RESUMO
Cathepsins, a family of lysosomal peptidases, play a crucial role in maintaining cellular homeostasis by regulating protein turnover and degradation as well as many specific regulatory actions that are important for proper cell function and human health. Alterations in the activity and expression of cathepsins have been observed in many diseases such as cancer, inflammation, neurodegenerative disorders, bone remodelling-related conditions and others. These changes are not exclusively harmful, but rather appear to be a compensatory response on the lack of one cathepsin in order to maintain tissue integrity. The upregulation of specific cathepsins in response to the inhibition or dysfunction of other cathepsins suggests a fine-tuned system of proteolytic balance and understanding the compensatory role of cathepsins may improve therapeutic potential of cathepsin's inhibitors. Selectively targeting one cathepsin or modulating their activity could offer new treatment strategies for a number of diseases. This review emphasises the need for comprehensive research into cathepsin biology in the context of disease. The identification of the specific cathepsins involved in compensatory responses, the elucidation of the underlying molecular mechanisms and the development of targeted interventions could lead to innovative therapeutic approaches.
RESUMO
Glioblastoma (GBM) is the most common and deadly primary brain tumor in adults. Understanding GBM pathobiology and discovering novel therapeutic targets are critical to finding efficient treatments. Upregulation of the lysosomal cysteine carboxypeptidase cathepsin X has been linked to immune dysfunction and neurodegenerative diseases, but its role in cancer and particularly in GBM progression in patients is unknown. In this study, cathepsin X expression and activity were found to be upregulated in human GBM tissues compared to low-grade gliomas and nontumor brain tissues. Cathepsin X was localized in GBM cells as well as in tumor-associated macrophages and microglia. Subsequently, potent irreversible (AMS36) and reversible (Z7) selective cathepsin X inhibitors were tested in vitro. Selective cathepsin X inhibitors decreased the viability of patient-derived GBM cells as well as macrophages and microglia that were cultured in conditioned media of GBM cells. We next examined the expression pattern of neuron-specific enzyme γ-enolase, which is the target of cathepsin X. We found that there was a correlation between high proteolytic activity of cathepsin X and C-terminal cleavage of γ-enolase and that cathepsin X and γ-enolase were colocalized in GBM tissues, preferentially in GBM-associated macrophages and microglia. Taken together, our results on patient-derived material suggest that cathepsin X is involved in GBM progression and is a potential target for therapeutic approaches against GBM.
Assuntos
Neoplasias Encefálicas/metabolismo , Catepsina Z/metabolismo , Glioblastoma/metabolismo , Fosfopiruvato Hidratase/metabolismo , Microambiente Tumoral , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Catepsina Z/antagonistas & inibidores , Catepsina Z/genética , Regulação Neoplásica da Expressão Gênica , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Microglia/efeitos dos fármacos , Microglia/metabolismo , Regulação para CimaRESUMO
New therapeutic targets that could improve current antitumor therapy and overcome cancer resistance are urgently needed. Promising candidates are lysosomal cysteine cathepsins, proteolytical enzymes involved in various critical steps during cancer progression. Among them, cathepsin X, which acts solely as a carboxypeptidase, has received much attention. Our results indicate that the triazole-based selective reversible inhibitor of cathepsin X named Z9 (1-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)-2-((4-isopropyl-4H-1,2,4-triazol-3-yl)thio)ethan-1-one) significantly reduces tumor progression, both in vitro in cell-based functional assays and in vivo in two independent tumor mouse models: the FVB/PyMT transgenic and MMTV-PyMT orthotopic breast cancer mouse models. One of the mechanisms by which cathepsin X contributes to cancer progression is the compensation of cathepsin-B activity loss. Our results confirm that cathepsin-B inhibition is compensated by an increase in cathepsin X activity and protein levels. Furthermore, the simultaneous inhibition of both cathepsins B and X with potent, selective, reversible inhibitors exerted a synergistic effect in impairing processes of tumor progression in in vitro cell-based assays of tumor cell migration and spheroid growth. Taken together, our data demonstrate that Z9 impairs tumor progression both in vitro and in vivo and can be used in combination with other peptidase inhibitors as an innovative approach to overcome resistance to antipeptidase therapy.
