ABSTRACT
OBJECTIVE: Irinotecan (IR/CPT-11) is a semisynthetic, water-soluble derivative of the alkaloid camptothecin. It is a topoisomerase I group antineoplastic drug commonly used for the treatment of many cancer types, although it has side effects in tissues such as the testis. Curcumin (CRC) is a polyphenol compound produced from the Indian saffron root; it is used as food colouring and food flavouring. This study examined the testis-specific side effects of IR and the ability of CRC to protect against these side effects. MATERIALS AND METHODS: Forty male Sprague-Dawley rats were used in our study (n = 10). The rats were randomly divided into the following four groups: control, IR, IR + CRC, and CRC. IR 10 mg/kg/day was administered intraperitoneally and CRC 100 mg/kg was administered orally. Blood and testicular samples were collected from rats in all four groups on day 30 after drug administration. Histological, biochemical, and spermatological analyses were conducted. RESULTS: Testis tissue and blood samples were collected from the four groups. Tissue samples from the control and CRC groups demonstrated normal histological appearance on light microscopy. The IR group exhibited the following findings: vascular congestion in the tunica albuginea layer; tubular degeneration and vascular congestion in the interstitial area; oedema, vacuolisation, and luminised cells in the seminiferous tubule; and cells that temporarily stopped dividing at any stage of division in the seminiferous tubule epithelium. In the IR+CRC group, histopathological damage was significantly reduced by CRC treatment. Biochemical analysis showed that the level of thiobarbituric acid reactive substance (TBARS) was significantly increased in the IR group, compared with the other groups. CRC treatment significantly decreased this IR-mediated increase in TBARS level, and the TBARS level in the IR + CRC group approached the level observed in the control group. IR treatment caused significant decreases in glutathione (GSH), superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) levels. However, CRC administration tended to ameliorate the decreases in GSH, SOD, CAT, and GPx levels. CONCLUSIONS: In this study, IR had some toxic effects in rat testis tissue; these effects were ameliorated by CRC treatment. Further studies are warranted to confirm our results.
Subject(s)
Curcumin/pharmacology , Irinotecan/toxicity , Protective Agents/pharmacology , Testis/drug effects , Animals , Male , Rats , Rats, Sprague-Dawley , Thiobarbituric Acid Reactive Substances/metabolism , Topoisomerase I Inhibitors/toxicityABSTRACT
Homogentisic acid (HGA) is the most abundant phenolic compound in strawberry tree (Arbutus unedo L.) honey and an intermediate in the metabolism of phenylalanine and tyrosine. Since HGA exerts its dual nature (pro-oxidant and antioxidant), which depends on the concentration and cell type, the aim of study was to determine whether HGA possess cytoprotective effects and could counteract the cyto- and genotoxic effects of the antineoplastic drug irinotecan (IRI). Tested concentrations corresponded to HGA content in average daily dose of strawberry tree honey as well as five- and ten-fold higher concentrations. Cyto- and genoprotective effects were tested on human peripheral blood lymphocytes using chromosomal aberrations assay and cytokinesis-block micronucleus cytome assay. HGA, even at concentrations 10-fold higher than the one present in the daily amount of consumed strawberry tree honey, posed a non-significant cytotoxic threat to lymphocytes, had a negligible potential for causing cytogenetic damage in treated cells, and did not significantly impair their proliferation. Results of the chromosomal aberration assay and CBMN Cyt assay also showed that HGA efficiently counteracted the detrimental cytogenetic effects of IRI in vitro. The finding on cyto- and genoprotective effects of HGA merits further research in order to better explain the safety profile of this compound and to assess its potency for the development of novel nutraceutical products.
Subject(s)
Homogentisic Acid/pharmacology , Irinotecan/toxicity , Lymphocytes/drug effects , Topoisomerase I Inhibitors/toxicity , Chromosome Aberrations , Humans , In Vitro TechniquesABSTRACT
Novel nontoxic derivatives of SN38 with favorable antineoplastic properties were characterized in water solution using NMR. The phenomena observed by NMR were linked to basic pharmacological properties, such as solubility, bioavailability, chemical and stereochemical stability, and binding to natural DNA oligomers through the terminal G-C base pair, which is commonly considered a biological target of Topo I inhibitors. Compound 1, with bulky substituents at both C5(R) and C20(S) on the same side of a camptothecin core, manifests self-association, whereas diastereomers 2, with bulky C5(S) and C20(S) substituents are mostly monomeric in solution. The stereogenic center at C5 is stable in water solution at pH 5-6. The compound with an (N-azetidinyl)methyl substituent at C9 can undergo the retro Mannich reaction after a prolonged time in water solution. Both diastereomers exhibit different abilities in terms of binding to DNA oligomers: compound 1 is strongly bound, whereas the binding of compound 2 is rather weak. Molecular modeling produced results consistent with NMR experiments. These complementary data allow linking of the observed phenomena in NMR experiments to basic preliminary information on the pharmacodynamic character of compounds and are essential for planning further development research.
