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We report the case of a 42-year-old female diagnosed with acute promyelocytic leukemia (APL), who developed differentiation syndrome (DS) on day 14 during induction therapy with all-trans retinoic acid (ATRA) and arsenic trioxide (ATO) with sudden-onset dyspnea, abdominal pain, tachycardia, and fever. Her laboratory findings were remarkable for acute kidney injury (AKI), worsening leukocytosis, thrombocytopenia, and lactic acidosis. She was also found to have flash pulmonary edema and a pericardial effusion. Despite immediate dexamethasone and methylprednisolone administration along with cessation of induction therapy, she continued to worsen and suffered a non-shockable cardiac arrest. Return of spontaneous circulation (ROSC) was achieved, but she was in profound shock requiring multiple vasopressors. The patient suffered repeat cardiac arrest later that day and passed away within 24 hours. DS is a potentially life-threatening complication in APL treatment, occurring in about 25% of APL patients and posing significant treatment challenges.

Arsenic trioxide (ATO) is an approved therapy for the treatment of acute promyelocytic leukemia, but the extension of arsenic-based therapies to other types of malignancies, notably tumor-forming cancers, has been slow. Nanodelivery vehicles offer a means of effectively delivering ATO to tumors. Very recently, there has been a series of developments in the formulation of arsenic-based nanomedicines that are not simply loaded with ATO. Realgar nanoparticles are comprised of molecular As4S4 units. Current studies suggest that realgar nanoparticles ultimately act in a manner similar to ATO, but with greatly attenuated toxic side effects. A drastically different approach is taken with arsenene nanosheets, a 2-dimensional form of elemental As. The electronic properties of this material allow it to mediate both photothermal therapy and photodynamic therapy. The exploration of these nanomaterials is still in its infancy but is poised to allow arsenic-based therapy to make yet another significant impact on cancer treatment.

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Acute myeloid leukemia (AML) is the most frequently diagnosed type of acute leukemia in adults. It commonly affects people aged 60 or older, as incidence increases with age, and it is characterized by the accumulation of immature hematopoietic progenitor cells in the bone marrow. Despite recent treatment advances and improvements for certain subtypes, as acute promyelocyte leukemia (APL), AML remains difficult to cure. While many patients reach remission after induction treatment, relapses are common and 5-year overall survival remains dismal. Long non-coding RNAs (lncRNAs) are involved in various regulatory cellular functions and, like coding genes, they are frequently dysregulated in cancer. In this thesis, the aim was to elucidate the functional implications of lncRNAs in the biology and treatment response of AML and normal hematopoiesis in order to improve understanding of AML pathology. In Paper I, whole-transcriptome sequencing identified the novel lncRNA MALNC. Clinical correlation analyses and CRISPR-knockout cell models were used to functionally explore its implications in AML. It was identified that enhanced MALNC expression is specifically associated with the AML-subtypes APL and AML with co-mutant NPM1/IDH2R140. Further, it was shown that MALNC is implicated in key factors of leukemogenesis, like differentiation and proliferation, and that MALNC expression associates with better overall survival in AML patients. Moreover, knockout of the MALNC gene sensitized AML cells to arsenic trioxide (ATO), all-trans retinoic acid (ATRA)-ATO combination and venetoclax treatment. In Paper II, three high-throughput functional CRISPR interference screens were performed to identify lncRNAs implicated in proliferation, differentiation or venetoclax response. Several novel lncRNAs were identified to potentially play a positive or negative role in these processes and furthermore were found to implicate AML prognosis. In Paper III, the lncRNA NEAT1 was studied in respect t

Acute myeloid leukemia (AML) is the most frequently diagnosed type of acute leukemia in adults. It commonly affects people aged 60 or older, as incidence increases with age, and it is characterized by the accumulation of immature hematopoietic progenitor cells in the bone marrow. Despite recent treatment advances and improvements for certain subtypes, as acute promyelocyte leukemia (APL), AML remains difficult to cure. While many patients reach remission after induction treatment, relapses are common and 5-year overall survival remains dismal. Long non-coding RNAs (lncRNAs) are involved in various regulatory cellular functions and, like coding genes, they are frequently dysregulated in cancer. In this thesis, the aim was to elucidate the functional implications of lncRNAs in the biology and treatment response of AML and normal hematopoiesis in order to improve understanding of AML pathology. In Paper I, whole-transcriptome sequencing identified the novel lncRNA MALNC. Clinical correlation analyses and CRISPR-knockout cell models were used to functionally explore its implications in AML. It was identified that enhanced MALNC expression is specifically associated with the AML-subtypes APL and AML with co-mutant NPM1/IDH2R140. Further, it was shown that MALNC is implicated in key factors of leukemogenesis, like differentiation and proliferation, and that MALNC expression associates with better overall survival in AML patients. Moreover, knockout of the MALNC gene sensitized AML cells to arsenic trioxide (ATO), all-trans retinoic acid (ATRA)-ATO combination and venetoclax treatment. In Paper II, three high-throughput functional CRISPR interference screens were performed to identify lncRNAs implicated in proliferation, differentiation or venetoclax response. Several novel lncRNAs were identified to potentially play a positive or negative role in these processes and furthermore were found to implicate AML prognosis. In Paper III, the lncRNA NEAT1 was studied in respect t

Introduction. Acute promyelocytic leukemia (APL) is a rare subtype of acute myeloid leukemia (AML), characterized by a particularly aggressive clinical behavior, that in the absence of treatment is usually fatal. The objective of this work was to determine the clinical and cytogenetic characteristics of a cohort of patients with APL, in order to evaluate their relationship with the outcome and prognosis of these patients. Methodology. An observational, descriptive, retrospective study of patients older than 15 years with a diagnosis of APL treated at the Hospital Universitario San Vicente Fundación, between 2012 and 2020, was carried out. Results. A total of 32 patients were included. The mean age at diagnosis was 37 years, 84.4% of the patients had the t(15;17) in the karyotype, and 93.75% had positive FISH. 12.5% of cases had a complex karyotype. Mortality in the first 30 days was 15.6%, with bleeding being the most common cause of death. All patients who survived achieved complete remission (84.3%). In an average follow-up of 24 months, 14.8% of cases relapsed. In the bivariate analysis, a relationship was found between the male sex and having a complex karyotype (p<0.015). No relationship was found between complex karyotype and early mortality (p=0.358), nor between complex karyotype and relapse (p=0.052). Conclusions. We present the clinical and cytogenetic characteristics of a cohort of patients with APL in Colombia. Central nervous system bleeding was the main cause of early mortality, with all surviving patients achieving complete remission on induction therapy. Mortality, complete remission and relapse rates were similar to those reported by other Latin American series, but lower than studies from European countries. Contrary to what has been reported in other studies, no relationship was found between complex karyotype and early mortality or relapse., Introducción. La leucemia promielocítica aguda (LPA) es un subtipo poco frecuente de leucemia mieloide aguda (LMA), que se caracteriza por un comportamiento clínico particularmente agresivo, y en ausencia de tratamiento, su curso generalmente es fatal. El objetivo de este trabajo fue determinar las características clínicas y citogenéticas de una cohorte de pacientes con LPA, con la finalidad de evaluar su relación con las complicaciones, el pronóstico y el desenlace de estos pacientes. Metodología. Se realizó un estudio observacional, descriptivo, retrospectivo de los pacientes mayores de 15 años con diagnóstico de LPA, atendidos en el Hospital Universitario San Vicente Fundación, entre los años 2012 a 2020. Resultados. Un total de 32 pacientes fueron incluidos. La edad media del diagnóstico fue 37 años. El 84,4% de los pacientes tenía la traslocación (15;17) en el cariotipo, y el 93,75% tenían FISH positivo. El 12,5% de los casos tenían cariotipo complejo. La mortalidad en los primeros 30 días fue del 15,6%, siendo el sangrado la causa de muerte más frecuente. Todos los pacientes que sobrevivieron alcanzaron la remisión completa (84,3%). En un promedio de seguimiento de 24 meses, el 14,8% de los casos recayeron. En el análisis bivariado se encontró relación entre sexo masculino y tener cariotipo complejo (p=0,015). No se encontró relación entre cariotipo complejo y mortalidad temprana (p=0,358), tampoco entre cariotipocomplejo y recaída (p=0,052). Conclusiones. Se presentan las características clínicas y citogenéticas de una cohorte de pacientes con LPA en Colombia. El sangrado en el sistema nervioso central fue la principal causa de mortalidad temprana, todos los pacientes que sobrevivieron alcanzaron la remisión completa con la terapia de inducción. Las tasas de mortalidad, remisión completa y recaída fueron similares a las reportadas por otras series latinoamericanas, pero inferiores a estudios provenientes de países europeos. Contrario a lo reportado en otros e

