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A biotransformation pair consisting of vinblastine: vincristine present in the Catharanthus roseus plant is of immense pharmacological significance. In this study, we successfully transformed vinblastine into vincristine outside the plant using Pseudomonas aeruginosa 8485 and Pseudomonas fluorescens 2421 and evaluated the antiangiogenic potential of thus produced vincristine through the CAM assay. The toxicity assay showed that both Pseudomonas spp. can tolerate varying concentrations (25-100 µl of 1 mg/ml) of vinblastine. The biotransformation was performed in a liquid nutrient broth medium containing vinblastine (25-100 µl), and Pseudomonas spp. inoculums (50-150 µl) by incubating at 30 °C and 37 °C, respectively for 8 days. The process was optimized for substrate and culture concentrations, pH, temperature, and rotation speed (rpm) for the highest conversion. Analysis using LC-MS/MS confirmed the presence of vincristine as a product of the vinblastine biotransformation by two Pseudomonas spp. P. fluorescens 2421 showed a faster conversion rate with 95% of vinblastine transformed within 24 h than P. aeruginosa 8485, which demonstrated a conversion rate of 92% on the 8 th day. From LC-MS/MS analysis, the optimal conditions for the reaction were determined as vinblastine (25 µl), microbial inoculums (150 µl or 200 × 10 6 and 210 × 10 6  CFU/ml), pH 7.4, rotation speed of 180 rpm, and temperatures of 30 °C and 37 °C with incubation time of 8 days. The vincristine produced exhibited potent antiangiogenic activity in the CAM assay reducing the thickness and branching of blood vessels in a dose-dependent manner. The study concludes that both Pseudomonas spp. showed promise for vincristine production from vinblastine, without compromising its antiangiogenic properties., (© 2024. The Author(s).)

The efficacy of many cancer drugs is hindered by P-glycoprotein (Pgp), a cellular pump that removes drugs from cells. To improve chemotherapy, drugs capable of evading Pgp must be developed. Despite similarities in structure, vinca alkaloids (VAs) show disparate Pgp-mediated efflux ratios. ATPase activity and binding affinity studies show at least two binding sites for the VAs: high- and low-affinity sites that stimulate and inhibit the ATPase activity rate, respectively. The affinity for ATP from the ATPase kinetics curve for vinblastine (VBL) at the high-affinity site was 2- and 9-fold higher than vinorelbine (VRL) and vincristine (VCR), respectively. Conversely, VBL had the highest K m (ATP) for the low-affinity site. The dissociation constants (K D s) determined by protein fluorescence quenching were in the order VBL < VRL< VCR. The order of the K D s was reversed at higher substrate concentrations. Acrylamide quenching of protein fluorescence indicate that the VAs, either at 10 µM or 150 µM, predominantly maintain Pgp in an open-outward conformation. When 3.2 mM AMPPNP was present, 10 µM of either VBL, VRL, or VCR cause Pgp to shift to an open-outward conformation, while 150 µM of the VAs shifted the conformation of Pgp to an intermediate orientation, between opened inward and open-outward. However, the conformational shift induced by saturating AMPPNP and VCR condition was less than either VBL or VRL in the presence of AMPPNP. At 150 µM, atomic force microscopy (AFM) revealed that the VAs shift Pgp population to a predominantly open-inward conformation. Additionally, STDD NMR studies revealed comparable groups in VBL, VRL, and VCR are in contact with the protein during binding. Our results, when coupled with VAs-microtubule structure-activity relationship studies, could lay the foundation for developing next-generation VAs that are effective as anti-tumor agents. A model that illustrates the intricate process of Pgp-mediated transport of the VAs is presented., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 American Pharmacists Association. Published by Elsevier Inc. All rights reserved.)

Previous studies have suggested that the incidence of vincristine-induced peripheral neuropathy (VIPN) is potentially linked with cytochrome P450 (CYP)3A5, a polymorphic enzyme that metabolizes vincristine in vitro, and with concurrent use of azole antifungals such as ketoconazole. The assumed mechanism for these interactions is through modulation of CYP3A-mediated metabolism, leading to decreased vincristine clearance and increased susceptibility to VIPN. Given the controversy surrounding the contribution of these mechanisms, we directly tested these hypotheses in genetically engineered mouse models with a deficiency of the entire murine Cyp3a locus [Cyp3a(-/-) mice] and in humanized transgenic animals with hepatic expression of functional and nonfunctional human CYP3A5 variants. Compared with wild-type mice, the systemic exposure to vincristine was increased by only 1.15-fold (95% confidence interval, 0.84-1.58) in Cyp3a(-/-) mice, suggesting that the clearance of vincristine in mice is largely independent of hepatic Cyp3a function. In line with these observations, we found that Cyp3a deficiency or pretreatment with the CYP3A inhibitors ketoconazole or nilotinib did not influence the severity and time course of VIPN and that exposure to vincristine was not substantially altered in humanized CYP3A5*3 mice or humanized CYP3A5*1 mice compared with Cyp3a(-/-) mice. Our study suggests that the contribution of CYP3A5-mediated metabolism to vincristine elimination and the associated drug-drug interaction potential is limited and that plasma levels of vincristine are unlikely to be strongly predictive of VIPN. SIGNIFICANCE STATEMENT: The current study suggests that CYP3A5 genotype status does not substantially influence vincristine disposition and neurotoxicity in translationally relevant murine models. These findings raise concerns about the causality of previously reported relationships between variant CYP3A5 genotypes or concomitant azole use with the incidence of vincristine neurotoxicity., (Copyright © 2024 by The American Society for Pharmacology and Experimental Therapeutics.)