Subject(s)
Antineoplastic Agents/chemistry , DNA/chemistry , Irinotecan/analogs & derivatives , Molecular Docking Simulation , Topoisomerase I Inhibitors/chemistry , Antineoplastic Agents/toxicity , Irinotecan/toxicity , Topoisomerase I Inhibitors/toxicityABSTRACT
The anticancer drug irinotecan shows serious dose-limiting gastrointestinal toxicity regardless of intravenous dosing. Although enzymes and transporters involved in irinotecan disposition are known, quantitative contributions of these mechanisms in complex in vivo disposition of irinotecan are poorly understood. We explained intestinal disposition and toxicity of irinotecan by integrating 1) in vitro metabolism and transport data of irinotecan and its metabolites, 2) ex vivo gut microbial activation of the toxic metabolite SN-38, and 3) the tissue protein abundance data of enzymes and transporters relevant to irinotecan and its metabolites. Integration of in vitro kinetics data with the tissue enzyme and transporter abundance predicted that carboxylesterase (CES)-mediated hydrolysis of irinotecan is the rate-limiting process in the liver, where the toxic metabolite formed is rapidly deactivated by glucuronidation. In contrast, the poor SN-38 glucuronidation rate as compared with its efficient formation by CES2 in the enterocytes is the key mechanism of the intestinal accumulation of the toxic metabolite. The biliary efflux and organic anion transporting polypeptide-2B1-mediated enterocyte uptake can also synergize buildup of SN-38 in the enterocytes, whereas intestinal P-glycoprotein likely facilitates SN-38 detoxification in the enterocytes. The higher SN-38 concentration in the intestine can be further nourished by ß-d-glucuronidases. Understanding the quantitative significance of the key metabolism and transport processes of irinotecan and its metabolites can be leveraged to alleviate its intestinal side effects. Further, the proteomics-informed quantitative approach to determine intracellular disposition can be extended to determine susceptibility of cancer cells over normal cells for precision irinotecan therapy. SIGNIFICANCE STATEMENT: This work provides a deeper insight into the quantitative relevance of irinotecan hydrolysis (activation), conjugation (deactivation), and deconjugation (reactivation) by human or gut microbial enzymes or transporters. The results of this study explain the characteristic intestinal exposure and toxicity of irinotecan. The quantitative tissue-specific in vitro to in vivo extrapolation approach presented in this study can be extended to cancer cells.
Subject(s)
Gastrointestinal Microbiome/drug effects , Hepatobiliary Elimination , Inactivation, Metabolic/drug effects , Irinotecan , Organic Anion Transporters/metabolism , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/toxicity , Carboxylesterase/metabolism , Cell Line, Tumor , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Glucuronidase/metabolism , Hepatobiliary Elimination/drug effects , Hepatobiliary Elimination/physiology , Humans , Irinotecan/analogs & derivatives , Irinotecan/pharmacokinetics , Irinotecan/toxicity , Liver/enzymology , Topoisomerase I Inhibitors/pharmacokinetics , Topoisomerase I Inhibitors/toxicityABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Irinotecan (CPT-11) is a valuable chemotherapeutic compound, but its use is associated with severe diarrhea in some patients. The CPT-11 prodrug is converted into the active 7-ethyl-10-hydroxycamptothecin (SN-38) metabolite, which can then be retained for extended periods in the intestine, leading to the onset of diarrhea and related symptoms. Banxia Xiexin Decoction (BXD) is commonly employed for the treatment of gastroenteritis in traditional Chinese medicine (TCM), and in clinical settings, it is used to prevent diarrhea in patients undergoing CPT-11 treatment. To date, however, there have been no studies specifically examining which components of BXD can alleviate the gastrointestinal symptoms associated with CPT-11 administration. AIM: This study aimed to identify the main herbal components of BXD associated with protection against CPT-11-induced intestinal toxicity in a murine model system. MATERIALS AND METHODS: SN-38 levels were measured by UPLC-ESI-MS/MS in samples collected from mice subjected to CPT-11-induced diarrhea that had been administered BXD or different components thereof. Pearson correlation and Grey relational analyses were then used to explore spectrum-effect relationships between reductions in intestinal SN-38 levels and specific chemical fingerprints in samples from mice administered particular combinations of BXD component herbs. RESULTS: We found that different herbal combinations were associated with significant differences in intestinal SN-38 reductions in treated mice. Our spectrum-effect analysis revealed that BXD components including chrysin 6-C-arabinoside-8-C-glucoside, coptisine, hydroxyl oroxylin A 7-O-glucuronide (hydroxyl wogonoside), baicalin, an isomer of 5,6,7-trihydroxyl-flavanone-7-O-glucuronide, berberine, palmatine, and chrysin-7-O-glucuronide were all directly linked with reductions in intestinal SN-38 levels. We therefore speculate that these compounds are the primary bioactive components of BXD, suggesting that they offer protection against CPT-11-induced diarrhea. CONCLUSION: By utilizing UPLC to analyze SN-38 levels in mice treated with a variety of herbal combinations, we were able to effectively explore BXD spectrum-effect relationships and to thereby establish the components of this medicinal preparation that were bioactive and capable of preventing CPT-11-induced diarrhea in mice. This and similar spectrum-effect studies represent a robust means of exploring the mechanistic basis for the pharmacological activity of TCM preparations.
