Arsenic Trioxide Treatment Research Articles

Additivity, antagonism, and synergy in arsenic trioxide-induced growth inhibition of C6 glioma cells: Effects of genistein, quercetin and buthionine-sulfoximine

Arsenic trioxide (ATO) induces clinical remission in acute promyelocytic leukemia and growth inhibition in various cancer cell lines in vitro. Recently, genistein and quercetin were reported to potentiate ATO-provoked apoptosis in leukemia and hepatocellular carcinoma cells. Genistein acted via enhanced ROS generation and quercetin via glutathione depletion. Searching for potential strategies for the treatment of malignant gliomas in this study the capacity of these flavonoids to sensitize rat C6 astroglioma cells for the cytotoxic action of ATO was investigated. ATO inhibited cell growth in a concentration- and time-dependent manner. This effect was accompanied neither by enhanced radical generation nor lipid peroxidation and was not attributed to apoptosis. ATO treatment concentration-dependently increased glutathione levels. Genistein enhanced radical generation. Combined with ATO it inhibited cell growth additively. Additivity was also obtained after cotreatment with ATO and H2O2. Quercetin acted antagonistically on ATO-induced growth inhibition. Quercetin increased glutathione levels. In contrast, buthionine-sulfoximine (BSO) depleted cellular glutathione and acted synergistically with ATO. In conclusion, in C6 cells neither genistein nor quercetin are suited as sensitizing agent, in contrast to BSO. Depletion of cellular glutathione content rather than an increase of ROS generation plays a central role in the enhancement of ATO-toxicity in C6 cells.

Combined arsenic trioxide-cisplatin treatment enhances apoptosis in oral squamous cell carcinoma cells

Oral squamous cell carcinoma (OSCC) accounts for the majority of oral cancers. Despite recent advances in OSCC diagnostics and therapeutics, the overall survival rate still remains low. Here, we assessed the efficacy of a combinatorial arsenic trioxide (ATO) and cisplatin (CDDP) treatment in human OSCC cells. The combinatorial effect of ATO/CDDP on the growth and apoptosis of OSCC cell lines HSC-2, HSC-3, and HSC-4 was evaluated using MTT and annexin V assays, respectively. Chou-Talalay analyses were preformed to evaluate the combinatorial effects of ATO/CDDP on the dose-reduction index (DRI). To clarify the mechanism underlying the ATO/CDDP anticancer effect, we also examined the involvement of reactive oxygen species (ROS) in ATO/CDDP-induced apoptosis. Combination index (CI) analyses revealed that a synergistic interaction of ATO and CDDP elicits a wide range of effects in HSC-2 cells, with CI values ranging from 0.78 to 0.90, where CI < 1 defines synergism. The CI values in HSC-3 and HSC-4 cells ranged from 0.34 to 0.45 and from 0.60 to 0.92, respectively. In addition, ATO/CDDP yielded favorable DRI values ranging from 1.6-fold to 7.71-fold dose reduction. Compared to mono-therapy, ATO/CDDP combinatorial therapy significantly augmented the loss of mitochondrial potential, caspase-3/7 activity and subsequent apoptosis. These changes were all abrogated by the antioxidant N-acetylcysteine. This study provides the first evidence for a synergistic ATO/CDDP anticancer (apoptotic) activity in OSCC cells with a favorable DRI, thereby highlighting its potential as a combinational therapeutic regime in OSCC.

Treatment of an Acute Promyelocytic Leukemia Relapse Using Arsenic Trioxide and All-Trans-Retinoic in a 6-Year-Old Child

In adult therapy, arsenic trioxide (ATO) and all-trans-retinoic acid (ATRA) are recognized as active treatment of relapsed acute promyelocytic leukemia (APL). The efficacy of this combination in pediatric APL has not yet been well established. We report the case of a 6-year-old girl with relapsed APL, with a PML-RARα mutation, treated with a combination of ATO and ATRA. Over a period of 5 months, she received in total, 75 doses of intravenous ATO and 40 doses of oral ATRA. Currently, 22 months after relapse, she is still in complete remission. Here, we describe treatment of a relapsed APL in a child with limited treatment of ATO and ATRA and review the literature.

Enhanced autophagy follows LMP1 expression disruption in EBV+ P3HR1 cells treated with arsenic trioxide.