Assuntos
Catepsina B/antagonistas & inibidores , Catepsinas/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Carga Tumoral/efeitos dos fármacos , Animais , Catepsina B/metabolismo , Catepsinas/genética , Catepsinas/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Inibidores Enzimáticos/química , Humanos , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Camundongos Transgênicos , Invasividade Neoplásica , Infiltração de Neutrófilos/efeitos dos fármacosRESUMO
Modern anticancer therapies favor a targeted approach. Tyrosine kinase inhibitors (TKIs) are drugs that target molecular pathways involved in various types of malignancies. Although TKIs are safe and well tolerated, they remain not completely selective; e.g., endocrine-mediated adverse events have been observed with their use. In the present study, the effects of seven TKIs were determined on the activities of androgen receptor, estrogen receptor α (ERα), glucocorticoid receptor and thyroid receptor in vitro using stably transfected cell lines expressing firefly luciferase reporter gene: AR-EcoScreen, hERα-HeLa9903, MDA-kb2, and GH3.TRE-Luc cells, respectively. Antiandrogenic activity was seen for erlotinib, estrogenic activity for imatinib, antiestrogenic activity for dasatinib, erlotinib, nilotinib, regorafenib and sorafenib, glucocorticoid activity for erlotinib and ibrutinib, antiglucocorticoid activity for regorafenib and sorafenib, and antithyroid activity for ibrutinib. Additionally, synergism was seen for 1-5 µM dasatinib and 500 nM hydrocortisone combination for glucocorticoid activity in MDA-kb2 cells. The estrogenic activity of imatinib was confirmed as mediated through ERα, and interference of the TKIs with the reporter gene assays was ruled out in a cell-lysate-based firefly luciferase enzyme inhibition assay. Imatinib in combination with 4-hydroxytamoxifen showed concentration-dependent effects on the metabolic activity of ERα-expressing AN3CA, MCF-7, and SKOV3 cells, and on cell proliferation and adhesion of MCF-7 cells. These findings contribute to the understanding of the endocrine effects of TKIs, in terms of toxicity and effectiveness, and define the need to further evaluate the endocrine disrupting activities of TKIs to safeguard human and environmental health.
Assuntos
Antineoplásicos/farmacologia , Antitireóideos/farmacologia , Receptor alfa de Estrogênio/antagonistas & inibidores , Proteínas Tirosina Quinases/antagonistas & inibidores , Receptores de Glucocorticoides/antagonistas & inibidores , Antagonistas de Receptores de Andrógenos , Animais , Linhagem Celular , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Hormônios TireóideosRESUMO
Endocrine disrupting chemicals (EDCs) are associated with cancer development and progression due to their promotion of increased cell invasiveness and metastasis formation. However, the effects of EDCs on cell adhesion mediated through integrins have not been well studied to date. Their actions are implicated by binding sites for hormones on the vitronectin receptor (VTNR; or integrin αvß3), which is involved in tumor angiogenesis and metastasis. VTNR-expressing human umbilical vein endothelial cells (HUVECs) were used to determine the effects of EDCs and endogenous hormones on cell adhesion to vitronectin-coated surfaces, and on VTNR activation. Cell adhesion was significantly increased for bisphenol A, triclocarban, and triclosan (10, 100 nM; p < 0.05), with similar trends for bisphenols AF and S (10, 100 nM; p > 0.05). No changes in cell adhesion were seen for 5α-dihydrotestosterone, 17ß-estradiol, triiodothyronine, imatinib and paroxetine. These data indicate that EDC-mediated increases in HUVEC adhesion to vitronectin are not mediated through androgenic, estrogenic, or thyroid activities, nor through activation of VTNR. Although these effects of EDCs on HUVEC adhesion require further investigation of the underlying mechanism(s) of action to define their biological relevance, the low-dose effects and nonmonotonic responses revealed here define the need for further investigation of these EDCs.
