Assim como o número de casos de câncer vem aumentando em nível global, a busca por abordagens terapêuticas visando uma maior eficácia com um menor poder de causar efeitos prejudiciais aos pacientes também vem crescendo. As antraciclinas e antracenodionas, as quais tem como exemplos, doxorrubicina (DOX) e mitoxantrona (MTX), respectivamente, são fármacos utilizados na quimioterapia em diversas neoplasias incluindo tumores sólidos e não sólidos tais como de mama, leucemias, linfomas, sarcomas etc. Embora sejam eficazes ao que se propõem, o tratamento com estas moléculas pode acarretar em efeitos secundários, tais como arritmias e insuficiência cardíaca. Estas drogas além de interagirem com o ferro e apresentarem capacidade de gerar espécies reativas de oxigénio (ROS), apresentam como principal mecanismo a inibição da enzima topoisomerase 2 (Top2). Os inibidores de PARP-1 emergiram como uma nova alternativa para tratar determinados tipos de neoplasias em que a letalidade sintética possa ser explorada. Além disto, já foi relatado que a toxicidade cardíaca induzida por DOX seja influenciada pela atividade de PARP-1. O objetivo desta tese foi, portanto, avaliar a influência da inibição de PARP-1 na toxicidade cardíaca de DOX e MTX em células cardíacas. Células foram incubadas durante 24h com DOX ou MTX na presença ou na ausência de inibidor de PARP-1. Ensaios de viabilidade, apoptose e genotoxicidade e foram realizados. Além disso, a fosforilação de proteínas envolvidas na resposta a danos no DNA (ATM, MRE-11 e H2AX) foram avaliadas por western blot e imunofluorescência. Os resultados demonstraram que a inibição de PARP-1, apesar de diminuir a concentração de ROS, diminui a viabilidade de células H9c2 tratadas com DOX ou MTX por aumentar a geração de quebras duplas no DNA induzida por estes fármacos. / As the number of people with cancer are globally increasing, the search for therapeutic approaches that increases efficiency decreasing harmful effects to patients is also growing, giving rise to cardio-oncology. Anthracyclines, e.g., doxorubicin (DOX), and anthracenediones, e.g., mitoxantrone (MTX), are drugs used in the chemotherapy of several cancer types, including solid and non-solid malignancies such as breast cancer, leukemia, lymphomas, and sarcomas. Although they are effective in tumor therapy, treatment with these two drugs may lead to side effects such as arrhythmia and heart failure. These drugs interact with iron to generate reactive oxygen species (ROS), target topoisomerase 2 (Top2), and impair mitochondria. PARP-1 inhibitors have emerged as a new alternative for treating certain types of malignancies in which the synthetic lethality can be exploited. Furthermore, it has been reported that DOX-induced cardiac cardiotoxicity is influenced by PARP-1 activity. The main goal of this thesis was, therefore, to evaluate PARP-1 inhibition influence in cardiac toxicity of DOX and MTX in cardiac cells. Cells were incubated for 24h with MTX or DOX in presence or absence of PARP-1 inhibitor. Viability, oxidative stress and genotoxicity assays have been conducted. Furthermore, phosphorylation of proteins involved in response to DNA damage (ATM, H2AX and MRE-11) were evaluated by western blot and immunofluorescence. Results demonstrated that inhibition of PARP-1, although decreasing ROS generation, decreases H9c2 cells viability after DOX or MTX by increasing DNA double strand break generation induced by these drugs.

Thèse (M.Sc.)--Université Laval, 2005. / Bibliogr.: f. 75-90. Publié aussi en version électronique.

No description available.

Thèse (M.Sc.) -- Université Laval, 1997. / Bibliogr.: f. 128-151. Publié aussi en version électronique.

Thèse (Ph. D.)--Université Laval, 2003. / Bibliogr. Publié aussi en version électronique.

Thèse (Ph. D.)--Université Laval, 2003. / Bibliogr.: f. [166]-176. Publié aussi en version électronique.

