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We synthesized new pyrrole and indole derivatives as human carbonic anhydrase (hCA) inhibitors with the potential to inhibit the Wnt/ß-catenin signaling pathway. The presence of both N1-(4-sulfonamidophenyl) and 3-(3,4,5-trimethoxyphenyl) substituents was essential for strong hCA inhibitors. The most potent hCA XII inhibitor 15 (Ki = 6.8 nM) suppressed the Wnt/ß-catenin signaling pathway and its target genes MYC, Fgf20, and Sall4 and exhibited the typical markers of apoptosis, cleaved poly(ADP-ribose)polymerase, and cleaved caspase-3. Compound 15 showed strong inhibition of viability in a panel of cancer cells, including colorectal cancer and triple-negative breast cancer cells, was effective against the NCI/ADR-RES DOX-resistant cell line, and restored the sensitivity to doxorubicin (DOX) in HT29/DX and MDCK/P-gp cells. Compound 15 is a novel dual-targeting compound with activity against hCA and Wnt/ß-catenin. It thus has a broad targeting spectrum and is an anticancer agent with specific potential in P-glycoprotein overexpressing cell lines.
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Anhidrasas Carbónicas , Neoplasias , Humanos , Relación Estructura-Actividad , Resistencia a Múltiples Medicamentos , Vía de Señalización Wnt , Resistencia a Antineoplásicos , Anhidrasas Carbónicas/metabolismo , Inhibidores de Anhidrasa Carbónica/farmacología , Anhidrasa Carbónica IX , Estructura Molecular , BencenosulfonamidasRESUMEN
Despite intensive efforts, no inhibitors of the Wnt/ß-catenin signaling pathway have been approved so far for the clinical treatment of cancer. We synthesized novel N-(heterocyclylphenyl)benzenesulfonamides as ß-catenin inhibitors. Compounds 5-10 showed strong inhibition of the luciferase activity. Compounds 5 and 6 inhibited the MDA-MB-231, HCC1806, and HCC1937 TNBC cells. Compound 9 induced in vitro cell death in SW480 and HCT116 cells and in vivo tumorigenicity of a human colorectal cancer line HCT116. In a co-immunoprecipitation study in HCT116 cells transfected with Myc-tagged T-cell factor 4 (Tcf-4), compound 9 abrogated the association between ß-catenin and Tcf-4. The crystallographic analysis of the ß-catenin Armadillo repeats domain revealed that compound 9 and Tcf-4 share a common binding site within the hotspot binding region close to Lys508. To our knowledge, compound 9 is the first small molecule ligand of this region to be reported. These results highlight the potential of this novel class of ß-catenin inhibitors as anticancer agents.
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A key mechanism driving colorectal cancer (CRC) development is the upregulation of MYC and its targets, including ornithine decarboxylase (ODC), a master regulator of polyamine metabolism. Elevated polyamines promote tumorigenesis in part by activating DHPS-mediated hypusination of the translation factor eIF5A, thereby inducing MYC biosynthesis. Thus, MYC, ODC and eIF5A orchestrate a positive feedback loop that represents an attractive therapeutic target for CRC therapy. Here we show that combined inhibition of ODC and eIF5A induces a synergistic antitumor response in CRC cells, leading to MYC suppression. We found that genes of the polyamine biosynthesis and hypusination pathways are significantly upregulated in colorectal cancer patients and that inhibition of ODC or DHPS alone limits CRC cell proliferation through a cytostatic mechanism, while combined ODC and DHPS/eIF5A blockade induces a synergistic inhibition, accompanied to apoptotic cell death in vitro and in mouse models of CRC and FAP. Mechanistically, we found that this dual treatment causes complete inhibition of MYC biosynthesis in a bimodal fashion, by preventing translational elongation and initiation. Together, these data illustrate a novel strategy for CRC treatment, based on the combined suppression of ODC and eIF5A, which holds promise for the treatment of CRC.