Acute myeloid leukemia (AML) is the most frequently diagnosed type of acute leukemia in adults. It commonly affects people aged 60 or older, as incidence increases with age, and it is characterized by the accumulation of immature hematopoietic progenitor cells in the bone marrow. Despite recent treatment advances and improvements for certain subtypes, as acute promyelocyte leukemia (APL), AML remains difficult to cure. While many patients reach remission after induction treatment, relapses are common and 5-year overall survival remains dismal. Long non-coding RNAs (lncRNAs) are involved in various regulatory cellular functions and, like coding genes, they are frequently dysregulated in cancer. In this thesis, the aim was to elucidate the functional implications of lncRNAs in the biology and treatment response of AML and normal hematopoiesis in order to improve understanding of AML pathology. In Paper I, whole-transcriptome sequencing identified the novel lncRNA MALNC. Clinical correlation analyses and CRISPR-knockout cell models were used to functionally explore its implications in AML. It was identified that enhanced MALNC expression is specifically associated with the AML-subtypes APL and AML with co-mutant NPM1/IDH2R140. Further, it was shown that MALNC is implicated in key factors of leukemogenesis, like differentiation and proliferation, and that MALNC expression associates with better overall survival in AML patients. Moreover, knockout of the MALNC gene sensitized AML cells to arsenic trioxide (ATO), all-trans retinoic acid (ATRA)-ATO combination and venetoclax treatment. In Paper II, three high-throughput functional CRISPR interference screens were performed to identify lncRNAs implicated in proliferation, differentiation or venetoclax response. Several novel lncRNAs were identified to potentially play a positive or negative role in these processes and furthermore were found to implicate AML prognosis. In Paper III, the lncRNA NEAT1 was studied in respect t

Acute myeloid leukemia (AML) is the most frequently diagnosed type of acute leukemia in adults. It commonly affects people aged 60 or older, as incidence increases with age, and it is characterized by the accumulation of immature hematopoietic progenitor cells in the bone marrow. Despite recent treatment advances and improvements for certain subtypes, as acute promyelocyte leukemia (APL), AML remains difficult to cure. While many patients reach remission after induction treatment, relapses are common and 5-year overall survival remains dismal. Long non-coding RNAs (lncRNAs) are involved in various regulatory cellular functions and, like coding genes, they are frequently dysregulated in cancer. In this thesis, the aim was to elucidate the functional implications of lncRNAs in the biology and treatment response of AML and normal hematopoiesis in order to improve understanding of AML pathology. In Paper I, whole-transcriptome sequencing identified the novel lncRNA MALNC. Clinical correlation analyses and CRISPR-knockout cell models were used to functionally explore its implications in AML. It was identified that enhanced MALNC expression is specifically associated with the AML-subtypes APL and AML with co-mutant NPM1/IDH2R140. Further, it was shown that MALNC is implicated in key factors of leukemogenesis, like differentiation and proliferation, and that MALNC expression associates with better overall survival in AML patients. Moreover, knockout of the MALNC gene sensitized AML cells to arsenic trioxide (ATO), all-trans retinoic acid (ATRA)-ATO combination and venetoclax treatment. In Paper II, three high-throughput functional CRISPR interference screens were performed to identify lncRNAs implicated in proliferation, differentiation or venetoclax response. Several novel lncRNAs were identified to potentially play a positive or negative role in these processes and furthermore were found to implicate AML prognosis. In Paper III, the lncRNA NEAT1 was studied in respect t

Acute myeloid leukemia (AML) is the most frequently diagnosed type of acute leukemia in adults. It commonly affects people aged 60 or older, as incidence increases with age, and it is characterized by the accumulation of immature hematopoietic progenitor cells in the bone marrow. Despite recent treatment advances and improvements for certain subtypes, as acute promyelocyte leukemia (APL), AML remains difficult to cure. While many patients reach remission after induction treatment, relapses are common and 5-year overall survival remains dismal. Long non-coding RNAs (lncRNAs) are involved in various regulatory cellular functions and, like coding genes, they are frequently dysregulated in cancer. In this thesis, the aim was to elucidate the functional implications of lncRNAs in the biology and treatment response of AML and normal hematopoiesis in order to improve understanding of AML pathology. In Paper I, whole-transcriptome sequencing identified the novel lncRNA MALNC. Clinical correlation analyses and CRISPR-knockout cell models were used to functionally explore its implications in AML. It was identified that enhanced MALNC expression is specifically associated with the AML-subtypes APL and AML with co-mutant NPM1/IDH2R140. Further, it was shown that MALNC is implicated in key factors of leukemogenesis, like differentiation and proliferation, and that MALNC expression associates with better overall survival in AML patients. Moreover, knockout of the MALNC gene sensitized AML cells to arsenic trioxide (ATO), all-trans retinoic acid (ATRA)-ATO combination and venetoclax treatment. In Paper II, three high-throughput functional CRISPR interference screens were performed to identify lncRNAs implicated in proliferation, differentiation or venetoclax response. Several novel lncRNAs were identified to potentially play a positive or negative role in these processes and furthermore were found to implicate AML prognosis. In Paper III, the lncRNA NEAT1 was studied in respect t

Patients with triple negative breast cancers (TNBCs)—highly aggressive tumors that do not express estrogen, progesterone, and human epidermal growth factor 2 receptors—have limited treatment options. Fewer than 30% of women with metastatic TNBC survive five years after their diagnosis, with a mortality rate within three months after a recurrence of 75%. Although TNBCs show a higher response to platinum therapy compared to other breast cancers, drug resistance remains a major obstacle; thus, platinum drugs with novel mechanisms are urgently needed. Arsenoplatins (APs) represent a novel class of anticancer agents designed to contain the pharmacophores of the two FDA approved drugs cisplatin and arsenic trioxide (As2O3) as one molecular entity. Here, we present the syntheses, crystal structures, DFT calculations, and antiproliferative activity of iodide analogs of AP-1 and AP-2, i.e., AP-5 and AP-4, respectively. Antiproliferative studies in TNBC cell lines reveal that all AP family members are more potent than cisplatin and As2O3 alone. DFT calculations demonstrate there is a low energy barrier for hydrolysis of the platinum-halide bonds in arsenoplatins, possibly contributing to their higher cytotoxicities compared to cisplatin.

Patients with triple negative breast cancers (TNBCs)—highly aggressive tumors that do not express estrogen, progesterone, and human epidermal growth factor 2 receptors—have limited treatment options. Fewer than 30% of women with metastatic TNBC survive five years after their diagnosis, with a mortality rate within three months after a recurrence of 75%. Although TNBCs show a higher response to platinum therapy compared to other breast cancers, drug resistance remains a major obstacle; thus, platinum drugs with novel mechanisms are urgently needed. Arsenoplatins (APs) represent a novel class of anticancer agents designed to contain the pharmacophores of the two FDA approved drugs cisplatin and arsenic trioxide (As2O3) as one molecular entity. Here, we present the syntheses, crystal structures, DFT calculations, and antiproliferative activity of iodide analogs of AP-1 and AP-2, i.e., AP-5 and AP-4, respectively. Antiproliferative studies in TNBC cell lines reveal that all AP family members are more potent than cisplatin and As2O3 alone. DFT calculations demonstrate there is a low energy barrier for hydrolysis of the platinum-halide bonds in arsenoplatins, possibly contributing to their higher cytotoxicities compared to cisplatin.

