Cisplatin-sensitive A2780 Research Articles

Intracellular GSH Alterations and Its Relationship to Level of Resistance following Exposure to Cisplatin in Cancer Cells.

One of the major complications in cancer chemotherapy with cisplatin as one of the important medicines in treatment regimens of different cancers is the development of resistance. One of the most described cellular defense mechanisms involved in resistance is glutathione (GSH), thus in this study, the effects of cisplatin on the total intracellular GSH level (GSHi) in some sensitive and resistant variants of human cell lines (hepatocarcinoma HepG2, skin A375, cisplatin sensitive glioblastoma U373MG and cisplatin resistant glioblastoma U373MGCP, cisplatin sensitive ovary A2780S and cisplatin resistant A2780CP cells) were studied. MTT assay was performed to measure cytotoxicity of cisplatin (33.3 µM for 1 hour). Following cisplatin exposure, GSHi (per million cells) was evaluated using a photometrical assay up to 90 minutes. Our results indicate that there are significant differences between GSHi content of A2780CP and U373MGCP cells compared to other cell lines. Moreover, IC50 of cisplatin in different cells seems to have a relation with mean of GSH level in 90 minutes (GSH (mean)90). As a conclusion, it seems that resistance to cisplatin in different cell lines is more related with the diverse patterns of GSHi variations following cisplatin exposure than its original level, and/or its cellular increase or decrease. It is also suggested that GSH (mean)90 may be used as a factor for the prediction of cellular resistance to cisplatin.

Investigation on metabolism of cisplatin resistant ovarian cancer using a genome scale metabolic model and microarray data.

Many cancer cells show significant resistance to drugs that kill drug sensitive cancer cells and non-tumor cells and such resistance might be a consequence of the difference in metabolism. Therefore, studying the metabolism of drug resistant cancer cells and comparison with drug sensitive and normal cell lines is the objective of this research. Metabolism of cisplatin resistant and sensitive A2780 epithelial ovarian cancer cells and normal ovarian epithelium has been studied using a generic human genome-scale metabolic model and transcription data. The results demonstrate that the most different metabolisms belong to resistant and normal models, and the different reactions are involved in various metabolic pathways. However, large portion of distinct reactions are related to extracellular transport for three cell lines. Capability of metabolic models to secrete lactate was investigated to find the origin of Warburg effect. Computational results introduced SLC25A10 gene, which encodes mitochondrial dicarboxylate transporter involved in exchanging of small metabolites across the mitochondrial membrane that may play key role in high growing capacity of sensitive and resistant cancer cells. The metabolic models were also used to find single and combinatorial targets that reduce the cancer cells growth. Effect of proposed target genes on growth and oxidative phosphorylation of normal cells were determined to estimate drug side-effects. The deletion results showed that although the cisplatin did not cause resistant cancer cells death, but it shifts the cancer cells to a more vulnerable metabolism.

Abstract 2358: Dissecting the cellular response to cisplatin from RNA transcription to translation

Abstract Platinum-based antineoplastic drugs, such as cisplatin, are DNA damaging agents typically utilized in cancer treatment. In ovarian cancer, however, these drugs have a relatively low therapeutic index (which is a measure of clinical benefit over toxicity of these drugs), thus even a small change (reduction) in the cellular sensitivity to these chemotherapeutic agents leads to clinical treatment failure, also known as ‘drug resistance’. To increase the clinical efficacy of platinum-based antineoplastic drugs, we need to develop therapies that are less likely to result in drug resistance, or we need to modify the current therapies to prevent or reduce drug resistance. To gain mechanistic insights into the process of drug resistance that may help us to develop new or modified therapies that are less prone to clinical drug resistance, we have focused our studies on the comprehensive analysis of two still poorly studied cisplatin-associated processes at genome-wide scale: the processes of RNA nascent transcription and RNA translation. In particular, we investigate by global run-on combined with next-generation sequencing (GRO-seq) how cisplatin-sensitive A2780 ovarian cancer cells and its cisplatin-resistance derivative, A2780cis cells, respond to acute cisplatin treatment at the level of the nascent transcriptome. We also investigate, in this case by ribosome profiling, how the same cells respond to acute cisplatin treatment at the level of the ribosome-associated transcriptome (also known as translatome). These two massive analyses, complemented with conventional profiling of steady-state RNA (profiled by RNA-seq) provide the first multidimensional analysis of the actions induced by any chemotherapeutic agent at the level of the three basic states of the transcriptome (nascent RNA, steady-state RNA, and translated RNA). Interestingly, this analysis reveals an unexpected large fraction of uncoupled events between these three transcriptomic states, which challenges the conventional one-dimensional approach based exclusively on RNA-seq for the study of drug resistance. Note: This abstract was not presented at the meeting. Citation Format: Carlos Mackintosh, Sergiy Konovalov, Ivan Garcia-Bassets. Dissecting the cellular response to cisplatin from RNA transcription to translation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2358. doi:10.1158/1538-7445.AM2014-2358