Assuntos
Adesão Celular/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Integrina alfaVbeta3/efeitos dos fármacos , Compostos Benzidrílicos/toxicidade , Carbanilidas/toxicidade , Fluorocarbonos/toxicidade , Hormônios/toxicidade , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Fenóis/toxicidade , Sulfonas/toxicidade , Triclosan/toxicidade , Vitronectina/metabolismoRESUMO
Cysteine cathepsins are primarily involved in the degradation and recycling of proteins in endo-lysosomal compartments but are also gaining recognition as pivotal proteolytic contributors to various immune functions. Through their extracellular proteolytic activities within the hematopoietic stem cell niche, they are involved in progenitor cell mobilization and differentiation. Cysteine cathepsins, such as cathepsins L and S contribute to antigen-induced adaptive immunity through major histocompatibility complex class II antigen presentation whereas cathepsin X regulates T-cell migration. By regulating toll-like receptor signaling and cytokine secretion cysteine cathepsins activate innate immune cells and affect their functional differentiation. Cathepsins C and H are expressed in cytotoxic T lymphocytes and natural killer cells and are involved in processing of pro-granzymes into proteolytically active forms. Cytoplasmic activities of cathepsins B and L contribute to the maintenance of homeostasis of the adaptive immune response by regulating cell death of T and B lymphocytes. The expression pattern, localization, and activity of cysteine cathepsins is tightly connected to their function in immune cells. Furthermore, cysteine cathepsins together with their endogenous inhibitors, serve as mediators in the interplay between cancer and immune cells that results in immune cell anergy. The aim of the present article is to review the mechanisms of dysregulation of cysteine cathepsins and their inhibitors in relation to immune dysfunction to address new possibilities for regulation of their function.
Assuntos
Diferenciação Celular/imunologia , Cisteína Proteases/metabolismo , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Imunomodulação , Animais , Diferenciação Celular/genética , Anergia Clonal/imunologia , Cisteína Proteases/química , Cisteína Proteases/genética , Inibidores de Cisteína Proteinase/farmacologia , Humanos , Tolerância Imunológica , Imunomodulação/efeitos dos fármacos , Imunossenescência/efeitos dos fármacos , Família Multigênica , Organogênese/genética , Organogênese/imunologia , Relação Estrutura-Atividade , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Over the last 2 decades, several coronaviruses (CoVs) have crossed the species barrier into humans, causing highly prevalent and severe respiratory diseases, often with fatal outcomes. CoVs are a large group of enveloped, single-stranded, positive-sense RNA viruses, which encode large replicase polyproteins that are processed by viral peptidases to generate the nonstructural proteins (Nsps) that mediate viral RNA synthesis. Papain-like peptidases (PLPs) and chymotrypsin-like cysteine 3C-like peptidase are essential for coronaviral replication and represent attractive antiviral drug targets. Furthermore, CoVs utilize the activation of their envelope spike glycoproteins by host cell peptidases to gain entry into cells. CoVs have evolved multiple strategies for spike protein activation, including the utilization of lysosomal cysteine cathepsins. In this review, viral and host peptidases involved in CoV cell entry and replication are discussed in depth, with an emphasis on papain-like cysteine cathepsins. Furthermore, important findings on cysteine peptidase inhibitors with regard to virus attenuation are highlighted as well as the potential of such inhibitors for future treatment strategies for CoV-related diseases.