Assim como o número de casos de câncer vem aumentando em nível global, a busca por abordagens terapêuticas visando uma maior eficácia com um menor poder de causar efeitos prejudiciais aos pacientes também vem crescendo. As antraciclinas e antracenodionas, as quais tem como exemplos, doxorrubicina (DOX) e mitoxantrona (MTX), respectivamente, são fármacos utilizados na quimioterapia em diversas neoplasias incluindo tumores sólidos e não sólidos tais como de mama, leucemias, linfomas, sarcomas etc. Embora sejam eficazes ao que se propõem, o tratamento com estas moléculas pode acarretar em efeitos secundários, tais como arritmias e insuficiência cardíaca. Estas drogas além de interagirem com o ferro e apresentarem capacidade de gerar espécies reativas de oxigénio (ROS), apresentam como principal mecanismo a inibição da enzima topoisomerase 2 (Top2). Os inibidores de PARP-1 emergiram como uma nova alternativa para tratar determinados tipos de neoplasias em que a letalidade sintética possa ser explorada. Além disto, já foi relatado que a toxicidade cardíaca induzida por DOX seja influenciada pela atividade de PARP-1. O objetivo desta tese foi, portanto, avaliar a influência da inibição de PARP-1 na toxicidade cardíaca de DOX e MTX em células cardíacas. Células foram incubadas durante 24h com DOX ou MTX na presença ou na ausência de inibidor de PARP-1. Ensaios de viabilidade, apoptose e genotoxicidade e foram realizados. Além disso, a fosforilação de proteínas envolvidas na resposta a danos no DNA (ATM, MRE-11 e H2AX) foram avaliadas por western blot e imunofluorescência. Os resultados demonstraram que a inibição de PARP-1, apesar de diminuir a concentração de ROS, diminui a viabilidade de células H9c2 tratadas com DOX ou MTX por aumentar a geração de quebras duplas no DNA induzida por estes fármacos. / As the number of people with cancer are globally increasing, the search for therapeutic approaches that increases efficiency decreasing harmful effects to patients is also growing, giving rise to cardio-oncology. Anthracyclines, e.g., doxorubicin (DOX), and anthracenediones, e.g., mitoxantrone (MTX), are drugs used in the chemotherapy of several cancer types, including solid and non-solid malignancies such as breast cancer, leukemia, lymphomas, and sarcomas. Although they are effective in tumor therapy, treatment with these two drugs may lead to side effects such as arrhythmia and heart failure. These drugs interact with iron to generate reactive oxygen species (ROS), target topoisomerase 2 (Top2), and impair mitochondria. PARP-1 inhibitors have emerged as a new alternative for treating certain types of malignancies in which the synthetic lethality can be exploited. Furthermore, it has been reported that DOX-induced cardiac cardiotoxicity is influenced by PARP-1 activity. The main goal of this thesis was, therefore, to evaluate PARP-1 inhibition influence in cardiac toxicity of DOX and MTX in cardiac cells. Cells were incubated for 24h with MTX or DOX in presence or absence of PARP-1 inhibitor. Viability, oxidative stress and genotoxicity assays have been conducted. Furthermore, phosphorylation of proteins involved in response to DNA damage (ATM, H2AX and MRE-11) were evaluated by western blot and immunofluorescence. Results demonstrated that inhibition of PARP-1, although decreasing ROS generation, decreases H9c2 cells viability after DOX or MTX by increasing DNA double strand break generation induced by these drugs.

Assim como o número de casos de câncer vem aumentando em nível global, a busca por abordagens terapêuticas visando uma maior eficácia com um menor poder de causar efeitos prejudiciais aos pacientes também vem crescendo. As antraciclinas e antracenodionas, as quais tem como exemplos, doxorrubicina (DOX) e mitoxantrona (MTX), respectivamente, são fármacos utilizados na quimioterapia em diversas neoplasias incluindo tumores sólidos e não sólidos tais como de mama, leucemias, linfomas, sarcomas etc. Embora sejam eficazes ao que se propõem, o tratamento com estas moléculas pode acarretar em efeitos secundários, tais como arritmias e insuficiência cardíaca. Estas drogas além de interagirem com o ferro e apresentarem capacidade de gerar espécies reativas de oxigénio (ROS), apresentam como principal mecanismo a inibição da enzima topoisomerase 2 (Top2). Os inibidores de PARP-1 emergiram como uma nova alternativa para tratar determinados tipos de neoplasias em que a letalidade sintética possa ser explorada. Além disto, já foi relatado que a toxicidade cardíaca induzida por DOX seja influenciada pela atividade de PARP-1. O objetivo desta tese foi, portanto, avaliar a influência da inibição de PARP-1 na toxicidade cardíaca de DOX e MTX em células cardíacas. Células foram incubadas durante 24h com DOX ou MTX na presença ou na ausência de inibidor de PARP-1. Ensaios de viabilidade, apoptose e genotoxicidade e foram realizados. Além disso, a fosforilação de proteínas envolvidas na resposta a danos no DNA (ATM, MRE-11 e H2AX) foram avaliadas por western blot e imunofluorescência. Os resultados demonstraram que a inibição de PARP-1, apesar de diminuir a concentração de ROS, diminui a viabilidade de células H9c2 tratadas com DOX ou MTX por aumentar a geração de quebras duplas no DNA induzida por estes fármacos. / As the number of people with cancer are globally increasing, the search for therapeutic approaches that increases efficiency decreasing harmful effects to patients is also growing, giving rise to cardio-oncology. Anthracyclines, e.g., doxorubicin (DOX), and anthracenediones, e.g., mitoxantrone (MTX), are drugs used in the chemotherapy of several cancer types, including solid and non-solid malignancies such as breast cancer, leukemia, lymphomas, and sarcomas. Although they are effective in tumor therapy, treatment with these two drugs may lead to side effects such as arrhythmia and heart failure. These drugs interact with iron to generate reactive oxygen species (ROS), target topoisomerase 2 (Top2), and impair mitochondria. PARP-1 inhibitors have emerged as a new alternative for treating certain types of malignancies in which the synthetic lethality can be exploited. Furthermore, it has been reported that DOX-induced cardiac cardiotoxicity is influenced by PARP-1 activity. The main goal of this thesis was, therefore, to evaluate PARP-1 inhibition influence in cardiac toxicity of DOX and MTX in cardiac cells. Cells were incubated for 24h with MTX or DOX in presence or absence of PARP-1 inhibitor. Viability, oxidative stress and genotoxicity assays have been conducted. Furthermore, phosphorylation of proteins involved in response to DNA damage (ATM, H2AX and MRE-11) were evaluated by western blot and immunofluorescence. Results demonstrated that inhibition of PARP-1, although decreasing ROS generation, decreases H9c2 cells viability after DOX or MTX by increasing DNA double strand break generation induced by these drugs.