Study Question: What is the impact of low- or moderate-risk gonadotoxic chemotherapy received prior to testicular tissue freezing (TTF), and of the cancer itself, on spermatogonia quantity in testicular tissue from (pre)pubertal boys?, Summary Answer: Vincristine, when associated with alkylating agents, has an additional adverse effect on spermatogonia quantity, while carboplatin has no individual contribution to spermatogonia quantity, in testicular tissue of (pre)pubertal boys, when compared to patients who have received non-alkylating chemotherapy., What Is Known Already: The improved survival rates after cancer treatment necessitate the inclusion of fertility preservation procedures as part of the comprehensive care for patients, taking into consideration their age. Sperm cryopreservation is an established procedure in post-pubertal males while the TTF proposed for (pre)pubertal boys remains experimental. Several studies exploring testicular tissue of (pre)pubertal boys after TTF have examined the tubular fertility index (TFI, percentage of seminiferous tubule cross-sections containing spermatogonia) and the number of spermatogonia per seminiferous tubule cross-section (S/T). All studies have demonstrated that TFI and S/T always decrease after the introduction of chemotherapeutic agents, especially those which carry high gonadotoxic risks such as alkylating agents., Study Design, Size, Duration: Testicular tissue samples from 79 (pre)pubertal boys diagnosed with cancer (from 6 months to 16 years of age) were cryopreserved between May 2009 and June 2014. Their medical diagnoses and previous chemotherapy exposures were recorded. We examined histological sections of (pre)pubertal testicular tissue to elucidate whether the chemotherapy or the primary diagnosis affects mainly TFI and S/T., Participants/materials, Setting, Methods: (Pre)pubertal boys with cancer diagnosis who had been offered TTF prior to conditioning treatment for hematopoietic stem cell transplantation were included in the study. All the patients had previously received chemotherapy with low- or moderate-risk for future fertility. We have selected patients for whom the information on the chemotherapy received was complete. The quantity of spermatogonia and quality of testicular tissue were assessed by both morphological and immunohistochemical analyses., Main Results and the Role of Chance: A significant reduction in the number of spermatogonia was observed in boys treated with alkylating agents. The mean S/T values in boys exposed to alkylating agents were significantly lower compared to boys exposed to non-alkylating agents (P = 0.018). In contrast, no difference was observed for patients treated with carboplatin as the sole administered alkylating agent compared to the group of patients exposed to non-alkylating agents. We observed an increase of S/T with age in the group of patients who did not receive any alkylating agent and a decrease of S/T with age when patients received alkylating agents included in the cyclophosphamide equivalent dose (CED) formula (r = 0.6166, P = 0.0434; r = -0.3759, P = 0.0036, respectively). The TFI and S/T decreased further in the group of patients who received vincristine in combination with alkylating agents (decrease of 22.4%, P = 0.0049 and P < 0.0001, respectively), but in this group the CED was also increased significantly (P < 0.0001). Multivariate analysis, after CED adjustment, showed the persistence of a decrease in TFI correlated with vincristine administration (P = 0.02)., Limitations, Reasons for Caution: This is a descriptive study of testicular tissues obtained from (pre)pubertal boys who were at risk of infertility. The study population is quite heterogeneous, with a small number of patients in each sub-group. Our results are based on comparisons between patients receiving alkylating agents compared to patients receiving non-alkylating agents rather than chemotherapy-naive patients. The French national guidelines for fertility preservation in cancer patients recommend TTF before highly gonadotoxic treatment. Therefore, all the patients had received low- or moderate-risk gonadotoxic chemotherapy before TTF. Access to testicular tissue samples from chemotherapy-naive patients with comparable histological types of cancer was not possible. The functionality of spermatogonia and somatic cells could not be tested by transplantation or in vitro maturation due to limited sample sizes., Wider Implications of the Findings: This study summarizes the spermatogonial quantity of (pre)pubertal boys prior to TTF. We confirmed a negative correlation between the cumulative exposure to alkylating agents and spermatogonial quantity. In addition, the synergistic use of vincristine in combination with alkylating agents showed a cumulative deleterious effect on the TFI. For patients for whom fertility preservation is indicated, TTF should be proposed for chemotherapy with a predicted CED above 4000 mg/m2. However, the data obtained from vincristine and carboplatin use should be confirmed in a subsequent study including more patients., Study Funding/competing Interest(s): This study had financial support from a French national research grant PHRC No. 2008/071/HP obtained by the French Institute of Cancer and the French Healthcare Organization. The sponsors played no role in the study. The authors declare no conflicts of interest., Trial Registration Number: N/A., (© The Author(s) 2023. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Vincristine (VCR) is a vital component in numerous treatment regimens for pediatric blood cancer. VCR-induced peripheral neuropathy (VIPN) represents a type of VCR toxicity influenced by multiple factors, including age, race, genetic traits, dosage, interactions, and administration regimen. However, the dose-response relationship of VIPN remains elusive. VCR is primarily metabolized by cytochrome P450 (CYP) 3 A to generate the major metabolite M1. To date, there is a lack of literature documenting the pharmacokinetics (PK) characteristics of VCR and M1 in Chinese children within a 96 h timeframe. To address the gap, a developed LC-MS/MS method was successfully employed in the PK study of VCR and M1 in Chinese pediatric acute lymphoblastic leukemia (ALL) patients. M1 was obtained through in vitro metabolism experiments, and mixed plasma samples of M1 and VCR were prepared. Plasma samples were pre-processed using the solid-phase extraction (SPE) technique. Samples were loaded into ProElut C18 Cartridges, washed with 5% methanol aqueous solution, and eluted with methanol. The eluent was concentrated and reconstituted for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. The standard calibration curves for VCR and M1 were 0.1-50 ng/mL and 0.05-5 ng/mL, respectively, with linear coefficients exceeding 0.99. Accuracy and precision of quality control (QC) samples fell within 115%. The analytical approach satisfied the quantitative demands for VCR and M1 in plasma samples within 96 h. VCR was metabolized to M1 at a relatively constant proportion (5.37%-18.06%) of VCR in vivo. No significant differences were observed in PK parameters of VCR in Chinese children compared to other countries and races. Further investigation is required to identify the key factors influencing VIPN in Chinese children., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Nicotinamide adenine dinucleotide (NAD) + -biosynthetic and consuming enzymes are involved in various intracellular events through the regulation of NAD + metabolism. Recently, it has become clear that alterations in the expression of NAD + -biosynthetic and consuming enzymes contribute to the axonal stability of neurons. We explored soluble bioactive factor(s) that alter the expression of NAD + -metabolizing enzymes and found that cytokine interferon (IFN)-γ increased the expression of nicotinamide nucleotide adenylyltransferase 2 (NMNAT2), an NAD + -biosynthetic enzyme. IFN-γ activated signal transducers and activators of transcription 1 and 3 (STAT1/3) followed by c-Jun N-terminal kinase (JNK) suppression. As a result, STAT1/3 increased the expression of NMNAT2 at both mRNA and protein levels in a dose- and time-dependent manner and, at the same time, suppressed activation of sterile alpha and Toll/interleukin receptor motif-containing 1 (SARM1), an NAD + -consuming enzyme, and increased intracellular NAD + levels. We examined the protective effect of STAT1/3 signaling against vincristine-mediated cell injury as a model of chemotherapy-induced peripheral neuropathy (CIPN), in which axonal degeneration is involved in disease progression. We found that IFN-γ-mediated STAT1/3 activation inhibited vincristine-induced downregulation of NMNAT2 and upregulation of SARM1 phosphorylation, resulting in modest suppression of subsequent neurite degradation and cell death. These results indicate that STAT1/3 signaling induces NMNAT2 expression while simultaneously suppressing SARM1 phosphorylation, and that both these actions contribute to suppression of axonal degeneration and cell death., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflicts of interest with respect to the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Vincristine (VCR), an alkaloid isolated from vinca, is a commonly used chemotherapeutic drug. However, VCR therapy can lead to dose-dependent peripheral neurotoxicity, mainly manifesting as neuropathic pain, which is one of the dominant reasons for limiting its utility. Experimentally, we discovered that VCR-induced neuropathic pain (VINP) was accompanied by astrocyte activation; the upregulation of phospho-CaMKII (p-CaMKII), Ca V 3.2, and Connexin-43 (Cx43) expression; and the production and release of inflammatory cytokines and chemokines in the spinal cord. Similar situations were also observed in astrocyte cultures. Interestingly, these alterations were all reversed by intrathecal injection of KN-93 (a CaMKII inhibitor) or L-Ascorbic acid (a Ca V 3.2 inhibitor). In addition, KN-93 and L-Ascorbic acid inhibited the increase in [Ca 2+ ] i associated with astrocyte activation. We also verified that knocking down or inhibiting Cx43 level via intrathecal injection of Cx43 siRNA or Gap27 (a Cx43 mimetic peptide) relieved pain hypersensitivity and reduced the release of inflammatory factors; however, they did not affect astrocyte activation or p-CaMKII and Ca V 3.2 expression. Besides, the overexpression of Cx43 through the transfection of the Cx43 plasmid did not affect p-CaMKII and Ca V 3.2 expressions in vitro. Therefore, CaMKII and Ca V 3.2 may activate astrocytes by increasing [Ca 2+ ] i , thereby mediating Cx43-dependent inflammation in VINP. Moreover, we demonstrated that the CaMKII signalling pathway was involved in VCR-induced inflammation, apoptosis, and mitochondrial damage. Collectively, our findings show a novel mechanism by which CaMKII and Ca V 3.2 mediate Cx43-dependent inflammation by activating astrocytes in neuropathic pain induced by VCR., (© 2021. The Author(s), under exclusive licence to Springer Nature B.V.)

Catharanthus roseus is a perennial herb of the Apocynaceae family, from which about 200 kinds of alkaloids have been characterized. Most alkaloids from C. roseus are terpenoid indole alkaloids (TIAs), such as vinblastine and vincristine, which are widely used in the clinic for their good antitumor activity. However, they were only biosynthesized in C. roseus, and their content in C. roseus is extremely low. The access to these valuable compounds is by plant extraction or chemical semisynthesis from their precursors catharanthine and vindoline. Since catharanthine and vindoline are also obtained from C. roseus, the supply of vinblastine and vincristine makes it difficult to meet market demands. Therefore, how to improve the yield of TIAs is an attractive issue. In this study, we compared the regulatory effect of two critical transcription factors, octadecanoid-derivative responsive Catharanthus AP2-domain protein 3 (ORCA3) and octadecanoid-derivative responsive Catharanthus AP2-domain protein 4 (ORCA4), on the biosynthesis of TIAs in C. roseus. The results showed that overexpressing both two transcription factors could increase the accumulation of TIAs. The effect was more significant when ORCA4 was overexpressed. To acquire C. roseus TIAs on a continuous and consistent basis, we then created and acquired C. roseus stem cells stably overexpressing ORCA4. This is the first time a recombinant C. roseus stem cell system with stable ORCA4 overexpression has been developed, which not only provides new ideas for future research in this area but also breaches new life into the industrial application of using plant cell culture to obtain natural products., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Masson SAS. All rights reserved.)

Since the discovery of the anticancer drugs vinblastine and vincristine, Catharanthus roseus has been intensively studied for biosynthesis of several terpene indole alkaloids (TIAs). Due to their low abundance in plant tissues at a simultaneously high demand, modes of production alternative to conventional extraction are mandatory. Plant cell fermentation might become one of these alternatives, yet decades of research have shown limited success to certain product classes, leading to the question: how to preserve the intrinsic ability to produce TIAs (metabolic competence) in cell culture? We used the strategy to use the developmental potency of mature embryos to generate such strains. Two cell strains (C1and C4) from seed embryos of Catharanthus roseus were found to differ not only morphologically, but also in their metabolic competence. This differential competence became manifest not only under phytohormone elicitation, but also upon feeding with alkaloid pathway precursors. The more active strain C4 formed larger cell aggregates and was endowed with longer mitochondria. These cellular features were accompanied by higher alkaloid accumulation in response to methyl jasmonate (MeJA) elicitation. The levels of catharanthine could be increased significantly, while the concurrent vindoline branch of the pathway was blocked, such that no bisindole alkaloids were detectable. By feeding vindoline to MeJA-elicited C4 cells, vincristine became detectable; however, only to marginal amounts. In conclusion, these results show that cultured cells are not "de-differentiated", but can differ in metabolic competence. In addition to elicitation and precursor feeding, the cellular properties of the "biomatter" are highly relevant for the success of plant cell fermentation., (© 2022. The Author(s).)

