Complexos ciclometalados de paládio (II) contendo oximas como agentes antitumorais em modelo celular de osteossarcoma: um estudo in vitro e in vivo
Data
2023-09-04
Autores
Pereira, Thales Hebert Regiani 
Orientadores
Trindade, Claudia Bincoletto
Tipo
Dissertação de mestrado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
A terapia antineoplásica tem por objetivo erradicar as células tumorais por meio da
indução de morte celular ou ainda a inibição do ciclo e da proliferação celular. A busca por
fármacos capazes de modular eventos associados à morte e/ou à sobrevivência celular é
de grande interesse. Nas últimas décadas, os ciclopaladados têm sido amplamente
estudados pela sua capacidade de interagir com alvos moleculares específicos envolvidos
no controle da morte celular. Neste projeto, avaliamos os possíveis efeitos antitumorais de
uma nova classe de complexos ciclopaladados de paládio (II) contendo oximas em um
modelo celular de osteossarcoma, o qual é altamente agressivo e resistente a muitos
fármacos. A cisplatina (CDDP) foi utilizada como controle positivo em todos os
experimentos. Demonstramos que os complexos organometálicos denominados paládiopiperidina-oxima (Pd-BPO) e paládio-morfolina-oxima (Pd-BMO) são mais citotóxicos do
que a CDDP em células de osteosarcoma SaOs-2 pelo ensaio de MTT. A coloração com
Anexina-FITC/7-AAD demonstrou um maior potencial para os complexos de paládiooxima na indução de apoptose tardia nestas células em relação a CDDP após 24 horas
de exposição, um evento confirmado também usando o inibidor de pan-caspase Z-VADFMK. Em comparação com a CDDP, apenas os complexos paládio-oxima erradicam a
clonogenicidade das células SaOs-2 após 7 dias de tratamento. O envolvimento do eixo
lisossomo-mitocôndria nas propriedades indutoras de morte celular dos complexos
também foi verificado. Os complexos induziram a permeabilização da membrana
lisossomal (PML) em um ensaio usando laranja de acridina (L/A) e citometria de fluxo.
Além disso, os mesmos mostraram maior eficiência em despolarizar a membrana
mitocondrial de células SaOs-2 em relação a CDDP e ao controle sem tratamento,
utilizando o TMRE. Para os estudos in vivo, o C. elegans foi usado e demonstrou que
ambos os complexos reduzem a locomoção e o bombeamento faríngeo após 24 horas de
tratamento na mesma extensão que a verificada com o tratamento com a CDDP.
Concluímos que ambos os complexos são mais eficientes do que a CDDP na indução de
morte em células SaOs-2. A toxicidade desses complexos para o C. elegans foi
semelhante à induzida pela CDDP
Antineoplastic therapy aims to eradicate tumor cells by inducing their death or inhibiting their cell cycle and proliferation. The search for new drugs capable of modulating events associated with cell death or survival is of great interest. In recent decades, cyclopalladated have been widely studied for their ability to interact with specific molecular targets involved in the control of cell death. In this project, we evaluated the possible antitumor effects of a new class of cyclopalladated complexes containing oximes, in a cellular model of osteosarcoma, which is highly aggressive and resistant to many drugs. Cisplatin (CDDP) was used as a positive control in all experiments. We demonstrated that the organometallic complexes called palladium-piperidine-oxime (Pd-BPO-1) and palladium-morpholine-oxime (Pd-BMO-2) are more cytotoxic than CDDP in SaOs-2 osteosarcoma cells by MTT assay. Annexin-FITC/7-AAD staining demonstrated a greater potential for palladium-oxime complexes in inducing apoptosis/late apoptosis in these cells compared to CDDP after 24 hours of exposure, an event confirmed also using the pan inhibitor -caspase Z-VAD-FMK. Compared to CDDP, only palladium-oxime complexes eradicate the clonogenicity of SaOs-2 cells after 7 days of treatment. The involvement of the lysosome-mitochondria axis in the cell death inducing properties of the complexes was also verified. Using LysoTrackerRed, we verified a decrease in the fluorescence intensity of this probe in cells treated with the complexes in relation to CDDP, indicating an increase in lysosomal pH. The complexes also induced lysosomal membrane permeabilization (LMP) in an assay using acridine orange (L/A) and confocal microscopy. The greater efficiency of the complexes in depolarizing the mitochondrial membrane of SaOs-2 cells compared to CDDP, and the untreated control, was also observed using the TMRE. For the in vivo studies, C. elegans was used and demonstrated that both complexes reduce locomotion and pharyngeal pumping after 24 hours of treatment to the same extent as that seen with CDDP treatment. We conclude that both complexes are more efficient than CDDP in inducing death in SaOS-2 cells. The toxicity of these complexes to C. elegans was similar to that induced by CDDP.
Antineoplastic therapy aims to eradicate tumor cells by inducing their death or inhibiting their cell cycle and proliferation. The search for new drugs capable of modulating events associated with cell death or survival is of great interest. In recent decades, cyclopalladated have been widely studied for their ability to interact with specific molecular targets involved in the control of cell death. In this project, we evaluated the possible antitumor effects of a new class of cyclopalladated complexes containing oximes, in a cellular model of osteosarcoma, which is highly aggressive and resistant to many drugs. Cisplatin (CDDP) was used as a positive control in all experiments. We demonstrated that the organometallic complexes called palladium-piperidine-oxime (Pd-BPO-1) and palladium-morpholine-oxime (Pd-BMO-2) are more cytotoxic than CDDP in SaOs-2 osteosarcoma cells by MTT assay. Annexin-FITC/7-AAD staining demonstrated a greater potential for palladium-oxime complexes in inducing apoptosis/late apoptosis in these cells compared to CDDP after 24 hours of exposure, an event confirmed also using the pan inhibitor -caspase Z-VAD-FMK. Compared to CDDP, only palladium-oxime complexes eradicate the clonogenicity of SaOs-2 cells after 7 days of treatment. The involvement of the lysosome-mitochondria axis in the cell death inducing properties of the complexes was also verified. Using LysoTrackerRed, we verified a decrease in the fluorescence intensity of this probe in cells treated with the complexes in relation to CDDP, indicating an increase in lysosomal pH. The complexes also induced lysosomal membrane permeabilization (LMP) in an assay using acridine orange (L/A) and confocal microscopy. The greater efficiency of the complexes in depolarizing the mitochondrial membrane of SaOs-2 cells compared to CDDP, and the untreated control, was also observed using the TMRE. For the in vivo studies, C. elegans was used and demonstrated that both complexes reduce locomotion and pharyngeal pumping after 24 hours of treatment to the same extent as that seen with CDDP treatment. We conclude that both complexes are more efficient than CDDP in inducing death in SaOS-2 cells. The toxicity of these complexes to C. elegans was similar to that induced by CDDP.
Descrição
Citação
PEREIRA, Thales Hebert Regiani. Complexos ciclometalados de paládio (II) contendo oximas como agentes antitumorais em modelo celular de osteossarcoma: um estudo in vitro e in vivo. 2023. 82 f. Dissertação (Mestrado em Farmacologia) - Escola Paulista de Medicina, Universidade Federal de São Paulo (UNIFESP). São Paulo, 2023.