Tetramethoxychalcone, a chalcone derivative, suppresses proliferation, blocks cell cycle progression, and induces apoptosis of human ovarian cancer cells.

In the present study, we investigated the in vitro antitumor functions of a synthetic chalcone derivative 4,3′,4′,5′- tetramethoxychalcone (TMOC) in ovarian cancer cells. We found that TMOC inhibited the proliferation and colony formation of cisplatin sensitive cell line A2780 and resistant cell line A2780/CDDP, as well as ovarian cancer cell line SKOV3 in a time- and dose-dependent manner. Treatment of A2780 cells with TMOC resulted in G0/G1 cell cycle arrest through the down-regulation of cyclin D1 and CDK4, and the up-regulation of p16, p21 and p27 proteins. We demonstrated that TMOC might induce cell apoptosis through suppressing Bcl-2 and Bcl-xL, but enhancing the expression of Bax and the cleavage of PARP-1. Treatment of TMOC also reduced the invasion and migration of A2780 cells. Finally, we found that TMOC inhibited the constitutive activation of STAT3 signaling pathway and induced the expression of the tumor suppressor PTEN regardless of the p53 status in cell lines. These data suggest that TMOC may be developed as a potential chemotherapeutic agent to effectively treat certain cancers including ovarian cancer.

Polyketides from the mangrove-derived endophytic fungus Acremonium strictum

Five new polyketide derivatives, 6′-hydroxypestalotiopsone C (1), acropyrone (2), bicytosporone D (3), waol acid (4), and pestalotiopene C (5), together with seven known metabolites (6–12), were obtained from extracts of the endophytic fungus Acremonium strictum, isolated from the mangrove tree Rhizophora apiculata. The structures of the isolated compounds were elucidated on the basis of comprehensive NMR and MS analysis. Compounds 6, 7, and 9 showed moderate cytotoxic activity against human cisplatin-sensitive (IC50 values 27.1, 76.2, and 8.3μM, respectively) and resistant A2780 cell lines (IC50 values 12.6, 30.1, and 19.0μM, respectively), whereas only 9 exhibited antibacterial activity against Staphylococcus aureus (MIC value 14.3μM).

Cisplatin-induced senescence in ovarian cancer cells is mediated by GRP78.

Glucose-regulated protein78 (GRP78), the most abundant and well-characterized glucose-regulated protein, is a major stress-inducible chaperone localized to the endoplasmic reticulum (ER). The purpose of the present study was to investigate the mechanisms of GRP78 involved in the senescence sensitivity of ovarian cancer cells to cisplatin. In the present study, we found that the chemotherapy-sensitive ovarian tumor sections showed strong staining for heterochromatin protein1-γ (HP1-γ), but weak staining for GRP78. Cisplatin-sensitive A2780 cells with low expression of GRP78 tended to undergo senescence easily when compared with the cisplatin-resistant C13K cells following a dose-gradient cisplatin exposure. Forced overexpression of GRP78 protected the cisplatin-sensitive A2780 cells from cisplatin-induced senescence through P53 and CDC2. Knockdown of GRP78 rescued the senescence sensitivity of cisplatin-resistant C13K cells to cisplatin through P21 and CDC2. Twisting of Ca2+ release from ER stores by GRP78 was established to be associated with the sensitivity of cisplatin-induced senescence in ovarian cancer cell lines. In conclusion, GRP78 may have anti-senescence effects on ovarian cancer cells involving multiple mechanisms. Intervention against GRP78 may reduce cisplatin resistance in ovarian cancer.