Acute promyelocytic leukemia is a variant of acute myeloid leukemia characterized by t(15;17) and PML/RAR alfa fusion gene. The discovery of the molecular pathogenesis has led to entitle all-trans retinoic acid (ATRA) as the first targeted therapy for acute leukemia. It is usually associated to anthracycline-based chemotherapy with high response rates, but potential long-term sequelae including therapy-related malignancies have been observed. Arsenic trioxide (ATO) was added to obviate these complications and investigational trials aimed to a new strategy with the incorporation of arsenic trioxide (ATO) into initial therapy instead of chemotherapy in selected patients. ATRA plus ATO without chemotherapy was the first attempt to treat low and intermediate-risk patients with APL. Our study aims to describe a monocentric cohort of patients with newly diagnosed APL effectively treated with ATO plus ATRA underlying its efficacy together with the high grade of tolerability of this association. From January 2009 to December 2019 23 APL patients were diagnosed and treated with ATO plus ATRA regimen: 14 males and 9 females patients with a median age of 45 years (range 18-72), for the majority intermediate risk (15 patients, 65%). The treatment was well tolerated and all patients achieved molecular remission after a median time of 3 months (range 1-6 months). All patients proceeded to consolidation phase as outpatients, they maintained complete molecular response at a median time of 44 months (range 15-127) except for 1 patient. All but one patient are alive and in response at a median follow-up of 48 months (range 9-141) without late effects. ATO plus ATRA regimen shows advantages in comparison to chemotherapy; in fact it allowed to treat patients in which chemotherapy could even not be applicable and it did not show secondary hematological diseases. The association of ATO to ATRA as chemo-free regimen enabled to treat APL even without chemotherapy.

Skeletni sistem je kompleksan organ koji se formira tokom embriogeneze, znatno uvećava tokom detinjstva, remodeluje tokom celog života i regeneriše nakon povreda. Formiranje kostiju je striktno kontrolisan proces, vremenski i prostorno. Prostorno gledano, granice skeletnog sistema su jasno ograničene u ljudskom telu. Van skeletna formacija kostiju ili heterotopna osifikacija (HO) je patološka pojava koja se javlja sporadično i predstavlja klinički problem jer dovodi do bola, otoka i potpune imobilizacije zglobova..., Skeletal system represents a very complex organ which forms during embryogenesis, develops during childhood, remodels during our lives and regenerates after injuries. Bone formation is strictly controlled, both spatially and temporally. Spatially, bones formation is restricted to the clearly defined skeleton boundaries. Ectopic bone formation or heterotopic ossification (HO) is a pathological bone formation which occurs periodically and is presented with a painful, swollen and motion restricted joints...

Survivin is the smallest and a well-studied member of the inhibitors of apoptosis proteins (IAPs) family, which is involved in the regulation of cell division, inhibition of both caspasedependent and -independent apoptosis in cancer cells and promotion of angiogenesis. Survivin is detectable during embryonic and foetal development but is undetectable in normal adult tissues. It is, however, expressed in transformed cell lines as well as in most common types of human cancers. Regulation of survivin remains poorly understood, and the discovery of the regulatory biomolecules, microRNAs (MiRs) present an interesting opportunity to investigate the regulation of this protein and its variants in cancers, especially breast cancer. Additionally, the expression of the survivin splice variants during cell cycle progression and apoptosis is not fully understood. The aims of this study were to investigate the role of arsenic trioxide on the expression of survivin splice variants and their specific microRNAs during cell cycle progression and apoptosis in human breast cancer MCF-7 cells. The study also aimed at ascertaining the toxicity and efficacy of using coal fly ash-derived β-cyclodextrin carbon nanospheres to deliver arsenic trioxide into the MCF-7 cells. Carbon nanospheres (CNSs) were synthesised using a chemical vapour deposition method while arsenic trioxide was deposited using wet impregnation method to form the arsenic trioxide-β-cyclodextrin carbon nanospheres (ATO-β-cyclodextrin-CNSs). The formation of the CNSs and the loading of arsenic trioxide to CNSs were confirmed using scanning electron microscopy/energy dispersive X-ray detection (SEM-EDX). The in vitro cytotoxicity effect of the β-cyclodextrin carbon nanospheres (CNSs), arsenic trioxide and arsenic trioxide-β-cyclodextrin CNSs against KMST-6 and MCF-7 cells was analysed using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5 diphenyltetrazolium bromide (MTT) Assay, Muse® Count and Viability Assay and light/fluorescence m

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Retinoic acid (RA) and arsenic target the t(15;17)(q24;q21) PML/RARA driver of acute promyelocytic leukemia (APL), their combination now curing over 95% patients. We report exome sequencing of 64 matched samples collected from patients at initial diagnosis, during remission, and following relapse after historical combined RA-chemotherapy treatments. A first subgroup presents a high incidence of additional oncogenic mutations disrupting key epigenetic or transcriptional regulators (primarily WT1) or activating MAPK signaling at diagnosis. Relapses retain these cooperating oncogenes and exhibit additional oncogenic alterations and/or mutations impeding therapy response (RARA, NT5C2). The second group primarily exhibits FLT3 activation at diagnosis, which is lost upon relapse together with most other passenger mutations, implying that these relapses derive from ancestral pre-leukemic PML/RARA-expressing cells that survived RA/chemotherapy. Accordingly, clonogenic activity of PML/RARA-immortalized progenitors ex vivo is only transiently affected by RA, but selectively abrogated by arsenic. Our studies stress the role of cooperating oncogenes in direct relapses and suggest that targeting pre-leukemic cells by arsenic contributes to its clinical efficacy.

Background: Pro-oxidant drugs have been proposed for treating certain cancers but the resistance developed by cancer cells to oxidative stress limits its potential use in clinics. To understand the mechanisms underlying resistance to oxidative stress, we found that the chronic exposure to an H2O2-generating system (ascorbate/menadione, Asc/Men) or catalase overexpression (CAT3 cells) increased the resistance of cancer cells to oxidative stress, likely by increasing the antioxidant status of cancer cells. Methods: Modulation of catalase expression was performed by either protein overexpression or protein down-regulation using siRNA against catalase and aminotriazole as pharmacological inhibitor. The former approach was done by transfecting cells with a plasmid construct containing human catalase cDNA (CAT3 cells, derived from MCF-7 breast cancer cell line) or by generating resistant cells through chronic exposure to an oxidant injury (Resox cells). Cell survival was monitored by using the MTT reduction assay and further calculation of IC50 values. Protein expression was done by Western blots procedures. The formation of reactive oxygen species was performed by flow cytometry. The transcriptional activity of human catalase promoter was assessed by using transfected cells with a plasmid containing the - 1518/+ 16 promoter domain. Results: Using Resox and CAT3 cells (derived from MCF-7 breast cancer cell line) as models for cancer resistance to pro-oxidative treatment, we found that arsenic trioxide (ATO) remarkably sensitized Resox and CAT3 cells to Asc/Men treatment. Since catalase is a key antioxidant enzyme involved in detoxifying Asc/Men (as shown by siRNA-mediated catalase knockdown) that is overexpressed in resistant cells, we hypothesized that ATO might regulate the expression levels of catalase. Consistently, catalase protein level is decreased in Resox cells when incubated with ATO likely by a decreased transcriptional activity of the catalase promoter. Conclu

Retinoic acid (RA) and arsenic target the t(15;17)(q24;q21) PML/RARA driver of acute promyelocytic leukemia (APL), their combination now curing over 95% patients. We report exome sequencing of 64 matched samples collected from patients at initial diagnosis, during remission, and following relapse after historical combined RA-chemotherapy treatments. A first subgroup presents a high incidence of additional oncogenic mutations disrupting key epigenetic or transcriptional regulators (primarily WT1) or activating MAPK signaling at diagnosis. Relapses retain these cooperating oncogenes and exhibit additional oncogenic alterations and/or mutations impeding therapy response (RARA, NT5C2). The second group primarily exhibits FLT3 activation at diagnosis, which is lost upon relapse together with most other passenger mutations, implying that these relapses derive from ancestral pre-leukemic PML/RARA-expressing cells that survived RA/chemotherapy. Accordingly, clonogenic activity of PML/RARA-immortalized progenitors ex vivo is only transiently affected by RA, but selectively abrogated by arsenic. Our studies stress the role of cooperating oncogenes in direct relapses and suggest that targeting pre-leukemic cells by arsenic contributes to its clinical efficacy.