Event Abstract Back to Event Enhanced autophagy follows LMP1 expression disruption in EBV+ P3HR1 cells treated with arsenic trioxide. Abderezak Zebboudj1*, Chafia Touil-Boukoffa1 and Mehdi Bourouba1 1 University of Sciences and Technologies Houari Boumediene (USTHB), FSB, Algeria Introduction : The Epstein Barr Virus (EBV) latency is associated with several malignancies among which nasopharyngeal carcinoma (NPC) and immunocompromised patients lymphoma. Expression and signaling of the major viral oncogene latent membrane protein 1 (LMP1) is essential to the immortalization of NPC cells and in vitro of B lymphocytes. Arsenic trioxide (ATO) treatment was reported to induce apoptosis in NPC cells via LMP1 down-regulation. Conversely, autophagy has been linked to LMP1 expression in B cells and survival. Here, we analyzed the anti-proliferative activity of ATO on P3HR1 EBV+ lymphoblastoid cells and assessed it's effect on apoptosis, autophagy and LMP1 expression. Materials and Methods : LMP1 expressing cells P3HR1 were treated with different concentrations of ATO. Apoptosis was measured by annexin V/PI double staining. DiOC6 labeling was assessed for mitochondrial potential (ΔΨm). Degree of autophagy was assessed based on LC3BI/LC3BII conversion. The anti-proliferative effect of ATO was studied using soft agar clonogenicity test. LMP1 expression and transcription was analyzed by western blot and Q-RT-PCR. Results : Our results show that ATO treatment leads to increased annexin V labeling and loss of mitochondrial potential in P3HR1 cells. We found that apoptosis is accompanied with enhanced autophagic activity as indicated by LC3B profile. Our results also indicate that ATO treatment induces disruption of LMP1 expression after 72H of treatment and inhibition of its transcription. Thus effects were associated with suppression of cells clonogenicity. Conclusion : Our study suggests that ATO cytotoxicity in P3HR1 cells occurs via induction of apoptosis and up-regulation of autophagic activity. These processes may occur as a consequence of LMP1 transcription repression and depletion of cellular LMP1 pools. Taken together, our data suggest that ATO may regulate negatively tumor cells survival by suppressing LMP1 anti-apoptotic signaling and enhancing autophagic cell death. Acknowledgements This work was performed and co-directed in collaboration with Dr Mounira Chelbi- Alix and Dr Sébastien Nisole team "INSERM UMR-S 1124, Université Paris Descartes,Paris, France". We thank Pr Mohamed Seghier "Institut Pasteur d'Algérie" for material and technical support. Keywords: ATO, apoptosis and autophagy, LMP1, ebv, Lymphoma, B-Cell Conference: The First International Congress of Immunology and Molecular Immunopathology (CIMIP2014), Tlemcen, Algeria, 17 Oct - 20 Oct, 2014. Presentation Type: Oral Poster Presentation Topic: Lymphoproliferative disorders Citation: Zebboudj A, Touil-Boukoffa C and Bourouba M (2014). Enhanced autophagy follows LMP1 expression disruption in EBV+ P3HR1 cells treated with arsenic trioxide.. Front. Immunol. Conference Abstract: The First International Congress of Immunology and Molecular Immunopathology (CIMIP2014). doi: 10.3389/conf.fimmu.2014.04.00007 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 10 Sep 2014; Published Online: 01 Dec 2014. * Correspondence: Mr. Abderezak Zebboudj, University of Sciences and Technologies Houari Boumediene (USTHB), FSB, Bab-Ezzouar, Algiers, 16111, Algeria, azebboudj@usthb.dz Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Abderezak Zebboudj Chafia Touil-Boukoffa Mehdi Bourouba Google Abderezak Zebboudj Chafia Touil-Boukoffa Mehdi Bourouba Google Scholar Abderezak Zebboudj Chafia Touil-Boukoffa Mehdi Bourouba PubMed Abderezak Zebboudj Chafia Touil-Boukoffa Mehdi Bourouba Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Realgar (As4S4) nanoparticles and arsenic trioxide (As2O3) induced autophagy and apoptosis in human melanoma cells in vitro.