Gemcitabine as a molecular targeting agent that blocks the Akt cascade in platinum-resistant ovarian cancer

BackgroundGemcitabine (2′, 2′ –difluorodeoxycytidine) is one of many nonplatinum drugs that exhibit activity in recurrent, platinum-resistant ovarian cancer. However, the molecular mechanisms by which Gemcitabine treatment inhibits the proliferation of platinum-resistant ovarian cancer cells still remain unclear. We investigated whether Gemcitabine increases the efficacy of Cisplatin in platinum-resistant ovarian cancer models in vitro and in vivo.MethodsWe used Cisplatin-resistant Caov-3 cells, A2780CP cells and Cisplatin-sensitive A2780 cells to examine the sensitivity of the cell viability of Cisplatin and Gemcitabine using a 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and the sensitivity of the invasive activity of Cisplatin and Gemcitabine using an invasion assay with Matrigel. We examined the Akt kinase activity and matrix metalloproteinase 9 (MMP9) expression following Cisplatin and Gemcitabine treatment using a Western blot analysis and the mRNA expression of vascular endothelial growth factor (VEGF) using semi-quantitative RT-PCR. Moreover, we evaluated the effects of Cisplatin and Gemcitabine on the intra-abdominal dissemination of ovarian cancer in vivo.ResultsGemcitabine significantly inhibited Cisplatin-induced Akt activation in the Caov-3 and A2780CP cells, but not in the A2780 cells. In the presence of Gemcitabine, Cisplatin-induced growth inhibition and apoptosis were significantly enhanced in the Caov-3 and A2780CP cells. Co-treatment with Cisplatin and Gemcitabine almost completely inhibited invasion of both types of cells through the Matrigel; however, neither Cisplatin nor Gemcitabine alone inhibited the invasion of both types of cells. Gemcitabine inhibited not only the Cisplatin-induced activation of Akt, but also the MMP9 and mRNA expression of VEGF. Moreover, treatment with Gemcitabine increased the efficacy of Cisplatin-induced growth inhibition of the intra-abdominal dissemination and production of ascites in the athymic nude mice inoculated with Caov-3 cells.ConclusionsWe herein demonstrated that Gemcitabine inhibits the Akt kinase activity and angiogenetic activity following treatment with Cisplatin in platinum-resistant ovarian cancer cells. These results provide a rationale for using Gemcitabine in clinical regimens containing molecular targeting agents against platinum-resistant ovarian cancers.

Overexpression of CXCR4 is significantly associated with cisplatin-based chemotherapy resistance and can be a prognostic factor in epithelial ovarian cancer

The chemokine receptor 4 (CXCR4) plays an important role in the growth, angiogenesis and metastasis of various cancers, including epithelial ovarian cancer (EOC). However, the correlation between CXCR4 and the clinical response of EOC patients to chemotherapy remains unknown. 124 EOC patients were recruited to assess the relationship between CXCR4 and the response to cisplatin-based chemotherapy. The results showed that patients with a higher CXCR4 expression had a significantly lower chemosensitivity, a poorer progression-free survival and a lower overall survival than those with lower CXCR4 expression. In addition, knockdown of CXCR4 by small interfering RNA suppressed cell proliferation and resulted in G1/S arrest, increased apoptosis and chemosensitivity in both cisplatin-sensitive A2780 cells and cisplatin-resistant cell A2780/cis in vitro. Our data suggest that CXCR4 is one of the key molecules in cisplatin-based chemotherapy for EOC patients and that CXCR4 inhibition is a potential strategy to address the chemoresistance of EOC. [BMB Reports 2014; 47(1): 33-38]

Gold(iii) complexes inhibit growth of cisplatin-resistant ovarian cancer in association with upregulation of proapoptotic PMS2 gene