Retinoic acid (RA) in association with chemotherapy or with arsenic trioxide (ATO) results in high cure rates of acute promyelocytic leukemia (APL). We show that RA-induced differentiation of human leukemic cell lines and primary blasts dramatically increases their sensitivity to endoplasmic reticulum (ER) stress-inducing drugs at doses that are not toxic in the absence of RA. In addition, we demonstrate that the PERK pathway, triggered in response to ER stress, has a major protective role. Moreover, low amounts of pharmacologically induced ER stress are sufficient to strongly increase ATO toxicity. Indeed, in the presence of ER stress, ATO efficiently induced apoptosis in RA-sensitive and RA-resistant APL cell lines, at doses ineffective in the absence of ER stress. Our findings identify the ER stress-related pathways as potential targets in the search for novel therapeutic strategies in AML.

Background Arsenic trioxide (As2O3), a drug that has been used in China for approximately two thousand years, induces cell death in a variety of cancer cell types, including neuroblastoma (NB). The tyrosine kinase receptor (Trk) family comprises three members, namely TrkA, TrkB and TrkC. Various studies have confirmed that TrkA and TrkC expression is associated with a good prognosis in NB, while TrkB overexpression can lead to tumor cell growth and invasive metastasis. Previous studies have shown that As2O3 can inhibit the growth and proliferation of a human NB cell line and can also affect the N-Myc mRNA expression. It remains unclear whether As2O3 regulates Trks for the purposes of treating NB. Methods The aim of the present study was to investigate the effect of As2O3 on Trk expression in NB cell lines and its potential therapeutic efficacy. SK-N-SH cells were grown with increasing doses of As2O3 at different time points. We cultured SK-N-SH cells, which were treated with increasing doses of As2O3 at different time points. Trk expression in the NB samples was quantified by immunohistochemistry, and the cell cycle was analyzed by flow cytometry. TrkA, TrkB and TrkC mRNA expression was evaluated by real-time PCR analysis. Results Immunohistochemical and real-time PCR analyses indicated that TrkA and TrkC were over-expressed in NB, and specifically during stages 1, 2 and 4S of the disease progression. TrkB expression was increased in stage 3 and 4 NB. As2O3significantly arrested SK-N-SH cells in the G2/M phase. In addition, TrkA, TrkB and TrkC expression levels were significantly upregulated by higher concentrations of As2O3 treatment, notably in the 48-h treatment period. Our findings suggested that to achieve the maximum effect and appropriate regulation of Trk expression in NB stages 1, 2 and 4S, As2O3 treatment should be at relatively higher concentrations for longer delivery times;however, for NB stages 3 and 4, an appropriate concentration and infusio

Gastrointestinal stromal tumors (GIST) arise within the interstitial cell of Cajal (ICC) lineage due to activating KIT/PDGFRA mutations. Both ICC and GIST possess primary cilia (PC), which coordinate PDGFRA and Hedgehog signaling, regulators of gastrointestinal mesenchymal development. Therefore, we hypothesized that Hedgehog signaling may be altered in human GIST and controls KIT expression. Quantitative RT-PCR, microarrays, and next generation sequencing were used to describe Hedgehog/PC-related genes in purified human ICC and GIST. Genetic and pharmacologic approaches were employed to investigate the effects of GLI manipulation on KIT expression and GIST cell viability. We report that Hedgehog pathway and PC components are expressed in ICC and GIST and subject to dysregulation during GIST oncogenesis, irrespective of KIT/PDGFRA mutation status. Using genomic profiling, 10.2% of 186 GIST studied had potentially deleterious genomic alterations in 5 Hedgehog-related genes analyzed, including in the PTCH1 tumor suppressor (1.6%). Expression of the predominantly repressive GLI isoform, GLI3, was inversely correlated with KIT mRNA levels in GIST cells and non-KIT/non-PDGFRA mutant GIST. Overexpression of the 83-kDa repressive form of GLI3 or small interfering RNA-mediated knockdown of the activating isoforms GLI1/2 reduced KIT mRNA. Treatment with GLI1/2 inhibitors, including arsenic trioxide, significantly increased GLI3 binding to the KIT promoter, decreased KIT expression, and reduced viability in imatinib-sensitive and imatinib-resistant GIST cells. These data offer new evidence that genes necessary for Hedgehog signaling and PC function in ICC are dysregulated in GIST. Hedgehog signaling activates KIT expression irrespective of mutation status, offering a novel approach to treat imatinib-resistant GIST.

The main aim of this project is to find a suitable ATO concentration to combine with already approved chemotherapeutic agents to find that synergistic effect in triple negative breast cancer MDA-MB 231 cell line, as a new strategy to treat the disease.

The main aim of this project is to find a suitable ATO concentration to combine with already approved chemotherapeutic agents to find that synergistic effect in triple negative breast cancer MDA-MB 231 cell line, as a new strategy to treat the disease.

Gastrointestinal stromal tumors (GIST) arise within the interstitial cell of Cajal (ICC) lineage due to activating KIT/PDGFRA mutations. Both ICC and GIST possess primary cilia (PC), which coordinate PDGFRA and Hedgehog signaling, regulators of gastrointestinal mesenchymal development. Therefore, we hypothesized that Hedgehog signaling may be altered in human GIST and controls KIT expression. Quantitative RT-PCR, microarrays, and next generation sequencing were used to describe Hedgehog/PC-related genes in purified human ICC and GIST. Genetic and pharmacologic approaches were employed to investigate the effects of GLI manipulation on KIT expression and GIST cell viability. We report that Hedgehog pathway and PC components are expressed in ICC and GIST and subject to dysregulation during GIST oncogenesis, irrespective of KIT/PDGFRA mutation status. Using genomic profiling, 10.2% of 186 GIST studied had potentially deleterious genomic alterations in 5 Hedgehog-related genes analyzed, including in the PTCH1 tumor suppressor (1.6%). Expression of the predominantly repressive GLI isoform, GLI3, was inversely correlated with KIT mRNA levels in GIST cells and non-KIT/non-PDGFRA mutant GIST. Overexpression of the 83-kDa repressive form of GLI3 or small interfering RNA-mediated knockdown of the activating isoforms GLI1/2 reduced KIT mRNA. Treatment with GLI1/2 inhibitors, including arsenic trioxide, significantly increased GLI3 binding to the KIT promoter, decreased KIT expression, and reduced viability in imatinib-sensitive and imatinib-resistant GIST cells. These data offer new evidence that genes necessary for Hedgehog signaling and PC function in ICC are dysregulated in GIST. Hedgehog signaling activates KIT expression irrespective of mutation status, offering a novel approach to treat imatinib-resistant GIST.

The main aim of this project is to find a suitable ATO concentration to combine with already approved chemotherapeutic agents to find that synergistic effect in triple negative breast cancer MDA-MB 231 cell line, as a new strategy to treat the disease.