Objective The purpose of this preliminary study was to compare racial background and CYP3A distribution in pediatric acute lymphoblastic leukemia patients as it relates to vincristine-related neurotoxicity. Methods Patients with B-precursor acute lymphoblastic leukemia treated at Children’s Hospital of Michigan were eligible to participate in this study. Determination of the CYP3A variant for each patient was done using Qiagen DNA Blood Mini Kit and polymerase chain reaction amplification. Patients were monitored during their leukemia treatment course for vincristine-related neurotoxicity. Results Fifty-four patients were enrolled. Twenty-nine Caucasian patients (81%) and 13 African-American patients (77%) experienced neurotoxicity. CYP3A genotyping was done for 52 patients. Two African-American and two Caucasian patients were homozygous A/A for the CYP3A5*3 polymorphism. Three of these patients (75%) experienced grade 2 neuropathy. Two Caucasian patients and one African-American patient were heterozygous A/G. Two of these patients (66.7%) experienced grade 2 or 3 neuropathy. Thirty-five patients (67.3%) were homozygous for the mutant inactive G/G allele for CYP3A5*3, eight African-American and 27 Caucasian patients. Of these, six of the African-American patients (75%) and 22 of the Caucasian patients (81.5%) experienced neuropathy. Conclusion The CYP3A5*3 genotype causes very low expression of the CYP3A5 protein and hence decreased vincristine metabolism. In this study, patients who expressed CYP3A5*3 had an increased incidence of vincristine-related neurotoxicity. Overall, a greater percentage of Caucasian patients had documented incidences of neurotoxicity. A larger sample size and more detailed gene analysis are needed for future studies. [ABSTRACT FROM AUTHOR]

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 1 HNMR 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.

Background: Roughly half of all diffuse large B-cell lymphomas (DLBCLs) are infiltrated by large numbers of regulatory T-cells (Tregs). Although the presence of 'effector' Tregs in particular is associated with an inferior prognosis in patients on standard rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) immunochemotherapy, the role of this cell type during lymphoma initiation and progression is poorly understood., Methods: Here, we use tissue microarrays containing prospectively collected DLBCL patient specimens, as well as data from publicly available cohorts to explore the mutational landscape of Treg-infiltrated DLBCL. We further take advantage of a model of MYC-driven lymphoma to mechanistically dissect the contribution of Tregs to lymphoma pathogenesis and to develop a strategy of Treg-selective interleukin-2 (IL-2) starvation to improve immune control of MYC-driven lymphoma., Results: We find that all genetic DLBCL subtypes, except for one characterized by co-occurring MYD88 / CD79 mutations, are heavily infiltrated by Tregs. Spectral flow cytometry and scRNA-sequencing reveal the robust expression of functional and immunosuppressive markers on Tregs infiltrating MYC-driven lymphomas; notably, we find that intratumoral Tregs arise due to local conversion from naïve CD4 + precursors on tumor contact. Treg ablation in Foxp3 iDTR mice, or by antibody-mediated Treg-selective blockade of IL-2 signaling, strongly reduces the lymphoma burden. We identify lymphoma B-cells as a major source of IL-2, and show that the effects of Treg depletion are reversed by the simultaneous depletion of Foxp3-negative CD4 + T-cells, but not CD8 + T-cells or natural killer (NK) cells. The inhibition of ATP hydrolyzation and adenosine production by Tregs at least partly phenocopies the effects of Treg depletion. Treg depletion further synergizes with pro-apoptotic CD40 activation to sustain durable responses., Conclusion: The combined data implicate Tregs as a potential therapeutic target in DLBCL, especially in combination with other immunotherapies., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Catharanthus roseus is the sole resource of vinblastine and vincristine, two TIAs of great interest for their powerful anticancer activities. Increasing the concentration of these alkaloids in various organs of the plant is one of the important goals in C. roseus breeding programs. Plant probiotic bacteria (PBB) act as biotic elicitors and can induce the synthesis of secondary products in plants. The purpose of this research is to study the effects of PBB on expression of the TIA biosynthetic pathway genes and the content of alkaloids in C. roseus., Methods and Results: The individual and combined effects of P. fluorescens strains 169 and A. brasilense strains Ab-101 was studied for expression of the TIA biosynthetic pathway genes (G10H, DAT, T16H and CrPRX) using qRT-PCR and the content of vinblastine and vincristine using HPLC method in roots of C. roseus. P. fluorescens. This drastically increased the content of vinblastine and vincristine alkaloids, compared to the control in the roots, to 174 and 589 (µg/g), respectively. Molecular analysis showed bacterium significantly increased the expression of more genes in the TIA biosynthetic pathway compared to the control. P. fluorescens increased the expression of the final gene of the biosynthetic pathway (CrPRX) 47.9 times compared to the control. Our findings indicate the correlation between transcriptional and metabolic outcomes. The same was true for A. brasilense., Conclusions: It can be concluded that seed treatments and seedling root treatments composed of naturally occurring probiotic bacteria are likely to be widely applicable for inducing enhanced alkaloid contents in medicinal plants., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Underlying mechanism of nanostructures upon monoterpene induction in Catharanthus roseus has not been explored yet. In the current study, Copper, Iron and Zinc nanoparticles were biosynthesized by Eriobotrya japonica seed extract and capped with reduced glutathione. Biosynthesized nanoparticles and their capped analogues were characterized by UV-visible spectrophotometer, FTIR, XRD and SEM. Selected concentration of nanostructures were used in plant tissue culture media which instigated the production of alkaloids, tannins and flavonoids without significantly affecting the growth index of propagated calli and shoots cultures of C. roseus. Accelerated vincristine production was noticed in propagated calli and shoots under copper and zinc nanostress (1645-1865 μg/ml respectively) with the least effect by iron nanostructure. Highest concentration of calcium was recorded in in vitro shoots under capped (3.42 mg/ml ± 7.16) and uncapped (4.41 mg/ml ± 20.44) Zn nanoparticles compared to control (2.82 mg/ml ± 13.41). Real time PCR depicts nano-zinc mediated increased expression of DAT and PRX1 genes of TIA pathway. Significant correlation among PRX1/DAT gene expression with vincristine production and calcium accumulation in the presence of nanostress validate by PCA. This study paved way the opportunities of metal biogenic nanomaterials as an ideal drug modulator in plant tissue culture studies., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Microtubule targeting agents (MTAs) are widely used cancer chemotherapeutics which conventionally exert their effects during mitosis, leading to mitotic or postmitotic death. However, accumulating evidence suggests that MTAs can also generate death signals during interphase, which may represent a key mechanism in the clinical setting. We reported previously that vincristine and other microtubule destabilizers induce death not only in M phase but also in G1 phase in primary acute lymphoblastic leukemia cells. Here, we sought to investigate and compare the pathways responsible for phase-specific cell death. Primary acute lymphoblastic leukemia cells were subjected to centrifugal elutriation, and cell populations enriched in G1 phase (97%) or G2/M phases (80%) were obtained and treated with vincristine. We found death of M phase cells was associated with established features of mitochondrial-mediated apoptosis, including Bax activation, loss of mitochondrial transmembrane potential, caspase-3 activation, and nucleosomal DNA fragmentation. In contrast, death of G1 phase cells was not associated with pronounced Bax or caspase-3 activation but was associated with loss of mitochondrial transmembrane potential, parylation, nuclear translocation of apoptosis-inducing factor and endonuclease G, and supra-nucleosomal DNA fragmentation, which was enhanced by inhibition of autophagy. The results indicate that microtubule depolymerization induces distinct cell death pathways depending on during which phase of the cell cycle microtubule perturbation occurs. The observation that a specific type of drug can enter a single cell type and induce two different modes of death is novel and intriguing. These findings provide a basis for advancing knowledge of clinical mechanisms of MTAs., Competing Interests: Conflict of interest The authors declare they have no conflicts of interest with the contents of this article., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Vincristine (VCR) is an effective drug against acute lymphoblastic leukemia (ALL), many solid tumors, but not acute myeloid leukemia. It has been hypothesized that resistance of myeloblasts to VCR is related to myeloperoxidase (MPO) and production of hypochlorous acid (HOCl). We investigated the relationship between VCR degradation and MPO expression and serum HOCl concentrations in pediatric patients with ALL, lymphoma and solid tumors. We studied the sera from 43 children, of which 23 were newly diagnosed and as yet untreated cancer patients, 10 on chemotherapy and 10 healthy control subjects. Patients’ sera were incubated with VCR alone or in the presence of taurine (T) or acetaminophen (APAP) and post-incubation VCR and HOCL concentrations were measured. Significant correlations between serum MPO expression, HOCl concentrations and VCR degradation were seen. In the chemotherapy group, MPO-positive patients produced high levels of HOCl and reciprocally low post-incubation VCR levels. HOCl and VCR concentrations in this group were significantly different than other groups studied. Both APAP and T inhibited VCR degradation in the sera of the chemotherapy group but not to the same degree. The effects seen here were consistent for both ALL and the lymphoma/solid tumor cases. Our results indicate that HOCl can increase VCR degradation in vitro in the serum and this effect is significantly more pronounced in pediatric patients undergoing chemotherapy. [Copyright &y& Elsevier]