Subject(s)
Diarrhea/prevention & control , Drugs, Chinese Herbal/pharmacology , Intestinal Diseases/prevention & control , Irinotecan/toxicity , Animals , Chromatography, High Pressure Liquid , Diarrhea/chemically induced , Drugs, Chinese Herbal/chemistry , Female , Intestinal Diseases/chemically induced , Mice , Mice, Inbred ICR , Tandem Mass Spectrometry , Topoisomerase I Inhibitors/toxicityABSTRACT
PURPOSE: Intestinal mucositis is an important adverse effect of antineoplastic therapy, which remains without adequate treatment. The present study aimed to carry out a complete evaluation of the histopathological changes during irinotecan-induced intestinal mucositis, using the protocol most found in the pharmacological reports nowadays to better understand irinotecan toxicity and support future studies on drug discovery. METHODS: Intestinal mucositis was induced by treating swiss mice for 4 days with irinotecan (75 mg/kg, i.p.). After 72 h post irinotecan, the mice were sacrificed and the small intestine and colon were excised to performed histological analysis by stained tissue with hematoxylin/eosin (H&E). RESULTS: Histoarchitecture loss, villus/crypt ratio reduction, atrophy of the muscular layer, hypertrophy in the submucosal and mucous layers, ruptures in the epithelium, as well as extent cellular infiltrate and presence of micro abscesses and the fusion of the crypts were observed in the histological analysis. Moreover, duodenum and colon had increased intraepithelial lymphocytes and mitotic figures. However, submucosal ganglia were decreased in the duodenum and increased in the colon. CONCLUSIONS: The data obtained in the present study provides new evidence that irinotecan-induced intestinal mucositis highly affects small intestine and colon, further contributing to establish criteria in light of the histopathological changes induced by irinotecan during intestinal mucositis and facilitating inter-study comparisons.
Subject(s)
Intestinal Mucosa/drug effects , Irinotecan/toxicity , Mucositis/chemically induced , Topoisomerase I Inhibitors/toxicity , Animals , Colon/drug effects , Colon/pathology , Female , Intestinal Mucosa/pathology , Intestine, Small/drug effects , Intestine, Small/pathology , Irinotecan/administration & dosage , Mice , Mucositis/pathology , Topoisomerase I Inhibitors/administration & dosageABSTRACT
BACKGROUND: Intravitreal injections of topotecan are used in the management of retinoblastoma with vitreous seeds. This study evaluated whether intravitreal topotecan was associated with retinal toxicity. METHODS: Retrospective cohort study of patients with retinoblastoma who were treated with intravitreal topotecan at Memorial Sloan Kettering Cancer Center between December 2014 and May 2019. Electroretinogram (ERG) responses under anaesthesia were measured immediately before treatment with intravitreal topotecan and at the next visitor approximately one-month. Ocular toxicity was defined by a decrease in the ERG response at 30 Hz at follow-up. RESULTS: Ocular toxicity was evaluated by ERG on 50 evaluable injections administered to 28 eyes. 22 (44.0%) injections were performed with concurrent intravitreal melphalan. The median time to ERG measurement following an injection was 27 days. By using a paired t-test, intravitreal topotecan combined with melphalan (n=22) at a dose of 25 µg or 30 µg was associated with a significant decrease in ERG amplitude at follow-up (p=0.046, 95% CI -20.4 µV to -0.2 µV). Among eyes that only received topotecan (n=28) at doses of 20 µg or 30 µg, there was not a significant difference in ERG amplitude measured (p=0.85, 95% CI -7.0 µV to 5.8 µV). CONCLUSION: Intravitreal topotecan combined with intravitreal melphalan was associated with a decrease in ERG amplitude; there was not a significant decrease in ERG amplitude observed in patients who received topotecan alone. These findings suggest that intravitreal topotecan injections at doses of 20 µg or 30 µg are not associated with retinal toxicity in patients with retinoblastoma.
