Planejamento e síntese de novos inibidores e degradadores (PROteolysis TArgeting Chimera - PROTAC) de histonas deacetilases (HDACs) 1 e 2 para anemia falciforme
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Data
2023-06-06
Autores
Pavan, Aline Renata
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
A anemia falciforme (AF) é uma hemoglobinopatia causada por uma mutação pontual no gene da β-globina. Essa mutação resulta na formação de uma hemoglobina mutada (HbS) que polimeriza e causa mudanças irreversíveis na morfologia das hemácias, a qual adquire a forma de foice e é responsável pela fisiopatologia da doença. Dentre as estratégias descritas para tratamento da AF está a indução de expressão de γ-globina e produção de hemoglobina fetal (HbF), a qual é capaz de diminuir os sintomas da doença por reduzir as taxas de polimerização, dentre outros efeitos, e foi a estratégia selecionada para o presente trabalho. As enzimas epigenéticas histonas deacetilases (HDACs) 1 e 2 foram os alvos explorados para desenvolvimento de novos indutores de expressão de γ -globina e produção de HbF. Sendo assim, este trabalho foi dividido em duas partes: 1) Planejamento, síntese e avaliação farmacológica de novos inibidores de HDAC 1 e 2; 2) Planejamento, síntese e avaliação farmacológica de novas PROTACs para degradação de HDAC 1 e 2. Na primeira parte foram sintetizados 24 intermediários e 7 produtos finais inéditos com rendimentos que variaram entre 29 e 85%. Os dois compostos mais promissores ((21) e (31)) apresentaram valores de IC50 variando entre 17 e 950 nM e importante seletividade contra as HDACs 1 e 2. Em um ensaio preliminar utilizando-se células eritroleucêmicas (K562) ambos os compostos foram capazes de aumentar em, aproximadamente, 7x a expressão de gama-globina a 1 μM após o tempo de tratamento de 72h e 96 h sendo superior ao fármaco utilizado na terapia, a hidroxiureia. Na segunda parte, 40 intermediários e 13 moléculas inéditas foram sintetizadas com rendimentos que variaram entre 30 e 65%. Todos os compostos foram avaliados quanto a degradação das HDACs 1-3 e, de forma geral, os compostos apresentaram degradação seletiva contra HDAC-1. Destacam-se os compostos ARP-26, ARP-37 e ARP-49, com valores de degradação que variaram de 41 a 60% contra HDAC-1. Os valores de DC50 dos compostos ARP-26 e ARP-49 foram determinados, e apresentaram aproximadamente 50% de degradação à 2,5 μM. Sendo assim, foi possível sintetizar e obter novos inibidores de HDAC seletivos contra HDAC-1 e HDAC-2 e capazes de induzir a expressão de γ-globina a níveis acima do que o fármaco atualmente no mercado e que podem ser úteis como candidatos à fármacos para o tratamento da anemia falciforme. Foi possível também sintetizar e obter PROTACs inéditas com seletividade de degradação contra HDAC-1.
Sickle cell anemia (SCA) is a hereditary hemoglobinopathies caused by a single mutation in the gene of β-globin. Due to this mutation, a mutated hemoglobin is formed (HbS) leading to polymerization and causing irreversible changes in the morphology of red blood cells, which get a sickle shape e is responsible for the pathophysiology of the disease. Among the strategies described in the literature for the treatment of SCA is the induction of the expression of γ-globin and production of fetal hemoglobin (HbF), which is able to diminish the symptoms by reducing the polymerization and was the chosen strategy in this work. Epigenetic enzymes called histone deacetylases (HDACs) 1 and 2 were explored as targets for the development of new γ-globin and HbF inducers. Therefore, this work was divided into two parts: 1) Design, synthesis and pharmacological evaluation of new HDAC 1 and 2 inhibitors; 2) Design, synthesis and pharmacological evaluation of new PROTACs for degradation of HDAC 1 and 2. In the part 1, it was synthesized 24 intermediates and 7 final products in yields ranging from 29 and 85%. Two of the most promisor compounds presented IC50 values ranging from 17 to 950 nM and interesting selectivity towards HDACs 1 and 2. Preliminary assay in erythroleukemic cells (K562) demonstrated that both compounds were able to increase γ-globin expression by, approximately, 7- fold at 1 μM in a period of time of 72 and 96 hours, being most promising than the drug used in therapy, hydroxyurea (increase by 2.5-fold at 30 μM). In the part 2, it was synthesized 45 intermediates and 13 new final PROTACs with yields ranging from 30 to 65%. All compounds were evaluated towards HDAC 1-3 degradation and, in general, all of them presented selective degradation of HDAC-1. Compounds ARP-26, ARP-37 and ARP-49 presented the better degradation levels ranging from 41 to 60% against HDAC-1. DC50 values from compounds ARP-26 and ARP-49 were determined and shown approximately 50% of degradation at 2.5 μM. Therefore, in this work was possible to synthesize and characterize new HDAC inhibitors with interesting selectivity against HDAC- 1 and 2 with ability of induction of γ-globin in levels that exceed the results seen for the standard drug. It was also possible to synthesize and characterize new PROTACs with degradation selectivity against HDAC-1.
Sickle cell anemia (SCA) is a hereditary hemoglobinopathies caused by a single mutation in the gene of β-globin. Due to this mutation, a mutated hemoglobin is formed (HbS) leading to polymerization and causing irreversible changes in the morphology of red blood cells, which get a sickle shape e is responsible for the pathophysiology of the disease. Among the strategies described in the literature for the treatment of SCA is the induction of the expression of γ-globin and production of fetal hemoglobin (HbF), which is able to diminish the symptoms by reducing the polymerization and was the chosen strategy in this work. Epigenetic enzymes called histone deacetylases (HDACs) 1 and 2 were explored as targets for the development of new γ-globin and HbF inducers. Therefore, this work was divided into two parts: 1) Design, synthesis and pharmacological evaluation of new HDAC 1 and 2 inhibitors; 2) Design, synthesis and pharmacological evaluation of new PROTACs for degradation of HDAC 1 and 2. In the part 1, it was synthesized 24 intermediates and 7 final products in yields ranging from 29 and 85%. Two of the most promisor compounds presented IC50 values ranging from 17 to 950 nM and interesting selectivity towards HDACs 1 and 2. Preliminary assay in erythroleukemic cells (K562) demonstrated that both compounds were able to increase γ-globin expression by, approximately, 7- fold at 1 μM in a period of time of 72 and 96 hours, being most promising than the drug used in therapy, hydroxyurea (increase by 2.5-fold at 30 μM). In the part 2, it was synthesized 45 intermediates and 13 new final PROTACs with yields ranging from 30 to 65%. All compounds were evaluated towards HDAC 1-3 degradation and, in general, all of them presented selective degradation of HDAC-1. Compounds ARP-26, ARP-37 and ARP-49 presented the better degradation levels ranging from 41 to 60% against HDAC-1. DC50 values from compounds ARP-26 and ARP-49 were determined and shown approximately 50% of degradation at 2.5 μM. Therefore, in this work was possible to synthesize and characterize new HDAC inhibitors with interesting selectivity against HDAC- 1 and 2 with ability of induction of γ-globin in levels that exceed the results seen for the standard drug. It was also possible to synthesize and characterize new PROTACs with degradation selectivity against HDAC-1.
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Palavras-chave
Anemia falciforme, Hemoglobinopatia, Epigenética, Síntese orgânica, Hemoglobina