Various gold complexes have been known to overcome cisplatin resistance in cancer cells. Yet, their in vivo anti-tumor efficacies and detailed action mechanisms in overcoming this resistance remain largely unexplored. In this work we have established a xenograft model simultaneously consisting of both cisplatin-sensitive and cisplatin-resistant tumors by inoculating human ovarian cancer cells A2780 and its cisplatin-resistant variant A2780cis into different flanks of the same nude mouse. Towards this model, a gold(III) porphyrin complex [AuIII(TPP)]Cl (gold-1a, wherein [TPP]2− = meso-tetraphenylporphyrinato ligand) was found to effectively inhibit the growth of both kinds of tumors, while cisplatin failed to suppress the growth of A2780cis tumors under similar conditions. In both A2780 and A2780cis cells, gold-1a was found to transcriptionally upregulate postmeiotic segregation increased 2 (PMS2) which has DNA mismatch repair and proapoptotic functions. Suppression of PMS2 by RNA interference in A2780cis cells partially rescued the gold-1a-induced death of the cells, indicating that gold-1a inhibited growth of cisplatin-resistant ovarian cancer in association with upregulation of this gene. Two other stable gold(III) analogues including gold(III) octaethylporphyrin (2) and gold(III)-NHC (3) complexes also displayed similar anti-cancer activities on A2780cis cells and capability in PMS2 regulation. In contrast, a gold(I) phosphine complex (4), a gold(I) thiourea complex (5), KAuIIICl4 and cisplatin all displayed a preferential cytotoxicity only towards the cisplatin-sensitive A2780 cells. Taken together, this work has demonstrated the prospect of gold(III) complexes for the treatment of cisplatin-resistant/relapsed ovarian cancers.

Synthesis and In Vitro Pharmacological Behavior of Platinum(II) Complexes Containing 1,2-Diamino-1-(4-fluorophenyl)-2-alkanol Ligands

In continuation of our effort to optimize the pharmacological profile of [1,2-diamino-1,2-bis(4-fluorophenyl)ethane]dichloridoplatinum(II) complexes, we synthesized [1,2-diamino-1-(4-fluorophenyl)alkanol]dichloridoplatinum(II) analogs. The aim of this study was to evaluate the influence of hydroxyl groups at the C2 moiety on aqueous solubility, lipophilicity, cellular platinum accumulation, and cytotoxicity against MDA-MB-231, U-937, RAJI, and SC-1 cells as well as against cisplatin-sensitive and cisplatin-resistant A2780 and A2780cisR ovarian carcinoma cells. As expected, the OH groups improved the water solubility and decreased the lipophilicity of the neutral ligands, resulting in complexes with favorable pharmacokinetic properties. The cellular uptake of the compounds in MDA-MB-231 and U-937 cells proved to depend on the configuration and showed only a slight correlation with lipophilicity. The most active complexes were R,R/S,S configured, which points to a carrier-mediated mode of action. [threo-1,2-Diamino-1-(4-fluorophenyl)propan]dichloridoplatinum(II) and [threo-2,3-diamino-3-(4-fluorophenyl)propan-1-ol]dichloridoplatinum(II) possessed only low cross-resistance to cisplatin and were up to 10-fold more active in lymphoma cell lines.

Role of CFTR in oxidative stress and suicidal death of renal cells during cisplatin-induced nephrotoxicity

The clinical use of the antineoplastic drug cisplatin is limited by its deleterious nephrotoxic side effect. Cisplatin-induced nephrotoxicity is associated with an increase in oxidative stress, leading ultimately to renal cell death and irreversible kidney dysfunction. Oxidative stress could be modified by the cystic fibrosis transmembrane conductance regulator protein (CFTR), a Cl− channel not only involved in chloride secretion but as well in glutathione (GSH) transport. Thus, we tested whether the inhibition of CFTR could protect against cisplatin-induced nephrotoxicity. Using a renal proximal cell line, we show that the specific inhibitor of CFTR, CFTRinh-172, prevents cisplatin-induced cell death and apoptosis by modulating the intracellular reactive oxygen species balance and the intracellular GSH concentration. This CFTRinh-172-mediated protective effect occurs without affecting cellular cisplatin uptake or the formation of platinum-DNA adducts. The protective effect of CFTRinh-172 in cisplatin-induced nephrotoxicity was also investigated in a rat model. Five days after receiving a single cisplatin injection (5 mg/kg), rats exhibited renal failure, as evidenced by the alteration of biochemical and functional parameters. Pretreatment of rats with CFTRinh-172 (1 mg/kg) prior to cisplatin injection significantly prevented these deleterious cisplatin-induced nephrotoxic effects. Finally, we demonstrate that CFTRinh-172 does not impair cisplatin-induced cell death in the cisplatin-sensitive A549 cancer cell line. In conclusion, the use of a specific inhibitor of CFTR may represent a novel therapeutic approach in the prevention of nephrotoxic side effects during cisplatin treatment without affecting its antitumor efficacy.