ImportanceTumor resistance is an emerging problem for Smoothened (SMO) inhibitor-treated metastatic basal cell carcinoma (BCC). Arsenic trioxide and itraconazole antagonize the hedgehog (HH) pathway at sites distinct from those treated by SMO inhibitors.ObjectiveTo determine whether administration of intravenous arsenic trioxide and oral itraconazole in patients with metastatic BCC is associated with a reduction in GLI1 messenger RNA expression in tumor and/or normal skin biopsy samples.Design, setting, and participantsFive men with metastatic BCC who experienced relapse after SMO inhibitor treatment underwent intravenous arsenic trioxide treatment for 5 days, every 28 days, and oral itraconazole treatment on days 6 to 28. Data were collected from April 10 to November 14, 2013. Follow-up was completed on October 3, 2015, and data were analyzed from June 5 to October 6, 2015.Main outcomes and measuresThe primary outcome was the change in messenger RNA levels of the GLI family zinc finger 1 (GLI1) gene (HH-pathway target gene) in biopsy specimens of normal skin or BCC before and after treatment. Secondary objectives were evaluation of tumor response and tolerability.ResultsOf the 5 patients (mean [SD] age, 52 [9] years; age range, 43-62 years), 3 completed 3 cycles of treatment and 2 discontinued treatment early owing to disease progression or adverse events. Adverse effects included grade 2 transaminitis and grade 4 leukopenia with a grade 3 infection. Overall, arsenic trioxide and itraconazole reduced GLI1 messenger RNA levels by 75% from baseline (P < .001). The best overall response after 3 treatment cycles was stable disease in 3 patients.Conclusions and relevanceTargeting the HH pathway with sequential arsenic trioxide and itraconazole treatment is a feasible treatment for metastatic BCC. Although some patients experienced stable disease for 3 months, none had tumor shrinkage, which may be owing to transient GLI1 suppression with sequential dosing. Continuous

The main aim of this project is to find a suitable ATO concentration to combine with already approved chemotherapeutic agents to find that synergistic effect in triple negative breast cancer MDA-MB 231 cell line, as a new strategy to treat the disease.

CLL remains an incurable disease in spite of the many new compounds being tested. Arsenic trioxide (ATO) induces apoptosis in all CLL cell types and could constitute an efficient therapy. To further explore this, we have studied the gene expression profile induced by ATO in CLL cells. ATO modulated many genes, largely involved in oxidative stress, being HMOX1 the most upregulated gene, also induced at the protein level. ATO also increased MMP-9, as we previously observed, both at the mRNA and protein level. Using specific inhibitors, qPCR analyses, and gene silencing approaches we demonstrate that upregulation of MMP-9 by ATO involved activation of the p38 MAPK/AP-1 signaling pathway. Moreover, gene silencing HMOX1 or inhibiting HMOX1 activity enhanced p38 MAPK phosphorylation and c-jun expression/activation, resulting in transcriptional upregulation of MMP-9. Overexpression of HMOX1 or enhancement of its activity, had the opposite effect. Cell viability analyses upon modulation of HMOX1 expression or activity demonstrated that HMOX1 had a proapoptotic role and enhanced the cytotoxic effect of ATO in CLL cells. We have therefore identified a new mechanism in which HMOX1 plays a central role in the response of CLL cells to ATO and in the regulation of the anti-apoptotic protein MMP-9. Thus, HMOX1 arises as a new therapeutic target in CLL and the combination of HMOX1 modulators with ATO may constitute an efficient therapeutic strategy in CLL.

The Giant mine in the Northwest Territories, Canada, operated from 1948 to 1999, producing more than 7 000 000 oz of gold. Chemical roasting of the ore began in 1949 and sulphur dioxide and arsenic vapour were vented directly to the atmosphere. The first air emission controls were introduced in 1951 and captured arsenic in the form of arsenic trioxide. When control of the mine property was transferred to the federal department of Indian Affairs and Northern Development in 1999, the roaster complex had within it free and poorly contained arsenic trioxide dust, asbestos, cyanide and other hazardous substances. As the complex progressively deteriorated over time, the risks to neighbouring communities, onsite workers and environment increased, with an increasing likelihood of building collapse and releases of As, asbestos and CN. The ten primary structures included the Allis-Chalmers roaster building, calcine plant, the Dorrco roaster building, the Cottrell precipitator building, the baghouse building, the silo, the weight scale house, the stack fan building, the stack and the exterior flues and other ancillary infrastructure. A project team, consisting of individuals from across governmental departments, communications officers, health and safety officers, managers, consultants and contractors, was critical to successfully meeting the objectives. Over a two-year period, the ten structures were successfully dismantled, the non-hazardous and hazardous wastes were segregated and more than 3 000 t of As-contaminated materials removed and containerised without adverse effect on onsite workers, neighbouring communities or the environment., The Giant mine in the Northwest Territories, Canada, operated from 1948 to 1999, producing more than 7 000 000 oz of gold. Chemical roasting of the ore began in 1949 and sulphur dioxide and arsenic vapour were vented directly to the atmosphere. The first air emission controls were introduced in 1951 and captured arsenic in the form of arsenic trioxide. When control of the mine property was transferred to the federal department of Indian Affairs and Northern Development in 1999, the roaster complex had within it free and poorly contained arsenic trioxide dust, asbestos, cyanide and other hazardous substances. As the complex progressively deteriorated over time, the risks to neighbouring communities, onsite workers and environment increased, with an increasing likelihood of building collapse and releases of As, asbestos and CN. The ten primary structures included the Allis-Chalmers roaster building, calcine plant, the Dorrco roaster building, the Cottrell precipitator building, the baghouse building, the silo, the weight scale house, the stack fan building, the stack and the exterior flues and other ancillary infrastructure. A project team, consisting of individuals from across governmental departments, communications officers, health and safety officers, managers, consultants and contractors, was critical to successfully meeting the objectives. Over a two-year period, the ten structures were successfully dismantled, the non-hazardous and hazardous wastes were segregated and more than 3 000 t of As-contaminated materials removed and containerised without adverse effect on onsite workers, neighbouring communities or the environment.

Gold at the Giant mine in the Northwest Territories, Canada, was found to be locked in minerals which required roasting at extremely high temperatures, releasing gases with highly toxic arsenic trioxide. Throughout the 1950s, controls were put in place that minimised emissions to the air, but resulted in the collection of 237 000 t of highly toxic arsenic trioxide dust which was stored in underground stopes and chambers. Ore processing ceased in 1999 and a remediation project was created in 2005. The frozen block method was selected for the containment of the arsenic trioxide, which involves freezing the rock around the chambers containing the dust and maintaining this frozen state through the use of thermosyphons, designed for either active or passive heat transfer. An impermeable frozen zone is created which will completely encapsulate the arsenic dust indefinitely. The Baker Creek channel has been altered significantly during operation of the mine, which has created a number of issues. Water and sediment quality are significantly impacted by historical spills, the discharge of treated mine water and offsite inputs upstream has affected habitat development and quality, and certain reaches of the creek pass close to pits which are at risk of flooding. Options range from re-routing the entire creek to a new route that avoids the mine site entirely to making modifications to certain reaches. About 9 500 000 t of tailings were originally deposited in the north, central and south ponds, of which about 7 000 000 t remain. The ultimate remedial concept is a cover to prevent human or wildlife contact, and to encourage vegetation growth and shed runoff/prevent infiltration of surface water. To increase the stability of the underground mine workings, filling of the voids using a paste backfill delivered through surface drill holes has begun. Waste rock will be used to seal up most of the lateral openings, including the portals located in the open pits, and may also be suita, Gold at the Giant mine in the Northwest Territories, Canada, was found to be locked in minerals which required roasting at extremely high temperatures, releasing gases with highly toxic arsenic trioxide. Throughout the 1950s, controls were put in place that minimised emissions to the air, but resulted in the collection of 237 000 t of highly toxic arsenic trioxide dust which was stored in underground stopes and chambers. Ore processing ceased in 1999 and a remediation project was created in 2005. The frozen block method was selected for the containment of the arsenic trioxide, which involves freezing the rock around the chambers containing the dust and maintaining this frozen state through the use of thermosyphons, designed for either active or passive heat transfer. An impermeable frozen zone is created which will completely encapsulate the arsenic dust indefinitely. The Baker Creek channel has been altered significantly during operation of the mine, which has created a number of issues. Water and sediment quality are significantly impacted by historical spills, the discharge of treated mine water and offsite inputs upstream has affected habitat development and quality, and certain reaches of the creek pass close to pits which are at risk of flooding. Options range from re-routing the entire creek to a new route that avoids the mine site entirely to making modifications to certain reaches. About 9 500 000 t of tailings were originally deposited in the north, central and south ponds, of which about 7 000 000 t remain. The ultimate remedial concept is a cover to prevent human or wildlife contact, and to encourage vegetation growth and shed runoff/prevent infiltration of surface water. To increase the stability of the underground mine workings, filling of the voids using a paste backfill delivered through surface drill holes has begun. Waste rock will be used to seal up most of the lateral openings, including the portals located in the open pits, and may also be suita