The aim of the present study was to compare the effect of realgar nanoparticles and arsenic trioxide (ATO) on viability, DNA damage, proliferation, autophagy and apoptosis in the human melanoma cell lines BOWES and A375. The application of various flow cytometric methods for measurements cell viability, DNA cell cycle, mitochondrial potential, lysosomal activity, and intracellular content of glutathione was used. In addition, quantitative PCR, western blotting and multiplex bead array analyses were applied for evaluation of redox stress, autophagic flux, and cell signaling alterations.The results showed that realgar treatment of studied cells caused modulation of cell proliferation, induced a block in G2/M phase of the cell cycle and altered phosphorylation of IκB, Akt, ERK1/2, p38, and JNK kinases, as well as decreased mitochondrial membrane potential. Additionally, it appeared that induction of cell death by both realgar and ATO was dose-dependent, when lower (0.3 µM) dosage increased lysosomal activity and induced autophagy and higher (1.25 µM) concentration resulted in the appearance of apoptosis, while pan-caspase inhibitor attenuated more efficiently realgar- than ATO-induced cell death. Furthermore, low concentrations of ATO and realgar nanoparticles increased the content of intracellular glutathione and elevated γ-H2AX expression confirmed DNA damage preferentially at higher concentrations of both drugs used. Further analysis revealed slight differences in time-dependent phosphorylation pattern due to both realgar and ATO treatments, while significant differences were noticed between cell lines. In conclusion, realgar nanoparticles and ATO treatment induced dose-dependent activation of autophagy and apoptosis in both melanoma cell lines, when autophagy flux was determined at lower drug concentrations and the switch to apoptosis occurred at higher concentrations of both arsenic forms.

Arsenic Trioxide Overcomes Rapamycin-Induced Feedback Activation of AKT and ERK Signaling to Enhance the Anti-Tumor Effects in Breast Cancer

Inhibitors of the mammalian target of rapamycin (mTORi) have clinical activity; however, the benefits of mTOR inhibition by rapamycin and rapamycin-derivatives (rapalogs) may be limited by a feedback mechanism that results in AKT activation. Increased AKT activity resulting from mTOR inhibition can be a result of increased signaling via the mTOR complex, TORC2. Previously, we published that arsenic trioxide (ATO) inhibits AKT activity and in some cases, decreases AKT protein expression. Therefore, we propose that combining ATO and rapamycin may circumvent the AKT feedback loop and increase the anti-tumor effects. Using a panel of breast cancer cell lines, we find that ATO, at clinically-achievable doses, can enhance the inhibitory activity of the mTORi temsirolimus. In all cell lines, temsirolimus treatment resulted in AKT activation, which was decreased by concomitant ATO treatment only in those cell lines where ATO enhanced growth inhibition. Treatment with rapalog also results in activated ERK signaling, which is decreased with ATO co-treatment in all cell lines tested. We next tested the toxicity and efficacy of rapamycin plus ATO combination therapy in a MDA-MB-468 breast cancer xenograft model. The drug combination was well-tolerated, and rapamycin did not increase ATO-induced liver enzyme levels. In addition, combination of these drugs was significantly more effective at inhibiting tumor growth compared to individual drug treatments, which corresponded with diminished phospho-Akt and phospho-ERK levels when compared with rapamycin-treated tumors. Therefore, we propose that combining ATO and mTORi may overcome the feedback loop by decreasing activation of the MAPK and AKT signaling pathways.

Interplay of Loss of ERK Dependence and Amplification of Apoptotic Signals in Arsenic Treated Rat Hepatocytes

The present study was designed to demonstrate the loss of ERK dependence on amplified apoptotic signals in arsenic treated rat hepatocytes. Arsenic trioxide treatment (10 μM) exhibited simultaneous activation of caspase 9, caspase 3 and caspase 8. Moreover, expression and secretion of Fas ligand was also prominent. Reduction in nuclear translocation of pERK was directly associated with caspase activation. Treatment with ERK inhibitor further augmented caspase activation. An experiment with Fas receptor blocked cells confirmed self amplification of apoptotic signals via Fas R/caspase 8 interaction.

Both GLS silencing and GLS2 overexpression synergize with oxidative stress against proliferation of glioma cells