Acrolein is an electrophilic α, β-unsaturated aldehyde. Additionally, acrolein is a metabolite of the antineoplastic alkylating agent cyclophosphamide and is implicated in off-target effects, including to bladder hemorrhagic cystitis and cyclophosphamide-induced cardiotoxicity, both of which have led to serious secondary iatrogenic injury during and following chemotherapy. At low concentrations acrolein inhibits cell proliferation without inducing apoptosis, while at high concentrations may result in secondary apoptosis promotion. This investigation assessed the role of the enzyme poly (ADP-ribose) polymerase (PARP) in acrolein induced toxicity using the established toxicological H9c2 (2-1) cardiomyoblast in vitro model. H9c2 (2-1) cells were plated in 24-well plates at 75,000 cells per well three days prior to testing, followed by acrolein dosing at concentrations between 10 µM and 1000µM for either 30 or 55 minutes. PARP activity was quantitatively measured in total cell lysates using a biotin-avidin-conjugated horseradish peroxidase-TMB reporter system in a 96-well microplate formate. The lowest effective dose of toxicity at 30 minute dosing was found at 25 μM (PARP Activity 1.65-fold control) which returned to baseline at 100 μM; concentrations at or above 250 μM results in significant PARP activity reductions (≤ 0.46-fold control). Biomarkers were further characterized for cytotoxicity (AST presence), and viability (MTT reduction) in order to facilitate mechanistic characterization of PARP-mediated acrolein cardiotoxicity. Investigation of a PARP inhibitor was assessed to explore the intervention for acrolein induced cardiac tissue damage.

Drug-induced cardiotoxicity has resulted in a thorough evaluation of patient doses, treatments, and rehabilitation. One of the most commonly prescribed chemotherapeutic agents is cyclophosphamide. The active metabolite, acrolein, is one of the most potent inducers of cardiomyopathy. In this study, research was conducted on the H9c2 (2-1) cardiomyocyte cell line derived from the embryonic myocardium of rattus norvegicus to assess its competency for evaluation of the change in poly (ADP-ribose) polymerase (PARP) activity. The application of this model to study the effects of acrolein on PARP activation was chosen as an ideal determinant of cell damage produced by nitrogen mustards. To verify the legitimacy of this model, cardiomyocytes were exposed to acrolein in varying concentrations and time durations with a subsequent protein concentration measurement determined through the BCA Protein Assay. After the normalization of samples through volume adjustments and verification of sufficient protein, other aliquots were subjected to a PARP Assay in order to measure PARP activity. PARP was activated at exposure concentrations of 75 μM in all trials, with an average detection of 0.00569 ± 0.001 mU/200ng protein. Other concentrations showed varying degrees of PARP activation, verifying the model’s competency. PARP activation implies the potential use of this model for further research into targeted molecular therapy of PARP inhibition. Therefore, this model has the ability to be used as an assessment tool for the combined use of PARP inhibitors and chemotherapeutic agents; it can be useful for future research investigating the use and efficacy of PARP inhibitors in reducing drug-induced cardiotoxicity.