Subject(s)
Electroretinography/drug effects , Retina/drug effects , Retinal Neoplasms/drug therapy , Retinoblastoma/drug therapy , Topoisomerase I Inhibitors/toxicity , Topotecan/toxicity , Antineoplastic Agents, Alkylating/administration & dosage , Antineoplastic Combined Chemotherapy Protocols , Female , Follow-Up Studies , Humans , Infant , Intravitreal Injections , Male , Melphalan/administration & dosage , Retinal Neoplasms/pathology , Retinoblastoma/pathology , Retrospective Studies , Topoisomerase I Inhibitors/administration & dosage , Topotecan/administration & dosageABSTRACT
It has recently been established that the marked sensitivity of ATM deficient cells to topoisomerase poisons like camptothecin (Cpt) results from unrestrained end-joining of DNA ends at collapsed replication forks that is mediated by the non-homologous end joining [NHEJ] pathway and results in the induction of copious numbers of genomic alterations, termed "toxic NHEJ". Ablation of core components of the NHEJ pathway reverses the Cpt sensitivity of ATM deficient cells, but inhibition of DNA-PKcs does not. Here, we show that complete ablation of DNA-PKcs partially reverses the Cpt sensitivity of ATM deficient cells; thus, ATM deficient cells lacking DNA-PKcs are more resistant to Cpt than cells expressing DNA-PKcs. However, the relative sensitivity of DNA-PKcs proficient ATM deficient cells is inversely proportional to DNA-PKcs expression levels. These data suggest that DNA-PK may phosphorylate an ATM target (that contributes to Cpt resistance), explaining partial rescue of Cpt sensitivity in cells expressing high levels of DNA-PKcs. Although crippling NHEJ function by mutagenic blockade of the critical ABCDE autophosphorylation sites in DNA-PKcs also sensitizes cells to Cpt, this sensitization apparently occurs by a distinct mechanism from ATM ablation because blockade of these sites actually rescues ATM deficient cells from toxic NHEJ. These data are consistent with autophosphorylation of the ABCDE sites (and not ATM mediated phosphorylation) in response to Cpt-induced damage. In contrast, blockade of S3205 (an ATM dependent phosphorylation site in DNA-PKcs) that minimally impacts NHEJ, increases Cpt sensitivity. In sum, these data suggest that ATM and DNA-PK cooperate to facilitate Cpt-induced DNA damage, and that ATM phosphorylation of S3205 facilitates appropriate repair at collapsed replication forks.
Subject(s)
Ataxia Telangiectasia Mutated Proteins/metabolism , DNA Damage , DNA End-Joining Repair , DNA Replication , DNA-Activated Protein Kinase/metabolism , Camptothecin/toxicity , DNA/metabolism , Humans , Phosphorylation , Protein Processing, Post-Translational , Topoisomerase I Inhibitors/toxicityABSTRACT
Delayed diarrhea is a common side effect of irinotecan administration, leading to a reduction in dose and thus a delay in anticancer therapy. Ganciclovir (GCV), an antiretroviral drug, is used to treat cytomegalovirus (CMV) infection. It is unclear whether GCV exhibits protective effects against irinotecan-induced intestinal dysfunction. Intraperitoneal administration of irinotecan with or without GCV for 4 days induced intestinal toxicity in mice to analyze diarrhea; beta-glucuronidase (ß-GLU) activity; fecal occult blood; hepatic function in blood samples, histopathological changes; and NOD-like receptor 3 (NLRP3), toll-like receptor 4 (TLR4), high-mobility group box 1 protein (HMGB1), phosphorylated nuclear factor kappa B (p-NF-κB), occludin, and zonular occludens (ZO-1) expression in colonic and ileal tissues. In addition, an irinotecan-treated mouse model was constructed and analyzed based on survival time. Expression levels of NLRP3, TLR4, HMGB1, p-NF-κB, occludin, and ZO-1 in normal colonic epithelial cells (NCM460 cells) stimulated with SN-38 were analyzed. GCV treatment reduced various indicators of irinotecan-induced intestinal dysfunction, including delayed-onset diarrhea, pathomorphological changes indicating hepatotoxicity, and proteins related to the HMGB1/TLR4 pathway that induced inflammation; the results were increased body weight, food intake, and expression of the protective proteins occludin and ZO-1. No changes in ß-GLU activity were observed. Changes in the expression of proteins related to the HMGB1/TLR4 pathway, occludin, and ZO-1 in SN-38-stimulated NCM460 cells were similar to changes in these proteins in vivo. In addition, administration of GCV improved mouse survival, indicating that the drug had long-term efficacy. Furthermore, GCV + irinotecan did not decrease the anti-tumor effect of irinotecan in vivo. In summary, GCV had intestine-protective and anti-inflammatory properties that possibly reduced irinotecan-induced intestinal dysfunction.
Subject(s)
Colonic Neoplasms/drug therapy , Diarrhea/drug therapy , Ganciclovir/pharmacology , Inflammation/drug therapy , Intestinal Diseases/drug therapy , Irinotecan/toxicity , NLR Family, Pyrin Domain-Containing 3 Protein/antagonists & inhibitors , Animals , Antiviral Agents/pharmacology , Apoptosis , Biomarkers , Cell Proliferation , Colonic Neoplasms/metabolism , Colonic Neoplasms/pathology , Diarrhea/chemically induced , Diarrhea/metabolism , Diarrhea/pathology , Gene Expression Regulation , Humans , Inflammation/chemically induced , Inflammation/metabolism , Inflammation/pathology , Intestinal Diseases/chemically induced , Intestinal Diseases/metabolism , Intestinal Diseases/pathology , Male , Mice , Mice, Inbred BALB C , Mice, Nude , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Topoisomerase I Inhibitors/toxicity , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
The transformation abilities of CD44s and CD44v6 in normal intestinal epithelial cells have not yet been reported. Herein, we established both CD44s and CD44v6 overexpressing stable clones from rat IEC-6 cells and demonstrated that the CD44v6 clones had higher saturation density and anchorage independence. Additionally, CD44v6 clones were more resistant to oxaliplatin and irinotecan which might be attributed to a significantly increased B-cell lymphoma 2 level and a reduced DNA damage response in these cells. Moreover, c-Met and vascular endothelial growth factor receptor 2 signalings were involved in modulating the saturation density in CD44v6 clones. Interestingly, higher activation of both AKT and extracellular-signal-regulated kinase (ERK) were detected in CD44v6 clones which might account in part for the cell density-independent nuclear localization of Yes-associated protein (YAP). To no surprise, increases of both saturation density and anchorage independence in CD44v6 clones were markedly diminished by PI3K, AKT, MEK, and ERK inhibitors as well as YAP knockdown. By contrast, overexpression of a constitutively active YAP robustly increased the aforementioned phenotypes in IEC-6 cells. Collectively, our results suggest that upregulation of CD44v6, but not CD44s, induces the transformation of normal intestinal epithelial cells possibly via activating the c-Met/AKT/YAP pathway which might also explain the important role of CD44v6 in the initiation of various carcinomas.