Assuntos
Antivirais/farmacologia , Infecções por Coronavirus/virologia , Coronavirus/efeitos dos fármacos , Inibidores de Cisteína Proteinase/farmacologia , Internalização do Vírus/efeitos dos fármacos , Animais , Infecções por Coronavirus/tratamento farmacológico , Humanos , Replicação Viral/efeitos dos fármacosRESUMO
Increased proteolytic activity of cysteine cathepsins has long been known to facilitate malignant progression, and it has also been associated with tumor-promoting roles of myeloid-derived suppressor cells (MDSCs). Consequently, cysteine cathepsins have gained much attention as potential targets for cancer therapies. However, cross-talk between tumor cells and MDSCs needs to be taken into account when studying the efficacy of cathepsin inhibitors as anti-cancer agents. Here, we demonstrate the potential of the MDA-MB-231 breast cancer cell line to generate functional MDSCs from CD14+ cells of healthy human donors. During this transition to MDSCs, the overall levels of cysteine cathepsins increased, with the largest responses for cathepsins L and X. We used small-molecule inhibitors of cathepsins L and X (i.e., CLIK-148, Z9, respectively) to investigate their functional impact on tumor cells and immune cells in this co-culture system. Interactions with peripheral blood mononuclear cells reduced MDA-MB-231 cell invasion, while inhibition of cathepsin X activity by Z9 restored invasion. Inhibition of cathepsin L activity using CLIK-148 resulted in significantly increased CD8+ cytotoxicity. Of note, inhibition of cathepsins L and X in separate immune or tumor cells did not promote these functional changes. Together, our findings underlie the importance of tumor cell-immune cell interactions in the evaluation of the anti-cancer potential of cysteine cathepsin inhibitors.
Assuntos
Catepsina L/metabolismo , Cisteína/metabolismo , Células Supressoras Mieloides/metabolismo , Neoplasias/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Humanos , Leucócitos Mononucleares/metabolismo , Invasividade Neoplásica/patologia , Neoplasias/patologia , Células PC-3RESUMO
Cathepsins are lysosomal peptidases involved in intracellular protein catabolism as well as in various other physiological and pathological processes. Several members of the family, most notably cathepsins B, S, K and L, are frequently overexpressed in cancer and have been associated with remodeling of the proteins of the extracellular matrix, a process leading to tumor cell migration, invasion and metastasis. In addition, lysosomal cathepsins play a role in innate and adaptive immunity, regulation of antigen presentation, Toll-like receptor signaling, cytokine secretion, apoptosis, autophagy, differentiation, migration and cytotoxicity. In cancer, the cells of innate immunity, such as myeloid cells, are often subverted to the regulatory immunosuppressive phenotype. Most studies indicate that lysosomal cathepsins reinforce the pro-tumoral activity of myeloid-derived suppressor cells and tumor-associated macrophages as well as of neutrophils. On the other hand, in cytotoxic natural killer cells, tumor cells suppress lysosomal peptidases in their activation of perforin and granzymes, thus diminishing their killing ability. With multifaceted actions, lysosomal peptidases constitute an important regulatory mechanism for fine-tuning the anti-tumor immune response.
Assuntos
Imunidade Inata , Lisossomos/metabolismo , Neoplasias/etiologia , Neoplasias/metabolismo , Peptídeo Hidrolases/metabolismo , Animais , Biomarcadores , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Imunidade , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/metabolismo , Neoplasias/patologia , Transdução de Sinais , Receptores Toll-Like/metabolismoRESUMO
Cysteine cathepsins are peptidases with housekeeping functions that play different specific roles in different tissues. Endogenous peptidase inhibitors, such as cystatins and thyropins are the ultimate way of controlling their activity. It appears, however, that cathepsin X, a monocarboxypeptidase, whose overexpression is associated with several pathological processes, is not under the control of endogenous inhibitors. Inhibitors belonging to various groups inhibit other cathepsins tested, but none decrease the carboxypeptidase activity of cathepsin X. This absence of inhibitor control is another feature that distinguishes cathepsin X from other members of the cysteine peptidases.