SARM1 is a toll/interleukin-1 receptor -domain containing protein, with roles proposed in both innate immunity and neuronal degeneration. Murine SARM1 has been reported to regulate the transcription of chemokines in both neurons and macrophages; however, the extent to which SARM1 contributes to transcription regulation remains to be fully understood. Here, we identify differential gene expression in bone-marrow-derived macrophages (BMDMs) from C57BL/6 congenic 129 ES cell-derived Sarm1 -/- mice compared with wild type (WT). However, we found that passenger genes, which are derived from the 129 donor strain of mice that flank the Sarm1 locus, confound interpretation of the results, since many of the identified differentially regulated genes come from this region. To re-examine the transcriptional role of SARM1 in the absence of passenger genes, here we generated three Sarm1 -/- mice using CRISPR/Cas9. Treatment of neurons from these mice with vincristine, a chemotherapeutic drug causing axonal degeneration, confirmed SARM1's function in that process; however, these mice also showed that lack of SARM1 has no impact on transcription of genes previously shown to be affected such as chemokines. To gain further insight into SARM1 function, we generated an epitope-tagged SARM1 mouse. In these mice, we observed high SARM1 protein expression in the brain and brainstem and lower but detectable levels in macrophages. Overall, the generation of these SARM1 knockout and epitope-tagged mice has clarified that SARM1 is expressed in mouse macrophages yet has no general role in macrophage transcriptional regulation and has provided important new models to further explore SARM1 function., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Chemotherapy for non-Hodgkin lymphoma (NHL) in the hemodialysis (HD) patient is a challenging situation. Because many drugs are predominantly eliminated by the kidneys, chemotherapy in the HD patient requires special considerations concerning dose adjustments to avoid overdose and toxicities. Conversely, some drugs are removed by HD and may expose the patient to undertreatment, therefore the timing of drug administration in relation to HD sessions must be carefully planned. Also, the metabolites of some drugs show different toxicities and dialysability as compared with the parent drug, therefore this must also be catered for. However, the pharmacokinetics of many chemotherapeutics and their metabolites in HD patients are unknown, and the fact that NHL patients are often treated with distinct multiagent chemotherapy regimens makes the situation more complicated. In a realm where uncertainty prevails, case reports and case series reporting on actual treatment and outcomes are extremely valuable and can aid physicians in decision making from drug selection to dosing. We carried out an exhaustive review of the literature and adopted 48 manuscripts consisting of 66 HD patients undergoing 71 chemotherapy regimens for NHL, summarized the data, and provide recommendations concerning dose adjustments and timing of administration for individual chemotherapeutics where possible. The chemotherapy regimens studied in this review include, but are not limited to, rituximab, cyclophosphamide + vincristine + prednisolone (CVP) and cyclophosphamide + doxorubicin + vincristine + prednisolone (CHOP)-like regimens, chlorambucil, ibrutinib, bendamustine, methotrexate, platinum compounds, cytarabine, gemcitabine, etoposide, ifosfamide, melphalan, busulfan, fludarabine, mogamulizumab, brentuximab vedotin, and 90 Y-ibritumomab tiuxetan., (© 2021 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Vincristine (VCR) is widely used in cancer therapies, although its benefits on cardiac fibrosis remain unknown. Here, we investigated VCR's efficacy on cardiac fibrosis and elucidated the underlying mechanism of action. Network pharmacology was employed to predict the mechanism of VCR action on cardiac fibrosis. We induced cardiac fibrosis in adult male Sprague-Dawley (SD) rats via isoproterenol (ISO) injection, followed by treatment with VCR or vehicle. After 10 days of treatment, VCR-treated rats exhibited a significantly lower heart/body weight ratio relative to those treated with the vehicle. Moreover, cardiac fibrosis was alleviated in VCR-treated rats relative to vehicle-treated rats. The results revealed the down-regulation of mature caspase-1, interleukin (IL)-1β, and IL-18 in VCR-treated rats relative to vehicle-treated rats. We also observed less colocalization between the nucleotide-binding domain, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3) and apoptosis-associated speck-like protein containing a CARD (ASC) in VCR-treated rats compared with vehicle-treated rats. We then cultured neonatal rat cardiac fibroblasts (NRCFs) and exposed them to lipopolysaccharide (LPS) and adenosine triphosphate (ATP) in the presence or absence of VCR. The results indicated that VCR mediated the down-regulation of caspase-1, IL-1β, and IL-18 and the colocalization of NLRP3 and ASC in LPS+ATP-stimulated cardiac fibroblasts (CFs). We found evidence that VCR attenuates cardiac fibrosis by directly suppressing the activation of the NLRP3 inflammasome. These findings provide novel insights into VCR's mechanism of action in alleviating cardiac fibrosis., (© 2021 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Background: Although R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) remains the standard chemotherapy regimen for diffuse large B cell lymphoma (DLBCL) patients, not all patients are responsive to the scheme, and there is no effective method to predict treatment response., Methods: We utilized 5hmC-Seal to generate genome-wide 5hmC profiles in plasma cell-free DNA (cfDNA) from 86 DLBCL patients before they received R-CHOP chemotherapy. To investigate the correlation between 5hmC modifications and curative effectiveness, we separated patients into training (n = 56) and validation (n = 30) cohorts and developed a 5hmC-based logistic regression model from the training cohort to predict the treatment response in the validation cohort., Results: In this study, we identified thirteen 5hmC markers associated with treatment response. The prediction performance of the logistic regression model, achieving 0.82 sensitivity and 0.75 specificity (AUC = 0.78), was superior to existing clinical indicators, such as LDH and stage., Conclusions: Our findings suggest that the 5hmC modifications in cfDNA at the time before R-CHOP treatment are associated with treatment response and that 5hmC-Seal may potentially serve as a clinical-applicable, minimally invasive approach to predict R-CHOP treatment response for DLBCL patients.

ABCB1 detoxifies cells by exporting diverse xenobiotic compounds, thereby limiting drug disposition and contributing to multidrug resistance in cancer cells. Multiple small-molecule inhibitors and inhibitory antibodies have been developed for therapeutic applications, but the structural basis of their activity is insufficiently understood. We determined cryo-EM structures of nanodisc-reconstituted, human ABCB1 in complex with the Fab fragment of the inhibitory, monoclonal antibody MRK16 and bound to a substrate (the antitumor drug vincristine) or to the potent inhibitors elacridar, tariquidar, or zosuquidar. We found that inhibitors bound in pairs, with one molecule lodged in the central drug-binding pocket and a second extending into a phenylalanine-rich cavity that we termed the "access tunnel." This finding explains how inhibitors can act as substrates at low concentration, but interfere with the early steps of the peristaltic extrusion mechanism at higher concentration. Our structural data will also help the development of more potent and selective ABCB1 inhibitors., Competing Interests: The authors declare no competing interest.

Retinoblastoma (Rb) is the most common primary malignant intraocular tumor in children which develops from the retinal stem cells. Systemic chemotherapy is the typical therapeutic treatment and though most children survive Rb, they often lose their vision, or the eye needs to be enucleated. Regarding to the pure availability of the target tumor by systemic chemotherapy, the local anticancer drug administration would be advantageous to increase the local drug concentration and minimize adverse side effects of chemotherapy. The present paper describes a new hydrogel implant enabled to deliver therapeutically active doses of low molecular weight hydrophilic antitumor drugs topotecan and vincristine. The hydrogel implant is proposed as bi-layered with an inner hydrophilic layer from 2-hydroxyethyl methacrylate (HEMA) serving as a reservoir of the chemotherapeutic agent and an outer hydrophobic layer from 2-ethoxyethyl methacrylate (EOEMA) acting as a barrier to protect the surrounding vascularized tissue against cytotoxicity of the delivered chemotherapeutics. The experiments with enucleated pig eyes demonstrated the ability of tested drugs to diffuse through sclera and reach the vitreous humor. HEMA-based hydrogels were examined in terms of sorption, release and transport properties, showing the possibility of adjusting the loading capacity and diffusion of the drugs by the degree of crosslinking. The EOEMA-based gels proved to be an inert for drug sorption and diffusion. A chorioallantoic membrane assay demonstrated excellent biocompatibility of unloaded hydrogels, and in vitro experiments confirmed significant cytotoxicity of drug-loaded hydrogels against a Rb cell line; 2 days for those topotecan-loaded and a minimum of 6 days for vincristine-loaded hydrogels. The bi-layered hydrogel implant can be considered promising for local administration of active agents to eye-globe for the treatment of Rb and also other ocular disorders., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objective: Spreading depolarization (SD) is a transient self-propagating wave of neuronal and glial depolarization coupled with large membrane ionic changes and a subsequent depression of neuronal activity. Spreading depolarization in the cortex is implicated in migraine, stroke, and epilepsy. Conversely, spreading depolarization in the striatum, a brain structure deeply involved in motor control and in Parkinson's disease (PD) pathophysiology, has been poorly investigated., Methods: We characterized the participation of glutamatergic and dopaminergic transmission in the induction of striatal spreading depolarization by using a novel approach combining optical imaging, measurements of endogenous DA levels, and pharmacological and molecular analyses., Results: We found that striatal spreading depolarization requires the concomitant activation of D1-like DA and N-methyl-d-aspartate receptors, and it is reduced in experimental PD. Chronic l-dopa treatment, inducing dyskinesia in the parkinsonian condition, increases the occurrence and speed of propagation of striatal spreading depolarization, which has a direct impact on one of the signaling pathways downstream from the activation of D1 receptors., Conclusion: Striatal spreading depolarization might contribute to abnormal basal ganglia activity in the dyskinetic condition and represents a possible therapeutic target. © 2019 International Parkinson and Movement Disorder Society., (© 2019 International Parkinson and Movement Disorder Society.)

