Enhancing the Effective Chemotherapy: The Combined Inhibition of Rhinacanthin-C, 5-Fluorouracil, and Etoposide on Oral Cancer Cells.

OBJECTIVE: To investigate the effects of rhinacanthin-C (Rh-C), 5-FU, and etoposide on growth inhibition, as well as the effects of a combination of these inhibitors on the oral cell lines SCC9 and HSC4. METHODS: Cancer cell growth inhibition and inhibition combination were determined using the SRB assay. Cell viability and early apoptosis were determined using flow cytometry on cells stained with Annexin 5 and PI. Western blotting was performed to study the molecular mechanism of these inhibitors on oral cancer cells. RESULTS: The results showed that etoposide, 5-FU, and Rh-C exhibited more potent anti-proliferative effects on HSC4 cells compared to SCC9 cells in a time- and concentration-dependent manner. The combination of Rh-C and 5-FU was more effective in inhibiting cell growth than the drugs used alone. The combination of 5-FU and Rh-C resulted in a decrease in live HSC4 cells, with the highest percentage of cell death observed at a ratio of 40:6 µM. Furthermore, the combination of 5-FU and Rh-C reduced P-Akt levels leading to a decrease in cell survival. CONCLUSIONS: HSC4 cells were found to be more sensitive to the inhibitory effect of these drugs compared to SCC9 cells. These findings suggest that the use of Rh-C as a complementary therapy with 5-FU may have the potential for the treatment of oral cancer. the underlying mechanisms responsible for this difference in sensitivity between the two cell lines need to be further investigated.

Rosemary (Rosmarinus officinalis L.) essential oil alleviates testis failure induced by Etoposide in male rats.

Rosemary (Rosmarinus officinalis L.) is a shrub used to treat hepatic, intestinal, renal, respiratory, and reproductive failures. Etoposide a plant-based compound derived from Podophyllum pelltatum, has been used for human malignancies treatment. However, it induces testis, and hepatic failures. In the present study, impact of rosemary essential oil against testis failure, lipid parameters, and hepatic enzymes in male rats has been studied. Forty male Wistar albino rats were grouped in a completely randomized design with Etoposide injection (ETO), rosemary supplementation (ROS), with Etoposide injection and rosemary supplement (ETO+ROS), and control rats with no Etoposide injection and no rosemary (CON). The experiment lasted for seven consecutive weeks including one week as acclimatization time. At the end of the experiment, rats were sacrificed by cervical dislocation, and blood samples were analyzed for serum alkaline phosphatase (ALP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), low-density lipoprotein-Cholesterol (LDL-C), high-density lipoprotein-cholesterol (HDL-C), total cholesterol (TC), total Protein (TP), glucose (GLU) and testosterone. The left testis was harvested for histological examination. Results showed that rats with Etoposide injection had higher ALT, AST, and ALP the control rats. No significant difference was found among treatments in terms of glucose concentration in blood. Rosemary supplemntaion decreased cholesterol and TG concentration and increased HDL concentration in male rats. Furthermore, administration of rosemary essential oil increased blood testosterone but decreased ALT and AST. The epithelial height of seminiferous tubules was decreased significantly in ET as compared with CON. Rosemary essential oil lessened the adverse effect of Etopside on epithelial height in rat testis as it is shown in ET+ROS. In conclusion, dietary supplementation of rosemary essential oil alleviated liver toxicity and functional testis damage induced by Etopside.

Comparative study of the ameliorative effects of omega-3 versus selenium on etoposide-induced changes in Sertoli cells and ectoplasmic specialization of adult rat testes: immunohistochemical and electron microscopic study.

