[Bioactive secondary metabolites from an endophytic fungus Aspergillus sp. CCH-1E from Catharanthus roseus].

This study focused on the bioactive secondary metabolites of an endophytic fungus Aspergillus sp. CCH-1E from Catharanthus roseus. The secondary metabolites from Aspergillus sp. CCH-1E were isolated by using various chromatographic methods [such as normal-phase and reversed-phase chromatography and high-performance liquid chromatography(HPLC)], and their structures were identified by various spectroscopic methods [e.g., ultraviolet(UV) spectroscopy, infrared(IR) spectroscopy, nuclear magnetic resonance(NMR) spectroscopy, and high-resolution electrospray ionization mass spectrometry(HR-ESI-MS)]. Twelve compounds were yielded and identified from Aspergillus sp. CCH-1E, which are chermesinone H(1), chermesinone I(2), chermesinone B(3), 8,11-didehydrochermesinone B(4), chermesinone C(5), chermesinone A(6), chevalone B(7), barbacenic acid(8), 3,6,8-trihydroxy-3,5,7-trimethyl-3,4-dihydroisocoumarin(9), 5-hydroxy-2-methoxy-7-methyl-1,4-naphthoquinone(10), 1-hydroxy-6,8-dimethoxy-3-methylanthracene-9,10-dione(11), and 7-drimen-9α,11,12-triol(12). Among them, compounds 1 and 2 are new compounds. The growth inhibition effects of all compounds were evaluated against non-small cell lung cancer cell lines A549 and NCI-H1650, as well as human cervical cancer cell line HeLa by using methylthiazolyldiphenyl-tetrazolium bromide(MTT). Compound 7 significantly inhibited the growth of three tumor cells with the IC_(50) values of 1.22-2.43 µmol·L~(-1), respectively. Compounds 1-6 showed moderate cell growth inhibition with the IC_(50) values of 16.24-35.28 µmol·L~(-1).

Quantitative Single-Cell Mass Spectrometry Provides a Highly Resolved Analysis of Natural Product Biosynthesis Partitioning in Plants.

Plants produce an extraordinary array of natural products (specialized metabolites). Notably, these structurally complex molecules are not evenly distributed throughout plant tissues but are instead synthesized and stored in specific cell types. Elucidating both the biosynthesis and function of natural products would be greatly facilitated by tracking the location of these metabolites at the cell-level resolution. However, detection, identification, and quantification of metabolites in single cells, particularly from plants, have remained challenging. Here, we show that we can definitively identify and quantify the concentrations of 16 molecules from four classes of natural products in individual cells of leaf, root, and petal of the medicinal plant Catharanthus roseus using a plate-based single-cell mass spectrometry method. We show that identical natural products show substantially different patterns of cell-type localization in different tissues. Moreover, we show that natural products are often found in a wide range of concentrations across a population of cells, with some natural products at concentrations of over 100 mM per cell. This single-cell mass spectrometry method provides a highly resolved picture of plant natural product biosynthesis partitioning at a cell-specific resolution.

Catharanthus roseus-mediated CuAl2O4 nanocomposites for evaluation of killing kinetics.

The present article portrayed on the killing kinetic of human pathogenic bacteria using bioinspired mesoporous CuAl2O4 nanocomposites (NCs). The NCs was fabricated using leaf extract of medicinal plant Catharanthus roseus (CR) as a green reducer and stabilizer. As bio-fabricated material was calcined at 800 °C and characterized by several analytical techniques like X-ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FT-IR), Ultraviolet-Visible Diffuse Reflectance Spectroscopy (UV-DRS), Energy Dispersive X-Ray Spectroscopy (EDS), X-Ray Photoelectron Spectroscopy (XPS), Raman, Brunauer-Emmett-Teller (BET), Scanning Electron Microscopy (SEM), and Transmission Electron Microscopy (TEM) to authenticate its structure, phase, chemical bonding, chemical state, size and morphology behaviors. XRD and TEM revealed a reduced crystallite and nanoscale sizes of biosynthesized NCs. Moreover, XRD study exposed a cubic-structure of material, while transmission electron microscopy rendered an average particles size in range 10-15 nm. However, BET profile advocates a mesoporous nature of the particles. An effective biological molecular docking modulation assessed by substituting natural inhibitor by bioinspired NCs, while the protein PDB ID 4Z8D FabH as a receptor site for the present investigation. After assessment of molecular docking examination, the antibacterial activity of bioinspired NCs were performed against Staphylococcus aureus, Bacillus subtillis, Klebsiella pneumoniae and Escherichia coli using agar-well method. The broth culture method was employed on different pathogenic strains by kinetic growth assays and colony forming unit.