In this study, it was aimed to determine the effects of Amlodipine, a calcium channel blocker and vincristine (VCR) an antineoplastic, on human neuroblastomas using different doses. The cytotoxicity assays of the study were performed using the MTT method depending on time and concentration. After obtaining the mixture (up to 85% for SH-SY5Y) and sufficient branches (cortex neurons), the cells were treated with amlodipine (10 µM) and vincristine (0.5, 1 and 2 µg) at different concentrations for 24 h. MTT assay was performed by the commercially available kit (Sigma Aldrich, USA). Cells were harvested, washed and stained with PI and Annexin V, respectively, according to the manufacturer's protocol (Biovision, USA). Than analyzes were carried out. The results were quite impressive. When amlodipine (10 µM) was administered alone there was little change compared to the control. However, all doses of amlodipine (10 µM) and vincristine (0.5, 1 and 2 µg) were greater than the deaths in the doses alone (0.5, 1 and 2 µg) of vincristine alone. (P < 0.05). As a result, the combination of vincristine and amlodipine is more effective than vincristine alone in reducing the viability of cancer cells.

The important anticancer drugs, vinblastine, vincristine and analogs, are composed of the monoterpenoid indole alkaloids (MIAs), catharanthine and vindoline, found uniquely in the medicinal plant, Catharanthus roseus. While 26 genes involved in the assembly of these two MIAs are known, two key reactions have eluded characterization to complete the documentation of the vinblastine pathway in this plant species. The assembly of these dimeric MIAs requires O-acetylstemmadenine oxidase (ASO) and a dual function geissoschizine synthase (GS) that reduces cathenamine to form geissoschizine, and that also reduces the ASO product to form a common intermediate for subsequent conversion by four separate hydrolases to catharanthine, tabersonine or vincadifformine, respectively. The in planta role of ASO is supported by identifying a single amino acid-substituted ASO mutant with very low enzyme activity and by virus-induced gene silencing of ASO to produce plants that accumulate O-acetylstemmadenine rather than catharanthine and vindoline found in wild-type (WT) plants. The in planta role of GS is supported by showing that a low GS-expressing mutant accumulating lower levels of catharanthine and vindoline also displays significantly lower tabersonine-forming activity in coupled enzyme assays than in the WT background. Gene expression analyses demonstrate that both ASO and GS are highly enriched in the leaf epidermis where the pathways for catharanthine and tabersonine biosynthesis are expressed. The full elucidation of this canonical pathway enables synthetic biology approaches for manufacturing a broad range of MIAs, including these dimers used in cancer treatment., (© 2018 The Authors The Plant Journal © 2018 John Wiley & Sons Ltd.)

P-glycoprotein affects the transport of numerous drugs including chemotherapeutic drugs vincristine sulfate (VCR) and docetaxel (DTX), and is one of the main causes for multidrug resistance. Our previous studies have shown that oxypeucedanin (OPD) can enhance the intestinal transit of puerarin and VCR. However, the underlying mechanism is unclear. This study investigated the potential mechanism by which OPD improves P-gp-mediated drug transport. Molecular docking was performed to predict the binding force between OPD and P-gp and the contribution of OPD on P-gp activity. We observed the effect of OPD on the transport of VCR in MDCK-MDR1 cell monolayer and also measured the plasma pharmacokinetic parameters of DTX in the presence and absence of OPD by LC-MS/MS. Moreover, we further investigated the reversal mechanism of OPD on P-gp-mediated drug transport by determining the intracellular accumulation of Rhodamine-123 (Rh123) and P-gp ATPase activity as well as protein expression and mRNA level of P-gp. Our molecular docking results revealed that the binding force between OPD and P-gp was much lower than that between P-gp and verapamil (a P-gp substrate). The transport study in vitro indicated that OPD increased the flux of VCR across MDCK-MDR1 cell monolayer. The in vivo pharmacokinetic parameters data showed OPD increased the absorption of DTX. OPD activated P-gp ATPase activity and enhanced intracellular accumulation of Rh123 in MDCK-MDR1 cells. Western blotting and qRT-PCR outcomes indicated that OPD suppressed P-gp protein expression as well as downregulated P-gp mRNA level. Thus, OPD reverse P-gp-mediated drug transport via inhibition of P-gp activity and P-gp protein expression as well as downregulation of P-gp mRNA level. Our results suggest that OPD could reverse P-gp-mediated drug resistance in tumor cells.

Ethnopharmacological Importance: Catharanthus roseus (L.) G. Don. is an important medicinal plant with rich sources of remarkable health benefits consisting more than 100 alkaloids and significant amounts of bioactive compounds, which have been widely used as a folk medicine for treatment of several pathologies., The Aim of the Study: In the present study, we isolated and cultured innately undifferentiated cambium meristematic cells (CMCs), which were observed stable cell growth, enhancement of bioactive compounds from C.roseus., Materials and Methods: We attempted to determine the effect of association between time-course growth rates, bioactive compounds and terpenoids indole alkaloid (TIA) contents as well as antioxidant and anticancer efficacies of C. roseus CMC suspension culture treated by UV-C., Results: The bioactive compounds, vincristine contents, and antioxidant power were noticed significantly higher in 60 min exposure at 5 cm distances and with the directly collected sample (T7). A similar trend has also been noticed from the anticancer activity. Demonstration of TIA accumulation was found higher at 5 min exposure, at 20 cm distances and 48 h of incubation (T21) and the result of TIA contents had the highest correlation effects of anticancer activities., Conclusion: In the current study, we demonstrated that UV-C light could enhance the production of the essential compounds and bioactivities in the CMCs of C. roseus, and thus, C. roseus CMCs have the potential to serve as an industrial platform for the production of bioactive alkaloids and antioxidant, anticancer activity. Moreover, additional efforts should be made to irradiate CMC suspension cultures from C. roseus with UV-C to achieve better pharmacological profiles., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The medicinal plant Catharanthus roseus accumulates large numbers of terpenoid indole alkaloids (TIAs), including the pharmaceutically important vinblastine, vincristine, ajmalicine, and serpentine. The phytohormone ethylene or methyl jasmonate (MeJA) can markedly enhance alkaloid accumulation. The interaction between ethylene or MeJA in the regulation of TIA biosynthesis in C. roseus is unknown. Here, a metabolomics platform is reported that is based on liquid chromatography (LC) coupled with time-of-flight mass spectrometry to study candidate components for TIA biosynthesis, which is controlled by ethylene or MeJA in C. roseus. Multivariate analysis identified 16 potential metabolites mostly associated with TIA metabolic pathways and seven targeted metabolites, outlining the TIA biosynthesis metabolic networks controlled by ethylene or MeJA. Interestingly, ethylene and MeJA regulate the 2-C-methyl-d-erythritol 4-phosphate (MEP) and acetate-mevalonate (MVA) pathways through AACT and HMGS and through DXS, respectively, to induce TIA biosynthesis in C. roseus. Overall, both nontargeted and targeted metabolomics, as well as transcript analysis, were used to reveal that MeJA and ethylene control different metabolic networks to induce TIA biosynthesis.