Mitochondrial glutaminase (GA) plays an essential role in cancer cell metabolism, contributing to biosynthesis, bioenergetics, and redox balance. Humans contain several GA isozymes encoded by the GLS and GLS2 genes, but the specific roles of each in cancer metabolism are still unclear. In this study, glioma SFxL and LN229 cells with silenced isoenzyme glutaminase KGA (encoded by GLS) showed lower survival ratios and a reduced GSH-dependent antioxidant capacity. These GLS-silenced cells also demonstrated induction of apoptosis indicated by enhanced annexin V binding capacity and caspase 3 activity. GLS silencing was associated with decreased mitochondrial membrane potential (ΔΨm) (JC-1 dye test), indicating that apoptosis was mediated by mitochondrial dysfunction. Similar observations were made in T98 glioma cells overexpressing glutaminase isoenzyme GAB, encoded by GLS2, though some characteristics (GSH/GSSG ratio) were different in the differently treated cell lines. Thus, control of GA isoenzyme expression may prove to be a key tool to alter both metabolic and oxidative stress in cancer therapy. Interestingly, reactive oxygen species (ROS) generation by treatment with oxidizing agents: arsenic trioxide or hydrogen peroxide, synergizes with either KGA silencing or GAB overexpression to suppress malignant properties of glioma cells, including the reduction of cellular motility. Of note, negative modulation of GLS isoforms or GAB overexpression evoked lower c-myc and bcl-2 expression, as well as higher pro-apoptotic bid expression. Combination of modulation of GA expression and treatment with oxidizing agents may become a therapeutic strategy for intractable cancers and provides a multi-angle evaluation system for anti-glioma pre-clinical investigations. Silencing GLS or overexpressing GLS2 induces growth inhibition in glioma cell lines. Inhibition is synergistically enhanced after arsenic trioxide (ATO) or H2O2 treatment. Glutatione levels decrease in GLS-silenced cells but augment if GLS2 is overexpressed. ROS synergistically inhibit cell migration by GLS silencing or GLS2 overexpression. c-myc, bid, and bcl-2 mediate apoptosis resulting from GLS silencing or GLS2 overexpression.

Oxidation Of Peroxiredoxins By Thioredoxin Inactivation Is Mediated In Arsenic Trioxide-Induced Reactive Oxygen Species Formation and Its Cytotoxicity In Acute Promyelocytic Leukemia Cells

BackgroundsThe Arsenic trioxide (ATO) is an effective cancer therapeutic drug for acute promyelocytic leukemia (APL). ATO exerts its effect mainly raising oxidative stress. However, not only the mechanisms of reactive oxygen species (ROS) generation by ATO but involvement of redox enzymes including peroxiredoxin (PRX) and thioredoxin (TRX) remains elusive. Aim of current study is to elucidate the mechanism of redox enzymes to elevate ROS during ATO-induced apoptosis in APL-derived NB4 cells. MethodsNB4 cell line, which is one of the human acute promyelocytic leukemia cell lines, was cultured in RPMI-1640 medium supplemented with 10% FBS in CO2 humidified atmosphere at 37°C. NB4 cells were cultured with 2 μM arsenic trioxide to induce apoptosis for 16-48 hours. Apoptosis was measured by staining with 7-amino-actinomycin D (7-AAD) with flow cytometry. 2, 7-dichlrodihydro-fluorescein-diacetate (H2DCF-DA) and MitoSOX Red were used to detect cellular and mitochondrial ROS. SO2 form for PRX I, PRX II, and PRX III was detected by western blot assay using PRX SO2 form-specific antibody. Monomer/Dimer assay for PRX I, PRX II, PRX III, and TRX I was performed by western blot using non-reducing gel. ResultsIntracellular ROS of NB4 cells was increased significantly after 16 hour of ATO but decreased after 24 hour of ATO. Mitochondrial ROS of NB4 cells was increased significantly after 39 hour of ATO. Apoptosis of NB4 cell after ATO treatment was increased as time elapsed (24% on 16hr, 26% on 24hr, 48% on 39hr, and 60% on 48hr). Monomer, indicated active and reduced form, of peroxiredoxins was decreased and cysteine sulfinic acid (CP–SO2H) peroxiredoxins, indicated inactive and oxidized peroxiredoxins, was increased in NB4 cells after ATO treatment as time goes by. Similarily, monomer of thioredoxin-1 (active thioredoxin) was decreased and multimer of thioredoxin-1 (inactive thioredoxin) was increased in NB4 cells after ATO treatment as time elapsed. ConclusionsOur data showed inactivation of peroxiredoxins by oxidation was developed during ATO-induced ROS generation and APL cell apoptosis. These peroxiredoxins oxidation was probably due to increment of reduced thioredoxin in NB4 cells after ATO treatment. These findings suggest ATO-induced anti-leukemic activity is more likely due to a TRX system-mediated cellular redox changes. Our study may provide the insights for finding novel targets in the development of new therapies, which potentiate ATO-induced apoptosis in APL cells. Disclosures:No relevant conflicts of interest to declare.