Etoposide (Eto) is an anti-cancer drug that is associated with serious adverse effects on male reproductive function. Omega-3 polyunsaturated fatty acids (ω-3 PUFAs) and selenium (Se) are known as anti-inflammatory, anti-apoptotic and anti-oxidant agents. This work was designed to investigate changes in the biochemical parameters as well as alterations in Sertoli cell vimentin expression, ultrastructure and ectoplasmic specializations (ESs) following Eto treatment and to assess the ameliorative effect of ω-3 versus Se on these alterations. Eighty four adult male albino rats were used and classified into four groups: group I (control group), group II (Eto group) received Eto in a single intra-peritoneal (IP) dose (60 mg/kg B.W.), group III (Eto & ω-3 group) received the single IP dose of Eto as well as ω-3 (300 mg/kg B.W./day by intra-gastric intubation) starting 5 days before Eto injection till the time of sacrifice & group IV (Eto & Se group) received the single IP dose of Eto as well as Se (0.5 mg/kg B.W./day IP) starting 5 days before Eto injection till the time of sacrifice. The rats were subdivided into 2 subgroups (a) and (b) that were sacrificed 3 and 7 days after Eto injection respectively. Eto administration in group II induced increase in malondialdehyde (MDA), decrease in superoxide dismutase (SOD), collapse of Sertoli cell vimentin filaments and ultrastructural degenerative changes in both Sertoli cells and ESs. Se (group IV) reversed Eto toxic effects potently, while ω-3 (group III) had some limited protective effects.

Effect of Kaempferol and Its Glycoside Derivatives on Antioxidant Status of HL-60 Cells Treated with Etoposide.

Kaempferol is a well-known antioxidant found in many plants and plant-based foods. In plants, kaempferol is present mainly in the form of glycoside derivatives. In this work, we focused on determining the effect of kaempferol and its glycoside derivatives on the expression level of genes related to the reduction of oxidative stress-NFE2L2, NQO1, SOD1, SOD2, and HO-1; the enzymatic activity of superoxide dismutases; and the level of glutathione. We used HL-60 acute promyelocytic leukemia cells, which were incubated with the anticancer drug etoposide and kaempferol or one of its three glycoside derivatives isolated from the aerial parts of Lens culinaris Medik.-kaempferol 3-O-[(6-O-E-caffeoyl)-ß-d-glucopyranosyl-(1→2)]-ß-d-galactopyranoside-7-O-ß-d-glucuropyranoside (P2), kaempferol 3-O-[(6-O-E-p-coumaroyl)-ß-d-glucopyranosyl-(1→2)]-ß-d-galactopyranoside-7-O-ß-d-glucuropyranoside (P5), and kaempferol 3-O-[(6-O-E-feruloyl)-ß-d-glucopyranosyl-(1→2)]-ß-d-galactopyranoside-7-O-ß-d-glucuropyranoside (P7). We showed that none of the tested compounds affected NFE2L2 gene expression. Co-incubation with etoposide (1 µM) and kaempferol (10 and 50 µg/mL) leads to an increase in the expression of the HO-1 (9.49 and 9.33-fold at 10 µg/mL and 50 µg/mL, respectively), SOD1 (1.68-fold at 10 µg/mL), SOD2 (1.72-fold at 10-50 µg/mL), and NQO1 (1.84-fold at 50 µg/mL) genes in comparison to cells treated only with etoposide. The effect of kaempferol derivatives on gene expression differs depending on the derivative. All tested polyphenols increased the SOD activity in cells co-incubated with etoposide. We observed that the co-incubation of HL-60 cells with etoposide and kaempferol or derivative P7 increases the level of total glutathione in these cells. Taken together, our observations suggest that the antioxidant activity of kaempferol is related to the activation of antioxidant genes and proteins. Moreover, we observed that glycoside derivatives can have a different effect on the antioxidant cellular systems than kaempferol.

Neobavaisoflavone May Modulate the Activity of Topoisomerase Inhibitors towards U-87 MG Cells: An In Vitro Study.

Despite many advances in therapy, glioblastoma (GB) is still characterized by its poor prognosis. The main reason for this is unsuccessful treatment, which slightly extends the duration of remission; thus, new regimens are needed. One of many types of chemotherapeutics that are being investigated in this field is topoisomerase inhibitors, mainly in combination therapy with other drugs. On the other hand, the search for new anti-cancer substances continues. Neobavaisoflavone (NBIF) is a natural compound isolated from Psoralea corylifolia L., which possesses anti-oxidant, anti-inflammatory, and anti-cancer properties. The aim of this study was to evaluate the effect of NBIF in human U-87 MG glioblastoma cells in comparison to normal human NHA astrocytes, and to examine if it influences the activity of irinotecan, etoposide, and doxorubicin in this in vitro model. We demonstrated that NBIF decreases U-87 MG cells viability in a dose-dependent manner. Furthermore, we found that it inhibits cell growth and causes glutathione (GSH) depletion more intensely in U-87 MG cells than in astrocytes. This study also provides, for the first time, evidence of the potentialization of the doxorubicin effect by NBIF, which was shown by the reduction in the viability in U-87 MG cells.