The clinical characteristics, treatment options and outcomes in patients with acute promyelocytic leukaemia (APL) and hyperleucocytosis remain poorly defined. This study reviewed 242 consecutive patients with APL; 29 patients (12%) had a white blood cell count (WBC) ≥ 50 × 10(9) /l at presentation (median WBC 85·5 × 10(9) /l). Patients with hyperleucocytosis had inferior complete remission (CR) rates (69% vs. 88%; P = 0·004) and higher 4-week mortality (24% vs. 9%; P = 0·018) compared to patients without hyperleucocytosis. We noted a trend towards inferior 3-year disease-free survival (DFS) (69% vs. 80%; P = 0·057) and inferior 3-year overall survival (OS) (74% vs. 92%; P = 0·2) for patients with hyperleucocytosis. Leukapheresis was performed in 11 (38%) of the 29 patients with hyperleucocytosis. CR rate and 3-year OS were not significantly improved in patients who received leukapheresis. CR rate and 3-year OS for the 15 patients with hyperleucocytosis treated with all-trans retinoic acid (ATRA) plus arsenic trioxide (ATO) plus cytotoxic therapy (idarubicin or gemtuzumab ozogamicin) combinations were 100% and 100% vs. 57% and 35% for the 14 patients treated with non-ATRA/ATO combinations (P = 0·004 and P = 0·002). Leukapheresis does not improve the outcomes in patients with APL presenting with hyperleucocytosis. ATRA/ATO-based combinations are superior to other regimens in these patients.

The p53 gene is mutated in more than 50% of human tumors. Mutant p53 exerts an oncogenic function and is often highly expressed in cancer cells due to evasion of proteasome-dependent degradation. Thus, reactivating proteasome-dependent degradation of mutant p53 protein is an attractive strategy for cancer management. Previously, we found that arsenic trioxide (ATO), a drug for acute promyelocytic leukemia, degrades mutant p53 protein through a proteasome pathway. However, it remains unclear what is the E3 ligase that targets mutant p53 for degradation. In current study, we sought to identify an E3 ligase necessary for ATO-mediated degradation of mutant p53. We found that ATO induces expression of Pirh2 E3 ligase at the transcriptional level. We also found that knockdown of Pirh2 inhibits, whereas ectopic expression of Pirh2 enhances, ATO-induced degradation of mutant p53 protein. Furthermore, we found that Pirh2 E3 ligase physically interacts with and targets mutant p53 for polyubiquitination and subsequently proteasomal degradation. Interestingly, we found that ATO cooperates with HSP90 or HDAC inhibitor to promote mutant p53 degradation and growth suppression in tumor cells. Together, these data suggest that ATO promotes mutant p53 degradation in part via induction of the Pirh2-dependent proteasome pathway.

The p53 gene is mutated in more than 50% of human tumors. Mutant p53 exerts an oncogenic function and is often highly expressed in cancer cells due to evasion of proteasome-dependent degradation. Thus, reactivating proteasome-dependent degradation of mutant p53 protein is an attractive strategy for cancer management. Previously, we found that arsenic trioxide (ATO), a drug for acute promyelocytic leukemia, degrades mutant p53 protein through a proteasome pathway. However, it remains unclear what is the E3 ligase that targets mutant p53 for degradation. In current study, we sought to identify an E3 ligase necessary for ATO-mediated degradation of mutant p53. We found that ATO induces expression of Pirh2 E3 ligase at the transcriptional level. We also found that knockdown of Pirh2 inhibits, whereas ectopic expression of Pirh2 enhances, ATO-induced degradation of mutant p53 protein. Furthermore, we found that Pirh2 E3 ligase physically interacts with and targets mutant p53 for polyubiquitination and subsequently proteasomal degradation. Interestingly, we found that ATO cooperates with HSP90 or HDAC inhibitor to promote mutant p53 degradation and growth suppression in tumor cells. Together, these data suggest that ATO promotes mutant p53 degradation in part via induction of the Pirh2-dependent proteasome pathway.

The p53 gene is mutated in more than 50% of human tumors. Mutant p53 exerts an oncogenic function and is often highly expressed in cancer cells due to evasion of proteasome-dependent degradation. Thus, reactivating proteasome-dependent degradation of mutant p53 protein is an attractive strategy for cancer management. Previously, we found that arsenic trioxide (ATO), a drug for acute promyelocytic leukemia, degrades mutant p53 protein through a proteasome pathway. However, it remains unclear what is the E3 ligase that targets mutant p53 for degradation. In current study, we sought to identify an E3 ligase necessary for ATO-mediated degradation of mutant p53. We found that ATO induces expression of Pirh2 E3 ligase at the transcriptional level. We also found that knockdown of Pirh2 inhibits, whereas ectopic expression of Pirh2 enhances, ATO-induced degradation of mutant p53 protein. Furthermore, we found that Pirh2 E3 ligase physically interacts with and targets mutant p53 for polyubiquitination and subsequently proteasomal degradation. Interestingly, we found that ATO cooperates with HSP90 or HDAC inhibitor to promote mutant p53 degradation and growth suppression in tumor cells. Together, these data suggest that ATO promotes mutant p53 degradation in part via induction of the Pirh2-dependent proteasome pathway.

Small cell lung carcinoma (SCLC) is a very aggressive solid tumor and is often widely metastasized by the time of diagnosis. Despite good response to the initial chemotherapy, SCLC cells often develop multidrug resistance to conventionally used chemotherapeutic drugs, which cause almost all SCLC tumors to relapse. The 5-year survival rate for patients with limited disease is around 20% while it is only a few percent for patients with a disseminating disease. Arsenic trioxide (As2O3) is one of the oldest medicines used for treatment of different diseases and it is today used as first-line treatment for patients with relapsed or refractory acute promyelocytic leukemia. Here, we demonstrate that As2O3 is cytotoxic to SCLC cells and xenotransplanted SCLC tumors at clinically relevant concentrations and the effect is also sustained at hypoxic conditions. Areas of low oxygen tensions, hypoxia, are a common characteristic in solid tumors and are associated with aggressive tumor behavior, treatment resistance and poor outcome in several tumor forms. In response to hypoxia, tumor cells induce a transcriptional shift which is mainly regulated by the transcription factors hypoxia-inducible factor (HIF)-1 and HIF-2. HIF proteins consist of two subunits, an oxygen-regulated α-subunit and a constitutively expressed β-subunit. Previous reports have shown that the transcription factors are differentially regulated over time; HIF-1 primarily mediates the acute hypoxic response, whereas HIF-2 dominates during more chronic phases of hypoxia. We found that SCLC tumor specimens and cells lack expression of HIF-2α protein while HIF-1α is expressed at both acute and prolonged hypoxia. In addition, SCLC cells have a high adaptive capacity to hypoxia including a high proliferation rate and low cell death, even though we demonstrated a modest induction of well-known hypoxia-driven genes. We further show that knockdown of HIF1A using siRNA or shRNA, is not significantly affecting the cell via