Vincarostine A, a novel anti-malarial trimeric monoterpenoid indole alkaloid from Catharanthus roseus.

A novel trimeric monoterpenoid indole alkaloid, vincarostine A (1) consisting of an aspidosperma-iboga-aspidosperma type skeleton, was isolated from the whole plant of Catharanthus roseus. The structure including absolute stereochemistry was elucidated on the basis of 2D NMR data and CD spectrum. Vincarostine A (1) showed anti-malarial activity.

Catharanthus roseus-assisted bio-fabricated zinc oxide nanoparticles for promising antibacterial potential against Klebsiella pneumoniae.

This study emphasized on the synthesis of zinc oxide nanoparticles (ZnO NPs) in an environmentally friendly manner from the extract of Catharanthus roseus leaves and its antibacterial assessment against the pneumonia-causing pathogen Klebsiella pneumoniae. This simple and convenient phytosynthesis approach is found to be beneficial over conventional methods, wherein plants serve as excellent reducing, capping, and stabilizing agents that enables the formation of ZnO NPs without the use of harmful chemicals. The formation of ZnO NPs was confirmed through several characterization techniques such as UV-visible spectroscopy, XRD, FT-IR, SEM, HR-TEM, and EDX. XRD analysis revealed high polycrystallinity with crystallite size of approximately 13 nm. SEM and HR-TEM revealed the hexagonal structure of ZnO NPs with the particle size range of 20-50 nm. The EDX shows the elemental purity without any impurity. Furthermore, the antibacterial efficacy by the technique of disc diffusion exhibited clear inhibition zones in ZnO NPs-treated discs. In addition, 125 µg/mL of ZnO NP concentration showed minimum inhibition by the microbroth dilution method. The potent inhibitory activity was further validated with trypan blue dye exclusion and fluorescence microscopy. Finally, SEM examination confirmed the efficient antibacterial potential of ZnO NPs through disruption of the intact morphology of Klebsiella pneumoniae.

Vincazalidine A, a unique bisindole alkaloid from Catharanthus roseus.

A new dimeric indole alkaloid, vincazalidine A consisting of an aspidosperma type and a modified iboga type with 1-azatricyclo ring system consisting of one azepane and two piperidine rings coupled with an oxazolidine ring was isolated from Catharanthus roseus, and the structure including absolute stereochemistry was elucidated on the basis of spectroscopic data as well as DP4 statistical analysis. Vincazalidine A induced G2 arrest and subsequent apoptosis in human lung carcinoma cell line, A549 cells.

Isovincathicine from Catharanthus roseus induces apoptosis in A549 cells.

A dimeric indole alkaloid, isovincathicine consisting of an aspidosperma type and modified iboga with C-7-C-20 connection type skeletons was first isolated from Catharanthus roseus, and the structure including stereochemistry was elucidated on the basis of spectroscopic data as well as DP4 statistical analysis. Isovincathicine inhibited cell proliferation in A549 cells. We investigated the detailed mode of action of isovincathicine-induced inhibitory effects on cell proliferation in A549 cells. Flow cytometric analysis showed that isovincathicine-treated cells accumulated in the G2 phase after 24 h, and the percentage of cells showing cell death increased after 48 h. Western blotting also showed increased expression of BimEL, an apoptosis-related protein, and decreased expression of Mcl-1 and Bcl-xL. Isovincathicine was suggested to induce apoptosis in A549 cells by a mechanism is similar to that of vinblastine.

Combination of Machine Learning and Empirical Computation for the Structural Validation of Trirosaline, a Natural Trimeric Monoterpene Indole Alkaloid from Catharanthus roseus.

Chemical investigation of the emblematic Catharanthus roseus led to the discovery of trirosaline (1), the first example of a tris-ajmalicine-type monoterpene indole alkaloid and the first natural trimeric MIA ever reported from this deeply dug plant species. Its structure was primarily elucidated based on NMR and HRESIMS analyses, and the nature of its unique intermonomeric linkages was firmly confirmed based on a combination of empirical computation and ML-J-DP4 study. Its absolute configuration was mitigated by comparison of experimental and TDDFT-simulated electronic circular dichroism (ECD) spectra. A possible biosynthetic pathway for trirosaline (1) was postulated.