The Circadian Protein PER1 Modulates the Cellular Response to Anticancer Treatments.

The circadian clock driven by the daily light-dark and temperature cycles of the environment regulates fundamental physiological processes and perturbations of these sophisticated mechanisms may result in pathological conditions, including cancer. While experimental evidence is building up to unravel the link between circadian rhythms and tumorigenesis, it is becoming increasingly apparent that the response to antitumor agents is similarly dependent on the circadian clock, given the dependence of each drug on the circadian regulation of cell cycle, DNA repair and apoptosis. However, the molecular mechanisms that link the circadian machinery to the action of anticancer treatments is still poorly understood, thus limiting the application of circadian rhythms-driven pharmacological therapy, or chronotherapy, in the clinical practice. Herein, we demonstrate the circadian protein period 1 (PER1) and the tumor suppressor p53 negatively cross-regulate each other's expression and activity to modulate the sensitivity of cancer cells to anticancer treatments. Specifically, PER1 physically interacts with p53 to reduce its stability and impair its transcriptional activity, while p53 represses the transcription of PER1. Functionally, we could show that PER1 reduced the sensitivity of cancer cells to drug-induced apoptosis, both in vitro and in vivo in NOD scid gamma (NSG) mice xenotransplanted with a lung cancer cell line. Therefore, our results emphasize the importance of understanding the relationship between the circadian clock and tumor regulatory proteins as the basis for the future development of cancer chronotherapy.

Pathways Related to the Anti-Cancer Effects of Metabolites Derived from Cerrado Biome Native Plants: An Update and Bioinformatics Analysis on Oral Squamous Cell Carcinoma.

BACKGROUND: Oral cancer is a significant health problem worldwide. Oral squamous cell carcinoma (OSCC) is a malignant neoplasm of epithelial cells that mostly affects different anatomical sites in the head and neck and derives from the squamous epithelium or displays similar morphological characteristics. Generally, OSCC is often the end stage of several changes in the stratified squamous epithelium, which begin as epithelial dysplasia and progress by breaking the basement membrane and invading adjacent tissues. Several plant-based drugs with potent anti-cancer effects are considered inexpensive treatments with limited side effects for cancer and other diseases. OBJECTIVE: The aim of this review is to explore whether some Brazilian plant extracts or constituents exhibit anti-tumorigenic activity or have a cytotoxic effect on human oral carcinoma cells. METHODS: Briefly, OSCC and several metabolites derived from Brazilian plants (i.e., flavonoids, vinblastine, irinotecan, etoposide and paclitaxel) were used as keywords to search the literature on PubMed, GenBank and GeneCards. RESULTS: The results showed that these five chemical compounds found in Cerrado Biome plants exhibit anti-neoplastic effects. Evaluating the compounds revealed that they play a main role in the regulation of cell proliferation. CONCLUSION: Preserving and utilising the biodiversity of our planet, especially in unique ecosystems, such as the Cerrado Biome, may prove essential to preserving and promoting human health in modern contexts.

Inhibition of JAK-STAT Signaling with Baricitinib Reduces Inflammation and Improves Cellular Homeostasis in Progeria Cells.

Hutchinson-Gilford progeria syndrome (HGPS), a rare premature aging disorder that leads to death at an average age of 14.7 years due to myocardial infarction or stroke, is caused by mutations in the LMNA gene. Nearly 90% of HGPS cases carry the G608G mutation within exon 11 that generates a truncated prelamin A protein "progerin". Progerin accumulates in HGPS cells and induces premature senescence at the cellular and organismal levels. Children suffering from HGPS develop numerous clinical features that overlap with normal aging, including atherosclerosis, arthritis, hair loss and lipodystrophy. To determine whether an aberrant signaling pathway might underlie the development of these four diseases (atherosclerosis, arthritis, hair loss and lipodystrophy), we performed a text mining analysis of scientific literature and databases. We found a total of 17 genes associated with all four pathologies, 14 of which were linked to the JAK1/2-STAT1/3 signaling pathway. We report that the inhibition of the JAK-STAT pathway with baricitinib, a Food and Drug Administration-approved JAK1/2 inhibitor, restored cellular homeostasis, delayed senescence and decreased proinflammatory markers in HGPS cells. Our ex vivo data using human cell models indicate that the overactivation of JAK-STAT signaling mediates premature senescence and that the inhibition of this pathway could show promise for the treatment of HGPS and age-related pathologies.