Subject(s)
Cell Transformation, Neoplastic/pathology , Hyaluronan Receptors/genetics , Animals , Antineoplastic Agents/toxicity , Cell Line , Cell Transformation, Neoplastic/genetics , Epithelial Cells , Gene Expression Regulation/drug effects , Humans , Irinotecan/toxicity , Oxaliplatin/toxicity , Protein Isoforms , Rats , Topoisomerase I Inhibitors/toxicityABSTRACT
DNA damage response (DDR) processes, often caused by oxidative stress, are important in aging and -related disorders. We recently showed that G protein-coupled receptor (GPCR) kinase interacting protein 2 (GIT2) plays a key role in both DNA damage and oxidative stress. Multiple tissue analyses in GIT2KO mice demonstrated that GIT2 expression affects the GPCR relaxin family peptide 3 receptor (RXFP3), and is thus a therapeutically-targetable system. RXFP3 and GIT2 play similar roles in metabolic aging processes. Gaining a detailed understanding of the RXFP3-GIT2 functional relationship could aid the development of novel anti-aging therapies. We determined the connection between RXFP3 and GIT2 by investigating the role of RXFP3 in oxidative stress and DDR. Analyzing the effects of oxidizing (H2O2) and DNA-damaging (camptothecin) stressors on the interacting partners of RXFP3 using Affinity Purification-Mass Spectrometry, we found multiple proteins linked to DDR and cell cycle control. RXFP3 expression increased in response to DNA damage, overexpression, and Relaxin 3-mediated stimulation of RXFP3 reduced phosphorylation of DNA damage marker H2AX, and repair protein BRCA1, moderating DNA damage. Our data suggests an RXFP3-GIT2 system that could regulate cellular degradation after DNA damage, and could be a novel mechanism for mitigating the rate of age-related damage accumulation.
Subject(s)
DNA Damage , GTPase-Activating Proteins/metabolism , Gene Expression Regulation/drug effects , Oxidative Stress , Receptors, G-Protein-Coupled/metabolism , Camptothecin/toxicity , Computational Biology , Felodipine , GTPase-Activating Proteins/genetics , Gene Expression Regulation/physiology , Gene Regulatory Networks , HEK293 Cells , Humans , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, G-Protein-Coupled/genetics , Topoisomerase I Inhibitors/toxicityABSTRACT
Irinotecan (CPT-11) is a drug used against a wide variety of tumors, which can cause severe toxicity, possibly leading to the delay or suspension of the cycle, with the consequent impact on the prognosis of survival. The main goal of this work is to predict the toxicities derived from CPT-11 using artificial intelligence methods. The data for this study is conformed of 53 cycles of FOLFIRINOX, corresponding to patients with metastatic colorectal cancer. Supported by several demographic data, blood markers and pharmacokinetic parameters resulting from a non-compartmental pharmacokinetic study of CPT-11 and its metabolites (SN-38 and SN-38-G), we use machine learning techniques to predict high degrees of different toxicities (leukopenia, neutropenia and diarrhea) in new patients. We predict high degree of leukopenia with an accuracy of 76%, neutropenia with 75% and diarrhea with 91%. Among other variables, this study shows that the areas under the curve of CPT-11, SN-38 and SN-38-G play a relevant role in the prediction of the studied toxicities. The presented models allow to predict the degree of toxicity for each cycle of treatment according to the particularities of each patient.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Colorectal Neoplasms/drug therapy , Diarrhea/chemically induced , Irinotecan/pharmacokinetics , Irinotecan/toxicity , Leukopenia/chemically induced , Machine Learning , Models, Biological , Neutropenia/chemically induced , Topoisomerase I Inhibitors/pharmacokinetics , Topoisomerase I Inhibitors/toxicity , Aged , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Camptothecin/analogs & derivatives , Camptothecin/metabolism , Colorectal Neoplasms/secondary , Female , Fluorouracil/administration & dosage , Forecasting , Glucuronates/metabolism , Humans , Irinotecan/administration & dosage , Irinotecan/adverse effects , Leucovorin/administration & dosage , Male , Middle Aged , Oxaliplatin/administration & dosage , Topoisomerase I Inhibitors/administration & dosage , Topoisomerase I Inhibitors/adverse effectsABSTRACT
BACKGROUND: Histone acetylation plays an important role in DNA replication and repair because replicating chromatin is subject to dynamic changes in its structures. However, its precise mechanism remains elusive. In this report, we describe roles of the NuA4 acetyltransferase and histone H4 acetylation in replication fork protection in the fission yeast Schizosaccharomyces pombe. RESULTS: Downregulation of NuA4 subunits renders cells highly sensitive to camptothecin, a compound that induces replication fork breakage. Defects in NuA4 function or mutations in histone H4 acetylation sites lead to impaired recovery of collapsed replication forks and elevated levels of Rad52 DNA repair foci, indicating the role of histone H4 acetylation in DNA replication and fork repair. We also show that Vid21 interacts with the Swi1-Swi3 replication fork protection complex and that Swi1 stabilizes Vid21 and promotes efficient histone H4 acetylation. Furthermore, our genetic analysis demonstrates that loss of Swi1 further sensitizes NuA4 and histone H4 mutant cells to replication fork breakage. CONCLUSION: Considering that Swi1 plays a critical role in replication fork protection, our results indicate that NuA4 and histone H4 acetylation promote repair of broken DNA replication forks.