Induction of Apoptosis and Inhibition of Angiogenesis by PEGylated Liposomal Quercetin in Both Cisplatin-Sensitive and Cisplatin-Resistant Ovarian Cancers

The clinical efficiency of cisplatin against ovarian cancer is often limited by the development of drug resistance. In this work, we investigated PEGylated liposomal quercetin (Lipo-Que) on cisplatin-sensitive (A2780s) and cisplatin-resistant (A2780cp) human ovarian cancer models in vitro and in vivo to reveal whether a cisplatin-resistant ovarian cancer has susceptibility to quercetin (Que) and the mechanism of its antitumor activity. Lipo-Que was prepared using a solid dispersion method, and the obtained Lipo-Que is monodisperse with a mean diameter of 163 +/-10 nm. Besides, in vitro drug release assay showed a sustained release behavior of Lipo-Que. In vitro experiments suggested that Lipo-Que inhibited cell proliferation, induced apoptosis, and induced cell cycle arrest in both A2780s and A2780cp cells. Furthermore, antitumor activity of Lipo-Que was investigated in both cisplatin-sensitive and cisplatin-resistant human ovarian tumor xenograft models in nude mice. Lipo-Que significantly suppressed tumor growth in both models in comparison with free Que, blank liposomes (Lipo), or normal saline (NS). Furthermore, immunohistochemistry and immunofluorescence tests revealed that Lipo-Que induced apoptosis, decreased microvessel density, and inhibited proliferation of tumors in both A2780s and A2780cp tumor models. Therefore, our results suggest that Lipo-Que is an effective agent to inhibit tumor growth in both cisplatin-sensitive and cisplatin-resistant human ovarian cancers.

Evaluation of GPER agonist G-1 combined with navitoclax in platinum-resistant and -sensitive ovarian cancer cells.

e13563 Background: Most ovarian cancer patients are treated with platinum-based chemotherapy but eventually relapse with incurable disease. The G protein-coupled estrogen receptor GPER (GPR30) mediates Ca2+ mobilization in response to estrogen and G-1, a synthetic agonist. Large and sustained Ca2+ responses can lead to mitochondrial Ca2+ overload and apoptosis. Hence, we evaluated whether G-1 could induce apoptosis in cisplatin-sensitive A2780 and isogenic cisplatin–resistant CP70 (14-fold resistant), C30 (70-fold resistant) and C200 (157-fold resistant) human ovarian cancer cells. Bcl-2 and Bcl-xL protect mitochondria from Ca2+overload, and were overexpressed in these cisplatin-resistant cells; thus we also examined combining the Bcl-2 family inhibitor navitoclax with G-1. Methods: Cytoplasmic [Ca2+]c and mitochondrial [Ca2+]m were monitored using microscopy and fluorescent Ca2+ probes. Cell cycle, apoptosis and mitochondrial membrane potential (MMP) were assessed by flow cytometry of propidium iodide, Annexin V and DiIC1(5) -stained cells. The intracellular Ca2+ chelator BAPTA was used to block Ca2+mobilization. Results: Expression of the 53kDa GPER but not the 38 kDa isoform progressively increased with increasing cisplatin resistance. G-1 elicited sustained [Ca2+]c rises that correlated with 53 kDa GPER expression, followed by rises in [Ca2+]m. In all cells, 2.5 μM G-1 blocked cell cycle progression at G2/M, inhibited proliferation, and induced apoptosis (A2780 > C30 > CP70 ≥ C200). G-1 induced p53, caspase-3 and PARP cleavage, and MMP loss. BAPTA prevented G-1’s cell cycle and apoptotic effects in cells showing large Ca2+ mobilization responses but did not in cells with small Ca2+responses. Combining navitoclax with G-1 superadditively decreased cell viability and increased apoptosis. Conclusions: G-1 blocked cell cycle progression and induced apoptosis via a Ca2+-dependent pathway in cells expressing high 53 kDa GPER levels, but via a Ca2+-independent pathway in cells with low 53 kDa GPER expression. G-1 also interacted cooperatively with naviticlax. Therefore, G-1 plus navitoclax shows potential for therapeutic use in platinum-sensitive and -resistant ovarian cancer.

