RESUMO
Histone deacetylases (HDACs) are a group of enzymes that remove acetyl groups from histones, leading to the silencing of genes. Targeting specific isoforms of HDACs has emerged as a promising approach for cancer therapy, as it can overcome drawbacks associated with pan-HDAC inhibitors. HDAC6 is a unique HDAC isoform that deacetylates non-histone proteins and is primarily located in the cytoplasm. It also has two catalytic domains and a zinc-finger ubiquitin binding domain (Zf-UBD) unlike other HDACs. HDAC6 plays a critical role in various cellular processes, including cell motility, protein degradation, cell proliferation, and transcription. Hence, the deregulation of HDAC6 is associated with various malignancies. In this study, we report the design and synthesis of a series of HDAC6 inhibitors. We evaluated the synthesized compounds by HDAC enzyme assay and identified that compound 8g exhibited an IC50 value of 21 nM and 40-fold selective activity towards HDAC6. We also assessed the effect of compound 8g on various cell lines and determined its ability to increase protein acetylation levels by Western blotting. Furthermore, the increased acetylation of α-tubulin resulted in microtubule polymerization and changes in cell morphology. Our molecular docking study supported these findings by demonstrating that compound 8g binds well to the catalytic pocket via L1 loop of HDAC6 enzyme. Altogether, compound 8g represents a preferential HDAC6 inhibitor that could serve as a lead for the development of more potent and specific inhibitors.
Assuntos
Inibidores de Histona Desacetilases , Histona Desacetilases , Desacetilase 6 de Histona , Simulação de Acoplamento Molecular , Histona Desacetilases/metabolismo , Inibidores de Histona Desacetilases/química , Histonas/metabolismo , Ácidos Hidroxâmicos/químicaRESUMO
HDAC6 and Hsp90, existing as a cytosolic complex play an important role in maintaining the protein homeostasis. The interplay of HDAC6 and Hsp90 has attracted wide attention due to their important role and promise as therapeutic targets in malignant cancers. Therefore, the discovery of dual inhibitors targeting HDAC6 and Hsp90 is of high importance. In the present study, we describe the design, synthesis, and biological evaluation of bifunctional inhibitors against HDAC6 and Hsp90 interplay. In particular, compound 6e shows a significant inhibitory activity against both HDAC6 and Hsp90 with IC50 values of 106 nM and 61 nM, respectively. Compound 6e promotes the acetylation of HDAC6 substrate proteins such as α-tubulin and Hsp90 via HDAC6 inhibition, and also induces the degradation of Hsp90 clients such as Her2, EGFR, Met, Akt, and HDAC6 via Hsp90 inhibition. Compound 6e consequently furnishes potent antiproliferative effect on gefitinib-resistant H1975 non-small cell lung cancer (NSCLC) with a GI50 value of 1.7 µM. In addition, compound 6e successfully achieved significant tumor growth inhibition in H1975 NSCLC xenograft model without noticeable abnormal behavior, body weight changes, and apparent ocular toxicity. We conclude that compound 6e constitutes an excellent tool as well as a valuable lead for assessment of Hsp90 and HDAC6 dual inhibition with a single molecule.
Assuntos
Antineoplásicos , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Animais , Antineoplásicos/química , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Linhagem Celular Tumoral , Proteínas de Choque Térmico HSP90 , Desacetilase 6 de Histona , Inibidores de Histona Desacetilases/química , Humanos , Neoplasias Pulmonares/tratamento farmacológicoRESUMO
Heat shock protein 90 (Hsp90) plays an important role in cancer cell proliferation, survival, and migration by regulating the maturation and stabilization of numerous oncoproteins. Despite significant efforts in developing Hsp90 inhibitors, none of these have been approved for clinical use, mostly due to toxicity, such as liver, cardiac, and retinal toxicity. To avoid undesirable toxicity, we herein report a hydrogen peroxide-activated Hsp90 inhibitor, Boro-BZide (3), which is capable of selectively targeting cancer cells over normal cells. Boro-BZide (3) can be activated by high levels of hydrogen peroxide, releasing its parent active Hsp90 inhibitor. The mechanism of action was determined by a series of experiments including fluorescence polarization assay, cell viability assay, western blotting, high-pressure liquid chromatography (HPLC), and fluorescence-activated cell sorting (FACS) analysis. These efforts ultimately led to the identification of a novel hydrogen peroxide-activated Hsp90 prodrug with improved therapeutic index, which was less prone to furnish unwanted adverse effects. This hydrogen peroxide-responsive prodrug strategy will be beneficial for overcoming the toxicity hurdles of Hsp90 inhibitors for clinical application.