Subject(s)
DNA Replication , Histone Acetyltransferases/metabolism , Acetylation , Camptothecin/toxicity , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Histone Acetyltransferases/genetics , Histones/genetics , Histones/metabolism , Schizosaccharomyces/genetics , Schizosaccharomyces/metabolism , Schizosaccharomyces pombe Proteins/genetics , Schizosaccharomyces pombe Proteins/metabolism , Topoisomerase I Inhibitors/toxicityABSTRACT
Drug resistant tuberculosis (TB) is a major worldwide health problem. In addition to the bacterial mechanisms, human drug transporters limiting the cellular accumulation and the pharmacological disposition of drugs also influence the efficacy of treatment. Mycobacterium tuberculosis topoisomerase-I (MtTopo-I) is a promising target for antimicrobial treatment. In our previous work we have identified several hit compounds targeting the MtTopo-I by in silico docking. Here we expand the scope of the compounds around three scaffolds associated with potent MtTopo-I inhibition. In addition to measuring the effect of newly generated compounds on MtTopo-I activity, we characterized the compounds' antimicrobial activity, toxicity in human cells, and interactions with human multidrug transporters. Some of the newly developed MtTopo-I inhibitors have strong antimicrobial activity and do not harm mammalian cells. Moreover, our studies revealed significant human ABC drug transporter interactions for several MtTopo-I compounds that may modify their ADME-Tox parameters and cellular effects. Promising new drug candidates may be selected based on these studies for further anti-TB drug development.
Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Mycobacterium tuberculosis/enzymology , Topoisomerase I Inhibitors/metabolism , Topoisomerase I Inhibitors/pharmacology , ATP Binding Cassette Transporter, Subfamily B, Member 1/chemistry , ATP Binding Cassette Transporter, Subfamily G, Member 2/chemistry , Animals , Cell Line , Humans , Molecular Docking Simulation , Protein Binding , Protein Conformation , Topoisomerase I Inhibitors/toxicityABSTRACT
Camptothecin (CPT) is an important topoisomerase I enzyme (Topo I) targeting anti-cancer drug, but its oral administration is limited by poor bioavailability and severe side effects. In this study, a redox sensitive CPT prodrug loaded solid lipid nanoparticles (SLN) system for oral delivery was developed. First of all, CPT-palmitic acid conjugate via a cleavable disulfide bond linker (CPT-SS-PA) was synthesized and encapsulated into SLN. The drug release of SLN was evaluated in neutral environment, simulated gastrointestinal fluid and reductive solution. The results indicated that CPT-SS-PA SLN maintained chemical structural stability in simulated physiological environment but exhibited quick reduction-response release of CPT in the presence of dithiothreitol. Furthermore, in vitro cytotoxicity of CPT-SS-PA SLN was tested against cancer cell lines, and the cellular uptake behavior for oral delivery was checked by confocal laser scanning microscopy (CLSM) using Caco-2 cells model. From the data, CPT-SS-PA SLN revealed high anti-cancer activity and enhanced Caco-2 cell absorption. Finally, the oral bioavailability and intestinal safety of CPT-SS-PA SLN were preliminary evaluated by in vivo pharmacokinetic and histopathological study, respectively. This study demonstrated that CPT-SS-PA SLN could be developed as an effective CPT oral delivery system due to its enhanced oral bioavailability and reduced intestinal side effect.