RESUMO
Cathepsins B and X are lysosomal cysteine carboxypeptidases suggested as having a redundant role in cancer. They are involved in a number of processes leading to tumor progression but their role in the epithelial-mesenchymal transition (EMT) remains unknown. We have investigated the contribution of both cathepsins B and X in EMT using tumor cell lines differing in their expression of epithelial and mesenchymal markers and cell morphology. Higher levels of both cathepsins are shown to promote EMT and are associated with the mesenchymal-like cell phenotype. Moreover, simultaneous knockdown of the two peptidases triggers a reverse, mesenchymal to epithelial transition. Of the two cathepsins, cathepsin B appears to be the stronger promotor of EMT. Furthermore, we evaluated the involvement of cathepsin B and X in the transforming growth factor-ß1 (TGF-ß1) signaling pathway, one of the key signaling mechanisms triggering EMT in cancer. In MCF-7 cells the expression of cathepsin B was shown to depend on their activation with TGF-ß1 while, for cathepsin X, a TGF-ß1 independent mechanism of induction during EMT is indicated. EMT is thus shown to be another mechanism linking cathepsins B and X with tumor progression. With silencing of their expression or inhibition of enzymatic activity, the tumor cells could be reverted to less aggressive epithelial-like phenotype.
Assuntos
Neoplasias da Mama/genética , Catepsina B/genética , Catepsina Z/genética , Fator de Crescimento Transformador beta1/genética , Neoplasias da Mama/patologia , Movimento Celular/genética , Cisteína/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Humanos , Células MCF-7 , Transdução de Sinais/genéticaRESUMO
Cathepsin X, a cysteine carboxypeptidase, is upregulated in several types of cancer. Its molecular target in tumor cells is profilin 1, a known tumor suppressor and regulator of actin cytoskeleton dynamics. Cathepsin X cleaves off the C-terminal Tyr139 of profilin 1, affecting binding of poly-L-proline ligands and, consequently, tumor cell migration and invasion. Profilin 1 with mutations at the C-terminus, transiently expressed in prostate cancer cells PC-3, showed that Tyr139 is important for proper function of profilin 1 as a tumor suppressor. Cleaving off Tyr139 prevents the binding of clathrin, a poly-L-proline ligand involved in endocytosis. More profilin 1-clathrin complexes were present in PC-3 cells when cathepsin X was inhibited by its specific inhibitor AMS36 or silenced by siRNA. As a consequence, the endocytosis of FITC-labeled dextran and transferrin conjugate was significantly increased. These results constitute the first report of the regulation of clathrin-mediated endocytosis in tumor cells through proteolytic processing of profilin 1.
Assuntos
Catepsina Z/metabolismo , Clatrina/fisiologia , Endocitose/fisiologia , Profilinas/metabolismo , Tirosina/química , Linhagem Celular Tumoral , Expressão Gênica , Humanos , Invasividade Neoplásica , Metástase Neoplásica , Polimerização , Profilinas/química , Profilinas/genética , ProteóliseRESUMO
AIM: We hypothesized that, in stable angina patients, atorvastatin therapy lowers the cathepsin S (CTSS) concentrations, as assessed non-invasively according to a plasma analysis. In addition, the low-density lipoprotein (LDL) and high-density lipoprotein (HDL) size and subclasses in the plasma were analysed to establish the association between CTSS and lipoprotein metabolism and determine whether this association is atorvastatin-sensitive. METHODS: A total of 43 patients with stable angina received atorvastatin therapy (20 mg/day, 10 weeks). The plasma CTSS mRNA levels, CTSS protein concentrations and CTSS activity, as well as LDL and HDL size and subclasses, were analysed before and after treatment. RESULTS: Atorvastatin treatment did not change the plasma CTSS mRNA levels, although it lowered the plasma CTSS concentrations and activity. An increased plasma CTSS concentration and activity were found to be associated with a more atherogenic LDL subclass profile (a decreased dominant LDL size and increased percentage of small, dense LDL particles). The atorvastatin-induced CTSS-loweringeffect was concomitant with an improvement in the LDL subclass profile, and the changes were found to be interrelated. Concomitant, interrelated changes in the CTSS levels and LDL subclass profiles were found in the LDL phenotype B patients only (a dominant LDL diameter of ≤ 25.5 nm at the start of the study). In this subgroup, lowering of the plasma CTSS mRNA level also correlated with lowering of the proportion of small, dense LDL particles. CONCLUSIONS: Atorvastatin-induced CTSS-lowering and LDL subclass profile improvements in the plasma of LDL phenotype B patients with stable angina are concomitant and interrelated.