Roasting arsenopyrite-bearing gold ore for more than fifty years has resulted in nearly 300 000 tons of arsenic trioxide waste at the Giant mine, Yellowknife, NWT, Canada, mostly stored in underground chambers sealed with concrete bulkheads. Water samples from underground seepages are pH-neutral due to carbonate-bearing wall-rock and concrete. Some underground mine waters and surface soil pore waters at the Giant mine contain extremely high concentrations of As which have interacted with arsenic trioxide. These concentrations, several orders of magnitude higher than guidelines, occur in the absence of acid rock drainage. The planned remediation at Giant will reduce but not eliminate As release from the site. Specifically, the total As released (mostly in the form of dissolved As) will decrease from the current level of approximately 500 kg per year to less than 200 kg per year. Without this remediation, assuming that Giant mine were allowed to flood as is the case for most underground mines after closure, As release could increase to many thousands of kilograms per year. Interestingly, the very high cost of the proposed remediation plan is still outweighed by the monetary value of the gold extracted over the lifetime of Giant mine., Roasting arsenopyrite-bearing gold ore for more than fifty years has resulted in nearly 300 000 tons of arsenic trioxide waste at the Giant mine, Yellowknife, NWT, Canada, mostly stored in underground chambers sealed with concrete bulkheads. Water samples from underground seepages are pH-neutral due to carbonate-bearing wall-rock and concrete. Some underground mine waters and surface soil pore waters at the Giant mine contain extremely high concentrations of As which have interacted with arsenic trioxide. These concentrations, several orders of magnitude higher than guidelines, occur in the absence of acid rock drainage. The planned remediation at Giant will reduce but not eliminate As release from the site. Specifically, the total As released (mostly in the form of dissolved As) will decrease from the current level of approximately 500 kg per year to less than 200 kg per year. Without this remediation, assuming that Giant mine were allowed to flood as is the case for most underground mines after closure, As release could increase to many thousands of kilograms per year. Interestingly, the very high cost of the proposed remediation plan is still outweighed by the monetary value of the gold extracted over the lifetime of Giant mine.

While the anti-tumor efficacy of 2-deoxy-d-glucose (2-DG) is normally low in monotherapy, it may represent a valuable radio- and chemo-sensitizing agent. We here demonstrate that 2–10 mM 2-DG cooperates with arsenic trioxide (ATO) and other antitumor drugs to induce apoptosis in human myeloid leukemia cell lines. Using ATO and HL60 as drug and cell models, respectively, we observed that 2-DG/ATO combination activates the mitochondrial apoptotic pathway, as indicated by Bid-, and Bax-regulated cytochrome c and Omi/HtrA2 release, XIAP down-regulation, and caspase-9/-3 pathway activation. 2-DG neither causes oxidative stress nor increases ATO uptake, but causes inner mitochondria membrane permeabilization as well as moderate ATP depletion, which nevertheless do not satisfactorily explain the pro-apoptotic response. Surprisingly 2-DG causes cell line-specific decrease in LKB-1/AMPK phosphorylation/activation, and also causes Akt/mTOR/p70S6K and MEK/ERK activation, which is prevented by co-treatment with ATO. The use of kinase-specific pharmacologic inhibitors and/or siRNAs reveals that apoptosis is facilitated by AMPK inactivation and restrained by Akt and ERK activation, and that Akt and ERK activation mediates AMPK inhibition. Finally, 2-DG stimulates IGF-1R phosphorylation/activation, and co-treatment with IGF-1R inhibitor prevents 2-DG effects on Akt, ERK and AMPK, and facilitates 2-DG-provoked apoptosis. In summary 2-DG elicits IGF-1R-mediated AMPK inactivation and Akt and ERK activation, which facilitates or restrain apoptosis, respectively. 2-DG-provoked AMPK inactivation increases the apoptotic efficacy of ATO, while in turn ATO-provoked Akt and ERK inactivation may increase the efficacy of 2-DG as anti-tumor drug

The uncoupling proteins (UCPs) are mitochondrial carriers that modulate the energetic efficiency and, as a result, can lower superoxide levels. Here, we describe the discovery of a small-molecule inhibitor of the UCPs. Screening of potential UCP1 regulators led to the identification of chromane derivatives that inhibit its proton conductance. Members of the UCP family can act as a defense against oxidative stress and, thus, UCP2 plays a protective role in tumor cells. High UCP2 levels have been associated with chemoresistance. We demonstrate that chromanes also inhibit UCP2 and, in HT-29 human carcinoma cells, cause oxidative stress. The chromane derivatives can act synergistically with chemotherapeutic agents; for instance, they increase the toxicity of arsenic trioxide in HT-29 cells. These findings open a promising line in the development of novel anticancer agents, agents chromane derivatives chromanes findings open HT-29 cells HT-29 human carcinoma cells lower superoxide levels Members modulate potential UCP1 regulators protective role proton conductance tumor cells UCP family UCPs uncoupling proteins

Lonidamine is a safe, clinically useful anti-tumor drug, but its efficacy is generally low when used in monotherapy. We here demonstrate that lonidamine efficaciously cooperates with the anti-leukemic agent arsenic trioxide (ATO, Trisenox™) to induce apoptosis in HL-60 and other human leukemia cell lines, with low toxicity in non-tumor peripheral blood lymphocytes. Apoptosis induction by lonidamine/ATO involves mitochondrial dysfunction, as indicated by early mitochondrial permeability transition pore opening and late mitochondrial transmembrane potential dissipation, as well as activation of the intrinsic apoptotic pathway, as indicated by Bcl-XL and Mcl-1 down-regulation, Bax translocation to mitochondria, cytochrome c and Omi/HtrA2 release to the cytosol, XIAP down-regulation, and caspase-9 and -3 cleavage/activation, with secondary (Bcl-2-inhibitable) activation of the caspase-8/Bid axis. Lonidamine stimulates reactive oxygen species production, and lonidamine/ATO toxicity is attenuated by antioxidants. Lonidamine/ATO stimulates JNK phosphorylation/activation, and apoptosis is attenuated by the JNK inhibitor SP600125. In addition, lonidamine elicits ERK and Akt/mTOR pathway activation, as indicated by increased ERK, Akt, p70S6K and rpS6 phosphorylation, and these effects are reduced by co-treatment with ATO. Importantly, co-treatment with MEK/ERK inhibitor (U0126) and PI3K/Akt (LY294002) or mTOR (rapamycin) inhibitors, instead of ATO, also potentiates lonidamine-provoked apoptosis. These results indicate that: (i) lonidamine efficacy is restrained by drug-provoked activation of MEK/ERK and Akt/mTOR defensive pathways, which therefore represent potential therapeutic targets. (ii) Co-treatment with ATO efficaciously potentiates lonidamine toxicity via defensive pathway inhibition and JNK activation. And (iii) conversely, the pro-oxidant action of lonidamine potentiates the apoptotic efficacy of ATO as an anti-leukemic agent

Arsenic trioxide (As2O3, ATO) is a very efficacious, clinically established agent for the treatment of acute promyelocytic leukaemia, and also potentially useful against other haematological and non-haematological malignancies. Nonetheless, the relative resistance of many tumour cell types requires the generation of sensitizing strategies. One of the properties of ATO which might be exploited for therapeutic purposes is its sensitivity to the intracellular oxidant state, as revealed by increased apoptosis production under conditions of reduced glutathione (GSH) depletion and/or elevated reactive oxygen species (ROS) content. This review summarizes some studies from our laboratory demonstrating that experimental modulation of protein kinase activities (PI3K/Akt, JNK, MEK/ERK) potentiates ATO-provoked apoptosis in relatively resistant human acute myeloid leukaemia (U937, HL60) cell lines by mechanisms involving GSH depletion and/or increased ROS content. In a similar manner, co-treatment with dietary flavonoides such as genistein, normally considered as anti-oxidants, may potentiate apoptosis via generation of moderate oxidative stress and activation of ROS-inducible protein kinases. Finally, co-treatment with ATO may sensitize otherwise refractory leukaemia cells to TNFalpha-family cytokine-produced apoptosis, by mechanisms involving the interplay between the "intrinsic" (mitochondrial) and "extrinsic" (death receptor-mediated) pathways