Subject(s)
Camptothecin/administration & dosage , Drug Carriers , Lipids/chemistry , Nanoparticles , Palmitic Acids/administration & dosage , Prodrugs/administration & dosage , Topoisomerase I Inhibitors/administration & dosage , Administration, Oral , Animals , Biological Availability , Caco-2 Cells , Camptothecin/chemical synthesis , Camptothecin/pharmacokinetics , Camptothecin/toxicity , Drug Compounding , Drug Stability , Female , HT29 Cells , Hep G2 Cells , Humans , Intestinal Absorption , MCF-7 Cells , Mice, Inbred BALB C , Nanotechnology , Neoplasms/drug therapy , Neoplasms/pathology , Oxidation-Reduction , Palmitic Acids/chemical synthesis , Palmitic Acids/pharmacokinetics , Palmitic Acids/toxicity , Prodrugs/chemical synthesis , Prodrugs/pharmacokinetics , Prodrugs/toxicity , Technology, Pharmaceutical/methods , Topoisomerase I Inhibitors/chemical synthesis , Topoisomerase I Inhibitors/pharmacokinetics , Topoisomerase I Inhibitors/toxicityABSTRACT
A series of DNA minor groove binding fluorescent dimeric bisbenzimidazoles DBA(n) bearing linkers of various length were synthesized and their biochemical and antiviral activities were evaluated. Their antiviral activity was assessed in model cell systems infected with human herpes simplex virus (HSV-1) and cytomegalovirus (CMV). Compounds DBA(1) and DBA(7) demonstrated in vitro inhibitory properties towards HSV-1, and DBA(7) completely blocked the viral infection. Compound DBA(11) displayed the in vitro therapeutic activity towards both HSV-1 and CMV. All of the DBA(n) could fluoresce, were well soluble in water, not cytotoxic to a concentration of 240⯵M, penetrated well into cell nuclei by binding to DNA and could inhibit topo-I at low micromolecular concentrations.
Subject(s)
Antiviral Agents/chemistry , Benzimidazoles/chemistry , DNA/chemistry , Animals , Antiviral Agents/chemical synthesis , Antiviral Agents/toxicity , Benzimidazoles/chemical synthesis , Benzimidazoles/toxicity , Cattle , Cell Line, Tumor , Cell Nucleus/metabolism , Chlorocebus aethiops , Cytomegalovirus/drug effects , Fluorescence , Herpesvirus 1, Human/drug effects , Humans , Ligands , Solubility , Topoisomerase I Inhibitors/chemical synthesis , Topoisomerase I Inhibitors/chemistry , Topoisomerase I Inhibitors/toxicity , Vero CellsABSTRACT
Classic cytotoxic drugs remain indispensable instruments in antitumor therapy due to their effectiveness and a more prevalent insensitivity toward tumor resistance mechanisms. Herein we describe the favorable properties of 6-(N,N-dimethyl-2-aminoethoxy)-11-(3,4,5-trimethoxyphenyl)pyrido[3,4-c][1,9]phenanthroline (P8-D6), a powerful inducer of apoptosis caused by an equipotent inhibition of human topoisomerase I and II activities. A broad-spectrum effect against human tumor cell lines at nanomolar concentrations, as well as strong antileukemic effects, were shown to be superior to those of marketed topoisomerase-targeting drugs and dual topoisomerase inhibitors in clinical trials. The facile four-step synthesis, advantageous drugability properties, and initial inâ vivo data encourage the application of P8-D6 in appropriate animal tumor models and further drug development.
Subject(s)
Antineoplastic Agents/chemistry , Naphthalenes/chemical synthesis , Topoisomerase I Inhibitors/chemistry , Topoisomerase II Inhibitors/chemistry , Animals , Antineoplastic Agents/therapeutic use , Antineoplastic Agents/toxicity , Apoptosis/drug effects , Body Weight/drug effects , Cell Line, Tumor , Drug Screening Assays, Antitumor , Female , Humans , Mice , Mice, Nude , Naphthalenes/therapeutic use , Naphthalenes/toxicity , Neoplasms/drug therapy , Topoisomerase I Inhibitors/therapeutic use , Topoisomerase I Inhibitors/toxicity , Topoisomerase II Inhibitors/therapeutic use , Topoisomerase II Inhibitors/toxicity , Transplantation, HeterologousABSTRACT
PURPOSE: Clinical evidence of retinal pigment epithelium (RPE) alterations after intra-arterial (IAC) and intravitreal chemotherapy (IViC) of retinoblastoma has been reported. We, therefore, investigated the cellular toxic effects of melphalan, topotecan and carboplatin on the RPE in a cell culture model. METHODS: The effects of melphalan, carboplatin and topotecan on ARPE19 cell morphology were examined by phase contrast microscopy. Cell proliferation was quantified by BrdU incorporation, cell viability studied via MTS assays, and cell densities were estimated by Crystal Violet staining, and apoptosis induction studied via caspase 3/7-activity assays after a 24-hr incubation period. Staurosporine, media without fetal bovine serum, diluents of melphalan, carboplatin and topotecan were applied as positive and negative controls, respectively. RESULTS: We observed a concentration-dependent increase in the number and size of gaps in the ARPE19 cell layer with each drug. There was a significant decrease in proliferative activity and cell viability of RPE cells as well as an increase in apoptosis after 24 hrs culture in media supplemented with melphalan and topotecan. Carboplatin had comparable effects on cell proliferation and cell viability; however, no significant apoptotic impacts were observed. The three cytostatic drugs had insignificant effects on cell density measurements. CONCLUSIONS: Morphological monitoring and toxicity assays indicate a direct toxic effect of melphalan and the other two cytostatic drugs on ARPE19 cells. Thus, a direct toxic effect of melphalan in vivo after IAC or IViC on the RPE seems probable and may explain the clinical and angiographic RPE alterations observed in some retinoblastoma patients.