Ndfip Is a Novel Autophagy Regulator and It Is Deregulated In Acute Myeloid Leukemia

Abstract Dynamic protein turnover regulated through protein synthesis and degradation ensures cellular growth, proliferation and differentiation and has growing importance in cancer cells. Ndfip1 and Ndfip2 are known activators of the Nedd4 family of E3 ligases and regulate PTEN/Akt and MAP kinase signaling pathways. Autophagy is a major intracellular lysosomal degradation system, serving as a dynamic recycling platform, frequently deregulated in cancer. Acute myeloid leukemia (AML) has a constitutive Akt activation and autophagy suppression. Acute promyelocytic leukemia (APL) has increased protein accumulation and constitutive autophagy activation. Arsenic trioxide (ATO) is an effective treatment for APL and an established autophagy inductor. The crosstalks between ubiquitin proteasome system (UPS) and autophagy are an ongoing work and mostly unknown. We sought to investigate the expression levels of NDFIP1 and NDFIP2 in AML patients and to evaluate the role of both genes in autophagy. Bone marrow diagnostic samples from 27 patients with AML, 5 patients with APL and 15 healthy donors were analyzed, using qPCR. Functional studies were performed in vitro using the leukemic cell line U937 silenced for NDFIP1 (shNDFIP1) or NDFIP2 (shNDFIP2) through lentiviral particles. After puromycin selection and confirmation of transduction efficiency, cells were cultured upon ATO treatment 1µM or vehicle in the same concentration for 24h in order to induce autophagy. Acridine orange staining was used to quantify the percentage of acid vesicle organelles (AVO) and mitochondrial mass was estimated with Mitotracker Green, analyzed with flow cytometry. The expression of autophagic proteins (PIK3C3, Sqstm1, LC3 I/II) was investigated with western blot. NDFIP2 expression was significantly decreased in AML (P=0.04) and was increased in APL (P=0.018), compared with healthy donors. NDFIP2 expression also differed between AML and APL (P=0.002). NDFIP1 expression showed no differences among the subgroups analyzed. shNDFIP1 cells showed increased AVO percentage compared with the control. A slight AVO increase also occurred in shNDFIP2 cells. PIK3C3 expression was increased in shNDFIP1 and shNDFIP2 untreated cells. However, the mitochondrial mass was increased in both silenced cells. Moreover, shNDFIP1 and shNDFIP2 cells do not have modifications in LC3II or Sqstm1 expression. Interestingly, ATO treatment induced a significant increase of AVO only in control cells (P=0.01), but not in shNDFIP1 or shNDFIP2 cells. Increased AVO in control cells was accompanied by upregulation of PIK3C3 and LC3II and a decrease in Sqstm1. Conversely, ATO induced a downregulation of PIK3C3 and LC3II in silenced cells and upregulation of Sqstm1 in shNDFIP1 cells. The interplay between autophagy machinery and UPS is important under physiological conditions and has critical role in cancer. Autophagy is an essential adaptive response and modulates leukemic stemness. NDFIP is an endosomal protein with a role in vesicular trafficking. Better knowledge of UPS and autophagy intersections could lead to novel therapeutic targets for AML. Our results suggest that NDFIP is a novel autophagy regulator, as its silencing induces AVO accumulation, but without canonical autophagy activation, since the effector autophagy protein LC3II is decreased. NDFIP could be essential to autophagosome-lysosome fusion and its inhibition would block terminal stages of autophagy

Arsenic Trioxide Induces Apoptosis In Chronic Lymphocytic Leukemic Cells Through Down-Regulation Of Survivin Via The p53-Dependent Signaling Pathway

Arsenic trioxide induces cell apoptosis in a variety of tumor cells. However, the molecular mechanism underlying the killing effects of arsenic trioxide on tumor cells is not fully understood. Here, we demonstrate a signal pathway of arsenic trioxide treatment that leads to apoptosis in a B-cell chronic lymphocytic leukemia (WSU-CLL) cell line. Arsenic trioxide treatment significantly inhibited proliferation and induced apoptosis of WSU-CLL cells in a time- and dose-dependent manner. This anti-tumor effect of arsenic trioxide on WSU-CLL cells was mediated through the down-regulation of survivin and up-regulation of p53. Knock-down of intracellular survivin with siRNA potentiated the inhibitory effect of arsenic trioxide on proliferation in WSU-CLL cells. Also, knock-down of p53 by a specific siRNA prevented the down-regulation of survivin by arsenic trioxide and reduced the cytotoxic effect of arsenic trioxide to WSU-CLL cells. These results indicate that arsenic trioxide may be used to as novel therapeutic agent for chronic lymphocytic leukemia, which down-regulates survivin expression via a p53-dependent signaling pathway. Disclosures:No relevant conflicts of interest to declare.