New antiviral agents are urgently needed. Based on in vitro studies, arsenic trioxide (As2O3) seems to affect viral replication, although this has been studied only marginally in vivo. In this study the replication of coxsackievirus B3 (CVB3) was studied in Balb/c mice administered 1 mg As2O3/kg bw once daily during 7 days of infection and in Vero cells exposed for 3 or 5 days to 0.4, 2 or 4 µM As2O3. Viral RNA was measured by reverse-transcription PCR (RT-PCR) (in vitro and in vivo) and arsenic concentration was measured by inductively coupled plasma-mass spectrometry (ICP-MS) (in vivo). In vivo, As2O3 decreased viral RNA in the brain on days 3 (by 81%; p<0.05) and 7 (by 97%; p<0.01) and in the pancreas on day 7 (by 75%; p<0.05), two of the target organs of this infection. The results were confirmed in vitro, where As2O3 dose-dependently reduced viral RNA, with the effect being more pronounced in the surrounding culture medium than inside the infected cells, indicating an impaired virion release. Thus, As2O3 reduced CVB3 replication both in vitro and in vivo, indicating that As2O3 is a viable option in the pursuit of new therapeutic agents against viral infections.

Arsenic trioxide (As2O3, ATO) is a very efficacious, clinically established agent for the treatment of acute promyelocytic leukaemia, and also potentially useful against other haematological and non-haematological malignancies. Nonetheless, the relative resistance of many tumour cell types requires the generation of sensitizing strategies. One of the properties of ATO which might be exploited for therapeutic purposes is its sensitivity to the intracellular oxidant state, as revealed by increased apoptosis production under conditions of reduced glutathione (GSH) depletion and/or elevated reactive oxygen species (ROS) content. This review summarizes some studies from our laboratory demonstrating that experimental modulation of protein kinase activities (PI3K/Akt, JNK, MEK/ERK) potentiates ATO-provoked apoptosis in relatively resistant human acute myeloid leukaemia (U937, HL60) cell lines by mechanisms involving GSH depletion and/or increased ROS content. In a similar manner, co-treatment with dietary flavonoides such as genistein, normally considered as anti-oxidants, may potentiate apoptosis via generation of moderate oxidative stress and activation of ROS-inducible protein kinases. Finally, co-treatment with ATO may sensitize otherwise refractory leukaemia cells to TNFalpha-family cytokine-produced apoptosis, by mechanisms involving the interplay between the "intrinsic" (mitochondrial) and "extrinsic" (death receptor-mediated) pathways

The introduction of all-trans retinoic acid (ATRA) and, more recently, arsenic trioxide (ATO) into the therapy of acute promyelocytic leukemia (APL) has revolutionized the management and outcome of this disease. Several treatment strategies using these agents, usually in combination with chemotherapy, but also without or with minimal use of cytotoxic agents, have provided excellent therapeutic results. Cure of APL patients, however, is also dependent on peculiar aspects related to the management and supportive measures that are crucial to counteract life-threatening complications associated with the disease biology and molecularly targeted treatment. The European LeukemiaNet recently appointed an international panel of experts to develop evidence- and expert opinion-based guidelines on the diagnosis and management of APL. Together with providing current indications on genetic diagnosis, modern risk-adapted front-line therapy and salvage treatment, the review contains specific recommendations for the iden

Despite arsenic's (As) toxic potential, arsenic trioxide (As2O3) is used as a safe and effective treatment in acute promyelocytic leukaemia. However, it is unknown whether such therapy influences the balance of other trace elements in the body. In this study, mice were treated intraperitoneally daily with 1.0 mg As2O3/kg bw for 3, 5 or 7 days. As, and seven essential and nonessential trace elements with the potential to interact with As, were measured through inductively coupled plasma-mass spectrometry in serum, heart, lung, liver, pancreas, kidney, intestine and brain. As2O3 supplementation increased As in all target tissues on day 3, thereafter reaching an almost steady state. The major findings in other elements were a sequential decrease in serum zinc (on day 7 by 64%; P<0.001), and a decrease in selenium in the pancreas on day 3 (9%; P<0.05), in the intestine on day 3 (30%; P<0.001) and finally, in the brain on days 5 (12%; P<0.05) and 7 (15%; P<0.01). Changes in magnesium, iron, copper, cadmium and mercury were minor and inconsistent. This study suggests that supplementation with other trace elements may be beneficial when As2O3 treatment regimens are used in the clinic.

Recently it has been reported that arsenic trioxide (As203) is very effective in the treatment of acute promyelocytic leukemia (APL) by inducing apoptosis, but the molecular mechanism of its action on other leukemia remains unclear. In the present study, we tested the effect of As2 O3 at low concentration O.25-2.O μM on K562, a chronic myelogenous leukemia cell line. As2 03 inhibited the cell growth of K562 in a similar way to APL cell line NB4. Typical oncotic cell death, such as cytoplasmic swelling and swelling of organelles, was observed by morphological study and cell cycle was arrested at G2+M phases. During the treatment of As2 03, the CD95 and CD95 ligand (CD95L) expressions were upregulated, and caspase 8 and caspase 3 were activated, but bcl-2 expression was not changed. Treatment of the cells with anti-CD95 monoclonal antibody or ZVAD-fmk capable of blocking the CD95 signaling pathway ameliorated As2 03-induced oncosis. These results suggest that the induction of oncosis by As2 03 invoIves CD95/CD95L pathway in K562 and As2 03 may provide a novel therapy for leukemia other than APL.

The interactions of three heavy metal-containing compounds, cisplatin (CDDP), arsenic trioxide (As2O3), and mercury dichloride (HgCl2), with the multidrug resistance transporters MRP1 and MRP2 and the involvement of glutathione (GSH)-related processes herein were investigated. In Madin-Darby canine kidney cells stably expressing MRP1 or MRP2, viability, GSH content, calcein efflux and polarized GSH efflux were measured as a function of exposure to CDDP, As2O3 and HgCl2. In isolated Sf9-MRP1 and Sf9-MRP2 membrane vesicles, the interaction with MRP-associated ATPase activity was measured. In the latter model system adduct formation with GSH is not an issue. The data show that (1) CDDP interacts with both MRP1 and MRP2, and GSH appears to play no major role in this process, (2) As2O3 interacts with MRP1 and MRP2 in which process GSH seems to be essential, and (3) HgCl2 interacts with MRP1 and MRP2, either alone and/or as a metal-GSH complex. © 2008 Elsevier B.V. All rights reserved.

Recently, it has been reported that arsenic trioxide (As2 O3) is an effective anticancer agent for acute promyelocytic leukemia (APL). In the present study, we examined the anticancer effects of As2 O3 at low concentration (0.25～2.OμM) on two hunan leukemia/lymphoma cell lines, HL-60 and RL, in vitro. We found that As2 O3 inhibited the growth of HL-60 and RL similar to the reported APL cell line, NB4. Typical apoptosis was observed in morphologlcal study and DNA fragmentation assay, as well as a cell cycle arrest at subG1. To address the mechanism of As2 03-induced apoptosis, we also examined the effect of As2 O3 on the CD95/CD95L pathway and bcl-2 protein expression. The results showed that the CD95/CD95L expressions were upregulated; meanwhile, caspase 8 and caspase 3 were activated. However, the bcl-2 protein expression was downregulated. Using anti-CD95 monoclonal antibody to block the CD95 pathway, As2 O3 - induced apoptosis was ameliorated. These data suggest that in HL-60 and RL cell lines the CD95/CD95L pathway and downregulation of bc1-2 protein expression are invoIved in As2 O3-induced apoptosis.