Assuntos
Antineoplásicos/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Peróxido de Hidrogênio/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Peróxido de Hidrogênio/farmacologia , Células MCF-7 , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Chlorambucil is a nitrogen mustard-based DNA alkylating drug, which is widely used as a front-line treatment of chronic lymphocytic leukaemia (CLL). Despite its widespread application and success for the initial treatment of leukaemia, a majority of patients eventually develop acquired resistance to chlorambucil. In this regard, we have designed and synthesised a novel hybrid molecule, chloram-HDi that simultaneously impairs DNA and HDAC enzymes. Chloram-HDi efficiently inhibits the proliferation of HL-60 and U937 leukaemia cells with GI50 values of 1.24 µM and 1.75 µM, whereas chlorambucil exhibits GI50 values of 21.1 µM and 37.7 µM against HL-60 and U937 leukaemia cells, respectively. The mechanism behind its remarkably enhanced cytotoxicity is that chloram-HDi not only causes a significant DNA damage of leukaemia cells but also downregulates DNA repair protein, Rad52, resulting in the escalation of its DNA-damaging effect. Furthermore, chloram-HDi inhibits HDAC enzymes to induce the acetylation of α-tubulin and histone H3.
Assuntos
Antineoplásicos/farmacologia , Clorambucila/farmacologia , DNA de Neoplasias/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Antineoplásicos/síntese química , Antineoplásicos/química , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Clorambucila/síntese química , Clorambucila/química , Dano ao DNA , DNA de Neoplasias/química , Relação Dose-Resposta a Droga , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/química , Humanos , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
To develop novel CNS penetrant HDAC inhibitors, a new series of HDAC inhibitors having benzoheterocycle were designed, synthesized, and biologically evaluated. Among the synthesized compounds, benzothiazole derivative 9b exhibited a remarkable anti-proliferative activity (GI50 = 2.01 µM) against SH-SY5Y cancer cell line in a dose and time-dependent manner, better than the reference drug SAHA (GI50 = 2.90 µM). Moreover, compound 9b effectively promoted the accumulation of acetylated Histone H3 and α-tubulin through inhibition of HDAC1 and HDAC6 enzymes, respectively. HDAC enzyme assay also confirmed that compound 9b efficiently inhibited HDAC1 and HDAC6 isoforms with IC50 values of 84.9 nM and 95.9 nM. Furthermore, compound 9b inhibited colony formation capacity of SH-SY5Y cells, which is considered a hallmark of cell carcinogenesis and metastatic potential. The theoretical prediction, in vitro PAMPA-BBB assay, and in vivo brain pharmacokinetic studies confirmed that compound 9b had much higher BBB permeability than SAHA. In silico docking study demonstrated that compound 9b fitted in the substrate binding pocket of HDAC1 and HDAC6. Taken together, compound 9b provided a novel scaffold for developing CNS penetrant HDAC inhibitors and therapeutic potential for CNS-related diseases.
Assuntos
Peptídeos beta-Amiloides/química , Histona Desacetilase 1 , Desacetilase 6 de Histona , Inibidores de Histona Desacetilases , Simulação de Acoplamento Molecular , Proteínas de Neoplasias , Linhagem Celular Tumoral , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Histona Desacetilase 1/antagonistas & inibidores , Histona Desacetilase 1/química , Desacetilase 6 de Histona/antagonistas & inibidores , Desacetilase 6 de Histona/química , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/química , Humanos , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/química , Neoplasias/química , Neoplasias/tratamento farmacológico , Neoplasias/metabolismoRESUMO
Triple-negative breast cancers (TNBCs) are the most aggressive and metastatic subtype of breast cancers and exhibit poor clinical outcome due to the lack of drug target receptors such as estrogen receptors (ER), progesterone receptors (PR), and human epidermal growth factor receptor 2 (Her2). The limited effectiveness of therapeutic options and the poor prognosis of TNBC patients emphasize the urgent need for identifying new therapeutic agents. In this regard, heat shock protein 90 (Hsp90) has emerged as a promising therapeutic target for the treatment of TNBCs. Hsp90 is a molecular chaperone that regulates the folding, stability, and function of many oncogenic proteins. Hence, the inhibition of Hsp90 chaperone function leads to a simultaneous blockage of multiple signaling pathways in the proliferation and survival of cancers. In the present study, we performed the design, synthesis, and biological evaluation of Hsp90 inhibitors and found that a synthetic small molecule, DPide exerted a potent anticancer activity against TNBC cell line, MDAMB231 and nonsmall cell lung cancer (NSCLC) cell line, H1975 with GI50 values of 0.478 and 1.67 µM, respectively. Softagar colony formation assay also revealed that DPide suppressed the anchorageindependent growth of MDAMB231 cells. Western blot analysis indicated that the treatment of MDAMB231 cells with DPide induced the proteasomal degradation of EGFR, Her2, Met, Akt, cRaf, and Cdk4 and the consequent cleavage of PARP, leading to apoptotic cell death. DPide also inhibited the migration and MMP9 activity of MDAMB231 cells, suggesting that the metastatic potential of TNBCs could be suppressed by DPide. Collectively, DPide offers an effective therapeutic option for the treatment TNBCs.