Cucurbitacin IIa: a novel class of anti-cancer drug inducing non-reversible actin aggregation and inhibiting survivin independent of JAK2/STAT3 phosphorylation

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Cucurbitacin I Inhibits Cell Motility by Indirectly Interfering with Actin Dynamics

Cucurbitacins are plant natural products that inhibit activation of the Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway by an unknown mechanism. They are also known to cause changes in the organization of the actin cytoskeleton. actin depolymerization experiments, cucurbitacin I had no effect on the rate of actin filament disassembly at the nanomolar concentrations that inhibit cell migration. At elevated concentrations, the depolymerization rate was also unaffected, although there was a delay in the initiation of depolymerization. Therefore, cucurbitacin I targets some factor involved in cellular actin dynamics other than actin itself. Two candidate proteins that play roles in actin depolymerization are the actin-severing proteins cofilin and gelsolin. Cucurbitacin I possesses electrophilic reactivity that may lead to chemical modification of its target protein, as suggested by structure-activity relationship data. However, mass spectrometry revealed no evidence for modification of purified cofilin or gelsolin by cucurbitacin I.Cucurbitacin I results in accumulation of actin filaments in cells by a unique indirect mechanism. Furthermore, the proximal target of cucurbitacin I relevant to cell migration is unlikely to be the same one involved in activation of the JAK2/STAT3 pathway

Role of Mitochondrial Electron Transport Chain Complexes in Capsaicin Mediated Oxidative Stress Leading to Apoptosis in Pancreatic Cancer Cells

We evaluated the mechanism of capsaicin-mediated ROS generation in pancreatic cancer cells. The generation of ROS was about 4–6 fold more as compared to control and as early as 1 h after capsaicin treatment in BxPC-3 and AsPC-1 cells but not in normal HPDE-6 cells. The generation of ROS was inhibited by catalase and EUK-134. To delineate the mechanism of ROS generation, enzymatic activities of mitochondrial complex-I and complex-III were determined in the pure mitochondria. Our results shows that capsaicin inhibits about 2.5–9% and 5–20% of complex-I activity and 8–75% of complex-III activity in BxPC-3 and AsPC-1 cells respectively, which was attenuable by SOD, catalase and EUK-134. On the other hand, capsaicin treatment failed to inhibit complex-I or complex-III activities in normal HPDE-6 cells. The ATP levels were drastically suppressed by capsaicin treatment in both BxPC-3 and AsPC-1 cells and attenuated by catalase or EUK-134. Oxidation of mitochondria-specific cardiolipin was substantially higher in capsaicin treated cells. BxPC-3 derived ρ0 cells, which lack mitochondrial DNA, were completely resistant to capsaicin mediated ROS generation and apoptosis. Our results reveal that the release of cytochrome c and cleavage of both caspase-9 and caspase-3 due to disruption of mitochondrial membrane potential were significantly blocked by catalase and EUK-134 in BxPC-3 cells. Our results further demonstrate that capsaicin treatment not only inhibit the enzymatic activity and expression of SOD, catalase and glutathione peroxidase but also reduce glutathione level. Over-expression of catalase by transient transfection protected the cells from capsaicin-mediated ROS generation and apoptosis. Furthermore, tumors from mice orally fed with 2.5 mg/kg capsaicin show decreased SOD activity and an increase in GSSG/GSH levels as compared to controls. Taken together, our results suggest the involvement of mitochondrial complex-I and III in capsaicin-mediated ROS generation and decrease in antioxidant levels resulting in severe mitochondrial damage leading to apoptosis in pancreatic cancer cells

miR-337-3p and Its Targets STAT3 and RAP1A Modulate Taxane Sensitivity in Non-Small Cell Lung Cancers