Cost-Effectiveness Of Arsenic Trioxide In The Treatment Of Relapsed/Refractory Acute Promyelocytic Leukemia In Canada

Acute promyelocytic leukemia (APL) is a distinct and rare morphological, clinical and pathological variant of acute myeloid leukemia (AML). It represents approximately 10% to 15% of AML. APL is characterized by a high incidence of coagulopathy caused by disseminated intravascular coagulation and/or excessive fibrinolysis and is associated with a high early mortality. Current first-line treatments consist of all-trans retinoic acid (ATRA), anthracyclines and conventional chemotherapy (CT). Although considerable progress has been made in the first-line treatment of APL, about 20 to 30% of patients who achieved complete remission (CR) still relapseTrisenox® is a sterile injectable solution of arsenic trioxide (ATO) and has been approved in several countries, including Canada, for the induction of remission and consolidation in patients with APL who are refractory to, or have relapsed from, retinoid and anthracycline chemotherapy. At this time, ATO is recognized as the standard treatment for relapsed or refractory APL. However, it is not reimbursed yet by provincial public health care systems and was available through a special access program in Canada until product availability.The objective of this study was to assess, from a Canadian perspective, the economic impact of ATO in the treatment of patients with relapsed or refractory APL.A time-dependent Markov model was constructed to assess the cost-effectiveness of ATO compared to ATRA+CT in the treatment of relapsed/refractory APL. Because there was no head-to-head clinical trial available, data from the ATO treatment arm were taken from Soignet, 2001, while data for ATRA+CT were taken from Thomas, 2000. The comparative treatment was composed of ATRA + sequential CT including cytarabine, mitoxantrone or etoposide, followed by autologous hematopoietic stem cell transplantation (HSCT) in consolidation as described in Thomas, 2000. The Markov model comprises five health states: induction, second remission, treatment failure/relapse, post-failure, and death. The length of each Markov cycle was one month for the first 24-month study period then of one year. The model continued to run until all patients reached the absorbing state of death. All patients started in the induction state and could move to other health states thereafter. In case of treatment relapse/failure, patients were subsequently assigned to receive an allogeneic HSCT. The model also takes into account the incidence of treatment-induced grade 3-4 toxicity reported in both clinical trials (Soignet, 2001 and Thomas, 2000). Analyses were conducted from both a Canadian Ministry of Health (MoH) and a societal perspective over a lifetime horizon.In the treatment of relapsed/refractory APL, ATO is a cost-effective strategy over ATRA+CT, from both a health care system and a societal perspective. In fact, compared with ATRA+CT, ATO is associated with an incremental cost-effectiveness ratios (ICERs) of $20,443 per QALY and $22,219 per QALY, from a MoH and societal perspective respectively.Moreover, the results of the exhaustive sensitivity analysis confirm the robustness of the base-case results. In fact, according to the deterministic analysis results, ATO remained a cost-effective strategy compared with ATRA+CT from both perspectives. The ICERs vary between $9,785 and $40,732 / QALY from a MoH perspective and between $11,561 and $44,271 / QALY from a societal perspective. Results of the probabilistic sensitivity analysis indicated that, according to a willingness to pay of $50,000, ATO remains a cost-effective strategy in 99.37% and 98.98% of the simulations, from a MoH and a societal perspective respectively.In conclusion, this economic evaluation demonstrates that ATO+ATRA is a cost-effective strategy in the treatment of relapsed/refractory APL. Disclosures:Lachaine:Lundbeck Canada: Research Funding. Barakat:Lundbeck Canada: Employment.

Galectin-3 inhibition sensitizes human renal cell carcinoma cells to arsenic trioxide treatment

The anti-tumor effects of arsenic trioxide (ATO) were well established in acute promyelocytic leukemia, but not in renal cell carcinoma (RCC). Recent evidences indicate that galectin-3 (Gal-3) plays an anti-apoptotic role in chemotherapy induced tumor cell death. This study was intended to clarify the exact roles of Gal-3 performed in ATO-induced apoptosis in RCC cells. Weak apoptosis was observed in Gal-3-positive RCC cells (Caki-1, Caki-2, 786-0, and ACHN) following ATO treatment. However, ATO treatment upregulated Gal-3 expression concurrently caused a Synexin-cooperated translocation of Gal-3 from the nucleus to the cytoplasm. Gal-3-knockdown cells were more sensitive to ATO treatment as indicated by a strong mitochondria-dependent apoptosis following ATO treatment. Meanwhile, Gal-3 was found to inhibit ATO-induced apoptosis through enhancing Bcl-2 expression and stabilizing mitochondria. To confirm the results obtained from genetic method, we employed a Gal-3 inhibitor, modified citrus prectin (MCP), and co-treated the RCC cells with ATO. The cells showed an increased apoptosis in the syngeneic application of Gal-3 inhibition and ATO compared with ATO application alone. Based on these results, we conclude that Gal-3 inhibition sensitizes human renal cell carcinoma cells to ATO treatment through increasing mitochondria-dependent apoptosis. Our studies implicate synergetic application of ATO and Gal-3 inhibition as a potential strategy for RCC treatment.