Insight into the toxic effects of cis-dichloridoplatinum(II) complexes containing 7-azaindole halogeno derivatives in tumor cells

The cisplatin analogues cis-[PtCl2(3ClHaza)2] (1) and cis-[PtCl2(3IHaza)2] (2) (3ClHaza and 3IHaza are 3-chloro-7-azaindole and 3-iodo-7-azaindole, respectively) are quite toxic to ovarian tumor cells, with moderately better IC50 values than for cisplatin in the cisplatin-sensitive cell line A2780. We investigated potential factors which might be involved in the mechanism underlying the cytotoxic effects of 1 and 2 and compared these factors with those involved in the mechanism underlying the effects of conventional cisplatin. Our data indicate that the higher cytotoxicity of 1 and 2 originates mainly from their efficient cellular accumulation, different effects at the level of cell cycle regulation, and reduced propensity for DNA adduct repair. Studies of their reactivity toward cellular components reveal efficient binding to DNA, which is typically required for an active platinum drug. Further results suggest that 1 and 2 are capable of circumventing resistance to cisplatin induced by alterations in cellular accumulation and DNA repair. Hence, the latter two factors appear to be responsible for differences in the toxicity of 1 or 2, and cisplatin in tumor cells. The results of this work reinforce the idea that direct analogues of conventional cisplatin-containing halogeno-substituted 7-azaindoles offer much promise for the design of novel therapeutic agents.

Anti-Cancer Activities and Interaction of Imiquimod and Flex-Het, SHetA2, in Melanoma and Ovarian Cancer

New strategies are needed to treat cancers that do not respond to chemotherapy or resist chemotherapy after responding initially. The objective of this study was to evaluate non-cytotoxic drugs against two of these cancers, melanoma and ovarian cancer. Imiquimod is an immune stimulant that induces apoptosis in cancer cells. Flexible-Heteroarotinoids (Flex-Hets) are small molecules that regulate growth, differentiation and apoptosis in cancer cells with reduced effects on normal cells. Both imiquimod and SHetA2 inhibited growth and induced apoptosis in the B16 melanoma cell line and cisplatin-sensitive A2780 and cisplatin-resistant OVCAR-3 ovarian cancer cell lines. The growth inhibition was additive in A2780 and B16, and synergistic in OVCAR-3. Both compounds inhibited endothelial tube branching in vitro and exerted an additive effect when combined. Various combinations of imiquimod and SHetA2 did not cause significant differences in the overall survival in the syngeneic B16 murine melanoma model. SHetA2 induced complete tumor regression and a melanoma-free natural life-span in two mice. These cures occurred in one of ten mice treated with oral SHetA2 and one of ten mice intratumorally-injected with SHetA2. Exploratory modeling of the distribution of survival times suggested that the two surviving mice represent rare events. Histologic evaluation of the tumors revealed that imiquimod induced necrosis, SHetA2 induced differentiated architecture and increased cytoplasm, both agents reduced mitotic indices and angiogenesis and neither agent counteracted the effects of the other. No overt toxicities were observed. In conclusion, imiquimod and SHetA2 exhibit complementary anti-cancer activity in vitro and SHetA2 has promise as a single agent.