Background: Uveal melanoma is the most common primary intraocular malignancy in adults. So far, there have been no effective targeted therapeutic agents in patients with uveal melanoma. Artesunate is a semi-synthetic derivative of artemisinin extracted from traditional Chinese medicine Artemisia annua L for treatment of severe and multidrug-resistant malaria. Besides its antimalarial activity, artesunate is identified as an anti-cancer drug due to the inhibition of Wnt/β- catenin pathway in multiple types of cancer. However, the effect of artesunate on uveal melanoma remains unknown., Objective: We evaluated the anti-tumor effects of artesunate on uveal melanoma cells, and analyzed in terms of Wnt/β-catenin pathway, cell growth, cell death, cell migration, invasion and cancer stemlike cells (CSCs) properties., Methods: Primary (92.1, Mel270) and metastatic (Omm1 and Omm2.3) uveal melanoma cells were used. Immunofluorescence staining, dual luciferase reporter assay, Western blotting, MTS, soft agar cloning technique, Annexin V/PI analyses, wound healing scratch assay, in vitro transwell migration and invasion assays, aldehyde dehydrogenase (ALDH) analyses and melanosphere formation assay et al. were carried out., Results: Artesunate suppressed the phosphorylation of GSK3β at S9, and lowered the protein level of β-catenin and its downstream targets (c-Myc, cyclin D1). Artesunate potently inhibited cell viability and colony formation ability. Treatment with artesunate significantly induced apoptosis. In addition, artesunate significantly reduced the migration and invasion of uveal melanoma cells, impaired the traits of CSCs in vitro., Conclusion: Artesunate may be a potential interest for the therapy of uveal melanoma., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Nowadays, lung cancer, as a health problem in worldwide, has high mortality both in men and women. Despite advances in diagnosis and surgical techniques of lung cancer in recent decades, chemotherapy is still a fundamentally and extensively useful strategy. Vinca alkaloids are a class of important and widely used drugs in the treatment of lung cancer, targeting on the Vinca binding site at the exterior of microtubule plus ends. Either intrinsic or acquired resistance to chemotherapy of Vinca alkaloids has been a major obstacle to the treatment of lung cancer, which arose great interests in studies of understanding and overcoming resistance. In this review, we focused on the application and resistance mechanisms of the Vinca alkaloids such as vinblastine, vincristine, vinorelbine and vinflunine in lung cancer. We reviewed characteristic resistance mechanisms in lung cancer including over-expression of ATP-binding cassette (ABC) transporters P-glycoprotein and structural, functional or expression alterations of β-tubulin (βII, βIII, βIV) which may devote to the development of acquired resistance to the Vinca alkaloids; multidrug-resistance proteins (MRP1, MRP2, MRP3) and RLIP76 protein have also been identified that probably play a significant role in intrinsic resistance. Lung resistance-related protein (LRP) is contributed to lung cancer therapy resistance, but is not deal with the Vinca alkaloids resistance in lung cancer. Understanding the principle of the Vinca alkaloids in clinical application and mechanisms of drug resistance will support individualized lung cancer therapy and improve future therapies., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Purpose: Vincristine is known to interfere with OATP-mediated uptake of other compounds, hinting that vincristine itself could be a substrate of OATP transporters. The present study therefore aimed to investigate the role of OATP transporters in the hepatocellular disposition of vincristine., Methods: Vincristine uptake was studied in suspended rat and human hepatocytes as well as OATP-transfected Chinese hamster ovary (CHO) cells in the absence and presence of OATP transporter inhibitors. Membrane vesicles containing MDR1 or MRP1/2/3 were used to directly assess the role of these efflux transporters in vincristine disposition., Results: Uptake in suspended rat hepatocytes was temperature-dependent and could be inhibited by a range of OATP inhibitors. Furthermore, the MRP-inhibitor benzbromarone, but none of the tested MDR1 inhibitors, reduced vincristine efflux in rat and human suspended hepatocytes. OATP1B1-, OATP1B3- and OATP2B1- transfected CHO cells showed significantly increased vincristine uptake as compared to wild-type cells. Moreover, uptake in OATP-transfected CHO cells was reduced by OATP inhibitors. However, uptake studies in suspended human hepatocytes showed that only 10% of the total vincristine uptake process could be attributed to OATP-mediated transport. Studies with transporter-expressing membrane vesicles confirmed vincristine as an MDR1 substrate, while MRP1/2/3-mediated transport of vincristine could not be observed with this model system., Conclusions: Our findings show the involvement of OATP transporters in the disposition of vincristine in rat and human hepatocytes. However, in both species, hepatic uptake is overshadowed by a benzbromarone-sensitive efflux mechanism, possibly MRP3.

The goal of this research is to structure a hyaluronic acid modified nanostructured lipid carrier (HA-NLCs) for vincristine sulfate (VCR) delivery, and detect its efficiency to improve the oral bioavailability. Emulsion solvent evaporation method was used to prepare the HA-NLCs nanoparticles. The particle size, zeta potential and entrapment efficiency of VCR-NLCs and HA-NLCs were 187 ± 3.52 nm, -8.61 ± 1.29 mV, 33.12 ± 1.16% and 192 ± 4.41 nm, -32.82 ± 2.64 mV, 34.41 ± 2.21%, respectively. HA-NLCs could significantly improve the cellular uptake efficiency and cytotoxicity in MCF-7 cells than other VCR formulations. The expressions of apoptosis related protein Caspase-3, Caspase-9, Bax and Bcl-2 were estimated by western blot assay in MCF-7 cells, and HA-NLCs exhibited the strongest effect in promoting cell apoptosis. The pharmacokinetics of HA-NLCs was evaluated in Sprague-Dawley male rats and the relative oral bioavailability of HA-NLCs and VCR-NLCs was improved about 1.8-fold and two-fold compared with VCR solution, respectively. Therefore, these results indicated that HA-NLCs could significantly improve the oral bioavailability and was a promising vehicle for the oral delivery of VCR.

P-glycoprotein (P-gp) affects the transport of many drugs; including puerarin and vincristine. Our previous study demonstrated that imperatorin increased the intestinal absorption of puerarin and vincristine by inhibiting P-gp-mediated drug efflux. However; the underlying mechanism was not known. The present study investigated the mechanism by which imperatorin promotes P-gp-mediated drug transport. We used molecular docking to predict the binding force between imperatorin and P-gp and the effect of imperatorin on P-gp activity. P-gp efflux activity and P-gp ATPase activity were measured using a rhodamine 123 (Rh-123) accumulation assay and a Pgp-Glo™ assay; respectively. The fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH) was used to assess cellular membrane fluidity in MDCK-MDR1 cells. Western blotting was used to analyze the effect of imperatorin on P-gp expression; and P-gp mRNA levels were assessed by qRT-PCR. Molecular docking results demonstrated that the binding force between imperatorin and P-gp was much weaker than the force between P-gp and verapamil (a P-gp substrate). Imperatorin activated P-gp ATPase activity; which had a role in the inhibition of P-gp activity. Imperatorin promoted Rh-123 accumulation in MDCK-MDR1 cells and decreased cellular membrane fluidity. Western blotting demonstrated that imperatorin inhibited P-gp expression; and qRT-PCR revealed that imperatorin down-regulated P-gp (MDR1) gene expression. Imperatorin decreased P-gp-mediated drug efflux by inhibiting P-gp activity and the expression of P-gp mRNA and protein. Our results suggest that imperatorin could down-regulate P-gp expression to overcome multidrug resistance in tumors.

Purpose: Lymph cancers are heterogeneous malignancies of the hematopoietic and lymphoid tissues. Doxorubicin (DOX) and vincristine (VCR) are commonly used anti-cancer chemotherapeutic drugs, but their clinical uses are associated with dose-limiting systemic toxicity., Methods: In the present study, DOX and VCR were encapsulated into nanostructured lipid carriers (NLCs) and used them to treat B-cell lymphoma cells through the targeted delivery of DOX and VCR to lymph cancer animal model., Results: DOX and VCR encapsulated NLCs (DOX/VCR NLCs) demonstrated controlled drug release under physiological conditions. In addition, DOX/VCR NLCs exhibited the highest cytotoxicity and synergistic effect of two drugs in B-cell lymphoma cells and the best anti-tumor effect in vivo., Conclusion: DOX/VCR NLCs were proved to be more efficacious than the equivalent dose of free DOX and single drug (DOX or VCR) formulation in vitro and in vivo, and significantly reduced the drug-associated systemic toxicity.

Sustained release drug delivery systems remain a major clinical need for small molecule therapeutics in oncology. Here, mechanisms of small molecule interactions with silk protein films were studied with cationic oncology drugs, vincristine and doxorubicin, with a focus on hydrophobicity (non-ionic surfactant) and charge (pH and ionic strength). Interactions were primarily driven by charge interactions between the positively charged drugs and the negatively charged groups within the silk films. Exploiting chemical modifications of silk further modulated the drug interactions in a controlled fashion. Increasing anionic side groups via carboxylate- and sulfonate-modifications of tyrosine side chains in the silk protein using diazonium coupling chemistry, increased drug binding and altered drug release. The effects of silk film protein crystallinity, beta sheet content, on drug binding and release were also explored. Lower crystallinity supported more rapid drug binding when compared to higher crystalline silk films. The drug release kinetics were governed by the protonation state of vincristine and doxorubicin and were tunable based on silk crystallinity and chemistry. These studies depict an approach to characterize small molecule-silk protein interactions and methods to tune drug binding and release kinetics from this protein delivery matrix., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Endophytic fungi isolated from Catharanthus roseus were screened for the production of vincristine and vinblastine. Twenty-two endophytic fungi isolated from various tissues of C. roseus were characterized taxonomically by sequence analysis of the internal transcribed spacer (ITS) region of rDNA and grouped into 10 genera: Alternaria, Aspergillus, Chaetomium, Colletotrichum, Dothideomycetes, Eutypella, Eutypa, Flavodon, Fusarium and Talaromyces. The antiproliferative activity of these fungi was assayed in HeLa cells using the MTT assay. The fungal isolates Eutypella sp--CrP14, obtained from stem tissues, and Talaromyces radicus--CrP20, obtained from leaf tissues, showed the strongest antiproliferative activity, with IC50 values of 13.5 μg/ml and 20 μg/ml, respectively. All 22 endophytic fungi were screened for the presence of the gene encoding tryptophan decarboxylase (TDC), the key enzyme in the terpenoid indole alkaloid biosynthetic pathway, though this gene could only be amplified from T. radicus--CrP20 (NCBI GenBank accession number KC920846). The production of vincristine and vinblastine by T. radicus--CrP20 was confirmed and optimized in nine different liquid media. Good yields of vincristine (670 μg/l) in modified M2 medium and of vinblastine (70 μg/l) in potato dextrose broth medium were obtained. The cytotoxic activity of partially purified fungal vincristine was evaluated in different human cancer cell lines, with HeLa cells showing maximum susceptibility. The apoptosis-inducing activity of vincristine derived from this fungus was established through cell cycle analysis, loss of mitochondrial membrane potential and DNA fragmentation patterns.