Potentiating Biosynthesis of Alkaloids and Polyphenolic Substances in Catharanthus roseus Plant Using ĸ-Carrageenan.

Catharanthus roseus is a medicinal plant that produces indole alkaloids, which are utilized in anticancer therapy. Vinblastine and vincristine, two commercially important antineoplastic alkaloids, are mostly found in the leaves of Catharanthus roseus. ĸ-carrageenan has been proven as plant growth promoting substance for a number of medicinal and agricultural plants. Considering the importance of ĸ-carrageenan as a promoter of plant growth and phytochemical constituents, especially alkaloids production in Catharanthus roseus, an experiment was carried out to explore the effect of ĸ-carrageenan on the plant growth, phytochemicals content, pigments content, and production of antitumor alkaloids in Catharanthus roseus after planting. Foliar application of ĸ-carrageenan (at 0, 400, 600 and 800 ppm) significantly improved the performance of Catharanthus roseus. Phytochemical analysis involved determining the amount of total phenolics (TP), flavonoids (F), free amino acids (FAA), alkaloids (TAC) and pigments contents by spectrophotometer, minerals by ICP, amino acids, phenolic compounds and alkaloids (Vincamine, Catharanthine, Vincracine (Vincristine), and vinblastine) analysis uses HPLC. The results indicated that all examined ĸ-carrageenan treatments led to a significant (p ≤ 0.05) increase in growth parameters compared to the untreated plants. Phytochemical examination indicates that the spray of ĸ-carrageenan at 800 mg L-1 increased the yield of alkaloids (Vincamine, Catharanthine and Vincracine (Vincristine)) by 41.85 µg/g DW, total phenolic compounds by 3948.6 µg gallic/g FW, the content of flavonoids 951.3 µg quercetin /g FW and carotenoids content 32.97 mg/g FW as compared to the control. An amount of 400 ppm ĸ-carrageenan treatment gave the best contents of FAA, Chl a, Chl b and anthocyanin. The element content of K, Ca, Cu, Zn and Se increased by treatments. Amino acids constituents and phenolics compounds contents were altered by ĸ-carrageenan.

Do Fungicides Affect Alkaloid Production in Catharanthus roseus (L.) G. Don. Seedlings?

The presence of endophytes in plants is undeniable, but how significant their involvement is in the host plant biosynthetic pathways is still unclear. The results reported from fungicide treatments in plants varied. Fungicide treatment in Taxus was found to decrease the taxol content. In Ipomoea asarifolia, Pronto Plus and Folicur treatments coincided with the disappearance of ergot alkaloids from the plant. In Narcissus pseudonarcissus cv. Carlton, a mixture of fungicide applications decreased the alkaloids concentration and altered the carbohydrate metabolism. Jacobaea plants treated with Folicur reduced the pyrrolizidine alkaloids content. There have not been any studies into the involvement of endophytic fungi on alkaloids production of Catharanthus roseus until now. Though there is a report on the isolation of the endophytic fungi, Fusarium oxysporum from C. roseus, which was reported to produce vinblastine and vincristine in vitro. To detect possible collaborations between these two different organisms, fungicides were applied to suppress the endophytic fungi in seedlings and then measure the metabolomes by 1HNMR and HPLC analysis. The results indicate that endophytic fungi were not directly involved in alkaloids biosynthesis. Treatment with fungicides influenced both the primary and secondary metabolism of C. roseus. The systemic fungicides Pronto Plus and Folicur caused an increase in loganin and secologanin levels. In contrast, control samples had higher level of catharanthine and vindoline. This means that fungicide treatments cause changes in plant secondary metabolism.

[A Systematic Review of the Pharmacological and Phytochemical Profiles of Madagascar periwinkle as Potential Dietary Supplement].

Madagascar periwinkle (Catharanthus roseus) is a plant species known for its rich pharmacological and phytochemical properties. This systematic review aims to comprehensively evaluate the potential of Madagascar periwinkle as a dietary supplement. A thorough search of relevant databases yielded studies focusing on the pharmacological activities and phytochemical constituents of Madagascar periwinkle. The review highlights the diverse pharmacological effects of Madagascar periwinkle, including anti-cancer, anti-diabetic, anti-inflammatory, and antimicrobial properties, among others. Furthermore, the phytochemical analysis revealed the presence of various bioactive compounds such as alkaloids, flavonoids, terpenoids, and phenolics, which contribute to its medicinal properties. Despite the promising findings, further research is warranted to elucidate the mechanisms of action, safety profile, and potential interactions of Madagascar periwinkle as a dietary supplement. Overall, this systematic review provides valuable insights into the pharmacological and phytochemical profiles of Madagascar periwinkle, suggesting its potential as a natural dietary supplement with diverse health benefits.