Dual-targeted nano-in-nano albumin carriers enhance the efficacy of combined chemo/herbal therapy of lung cancer.

AIM: A Nano-in-Nano approach was exploited to facilitate incorporation of the chemotherapeutic drug etoposide (ETP) as nanosuspension, synergistically with berberine (BER) into hydrophilic albumin nanoparticles (HSA NPs). METHODS: For maximal tumor targeting, HSA was modified with mannose and phenyl-boronic acid. Furthermore, different crosslinkers were investigated for sustained release of water soluble BER from HSA NPs. RESULTS: The elaborated dual-targeted HSA NPs (216.2 nm) were spherical with high BER and ETP entrapment efficiency (69.5 and 87.6%, respectively) and loading (10.52 and 14.04%, respectively). The NPs exhibited sequential release pattern for both ETP and BER (51.55 and 34.33% over 72 h, respectively). Phenyl-boronic acid/mannose-HSA NPs demonstrated powerful cytotoxicity against A549 lung cancer cells (IC50: 12.4 µg/ml) correlated to enhanced cellular internalization. Dual-targeted NPs displayed 9.77-fold higher caspase-3 level and 3.5-fold lower VEGF level than positive control mice. CONCLUSION: Dual-targeted Nano-in-Nano albumin carriers could be beneficial for parenteral ETP/BER delivery to lung cancer.

Asteriscus graveolens Extract in Combination with Cisplatin/Etoposide/Doxorubicin Suppresses Lymphoma Cell Growth through Induction of Caspase-3 Dependent Apoptosis.

Chemotherapy drugs action against cancer is not selective, lead to adverse reactions and drug resistance. Combination therapies have proven more effective in defeating cancers. We hypothesize that plant extract/fraction contains many/several compounds and as such can target multiple pathways as cytotoxic agent and may also have chemo sensitizing activities. We designed a study in which, Asteriscus graveolens (Forssk.) Less (A. graveolens)-derived fraction that contains sesquiterpene lactone asteriscunolide isomers (AS) will be tested in combination with known chemotherapy drugs. Successful combination will permit to reduce chemotherapy drugs concentration and still get the same impact on cancer cells. Sesquiterpene lactone such as asteriscunolide isomers is a naturally occurring compound found in a variety of fruits, vegetables, and medicinal plants with anti-cancer properties. The experiments presented here showed that adding plant fraction containing AS permit reducing the concentration of cisplatin/etoposide/doxorubicin in order to reduce mouse BS-24-1 lymphoma cells (BS-24-1 cells) survival. It involved enhancing the production of Reactive Oxygen Species (ROS), activation of caspase-3 and inhibition of Topoisomerase I activity. Taken together, the results suggest that A. graveolens fraction sensitized BS-24-1 cells to cisplatin/etoposide/doxorubicin through induction of ROS and caspase-3-dependent apoptosis.

Role of Yin Yang-1 (YY1) in the transcription regulation of the multi-drug resistance (MDR1) gene.

Resistance to chemotherapy hinders the successful treatment of acute lymphoblastic leukemia (ALL). The multi-drug resistance-1 (MDR1/ABCB1) gene encodes P-glycoprotein (P-gp), which plays an important role in chemoresistance; however, its transcriptional regulation remains unclear. We investigated the role of YY1 in the regulation of MDR1 and its relation to ALL outcomes. Analysis of the MDR1 promoter revealed four putative YY1-binding sites, which we analyzed using a reporter system and ChIP analysis. YY1 silencing resulted in the inhibition of MDR1 expression and function. The clinical roles of YY1 and MDR1 expression were evaluated in children with ALL. Expression of both proteins was increased in ALL patients compared to controls. We identified a positive correlation between YY1 and MDR1 expression. High levels of YY1 were associated with decreased overall survival. Our results demonstrated that YY1 regulates the transcription of MDR1. Therefore, YY1 may serve as a useful prognostic and/or therapeutic target.

Protective Effects of Spatholobi Caulis Extract on Neuronal Damage and Focal Ischemic Stroke/Reperfusion Injury.