Subject(s)
Antineoplastic Agents/toxicity , Carboplatin/toxicity , Melphalan/toxicity , Retinal Pigment Epithelium/drug effects , Topoisomerase I Inhibitors/toxicity , Topotecan/toxicity , Apoptosis/drug effects , Caspase 3/metabolism , Caspase 7/metabolism , Cell Count , Cell Proliferation/drug effects , Cell Survival , Cells, Cultured , Dose-Response Relationship, Drug , Humans , Microscopy, Phase-Contrast , Retinal Pigment Epithelium/enzymology , Retinal Pigment Epithelium/pathologyABSTRACT
Methyleugenol is a substituted alkenylbenzene found in several herbs and spices. It is classified by the European Union's Scientific Committee on Food as a genotoxic carcinogen. We addressed the biological mechanism of the genotoxic properties of methyleugenol and its oxidative metabolites. Methyleugenol and the oxidative metabolites significantly enhanced the DNA damage in human colon carcinoma cells (HT29). Methyleugenol did not affect the protein status of γH2AX, a biomarker of DNA double-strand breaks, whereas its metabolites methyleugenol-2',3'-epoxide and 3'-oxomethylisoeugenol significantly increased the cellular phosphorylated H2AX level. Both of these metabolites also showed a significant induction of micronuclei in HT29 cells. Furthermore, we investigated whether topoisomerase interaction contribute to the observed effect on DNA integrity. Methyleugenol-2',3'-epoxide and 3'-oxomethylisoeugenol inhibited the activity of recombinant topoisomerase I. In HT29 cells, neither methyleugenol nor the metabolites affected the level of topoisomerase protein bound to DNA, excluding a topoisomerase poisoning mode of action. In addition, 3'-oxomethylisoeugenol potently diminished the level of camptothecin-stabilized topoisomerase I/DNA intermediates and camptothecin-induced DNA strand breaks. In conclusion, it could be suggested that 3'-oxomethylisoeugenol may also interact with classical or food-borne topoisomerase I poisons, diminishing their poisoning effectiveness.
Subject(s)
Carcinogens, Environmental/toxicity , Colonic Neoplasms/chemically induced , DNA Damage , DNA Topoisomerases, Type I/metabolism , Eugenol/analogs & derivatives , Mutagens/toxicity , Topoisomerase I Inhibitors/toxicity , Biomarkers, Tumor/agonists , Biomarkers, Tumor/metabolism , Biotransformation , Carcinogens, Environmental/analysis , Carcinogens, Environmental/metabolism , Carcinoma/chemically induced , Carcinoma/enzymology , Colonic Neoplasms/enzymology , Colonic Neoplasms/metabolism , Comet Assay , DNA Topoisomerases, Type I/chemistry , DNA Topoisomerases, Type I/genetics , Epoxy Compounds/analysis , Epoxy Compounds/metabolism , Epoxy Compounds/toxicity , Eugenol/analysis , Eugenol/metabolism , Eugenol/toxicity , Food Contamination , HT29 Cells , Histones/agonists , Histones/metabolism , Humans , Micronucleus Tests , Mutagens/analysis , Mutagens/metabolism , Neoplasm Proteins/agonists , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Oxidation-Reduction , Phosphorylation/drug effects , Protein Processing, Post-Translational/drug effects , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Spices/adverse effects , Spices/analysis , Topoisomerase I Inhibitors/analysis , Topoisomerase I Inhibitors/metabolismABSTRACT
CPT-11 is widely used for cancer therapy as a chemotherapeutic agent. Despite its good efficacy, a large number of side effects appeared during decades of clinical application. Delayed diarrhea, at dose limiting toxicity, happens after 24h of treatment and the rate of occurrence is up to 90%. Although many investments have been made on this negative impact, the real molecular mechanism of delayed diarrhea is poorly understood. In this study, we have discovered that CPT-11 promotes macrophage infiltration into intestinal tissues and activates the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome, resulting in a robust IL-1ß response and colonic inflammation similar to DSS (dextran sodium sulfate) induced experimental colitis. CPT-11 plus LPS primed mouse bone marrow-derived macrophages (BMDMs) and human acute monocytic leukemia cells (THP-1 cells) staying in a highly activated status, showing increased caspase-1 activity and releasing great amounts of IL-1ß and IL-18 as detected by ELISA and western blot. A further mechanism showed that JNK and NF-κB signaling pathways participated in inflammatory responses activated by CPT-11. These results prompted us to suggest that the NLRP3-IL-1ß signaling pathway might play an important role in CPT11-induced colitis. Our findings provide a basis for developing novel strategies that improve clinical implications of CPT-11.