Characteristics and prognosis analysis of additional chromosome abnormalities in newly diagnosed acute promyelocytic leukemia treated with arsenic trioxide as the front-line therapy

Currently, there are few studies that address the prognostic significance of baseline additional chromosomal abnormalities (ACAs) in newly diagnosed acute promyelocytic leukemia (APL) patients treated with arsenic trioxide (ATO) as the front-line therapy. A series of 271 consecutive APL patients has been cytogenetically investigated between 2004 and 2011 in our institution. The incidence of ACAs was 27% (46/172) in APL cases with t(15;17). Trisomy 8 was the most recurrent abnormality, accounting for 30% (14/46) of patients with ACAs, followed by +21 (7%, 3/46) and -7/7q (7%, 3/46). Nine cases (14.1%) were found to have additional balanced translocation aberrations, most of them are new and non-recurrent. Treatment protocols consisted of all-trans retinoic acid (ATRA) and chemotherapy with or without the ATO therapy. Overall, patients with and without ACAs had similar complete remission (CR) rates (94% and 98%, respectively, P=0.344). With a median follow-up of 41 months, univariate analysis showed that ACAs did not show any prognostic significance in relapse-free survival (RFS) and overall survival (OS). In addition, ATO treatment was an independent favorable predictor for RFS. Thus, this data provides insights into cytogenetic features of APL, and suggests that ATO-based combination therapy improved RFS in de novo APL patients, while ACAs had no impact on prognosis.

Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses.

Arsenic trioxide inhibits viability of pancreatic cancer stem cells in culture and in a xenograft model via binding to SHH-Gli

ObjectiveOverexpression of the sonic hedgehog (SHH) signaling pathway is an essential characteristic of pancreatic cancer stem cells (PCSCs) and arsenic trioxide (ATO) is described as a SHH inhibitor. This study evaluates whether ATO has the potential to inhibit viability of PCSCs via binding to SHH-Gli proteins.MethodsCell counting kit-8 and flow cytometry were used for analyzing apoptosis in cells in vitro. The animal model was an athymic nude mouse model bearing subcutaneous xenografts of SW1990 pancreatic cancer cells. The terminal deoxynucleotidyl transferase dUTP nick end labeling assay and immunohistochemistry were used for tumor tissue analysis. The interaction between Gli1 and ATO was examined by a confocal system and an ultraviolet absorption spectrum assay.ResultsATO induced apoptosis in pancreatic cancer cells, especially CD24+CD44+ cells in vitro. Combination treatment of ATO and low dose gemcitabine inhibited tumor growth by 60.9% (P = 0.004), and decreased the expression of CD24, CD44, and aldehyde dehydrogenase 1 family, member A1 significantly in vivo. ATO changed the structure of the recombinant Gli1 zinc finger peptides in a cell-free condition and the binding action of ATO to recombinant Gli1 was observed in cultured pancreatic cancer cells.ConclusionATO may have the potential to inhibit viability of PCSCs via binding to SHH-Gli proteins in vitro and in vivo.

ATRA and arsenic trioxide treatment for leukaemia

ATRA and arsenic trioxide treatment for leukaemia

In Vivo Effect of Arsenic Trioxide on Keap1-p62-Nrf2 Signaling Pathway in Mouse Liver: Expression of Antioxidant Responsive Element-Driven Genes Related to Glutathione Metabolism.