Background: The aim of the current work was to perform a clinical trial simulation (CTS) analysis to optimize a drug-drug interaction (DDI) study of vincristine in children who also received azole antifungals, taking into account challenges of conducting clinical trials in this population, and, to provide a motivating example of the application of CTS in the design of pediatric oncology clinical trials., Procedure: A pharmacokinetic (PK) model for vincristine in children was used to simulate concentration-time profiles. A continuous model for body surface area versus age was defined based on pediatric growth curves. Informative sampling time windows were derived using D-optimal design. The CTS framework was used to different magnitudes of clearance inhibition (10%, 25%, or 40%), sample size (30-500), the impact of missing samples or sampling occasions, and the age distribution, on the power to detect a significant inhibition effect, and in addition, the relative estimation error (REE) of the interaction effect., Results: A minimum group specific sample size of 38 patients with a total sample size of 150 patients was required to detect a clearance inhibition effect of 40% with 80% power, while in the case of a lower effect of clearance inhibition, a substantially larger sample size was required. However, for the majority of re-estimated drug effects, the inhibition effect could be estimated precisely (REE < 25%) in even smaller sample sizes and with lower effect sizes., Conclusion: This work demonstrated the utility of CTS for the evaluation of PK clinical trial designs in the pediatric oncology population., (© 2014 Wiley Periodicals, Inc.)

Background: A frequent mechanism of acquired multidrug resistance in human cancers is overexpression of ATP-binding cassette transporters such as the Multi-Drug Resistance Protein 1 (MDR-1). Nutlin-3, an MDM2-p53 antagonist, has previously been reported to be a competitive MDR-1 inhibitor., Methods: This study assessed whether the structurally diverse MDM2-p53 antagonists, MI-63, NDD0005, and RG7388 are also able to modulate MDR-1 function, particularly in p53 mutant neuroblastoma cells, using XTT-based cell viability assays, western blotting, and liquid chromatography-mass spectrometry analysis., Results: Verapamil and the MDM2-p53 antagonists potentiated vincristine-mediated growth inhibition in a concentration-dependent manner when used in combination with high MDR-1-expressing p53 mutant neuroblastoma cell lines at concentrations that did not affect the viability of cells when given alone. Liquid chromatography-mass spectrometry analyses showed that verapamil, Nutlin-3, MI-63 and NDD0005, but not RG7388, led to increased intracellular levels of vincristine in high MDR-1-expressing cell lines., Conclusions: These results show that in addition to Nutlin-3, other structurally unrelated MDM2-p53 antagonists can also act as MDR-1 inhibitors and reverse MDR-1-mediated multidrug resistance in neuroblastoma cell lines in a p53-independent manner. These findings are important for future clinical trial design with MDM2-p53 antagonists when used in combination with agents that are MDR-1 substrates.

Background: The ATP-binding cassette (ABC) transporters P-glycoprotein (P-gp)/ABCB1 and breast cancer resistance protein (BCRP)/ABCG2 are involved in the intestinal absorption and renal excretion of various substrate drugs. Their activities affect sub-therapeutic drug concentrations and excretion of natural transporter substrates. The new oral anti-HCV drug telaprevir has dramatically improved the efficacy of hepatitis-C virus (HCV) treatment, and recent studies have suggested a possible pharmacological interaction between telaprevir and P-gp. We studied the kinetics of in vitro interactions between telaprevir and P-gp and BCRP to understand the molecular basis of that interaction., Findings: The effect of telaprevir on P-gp- and BCRP-mediated transport was evaluated by an in vitro vesicle transporter assay using different transport substrates, and the kinetics of transporter inhibition was determined. The results showed that telaprevir could inhibit P-gp- and BCRP-mediated transport in the in vitro vesicle transport assay, with each IC50 values of ≈ 7 μmol/L and ≈ 30 μmol/L, respectively. Analyses of Lineweaver-Burk plots showed that telaprevir was likely to be a competitive inhibitor against P-gp and BCRP. Photoaffinity labeling experiments were employed to observe competitive inhibition by telaprevir using iodoarylazidoprazosin (IAAP) as a binding substrate for P-gp and BCRP. These experiments revealed that telaprevir inhibited [125I]-IAAP-binding with P-gp and BCRP., Conclusion: Telaprevir competitively inhibited P-gp and BCRP, and P-gp-mediated transport was more sensitive to telaprevir compared with BCRP-mediated transport. These data suggest that telaprevir represses the transporter functions of P-gp and BCRP via direct inhibition.

Understanding the interaction of anticancer drugs with model cell lines is important to elucidate the mode of action of these drugs as well as to develop cost effective and rapid screening methods. Raman spectroscopy has been demonstrated to be a valuable technique for high throughput, noninvasive analysis. The interaction of vincristine with a human lung adenocarcinoma cell line (A549) was investigated using Raman micro spectroscopy. The results were correlated with parallel measurements from the MTT cytotoxicity assay, which yielded an IC50 value of 0.10 ± 0.03 μM. The Raman spectral data acquired from vincristine treated A549 cells was analysed to understand its interaction with the nucleus in the cell and elucidate DNA intercalation. The dose dependent spectral changes in the nucleus are analysed by PLS-Jack knifing for the identification of the more significant changes associated with the mode of action of the drug. Results are correlated with a similar dose dependent expression analysis of the bcl-2 protein, an anti-apoptotic protein associated with DNA damage, in the vincristine treated A549 cells using flow cytometry. The results indicate the co-existence of two modes of action, microtubule binding at low doses and DNA intercalation at high doses.

Concomitant administration of herbal medicines with drugs that are P-glycoprotein (P-gp) substrates may produce significant herb-drug interactions. The purpose of this study was to evaluate the effects of Schisandra lignans extract (SLE) on P-gp thoroughly in vitro and in vivo, and to investigate the possible P-gp-based herb-drug interactions. In the in vitro experiments, the effect of SLE on the uptake and transport for P-gp substrates in Caco-2, LLC-PK1 and L-MDR1 cells were carefully investigated. Verapamil, a known P-gp inhibitor, was used as a positive control drug. Results shown that, 10 μM verapamil and SLE (0.5, 2.0, and 10.0 μg/ml) were observed to significantly enhance the uptake and inhibit the efflux ratio of P-gp substrates in Caco-2 and L-MDR1 cells. In vivo experiments showed that single-dose SLE at 500 mg/kg could increase the area under the plasma concentration time curve of digoxin and vincrisine significantly without affecting terminal elimination half-time. Long-term treatment with SLE for continuous 10 days could also increase the absorption of P-gp substrates with greatly down regulation of P-gp expression in rat intestinal and brain tissues. In conclusion, SLE was a strong P-gp inhibitor, which indicated a potential herb-drug interaction when SLE was co-administered with P-gp substrate drugs., (Copyright © 2013. Published by Elsevier GmbH.)

The purpose of this study was to determine the impact of CYP3A5 expression on inhibitory potency (Ki or IC50 values) of CYP3A inhibitors, using recombinant CYP3A4 and CYP3A5 (rCYP3A4 and rCYP3A5) and CYP3A5 genotyped human liver microsomes (HLMs). IC50 ratios between rCYP3A4 and rCYP3A5 (rCYP3A5/rCYP3A4) of ketoconazole (KTZ) and itraconazole (ITZ) were 8.5 and 8.8 for midazolam (MDZ), 4.7 and 9.1 for testosterone (TST), 1.3 and 2.8 for terfenadine, and 0.6 and 1.7 for vincristine, respectively, suggesting substrate- and inhibitor-dependent selectivity of the two azoles. Due to the difference in the IC50 values for CYP3A4 and CYP3A5, nonconcordant expression of CYP3A4 and CYP3A5 protein can significantly affect the observed magnitude of CYP3A-mediated drug-drug interactions in humans. Indeed, the IC50 values of KTZ and ITZ for CYP3A-catalyzed MDZ and TST metabolism were significantly higher in HLMs with CYP3A5*1/*1 and CYP3A5*1/*3 genotypes than in HLMs with the CYP3A5*3/*3 genotype, showing CYP3A5 expression-dependent IC50 values. Moreover, when IC50 values of KTZ and ITZ for different HLMs were kinetically simulated based on CYP3A5 expression level and enzyme-specific IC50 values, a good correlation between the simulated and the experimentally measured IC50 values was observed. Further simulation analysis revealed that both the Ki ratio (for inhibitors) and Vmax/Km ratio (for substrates) between CYP3A4 and CYP3A5 were major factors for CYP3A5 expression-dependent IC50 values. In conclusion, the present study demonstrates that CYP3A5 genotype and expression level have a significant impact on inhibitory potency for CYP3A-catalyzed drug metabolism, but that the magnitude of its effect is inhibitor-substrate pair specific.