Controlled release of cisplatin and cancer cell apoptosis with cisplatin encapsulated poly(lactic-co-glycolic acid) nanoparticles

The goal of the present study is to utilize cis-diamminedichloroplatinum (cisplatin) loaded polymer nanoparticles (NPs) to give a controlled, extended, and local drug therapy for the treatment of cancer. We have used biodegradable and biocompatible poly(lactic-co-glycolic acid) (PLGA) to prepare the NPs by adjusting the double emulsion technique using poly(vinylalcohol) as a surface active agent. The PLGA NPs were characterized for particle size and shape, controlled release of cisplatin, and degradation. Cisplatin solubility in deionized water was increased up to 4 mg/mL by simply changing the solution parameters. Cisplatin encapsulated NPs were incubated in phosphate buffered saline (PBS) at 37?C to study the release kinetics of cisplatin. Cisplatin was released in a sustained manner with less than 20% release during a 3-day period followed by 50% release during a 21-day period. A degradation study of PLGA NPs demonstrated the loss of spherical shape during a 21-day period. We also examined the cisplatin sensitive A2780 cell apoptosis when cells were incubated with cisplatin encapsulated PLGA NPs. A large number of cell apoptosis occurred as a result of cisplatin release from the PLGA NPs. These results suggest that cisplatin encapsulated PLGA NPs can be used to treat the cancer cells by injecting them into a localized site minimizing the side effects.

Phosphaplatins, next generation platinum antitumor agents: A paradigm shift in designing and defining molecular targets

Platinum(II)- and platinum(IV)-pyrophosphato-complexes (phosphaplatins) show excellent antitumor properties against a variety of human ovarian cancers, including cisplatin- and carboplatin-sensitive and resistant human cancers. To identify the lead compound of this class of non-DNA binding antitumor agents, stereoisomers of cis- and trans-1,2-cyclohexane-diamine(pyrophosphato)platinum(II) and of cis- and trans-1,2-cyclohexane-trans-dihydroxo-diamine(pyrophosphato)platinum(IV) were synthesized, and their in vitro efficacies were evaluated in a variety of human ovarian cancer cells including cisplatin- and carboplatin-resistant cancer cells. Some of these compounds exhibited IC50 values as low as 0.40μM, the lowest ever observed for any platinum anticancer drugs under identical conditions. More importantly, these compounds are highly active against cisplatin- and carboplatin-sensitive and resistant cancers; examples of these cancer cells include cisplatin-sensitive A2780, acquired cisplatin- and carboplatin-resistant A2780/C30, and inherent cisplatin-resistant OVCAR-10 cells. In vivo efficacy determined by using human ovarian xenografts indeed confirmed that phosphaplatins are quite effective in treating ovarian cancers. The superiority of phosphaplatins was observed in increased life span and tumor regression. These non-DNA binding compounds exhibit cytotoxicity in ovarian cancers primarily through apoptosis via cell cycle arrest both at S and G2 phases largely in a dose independent manner. Platinum(II) compounds showed reduced protein binding compared to either cisplatin or carboplatin. Furthermore, protein binding by phosphaplatins was largely limited to extracellular and cytosolic compartments. In vivo experiments revealed that platinum accumulations in kidney of mice were significantly less compared to cisplatin. The maximum tolerated dose was found be >60mg/kg via intravenous administration. Furthermore, rats treated with R,R-stereomer of the platinum(II) complex exhibited near normal white blood cell, platelet, and neutrophil counts indicating its non-toxic properties. Furthermore, the lead compound also stays in plasma for 24h giving ample time to reach desired targets. Taken together, these data indicate that phosphaplatins have the potential to treat resistant ovarian cancers, both acquired and de novo, without exhibiting severe toxic effects.

Discovery of dachshund 2 protein as a novel biomarker of poor prognosis in epithelial ovarian cancer