Assuntos
Angina Estável/sangue , Angina Estável/tratamento farmacológico , Biomarcadores/análise , Catepsinas/genética , HDL-Colesterol/sangue , LDL-Colesterol/sangue , Ácidos Heptanoicos/uso terapêutico , Pirróis/uso terapêutico , Angina Estável/genética , Anticolesterolemiantes/uso terapêutico , Atorvastatina , Western Blotting , Catepsinas/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Cathepsin X has been reported to be a tumor promotion factor in various types of cancer; however, the molecular mechanisms linking its activity with malignant processes are not understood. Here we present profilin 1, a known tumor suppressor, as a target for cathepsin X carboxypeptidase activity in prostate cancer PC-3 cells. Profilin 1 co-localizes strongly with cathepsin X intracellularly in the perinuclear area as well as at the plasma membrane. Selective cleavage of C-terminal amino acids was demonstrated on a synthetic octapeptide representing the profilin C-terminal region, and on recombinant profilin 1. Further, intact profilin 1 binds its poly-L-proline ligand clathrin significantly better than it does the truncated one, as shown using cathepsin X specific inhibitor AMS-36 and immunoprecipitation of the profilin 1/clathrin complex. Moreover, the polymerization of actin, which depends also on the binding of poly-L-proline ligands to profilin 1, was promoted by AMS-36 treatment of cells and by siRNA cathepsin X silencing. Our results demonstrate that increased adhesion, migration and invasiveness of tumor cells depend on the inactivation of the tumor suppressive function of profilin 1 by cathepsin X. The latter is thus designated as a target for development of new antitumor strategies.
Assuntos
Catepsina K/metabolismo , Profilinas/metabolismo , Neoplasias da Próstata , Actinas/metabolismo , Catepsina K/antagonistas & inibidores , Catepsina K/genética , Adesão Celular/genética , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Movimento Celular/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Ligantes , Masculino , Invasividade Neoplásica/genética , Profilinas/genética , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Ligação Proteica , RNA Interferente PequenoRESUMO
The motility of T cells depends on the dynamic spatial regulation of integrin-mediated adhesion and de-adhesion. Cathepsin X, a cysteine protease, has been shown to regulate T-cell migration by interaction with lymphocyte function associated antigen-1 (LFA-1). LFA-1 adhesion to the ICAM-1 is controlled by the association of actin-binding proteins with the cytoplasmic tail of the beta(2) chain of LFA-1. Cleavage by cathepsin X of the amino acid residues S(769), E(768) and A(767) from the C-terminal of the beta(2) cytoplasmic tail of LFA-1 is shown to promote binding of the actin-binding protein alpha-actinin-1. Furthermore, cathepsin X overexpression reduced LFA-1 clustering and induced an intermediate affinity LFA-1 conformation that is known to associate with alpha-actinin-1. Increased levels of intermediate affinity LFA-1 resulted in augmented cell spreading due to reduced attachment of T cells to the ICAM-1-coated surface. Gradual cleavage of LFA-1 by cathepsin X enables the transition between intermediate and high affinity LFA-1, an event that is crucial for effective T-cell migration.