III)/NaBH- mediated alkene functionalization reaction of anhydrovinblastine. Three analogs were examined across a panel of 15 human tumor cell lines, displaying remarkably potent cell growth inhibition activity (avg. IC = 200-300 pM), being 10-200-fold more potent than vinblastine (avg. IC = 6.1 nM). Significantly, the analogs also display further improved activity against the vinblastine-resistant HCT116/VM46 cell line that bears the clinically relevant overexpression of Pgp, exhibiting IC values on par with that of vinblastine against the sensitive HCT116 cell line, 100-200-fold greater than the activity of vinblastine against the resistant HCT116/VM46 cell line, and display a reduced 10-20-fold activity differential between the matched sensitive and resistant cell lines (vs 100-fold for vinblastine).
PubMed: 24223237
DOI: 10.1021/ml400281w -
Plant Physiology Jul 2013Catharanthus roseus is one of the most studied medicinal plants due to the interest in their dimeric terpenoid indole alkaloids (TIAs) vinblastine and vincristine, which...
Catharanthus roseus is one of the most studied medicinal plants due to the interest in their dimeric terpenoid indole alkaloids (TIAs) vinblastine and vincristine, which are used in cancer chemotherapy. These TIAs are produced in very low levels in the leaves of the plant from the monomeric precursors vindoline and catharanthine and, although TIA biosynthesis is reasonably well understood, much less is known about TIA membrane transport mechanisms. However, such knowledge is extremely important to understand TIA metabolic fluxes and to develop strategies aimed at increasing TIA production. In this study, the vacuolar transport mechanism of the main TIAs accumulated in C. roseus leaves, vindoline, catharanthine, and α-3',4'-anhydrovinblastine, was characterized using a tonoplast vesicle system. Vindoline uptake was ATP dependent, and this transport activity was strongly inhibited by NH4(+) and carbonyl cyanide m-chlorophenyl hydrazine and was insensitive to the ATP-binding cassette (ABC) transporter inhibitor vanadate. Spectrofluorimetry assays with a pH-sensitive fluorescent probe showed that vindoline and other TIAs indeed were able to dissipate an H(+) gradient preestablished across the tonoplast by either vacuolar H(+)-ATPase or vacuolar H(+)-pyrophosphatase. The initial rates of H(+) gradient dissipation followed Michaelis-Menten kinetics, suggesting the involvement of mediated transport, and this activity was species and alkaloid specific. Altogether, our results strongly support that TIAs are actively taken up by C. roseus mesophyll vacuoles through a specific H(+) antiport system and not by an ion-trap mechanism or ABC transporters.
Topics: Adenosine Triphosphate; Biological Transport; Catharanthus; Hydrogen-Ion Concentration; Indole Alkaloids; Ion Transport; Kinetics; Mesophyll Cells; Plants, Medicinal; Protons; Vacuoles; Vanadates; Vinblastine; Vinca Alkaloids
PubMed: 23686419
DOI: 10.1104/pp.113.220558