BackgroundThe Dachshund homolog 2 (DACH2) gene has been implicated in development of the female genital tract in mouse models and premature ovarian failure syndrome, but to date, its expression in human normal and cancerous tissue remains unexplored. Using the Human Protein Atlas as a tool for cancer biomarker discovery, DACH2 protein was found to be differentially expressed in epithelial ovarian cancer (EOC). Here, the expression and prognostic significance of DACH2 was further evaluated in ovarian cancer cell lines and human EOC samples.MethodsImmunohistochemical expression of DACH2 was examined in tissue microarrays with 143 incident EOC cases from two prospective, population-based cohorts, including a subset of benign-appearing fallopian tubes (n = 32). A nuclear score (NS), i.e. multiplier of staining fraction and intensity, was calculated. For survival analyses, cases were dichotomized into low (NS < = 3) and high (NS > 3) using classification and regression tree analysis. Kaplan Meier analysis and Cox proportional hazards modelling were used to assess the impact of DACH2 expression on survival. DACH2 expression was analysed in the cisplatin sensitive ovarian cancer cell line A2780 and its cisplatin resistant derivative A2780-Cp70. The specificity of the DACH2 antibody was tested using siRNA-mediated silencing of DACH2 in A2780-Cp70 cells.ResultsDACH2 expression was considerably higher in the cisplatin resistant A2780-Cp70 cells compared to the cisplatin-sensitive A2780 cells. While present in all sampled fallopian tubes, DACH2 expression ranged from negative to strong in EOC. In EOC, DACH2 expression correlated with several proteins involved in DNA integrity and repair, and proliferation. DACH2 expression was significantly higher in carcinoma of the serous subtype compared to non-serous carcinoma. In the full cohort, high DACH2 expression was significantly associated with poor prognosis in univariable analysis, and in carcinoma of the serous subtype, DACH2 remained an independent factor of poor prognosis.ConclusionsThis study provides a first demonstration of DACH2 protein being expressed in human fallopian tubes and EOC, with the highest expression in serous carcinoma where DACH2 was found to be an independent biomarker of poor prognosis. Future research should expand on the role of DACH2 in ovarian carcinogenesis and chemotherapy resistance.

Synthesis, characterization and antiproliferative activity on mesothelioma cell lines of bis(carboxylato)platinum(iv) complexes based on picoplatin

The synthesis and characterization of a series of picoplatin-based (picoplatin = [PtCl(2)(mpy)(NH(3))], mpy = 2-methylpyridine), Pt(iv) complexes with axial carboxylato ligands of increasing length are reported. The synthesis is based on the oxidation with hydrogen peroxide of picoplatin to give the cis,cis,trans-[PtCl(2)(mpy)(NH(3))(OH)(2)] intermediate and then its transformation into the dicarboxylato complexes cis,cis,trans-[PtCl(2)(mpy)(NH(3))(RCOO)(2)] (R = CH(3)(CH(2))(n), n = 0-4) with the corresponding anhydride. Pt(iv) complexes with n = 0-2 were selected to be tested on four malignant pleural mesothelioma (MPM) cell lines, on human mesothelial cells (HMC), and on the cisplatin-sensitive ovarian A2780 cell line along with cisplatin as a metallo-drug reference. In general, the longer the axial chain, the more cytotoxic and selective the Pt(IV) complex is. Pt(IV) analogs show good activity on the MPM cell lines, approaching or in some case bypassing that of cisplatin and represent quite promising drug candidates for the treatment of tumors whose chemoresistance is mainly based on glutathione overexpression, such as MPM.

Cisplatin Resistant Patterns in Ovarian Cell Line Using FTIR and Principle Component Analysis.

Cisplatin is a common chemotherapeutic agent that used for treatment of many solid cancers. Rapid identification of chemotherapy resistance is very important and may lead to effective treatment plan. Spectroscopy techniques, such as infrared spectroscopy, which are sensitive to biochemical composition of samples, have shown potentials to discriminate tissues. Developing in Fourier transform infrared (FTIR) as a diagnostic tool support conventional technique in investigating cell phenotype. By this goal three different cell lines, two cisplatin resistant OV2008-DDP (C13) and A2780-CP ovarian cell lines and one cisplatin sensitive A2780 cell line were investigated by FTIR spectroscopy. Data were subjected to principle component analysis (PCA) to obtain FTIR pattern for cisplatin resistance. Using FTIR spectroscopy on these cells in the range of 400-4000 cm(-1) was shown dramatic change in cells. Results shows that Cisplatin resistance pattern is characterized in spectrum with the alteration of conformation in secondary structure of proteins and a shift toward the high wave numbers of CH2 stretching vibration. The FTIR data set between 1000 and 3000 cm(-1) could be consumed as biochemical typicality spectra among resistant and sensitive cell lines while correctly classified by PCA model. Our work supports the promise of PCA analysis of FTIR data as a powerful combined approach for the development of automated methods to recognize resistant to cisplatin in experimental cell lines. One of the advantages of this tool is to investigate the resistant percent of cancer cells .Such technique may bring new tool in cancer diagnosis and stage definition in cancerous tissues.