Fields to fermentors: Brewing botanical chemotherapeutic precursors using genetically engineered yeast.

Plants are a rich source of chemotherapeutics and other essential medicines, but plant-based drug supply chains are unsustainable. Writing in Nature, Zhang et al.1 demonstrated a proof-of-concept alternate source of the anticancer drug vinblastine by engineering yeast to convert sugar and amino acids into its direct precursors, catharanthine and vindoline.

Biogenic Synthesis of Silver Nanoparticles Using Catharanthus roseus and Its Cytotoxicity Effect on Vero Cell Lines.

Background: Type 2 diabetes mellitus (DM2) is a chronic and sometimes fatal condition which affects people all over the world. Nanotherapeutics have shown tremendous potential to combat chronic diseases­including DM2­as they enhance the overall impact of drugs on biological systems. Greenly synthesized silver nanoparticles (AgNPs) from Catharanthus roseus methanolic extract (C. AgNPs) were examined primarily for their cytotoxic and antidiabetic effects. Methods: Characterization of C. AgNPs was performed by UV−vis spectroscopy, Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), and atomic force microscopy (AFM). The C. AgNPs were trialed on Vero cell line and afterwards on an animal model (rats). Results: The C. AgNPs showed standard structural and functional characterization as revealed by FTIR and XRD analyses. The zetapotential analysis indicated stability while EDX analysis confirmed the formation of composite capping with Ag metal. The cytotoxic effect (IC50) of C. AgNPs on Vero cell lines was found to be 568 g/mL. The animal model analyses further revealed a significant difference in water intake, food intake, body weight, urine volume, and urine sugar of tested rats after treatment with aqueous extract of C. AgNPs. Moreover, five groups of rats including control and diabetic groups (NC1, PC2, DG1, DG2, and DG3) were investigated for their blood glucose and glycemic control analysis. Conclusions: The C. AgNPs exhibited positive potential on the Vero cell line as well as on experimental rats. The lipid profile in all the diabetic groups (DG1-3) were significantly increased compared with both of the control groups (p < 0.05). The present study revealed the significance of C. AgNPs in nanotherapeutics.

A microbial supply chain for production of the anti-cancer drug vinblastine.

Monoterpene indole alkaloids (MIAs) are a diverse family of complex plant secondary metabolites with many medicinal properties, including the essential anti-cancer therapeutics vinblastine and vincristine1. As MIAs are difficult to chemically synthesize, the world's supply chain for vinblastine relies on low-yielding extraction and purification of the precursors vindoline and catharanthine from the plant Catharanthus roseus, which is then followed by simple in vitro chemical coupling and reduction to form vinblastine at an industrial scale2,3. Here, we demonstrate the de novo microbial biosynthesis of vindoline and catharanthine using a highly engineered yeast, and in vitro chemical coupling to vinblastine. The study showcases a very long biosynthetic pathway refactored into a microbial cell factory, including 30 enzymatic steps beyond the yeast native metabolites geranyl pyrophosphate and tryptophan to catharanthine and vindoline. In total, 56 genetic edits were performed, including expression of 34 heterologous genes from plants, as well as deletions, knock-downs and overexpression of ten yeast genes to improve precursor supplies towards de novo production of catharanthine and vindoline, from which semisynthesis to vinblastine occurs. As the vinblastine pathway is one of the longest MIA biosynthetic pathways, this study positions yeast as a scalable platform to produce more than 3,000 natural MIAs and a virtually infinite number of new-to-nature analogues.

Engineering Catharanthus roseus monoterpenoid indole alkaloid pathway in yeast.