NSCLC (non-small cell lung cancer) often exhibits resistance to paclitaxel treatment. Identifying the elements regulating paclitaxel response will advance efforts to overcome such resistance in NSCLC therapy. Using in vitro approaches, we demonstrated that over-expression of the microRNA miR-337-3p sensitizes NCI-H1155 cells to paclitaxel, and that miR-337-3p mimic has a general effect on paclitaxel response in NSCLC cell lines, which may provide a novel adjuvant strategy to paclitaxel in the treatment of lung cancer. By combining in vitro and in silico approaches, we identified STAT3 and RAP1A as direct targets that mediate the effect of miR-337-3p on paclitaxel sensitivity. Further investigation showed that miR-337-3p mimic also sensitizes cells to docetaxel, another member of the taxane family, and that STAT3 levels are significantly correlated with taxane resistance in lung cancer cell lines, suggesting that endogenous STAT3 expression is a determinant of intrinsic taxane resistance in lung cancer. The identification of a miR-337-3p as a modulator of cellular response to taxanes, and STAT3 and RAP1A as regulatory targets which mediate that response, defines a novel regulatory pathway modulating paclitaxel sensitivity in lung cancer cells, which may provide novel adjuvant strategies along with paclitaxel in the treatment of lung cancer and may also provide biomarkers for predicting paclitaxel response in NSCLC

Variegated clonality and rapid emergence of new molecular lesions in xenografts of acute lymphoblastic leukemia are associated with drug resistance

The use of genome-wide copy-number analysis and massive parallel sequencing has revolutionized the understanding of the clonal architecture of pediatric acute lymphoblastic leukemia (ALL) by demonstrating that this disease is composed of highly variable clonal ancestries following the rules of Darwinian selection. The current study aimed to analyze the molecular composition of childhood ALL biopsies and patient-derived xenografts with particular emphasis on mechanisms associated with acquired chemoresistance. Genomic DNA from seven primary pediatric ALL patient samples, 29 serially passaged xenografts, and six invivo selected chemoresistant xenografts were analyzed with 250Ksingle-nucleotide polymorphism arrays. Copy-number analysis of non-drug-selected xenografts confirmed a highly variable molecular pattern of variegated subclones. Whereas primary patient samples from initial diagnosis displayed a mean of 5.7 copy-number alterations per sample, serially passaged xenografts contained a mean of 8.2 and chemoresistant xenografts a mean of 10.5 copy-number alterations per sample, respectively. Resistance to cytarabine was explained by a new homozygous deletion of the DCK gene, whereas methotrexate resistance was associated with monoallelic deletion of FPGS and mutation of the remaining allele. This study demonstrates that selecting for chemoresistance in xenografted human ALL cells can reveal novel mechanisms associated with drug resistance

Convergence and divergence of bitterness biosynthesis and regulation in Cucurbitaceae

Differentiation of secondary metabolite profiles in closely related plant species provides clues for unravelling biosynthetic pathways and regulatory circuits, an area that is still underinvestigated. Cucurbitacins, a group of bitter and highly oxygenated tetracyclic triterpenes, are mainly produced by the plant family Cucurbitaceae. These compounds have similar structures, but differ in their antitumour activities and ecophysiological roles. By comparative analyses of the genomes of cucumber, melon and watermelon, we uncovered conserved syntenic loci encoding metabolic genes for distinct cucurbitacins. Characterization of the cytochrome P450s (CYPs) identified from these loci enabled us to unveil a novel multi-oxidation CYP for the tailoring of the cucurbitacin core skeleton as well as two other CYPs responsible for the key structural variations among cucurbitacins C, B and E. We also discovered a syntenic gene cluster of transcription factors that regulates the tissue-specific biosynthesis of cucurbitacins and may confer the loss of bitterness phenotypes associated with convergent domestication of wild cucurbits. This study illustrates the potential to exploit comparative genomics to identify enzymes and transcription factors that control the biosynthesis of structurally related yet unique natural products.National Key R & D Program for Crop Breeding [2016YFD0100307]; National Science Fund for Distinguished Young Scholars [31225025]; National Program on Key Basic Research Projects in China (the 973 Program) [2012CB113900]; leading talents of Guangdong province Program [00201515]; National Natural Science Foundation of China [31322047, 31401886, 31101550, 31672171]; Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences (CAAS-ASTIP-IVFCAAS); Chinese Ministry of Finance [1251610601001]; China Postdoctoral Science Foundation [2014M550902]; Discovery Grant from National Science and Engineering Research Council of Canada (NSERC); Canada Research Chair program; UK Biotechnological and Biological Sciences Research Council Institute Strategic Programme Grant 'Understanding and Exploiting Plant and Microbial Metabolism' [BB/J004561/1]; John Innes Foundation; Genomes to Natural Products National Institutes of Health Programme [U01GM110699]; Shenzhen Municipal Government; Dapeng District GovernmentSCI(E)[email protected]; [email protected]