Neuronal apoptotic cell death plays an important role in many neurological disorders, including Alzheimer's disease, Parkinson's disease, and ischemic stroke. Spatholobi Caulis (SC) has been widely used in traditional herbal medicine for the treatment of cancer, inflammation, viral infection, and anemia. However, the protective effects of SC extract (SCE) against apoptotic cell death in the brain have not been reported. We investigated the protective effects of SCE against neuronal injury etoposide-induced neurotoxicity and in rats subjected to focal transient ischemic stroke middle cerebral artery occlusion (MCAO) for 45 min, followed by 7 days of reperfusion. The in vitro study demonstrated that SCE protected cells against etoposide-induced cell viability loss in SH-SY5Y cells. Apoptotic phenotypes, such as cleaved PARP and caspase-3, and oxidative stress in etoposide-treated cells were ameliorated by SCE treatment. In MCAO-reperfusion injury, SCE promoted neuronal survival and level of brain-derived neurotrophic factor (BDNF) by reducing glial activation, oxidative stress, and apoptosis in the ipsilateral cortex. These results indicated that SCE exerted protective effects under etoposide treatment and in a MCAO-reperfusion model by reducing JNK and p38 MAPK activation. This study presents the first evidence that SCE has therapeutic potential for the treatment of ischemic stroke or neurological disorder-related cell death.

The Effect of Rosmarinus Officinalis and Chemotherapeutic Etoposide on Glioblastoma (U87 MG) Cell Culture.

AIM: To investigate whether high dose toxicities of etoposide can be overcome when used in combination with a natural compound named Rosmarinus Officinalis for glioblastoma (GBM). MATERIAL AND METHODS: The impact of Rosmarinus Officinalis in combination with etoposide on GBM U87 MG cells and Mouse Embryonic Fibroblast (MEF) cells was investigated. Both neutral red and 3-(4, 5-Dimethylthiazol-2-Yl)-2, 5-Diphenyltetrazolium Bromide (MTT) assays were employed to gauge cell viability. RESULTS: We observed that increased quantities of Rosmarinus Officinalis induced MEF cell proliferation while it inhibited the survival of GBM cells. Our results indicate that Rosmarinus Officinalis did not affect the cytotoxicity of etoposide on GBM cell cultures. In contrast, in the MEF cell cultures, Rosmarinus Officinalis induced proliferation and diminished the impact of etoposide. CONCLUSION: Rosmarinus Officinalis offers hope for developing new cancer treatment strategies. However, further studies are needed to verify these results.

Inhibitory Effects of Culinary Herbs and Spices on the Growth of HCA-7 Colorectal Cancer Cells and Their COX-2 Expression.

It is unclear if the anti-inflammatory properties of culinary herbs and spices (CHS) are linked to their ability to inhibit Colorectal cancer cell (CRC) growth. Furthermore, their therapeutic potential with regards to CRC is unknown. The aim of this study was to establish if the inhibition of HCA-7 CRC cell growth by a selection of culinary herbs and spices (CHS) is linked to the inhibition of the cells' cyclooxygenase-2 (COX-2 )expression, and to investigate their therapeutic potential. CHS inhibited the growth of Human colon adenocarcinoma-7 (HCA-7) cells; the order of potency was turmeric, bay leaf, ginger, sage, and rosemary; their combinations had a synergistic or additive effect on cell growth inhibition. CHS also inhibited COX-2 expression and activity; this action was comparable to that of the specific COX-2 inhibitor Celecoxib. Coincident with COX-2 inhibition was the accumulation of cells in the sub G1 phase of the HCA-7's cell cycle and, using bay leaf and turmeric, the cleavage of caspase 3 and poly (ADP-ribose) polymerase (PARP). This latter effect showed that the effect of these CHS on growth arrest was irreversible, and was comparable to that of the caspase activator Etoposide. This study provides evidence of a link between the inhibition of HCA-7 growth, and its COX-2 expression, by CHS, and their therapeutic potential.

Passage Variation of PC12 Cells Results in Inconsistent Susceptibility to Externally Induced Apoptosis.