Catharanthus roseus (Madagascar periwinkle), a medicinal plant possessing high pharmacological attributes, is widely recognized for the biosynthesis of anticancer monoterpenoid indole alkaloids (MIAs) - vinblastine and vincristine. The plant is known to biosynthesize more than 130 different bioactive MIAs, highly acclaimed in traditional and modern medicinal therapies. The MIA biosynthesis is strictly regulated at developmental and spatial-temporal stages and requires a well-defined cellular and sub-cellular compartmentation for completion of the entire MIAs biosynthesis. However, due to their cytotoxic nature, the production of vinblastine and vincristine occurs in low concentrations in planta and the absence of chemical synthesis alternatives projects a huge gap in demand and supply, leading to high market price. With research investigations spanning more than four decades, plant tissue culture and metabolic engineering (ME)-based studies were attempted to explore, understand, explain, improve and enhance the MIA biosynthesis using homologous and heterologous systems. Presently, metabolic engineering and synthetic biology are the two powerful tools that are contributing majorly in elucidating MIA biosynthesis. This review concentrates mainly on the efforts made through metabolic engineering of MIAs in heterologous microbial factories. KEY POINTS: • Yeast engineering provides alternative production source of phytomolecules • Yeast engineering also helps to discover missing plant pathway enzymes and genes.

Enhancement of vincristine under in vitro culture of Catharanthus roseus supplemented with Alternaria sesami endophytic fungal extract as a biotic elicitor.

Vincristine, one of the major vinca alkaloid of Catharanthus roseus (L.) G. Don. (Apocynaceae), was enhanced under in vitro callus culture of C. roseus using fungal extract of an endophyte Alternaria sesami isolated from the surface-sterilized root cuttings of C. roseus. Vindoline, a precursor molecule for vincristine production, was detected for the first time in the fungal endophyte A. sesami which was used as a biotic elicitor in this study to enhance vincristine content in the C. roseus callus. It was identified using high-performance liquid chromatography and mass spectroscopy techniques by matching retention time and mass data with reference molecule. Supplementing the heat sterilized A. sesami endophytic fungal culture extract into the callus culture medium of C. roseus resulted in the enhancement of vincristine content in C. roseus callus by 21.717% after 105-day culture.

Catharanthus roseus (L.) G. Don: A review of its ethnobotany, phytochemistry, ethnopharmacology and toxicities.

ETHNOPHARMACOLOGICAL RELEVANCE: Catharanthus roseus (L.) G. Don is a well known medicinal plant belonging to family Apocynaceae that have been traditionally used as medicine since ancient times. C. roseus is a well-recognized herbal medicine due to its anticancer bisindole alkaloids (vinblastine (111), vincristine (112) and vindesine (121)). In the Ayurvedic system of medicine, different parts of C. roseus are used in folklore herbal medicine for treatment of many types of cancer, diabetes, stomach disorders, kidney, liver and cardiovascular diseases. AIM OF THE STUDY: The main idea behind this communication is to update comprehensively and analyze critically the traditional applications, phytochemistry, pharmacological activities, and toxicity of various extracts and isolated compounds from C. roseus. MATERIALS AND METHODS: The presented data covers scientific works on C. roseus published across the world between 1967 and 2021 was searched from various international publishing houses using search engines as well as several traditional texts like Ayurveda and relevant books. Collected data from different sources was comprehensively summarized/analyzed for ethnomedicinal uses, phytochemistry, analytical chemistry, biological activities and toxicity studies of C. roseus. RESULTS AND DISCUSSION: C. roseus has a wide range of applications in the traditional system of medicine especially in cancer and diabetes. During phytochemical investigation, total of 344 compounds including monoterpene indole alkaloids (MIAs) (110), bisindole alkaloids (35), flavonoids (34), phenolic acids (9) and volatile constituents (156) have been reported in the various extracts and fractions of different plant parts of C. roseus. The extracts and isolated compounds of C. roseus have to exhibit many pharmacological activities such as anticancer/cytotoxic, antidiabetic, antimicrobial, antioxidant, larvicidal and pupicidal. The comparative toxicity of extracts and bioactive compounds investigated in dose dependent manner. The investigation of toxicity showed that the both extracts and isolated compounds are safe to a certain limit beyond that they cause adverse effects. CONCLUSION: This review is a comprehensive, critically analyzed summarization of sufficient baseline information of selected topics in one place undertaken till date on C. roseus for future works and drug discovery. The phytochemical investigation including biosynthetic pathways showed that the MIAs and bisindole alkaloids are major and characteristic class of compounds in this plant. The present data confirm that the extracts/fractions and their isolated alkaloids especially vinblastine (111) and vincristine (112) have a potent anticancer/cytotoxic and antidiabetic property and there is a need for further study with particular attention to the mechanisms of anticancer activity. In biosynthesis pathways of alkaloids especially bisindole alkaloids, some enzymes and rearrangement are unexposed therefore it is required to draw special attention. It also focuses on attracting the attention of scientific communities about the widespread biological activities of this species for its better utilization prospects in the near future.