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Proteínas de Ciclo Celular/química , Chaperoninas/química , Bibliotecas de Moléculas Pequenas/química , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Apoptose/efeitos dos fármacos , Sítios de Ligação/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Chaperoninas/antagonistas & inibidores , Chaperoninas/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Modelos Moleculares , Simulação de Acoplamento Molecular , Proteínas de Neoplasias/genética , Conformação Proteica/efeitos dos fármacos , Receptores de Estrogênio/genética , Receptores de Progesterona/genética , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/administração & dosagem , Bibliotecas de Moléculas Pequenas/síntese química , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Heat shock protein 90 (Hsp90) is a ubiquitous molecular chaperone that is responsible for the stabilization and maturation of many oncogenic proteins. Therefore, Hsp90 has emerged as an attractive target in the field of cancer chemotherapy. In this study, we report the design, synthesis, and biological evaluation of a series of Hsp90 inhibitors. In particular, compound 30f shows a significant Hsp90α inhibitory activity with IC50 value of 5.3 nM and an excellent growth inhibition with GI50 value of 0.42 µM against non-small cell lung cancer cells, H1975. Compound 30f effectively reduces the expression levels of Hsp90 client proteins including Her2, EGFR, Met, Akt, and c-Raf. Consequently, compound 30f promotes substantial cleavages of PARP, Caspase 3, and Caspase 8, indicating that 30f induces cancer cell death via apoptotic pathway. Moreover, cytochrome P450 assay indicates that compound 30f has weak inhibitory effect on the activities of five major P450 isoforms (IC50 > 5 µM for 1A2, 2C9, 2C19, 2D6, and 3A), suggesting that clinical interactions between 30f and the substrate drugs of the five major P450 isoforms are not expected. Compound 30f also inhibits the tumor growth in a mouse xenograft model bearing subcutaneous H1975 without noticeable abnormal behavior and body weight changes. The immunostaining and western immunoblot analysis of EGFR, Met, Akt in xenograft tissue sections of tumor further demonstrate a good agreement with the in vitro results.
Assuntos
Antineoplásicos/farmacologia , Benzamidas/farmacologia , Inibidores das Enzimas do Citocromo P-450/farmacologia , Desenho de Fármacos , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Resorcinóis/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Benzamidas/síntese química , Benzamidas/química , Proliferação de Células/efeitos dos fármacos , Inibidores das Enzimas do Citocromo P-450/síntese química , Inibidores das Enzimas do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/metabolismo , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Masculino , Camundongos Endogâmicos NOD , Camundongos SCID , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Resorcinóis/química , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
The molecular chaperone Hsp90 plays an important role in cancer cell survival and proliferation by regulating the maturation and stabilization of numerous oncogenic proteins. Due to its potential to simultaneously disable multiple signaling pathways, Hsp90 has emerged as an attractive therapeutic target for cancer treatment. In this study, the design, synthesis, and biological evaluation of a series of Hsp90 inhibitors are described. Among the synthetic compounds, 6,7-dihydrothieno [3,2-c]pyridin-5(4H)-yl amide 19 exhibits a remarkable binding affinity to the N-terminus of Hsp90 in a fluorescence polarization (FP) binding assay (IC50 = 50.3 nM). Furthermore, it effectively inhibits the proliferation of H1975 non-small cell lung cancer (NSCLC) and Skbr3 breast cancer cell lines with GI50 values of 0.31 µM and 0.11 µM, respectively. Compound 19 induces the degradation of the Hsp90 client proteins including EGFR, Her2, Met, c-Raf, and Akt, and consequently promotes apoptotic cancer cell death. Compound 19 also inhibits the growth of H1975 xenografts in NOD-scid IL2R gammanull mice without any apparent body-weight loss. The immunohistologic evaluation indicates that compound 19 decreases the expression of Akt in xenograft tumor tissue via an inhibition of the Hsp90 chaperon function. Additionally, the cytochrome P450 assay indicates that compound 19 has no effect on the activities of five major P450 isoforms (IC50 > 50 µM for 1A2, 2C9, 2C19, 2D6, and 3A), suggesting that clinical interactions between compound 19 and the substrate drugs of the five major P450 isoforms are not expected. Overall, compound 19 represents a new class of Hsp90 inhibitor with its 6,7-dihydrothieno[3,2-c]pyridin-5(4H)-yl-amide structure, and it has the therapeutic potential to overcome drug resistance in cancer chemotherapy.