The PC12 cell line is a widely used in vitro model for screening the neuroprotective activity of small molecule libraries. External insult due to serum deprivation or addition of etoposide induces cell death by apoptosis. While this screening method is commonly used in early stage drug discovery no protocol accounting for cell passage number effect on neuroprotective activity has been disclosed. We herein report that passage variation results in false-positive/false-negative identification of neuroprotective compounds; undifferentiated PC12 cells with high passage number are less sensitive to injury induced by serum-deprivation or etoposide treatment. In contrast, NGF differentiated PC12 cells of later passage number are more sensitive to injury induced by etoposide than lower passage number but only after 72 h. Passage number also affects the adherence phenotype of the PC12 cells, complicating screening assays. We report an optimized protocol for screening the neuroprotective activity of small molecules in PC12 cells, which accounts for passage number variations.

Quercetin alters the DNA damage response in human hematopoietic stem and progenitor cells via TopoII- and PI3K-dependent mechanisms synergizing in leukemogenic rearrangements.

Quercetin (Que) is an abundant flavonoid in the human diet and high-concentration food supplement with reported pro- and anti-carcinogenic activities. Topoisomerase II (TopoII) inhibition and subsequent DNA damage induction by Que was implicated in the mixed lineage leukemia gene (MLL) rearrangements that can induce infant and adult leukemias. This notion raised concerns regarding possible genotoxicities of Que in hematopoietic stem and progenitor cells (HSPCs). However, molecular targets mediating Que effects on DNA repair relevant to MLL translocations have not been defined. In this study we describe novel and potentially genotoxic Que activities in suppressing non-homologous end joining and homologous recombination pathways downstream of MLL cleavage. Using pharmacological dissection of DNA-PK, ATM and PI3K signalling we defined PI3K inhibition by Que with a concomitant decrease in the abundance of key DNA repair genes to be responsible for DNA repair inhibition. Evidence for the downstream TopoII-independent mutagenic potential of Que was obtained by documenting further increased frequencies of MLL rearrangements in human HSPCs concomitantly treated with Etoposide and Que versus single treatments. Importantly, by engaging a tissue engineered placental barrier, we have established the extent of Que transplacental transfer and hence provided the evidence for Que reaching fetal HSPCs. Thus, Que exhibits genotoxic effects in human HSPCs via different mechanisms when applied continuously and at high concentrations. In light of the demonstrated Que transfer to the fetal compartment our findings are key to understanding the mechanisms underlying infant leukemia and provide molecular markers for the development of safety values.

Isolation, Culture and Identification of Choriocarcinoma Stem-Like Cells from the Human Choriocarcinoma Cell-Line JEG-3.

BACKGROUND/AIMS: Cancer stem cells (CSCs) exhibit enhanced proliferative capacity and resistance to chemotherapy; however, choriocarcinoma CSCs have not yet been reported. In this study the human choriocarcinoma cell line JEG-3 was cultured in serum free media, and the characteristics of suspension and parental adherent JEG-3 cells were compared. METHODS: Cell proliferation, colony-formation, soft agar clonogenicity, and transwell invasion assays were performed in vitro, and tumor xenografts in BALB/c nude mice were used to evaluate stem cell properties. RESULTS: In serum-supplemented medium (SSM), JEG-3 cells were 4.51 ± 1.71% CD44+, 7.67 ± 2.67% CD133+, and 13.85 ± 2.95% ABCG2+. In serum-free medium (SFM), the expression of these markers increased to 53.08 ± 3.15%, 47.40 ± 2.67%, and 78.70 ± 7.16%, respectively. Moreover, suspension JEG-3 cells exhibited enhanced colony-formation capability as well as invasive and proliferative ability in vitro, alongside enhanced tumorigenic properties in vivo. Suspension JEG-3 cells also exhibited resistance to the chemotherapeutic drugs methotrexate, fluorouracil and etoposide. When seeded in serum supplemented medium, suspension JEG-3 cells readopted an adherent phenotype and continued to differentiate with no significant difference in the morphology between suspension and parent cells. CONCLUSION: In this study, choriocarcinoma stem-like cells (CSLCs) were isolated from the human choriocarcinoma JEG-3 cell line by SFM culture and characterized.

Cytotoxic activity of a synthetic deoxypodophyllotoxin derivative with an opened D-ring.