Arsenic is a Group I human carcinogen, and chronic arsenic exposure through drinking water is a major threat to human population. Liver is one of the major organs for the detoxification of arsenic. The present study was carried out in mice in vivo after arsenic treatment through drinking water at different doses and time of exposure. Arsenic toxicity is found to be mediated by reactive oxygen species. Nuclear factor (erythroid-2 related) factor 2 (Nrf2)/Keap1 (Kelch-like ECH-associated protein 1)/ARE (antioxidant response element)—driven target gene system protects cells against oxidative stress and maintains cellular oxidative homeostasis. Our result showed 0.4 ppm, 2 ppm, and 4 ppm arsenic trioxide treatment through drinking water for 30 days and 90 days induced damages in the liver of Swiss albino mice as evidenced by histopathology, disturbances in liver function, induction of heat shock protein 70, modulation of trace elements, alteration in reduced glutathione level, glutathione-s-transferase and catalase activity, malondialdehyde production, and induction of apoptosis. Cellular Nrf2 protein level and mRNA level increased in all treatment groups. Keap1 protein as well as mRNA level decreased concomitantly in arsenic treated mice. Our study clearly indicates the important role of Nrf2 in activating ARE driven genes related to GSH metabolic pathway and also the adaptive response mechanisms in arsenic induced hepatotoxicity.

Arsenic Trioxide Prevents Osteosarcoma Growth by Inhibition of GLI Transcription via DNA Damage Accumulation

The Hedgehog pathway is activated in various types of malignancies. We previously reported that inhibition of SMO or GLI prevents osteosarcoma growth in vitro and in vivo. Recently, it has been reported that arsenic trioxide (ATO) inhibits cancer growth by blocking GLI transcription. In this study, we analyzed the function of ATO in the pathogenesis of osteosarcoma. Real-time PCR showed that ATO decreased the expression of Hedgehog target genes, including PTCH1, GLI1, and GLI2, in human osteosarcoma cell lines. WST-1 assay and colony formation assay revealed that ATO prevented osteosarcoma growth. These findings show that ATO prevents GLI transcription and osteosarcoma growth in vitro. Flow cytometric analysis showed that ATO promoted apoptotic cell death. Comet assay showed that ATO treatment increased accumulation of DNA damage. Western blot analysis showed that ATO treatment increased the expression of γH2AX, cleaved PARP, and cleaved caspase-3. In addition, ATO treatment decreased the expression of Bcl-2 and Bcl-xL. These findings suggest that ATO treatment promoted apoptotic cell death caused by accumulation of DNA damage. In contrast, Sonic Hedgehog treatment decreased the expression of γH2AX induced by cisplatin treatment. ATO re-induced the accumulation of DNA damage attenuated by Sonic Hedgehog treatment. These findings suggest that ATO inhibits the activation of Hedgehog signaling and promotes apoptotic cell death in osteosarcoma cells by accumulation of DNA damage. Finally, examination of mouse xenograft models showed that ATO administration prevented the growth of osteosarcoma in nude mice. Because ATO is an FDA-approved drug for treatment of leukemia, our findings suggest that ATO is a new therapeutic option for treatment of patients with osteosarcoma.

SUMO1 negatively regulates the transcriptional activity of EVI1 and significantly increases its co-localization with EVI1 after treatment with arsenic trioxide

Aberrant expression of the proto-oncogene EVI1 (ecotropic virus integration site1) has been implicated not only in myeloid or lymphoid malignancies but also in colon, ovarian and breast cancers. Despite its importance in oncogenesis, the regulatory factors and mechanisms that potentiate the function of EVI1 and its consequences are partially known. Here we demonstrated that EVI1 is post-translationally modified by SUMO1 at lysine residues 533, 698 and 874. Although both EVI1 and SUMO1 were found to co-localize in nuclear speckles, the sumoylation mutant of EVI1 failed to co-localize with SUMO1. Sumoylation abrogated the DNA binding efficiency of EVI1 and also affected EVI1 mediated transactivation. The SUMO ligase PIASy was found to play a bi-directional role on EVI1, PIASy enhanced EVI1 sumoylation and augmented sumoylated EVI1 mediated repression. PIASy was also found to interact with EVI1 and impaired EVI1 transcriptional activity independent of its ligase activity. Arsenic trioxide (ATO) known to act as an antileukemic agent for acute promyelocytic leukemia (APL) not only enhanced EVI1 sumoylation but also enhanced the co-localization of EVI1 and SUMO1 in nuclear bodies distinct from PML nuclear bodies. ATO treatment also affected the Bcl-xL protein expression in EVI1 positive cell line. Thus, the results showed that arsenic treatment enhanced EVI1 sumoylation, deregulated Bcl-xL, which eventually may induce apoptosis in EVI1 positive cancer cells. The study for the first time explores and reports sumoylation of EVI1, which plays an essential role in regulating its function.