Salicylic acid (SA) has been reported to ameliorate various stresses in plants. In order to explore the role of SA under nickel (Ni) stress, thirty-days old plants of periwinkle (Catharanthus roseus L.) were supplied with eight treatments comprising basal application of Ni (0, 50, 100 and 150 mg kg(-1)) and foliar application of SA (0 and 10(-5)M) under net house conditions. Ni application significantly reduced the growth attributes including plant height, leaf-area index and fresh and dry weights of shoot and root. Increasing Ni concentration led to a gradual decrease in photosynthetic parameters and activities of nitrate reductase and carbonic anhydrase. The plants, undergoing Ni stress, exhibited a significant increase in the activity of superoxide dismutase, catalase and peroxidase together with an increase in electrolyte leakage and proline content. Total alkaloid content was also declined in Ni-treated plants. Foliar application of SA (10(-5)M) reduced the deleterious effects of Ni on plant growth, accelerating the restoration of growth processes. SA also improved the total alkaloid content under normal as well as adverse conditions. Foliar spray of SA significantly improved the content of anticancer alkaloids vincristine (by 22.2%) and vinblastine (by 50.0%) in plants treated with 150 mg kg(-1) of Ni., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The reactivation of latent human cytomegalovirus (HCMV) infection after transplantation is associated with high morbidity and mortality. In vivo, myeloid cells and their progenitors are an important site of HCMV latency, whose establishment and/or maintenance require expression of the viral transcript UL138. Using stable isotope labeling by amino acids in cell culture-based mass spectrometry, we found a dramatic UL138-mediated loss of cell surface multidrug resistance-associated protein-1 (MRP1) and the reduction of substrate export by this transporter. Latency-associated loss of MRP1 and accumulation of the cytotoxic drug vincristine, an MRP1 substrate, depleted virus from naturally latent CD14(+) and CD34(+) progenitors, all of which are in vivo sites of latency. The UL138-mediated loss of MRP1 provides a marker for detecting latent HCMV infection and a therapeutic target for eliminating latently infected cells before transplantation.

Background: P-glycoprotein (P-GP) and multidrug resistance-associated protein 2 (MRP2) are involved in transport of many drugs across blood-brain barrier (BBB). The function and expression of P-GP and MRP2 may be modulated by different pathologies. Acute liver failure (ALF) was reported to impair BBB function, resulting in the increased BBB permeability., Aims: We investigated whether ALF altered function and expression of P-GP and MRP2 in brain of thioacetamide-induced ALF rats., Methods: ALF was induced by intraperitoneal injection of thioacetamide (300 mg/kg) for 2 days with a 24-h interval. The rats were used for experiments at 6, 12 and 24 h after the second administration. P-GP and MRP2 function in brain were determined using the brain-to-plasma ratios of corresponding substrates (rhodamine 123 and vincristine for P-GP; sulfobromophthalein and dinitrophenyl-S-glutathione for MRP2). Evans blue was used for examining the BBB integrity. Western blot was accomplished to determine P-GP and MRP2 protein expression., Results: The brain-to-plasma ratios of rhodamine 123 and vincristine were significantly increased in ALF-6 h rats and almost returned to normal levels in ALF-24 h rats, whereas those of sulfobromophthalein and dinitrophenyl-S-glutathione were decreased in all ALF rats. Western blot results showed that ALF decreased brain P-GP levels at 6 and 12 h, whereas increased MRP2 levels at 6, 12 and 24 h. No significant difference of Evans blue concentrations in brain was found among the four groups., Conclusions: Function and expression of P-GP and MRP2 in brain of thioacetamide-induced ALF rats were oppositely altered., (© 2012 John Wiley & Sons A/S.)

We examined whether pretreatment of mouse brain blood vessel endothelial cell clone 4 (MBEC4) cells with sodium nitroprusside (SNP), a NO(x) donor, as an in vitro model of the bloodbrain barrier could affect P-glycoprotein (P-gp) functional activity. Uptake into the cells and MBEC4 plasma membrane vesicles (MPMVs) in the presence or absence of SNP pretreatment was used to investigate functional changes. Increased accumulation of [(3)H]vincristine, a widely used substrate for P-gp, into MBEC4 was observed upon SNP pretreatment, likely due to impaired P-gp function. To better understand the mechanism of the impairment, MPMVs were prepared and characterized in terms of purity and Na(+)-dependent glucose uptake. [(3)H]daunomycin uptake into MPMVs was diminished after SNP pretreatment in the presence of an ATP-regenerating system, indicating that the functional activity of P-gp was impaired after exposure to SNP. Under conditions of excess ATP, daunomycin uptake into the vesicles was still decreased after SNP pretreatment, indicating that SNP interacted directly with the transport system, but not with the ATP-regenerating system. Together, these results suggest that NO or NO(x) functionally impairs P-gp in the in vitro blood-brain barrier model with SNP pretreatment.

Objective: Under various drought conditions and nitrogen application, the content of vindoline, catharanthine, vincristine and vinblastine in the leaf of Catharanthus roseus were illustrated to improve the content of alkaloid theoretically., Method: Six groups were set in the experiment, which included: CK (natural control), CN (natural control + nitrogen), LK (low drought), LN (low drought + nitrogen), HK (high drought), HN (high drought + nitrogen) to discuss the change characteristics of total nitrogen, the activity of alkaline POD and TDC, the content of four alkaloids under the different conditions were measured., Result: Under LK condition, the activity of POD, TDC were enhanced. In the early stage of stress (0-21 d), vindoline, catharanthine, vincristine and vinblastine accumulated, and reduced in the later stage (28-35 d). For all groups, adding exogenous nitrogen could improve the total content of nitrogen, vindoline and vinblastine, meanwhile the activity of POD and TDC were enhanced as well. The LN, HN treatments were beneficial to accumulating catharanthine and vinblastine., Conclusion: Drought stress or additional nitrogen have an influence on both of the activities of POD and TDC, and the four alkaloids were affected as well. Thereinto, the LN condition was the most effective treatment for accumulating the four alkaloids (vindoline, catharanthine, vincristine and vinblastine), which were regulated by improve nitrogen content and enzymatic activity.

Previous studies showed that γ-linolenic acid (GLA, 18: 3 ω-6), arachidonic acid (AA, 20:4 ω-6), eicosapentaenoic acid (EPA, 20: 5 ω-3) and docosahexaenoic acid (DHA, 22:6 ω-3) have selective tumoricidal action. In the present study, it was observed that dihomo-gamma-linolenic acid (DGLA) and AA, EPA and DHA have cytotoxic action on both vincristine-sensitive (KB-3-1) and resistant (KB-Ch(R)-8-5) cancer cells in vitro that appeared to be a free-radical dependent process but not due to the formation of prostaglandins, leukotrienes and thromboxanes. Uptake of vincristine and fatty acids was higher while their efflux was lower in KB-3-1 cells compared with KB-Ch(R)-8-5 cells, suggesting that drug resistant cells have an effective efflux pump. GLA, DGLA, AA, EPA and DHA enhanced the uptake and decreased efflux in both drug-sensitive and drug-resistant cells and augmented the susceptibility of tumor cells especially, of drug-resistant cells to the cytotoxic action of vincristine. These results suggest that certain polyunsaturated fatty acids have tumoricidal action and are capable of enhancing the cytotoxic action of anti-cancer drugs specifically, on drug-resistant cells by enhancing drug uptake and reducing its efflux. Thus, polyunsaturated fatty acids either by themselves or in combination with chemotherapeutic drugs have the potential as anti-cancer molecules.

Multifunctional nanoassemblies (MNAs) were successfully developed for controlled delivery of water-soluble cationic vincristine sulfate (VCR) to overcome multidrug resistance (MDR). The incorporation of anionic small molecule of phosphatidylserine (PS) significantly enhanced the encapsulation efficiency of VCR in MNAs up to 94.4% by electrostatic interaction. Obvious sustained-release characteristics were found in VCR-loaded MNAs (VCR-MNAs) as the cumulative release of VCR was 83.2% at 96 h, and burst-release was effectively diminished. In vivo pharmacokinetics in rats following intravenous administration demonstrated that VCR-MNAs had higher AUC and longer t(1/2) than VCR solution (VCR-Sol). To investigate the MDR reversal effect and clarify the possible mechanism induced by MNAs, the cytotoxicity, cellular uptake and uptake mechanism experiments were performed in MCF-7 and P-glycoprotein over-expressing MCF-7/Adr cells, respectively. Compared with VCR-Sol, VCR-MNAs efficiently enhanced the cytotoxicity to 36.5-fold by increasing the cellular accumulation of VCR (12.6-fold higher) in MCF-7/Adr cells. The results of endocytosis inhibition experiment proved that VCR-MNAs were uptaken into the resistant cancer cells by clathrin- and caveolae-mediated endocytosis pathways, which escaped the efflux induced by P-gp transporter and thereby overcame the MDR of VCR., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The binding of vincristine with DNA has been investigated using Fourier transform infrared spectroscopy. Various changes in the double helical structure of DNA after addition of vincristine have been examined. It is evident from Fourier transform infrared results that vincristine-DNA interaction occurs through guanine and cytosine base pairs. External binding of vincristine with phosphate backbone of the DNA is also observed. Vincristine perturbs guanine band at 1714 cm(-1), cytosine band at 1488 cm(-1), and the phosphate vibrations at 1225 and 1086 cm(-1). The UV-visible spectra of vincristine-DNA complex show hypochromic and bathochromic shifts, indicating the intercalation of vincristine into the double helical structure of DNA. Both intercalative and external binding modes are observed for vincristine binding with DNA, with an estimated binding constant K = 1.0 × 10(3) M(-1).