Podophyllotoxin and its synthetic derivatives are valuable medicinal agents that have antitumor, insecticidal, and antifungal properties. We previously synthesized a deoxypodophyllotoxin derivative with an opened D-ring (DPD) exhibiting potent insecticidal activity. This article was firstly performed to identify the cytotoxicity of DPD toward human cancer cell lines (SGC7901, HeLa, and A549) and normal cell line (HEK293T) using MTT assay. DPD showed potent cytotoxicity against human cancer lines (HeLa and A549) and less cytotoxicity against normal cell lines HEK293T. DPD could also induce the cell cycle arrest at G2/M phase in HeLa cells and significantly increase the phosphorylation (Tyr 15) of CDC2 leading to inactivation of CDC2. The effects of DPD on cellular microtubule networks were detected using immunofluorescence technique in HeLa cells. The immunofluorescence results showed DPD influenced the arrangement and organization of cellular microtubule networks in HeLa cells. Microtubules are long, hollow cylinders made up of polymerized tubulin dimers. Total microtubules were separated after DPD treatment. Western blot results showed that the free polymerized tubulin dimers were obviously increased after DPD treatment. DPD may be a good drug candidate with the therapeutic potential to human cancer by affecting microtubule polymerization.

4,6-Substituted-1,3,5-triazin-2(1H)-ones as monocyclic catalytic inhibitors of human DNA topoisomerase IIα targeting the ATP binding site.

Human DNA topoisomerase IIα (htIIα) is a validated target for the development of novel anticancer agents. Starting from our discovered 4-amino-1,3,5-triazine inhibitors of htIIα, we investigated a library of 2,4,6-trisubstituted-1,3,5-triazines for novel inhibitors that bind to the htIIα ATP binding site using a combination of structure-based and ligand-based pharmacophore models and molecular docking. 4,6-substituted-1,3,5-triazin-2(1H)-ones 8, 9 and 14 were identified as novel inhibitors with activity comparable to the established drug etoposide (1). Compound 8 inhibits the htIIα decatenation in a superior fashion to etoposide. Cleavage assays demonstrated that selected compounds 8 and 14 do not act as poisons and antagonize the poison effect of etoposide. Microscale thermophoresis (MST) confirmed binding of compound 8 to the htIIα ATPase domain and compound 14 effectively inhibits the htIIα mediated ATP hydrolysis. The molecular dynamics simulation study provides further insight into the molecular recognition. The 4,6-disubstituted-1,3,5-triazin-2(1H)-ones represent the first validated monocyclic class of catalytic inhibitors that bind to the to the htIIα ATPase domain.

Fatal hyperammonemia and carbamoyl phosphate synthetase 1 (CPS1) deficiency following high-dose chemotherapy and autologous hematopoietic stem cell transplantation.

Fatal hyperammonemia secondary to chemotherapy for hematological malignancies or following bone marrow transplantation has been described in few patients so far. In these, the pathogenesis of hyperammonemia remained unclear and was suggested to be multifactorial. We observed severe hyperammonemia (maximum 475 µmol/L) in a 2-year-old male patient, who underwent high-dose chemotherapy with carboplatin, etoposide and melphalan, and autologous hematopoietic stem cell transplantation for a neuroblastoma stage IV. Despite intensive care treatment, hyperammonemia persisted and the patient died due to cerebral edema. The biochemical profile with elevations of ammonia and glutamine (maximum 1757 µmol/L) suggested urea cycle dysfunction. In liver homogenates, enzymatic activity and protein expression of the urea cycle enzyme carbamoyl phosphate synthetase 1 (CPS1) were virtually absent. However, no mutation was found in CPS1 cDNA from liver and CPS1 mRNA expression was only slightly decreased. We therefore hypothesized that the acute onset of hyperammonemia was due to an acquired, chemotherapy-induced (posttranscriptional) CPS1 deficiency. This was further supported by in vitro experiments in HepG2 cells treated with carboplatin and etoposide showing a dose-dependent decrease in CPS1 protein expression. Due to severe hyperlactatemia, we analysed oxidative phosphorylation complexes in liver tissue and found reduced activities of complexes I and V, which suggested a more general mitochondrial dysfunction. This study adds to the understanding of chemotherapy-induced hyperammonemia as drug-induced CPS1 deficiency is suggested. Moreover, we highlight the need for urgent diagnostic and therapeutic strategies addressing a possible secondary urea cycle failure in future patients with hyperammonemia during chemotherapy and stem cell transplantation.