Monoterpene Indole Alkaloids with Cav3.1 T-Type Calcium Channel Inhibitory Activity from Catharanthus roseus.

Catharanthus roseus is a well-known traditional herbal medicine for the treatment of cancer, hypertension, scald, and sore in China. Phytochemical investigation on the twigs and leaves of this species led to the isolation of two new monoterpene indole alkaloids, catharanosines A (1) and B (2), and six known analogues (3-8). Structures of 1 and 2 were established by 1H-, 13C- and 2D-NMR, and HREIMS data. The absolute configuration of 1 was confirmed by single-crystal X-ray diffraction analysis. Compound 2 represented an unprecedented aspidosperma-type alkaloid with a 2-piperidinyl moiety at C-10. Compounds 6-8 exhibited remarkable Cav3.1 low voltage-gated calcium channel (LVGCC) inhibitory activity with IC50 values of 11.83 ± 1.02, 14.3 ± 1.20, and 14.54 ± 0.99 µM, respectively.

Mining of the Catharanthus roseus Genome Leads to Identification of a Biosynthetic Gene Cluster for Fungicidal Sesquiterpenes.

Characterization of cryptic biosynthetic gene clusters (BGCs) from microbial genomes has been proven to be a powerful approach to the discovery of new natural products. However, such a genome mining approach to the discovery of bioactive plant metabolites has been muted. The plant BGCs characterized to date encode pathways for antibiotics important in plant defense against microbial pathogens, providing a means to discover such phytoalexins by mining plant genomes. Here is reported the discovery and characterization of a minimal BGC from the medicinal plant Catharanthus roseus, consisting of an adjacent pair of genes encoding a terpene synthase (CrTPS18) and cytochrome P450 (CYP71D349). These two enzymes act sequentially, with CrTPS18 acting as a sesquiterpene synthase, producing 5-epi-jinkoheremol (1), which CYP71D349 further hydroxylates to debneyol (2). Infection studies with maize revealed that 1 and 2 exhibit more potent fungicidal activity than validamycin. Accordingly, this study demonstrates that characterization of such cryptic plant BGCs is a promising strategy for the discovery of potential agrochemical leads. Moreover, despite the observed absence of 1 and 2 in C. roseus, the observed transcriptional regulation is consistent with their differential fungicidal activity, suggesting that such conditional coexpression may be sufficient to drive BGC assembly in plants.

Britannin, a sesquiterpene lactone induces ROS-dependent apoptosis in NALM-6, REH, and JURKAT cell lines and produces a synergistic effect with vincristine.

BACKGROUND: Britannin, a Sesquiterpene Lactone isolated from Inula aucheriana, has recently gained attraction in the therapeutic fields due to its anti-tumor properties. This study was designed to evaluate the effect of this agent on Acute Lymphoblastic Leukemia (ALL) cell lines, either as a monotherapy or in combination with Vincristine (VCR). METHODS AND RESULTS: To determine the anti-leukemic effects of Britannin on ALL-derived cell lines and suggest a mechanism of action for the agent, we used MTT assay, Annexin-V/PI staining, ROS assay, and real-time PCR analysis. Moreover, by using a combination index (CI), we evaluated the synergistic effect of Britannin on Vincristine. We found that unlike normal Peripheral Blood Mononuclear Cells (PBMCs) and L929 cells, Britannin reduced the viability of NALM-6, REH, and JURKAT cells. Among tested cells, NALM-6 cells had the highest sensitivity to Britannin, and this agent was able to induce p21/p27-mediated G1 cell cycle arrest and Reactive Oxygen Specious (ROS)-mediated apoptotic cell death in this cell line. When NALM-6 cells were treated with Nacetyl-L-Cysteine (NAC), a scavenger of ROS, Britannin could induce neither apoptosis nor reduce the survival of the cells suggesting that the cytotoxic effect of Britannin is induced through ROS-dependent manner. Moreover, we found that a low dose of Britannin enhanced the effect of Vincristine in NALM-6 cells by inducing apoptotic cell death via altering the expression of apoptotic-related genes. CONCLUSIONS: Overall, our results proposed a mechanism for the cytotoxic effect of Britannin, either as a single agent or in combination with Vincristine, in NALM-6 cells.