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Neoplasias Colorrectales , Factores de Iniciación de Péptidos , Poliaminas , Proteínas Proto-Oncogénicas c-myc , Animales , Ratones , Apoptosis , Proliferación Celular , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismo , Ornitina Descarboxilasa/farmacología , Poliaminas/metabolismo , Humanos , Factores de Iniciación de Péptidos/genética , Factores de Iniciación de Péptidos/metabolismo , Proteínas Proto-Oncogénicas c-myc/metabolismo , Factor 5A Eucariótico de Iniciación de TraducciónRESUMEN
The Hedgehog receptor, Patched1 (PTCH1), is a well-known tumour suppressor. While the tumour suppressor's activity is mostly ascribed to its function as a repressor of the canonical Smoothened/Gli pathway, its C-terminal domain (CTD) was reported to have additional non-canonical functions. One of them is the reduction of autophagic flux through direct interaction with the Unc-51, like the autophagy activating kinase (ULK) complex subunit autophagy-related protein-101 (ATG101). With the aim of investigating whether this function of PTCH1 is important in cancer cell fitness, we first identified frameshift mutations in the CTD of PTCH1 in cancer databases. We demonstrated that those mutations disrupt PTCH1 interaction with ATG101 and increase autophagic flux. Using deletion mutants of the PTCH1 CTD in co-immunoprecipitation studies, we established that the 1309-1447 region is necessary and sufficient for interaction with ATG101. We next showed that the three most common PTCH1 CTD mutations in endometrial, stomach and colon adenocarcinomas that cause frameshifts at S1203, R1308 and Y1316 lack the ability to interact with ATG101 and limit autophagic flux, determined by bafilomycin A1-sensitive accumulation of the autophagy markers LC3BII and p62. We next engineered PTCH1 indel mutations at S1223 by CRISPR/Cas9 in SW620 colon cancer cells. Comparison of two independent clones harbouring PTCH1 S1223fs mutations to their isogenic parental cell lines expressing wild-type PTCH1 showed a significant increase in basal and rapamycin-stimulated autophagic flux, as predicted by loss of ATG101 interaction. Furthermore, the PTCH1 CTD mutant cells displayed increased proliferation in the presence of rapamycin and reduced sensitivity to glycolysis inhibitors. Our findings suggest that loss of the PTCH1-ATG101 interaction by mutations in the CTD of PTCH1 in cancer might confer a selective advantage by stimulating autophagy and facilitating adaptation to nutrient deprivation conditions.
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BACKGROUND & AIMS: Common precursors for the liver, biliary tree, and pancreas exist at an early stage of development in the definitive endoderm forming the foregut. We have identified and characterised endodermal stem/progenitor cells with regenerative potential persisting in the adult human duodenum. METHODS: Human duodena were obtained from organ donors, and duodenal submucosal gland cells were isolated after removal of the mucosa layer. Cells were cultured on plastic or as organoids and were transplanted into severe combined immunodeficient (SCID) mouse livers. RESULTS: In situ studies of submucosal glands in the human duodenum revealed cells expressing stem/progenitor cell markers that had unique phenotypic traits distinguishable from intestinal crypt cells. Genetic signature studies indicated that the cells are closer to biliary tree stem cells and to definitive endodermal cells than to adult hepatocytes, supporting the interpretation that they are endodermal stem/progenitor cells. In vitro, human duodenal submucosal gland cells demonstrated clonal growth, capability to form organoids, and ability to acquire functional hepatocyte traits. In vivo, transplanted cells engrafted into the livers of immunocompromised mice and differentiated to mature liver cells. In an experimental model of fatty liver, human duodenal submucosal gland cells were able to rescue hosts from liver damage by supporting repopulation and regeneration of the liver. CONCLUSIONS: A cell population with clonal growth and organoid formation capability, which has liver differentiation potency in vitro and in vivo in murine experimental models, is present within adult duodenal submucosal glands. These cells can be isolated, do not require reprogramming, and thus could potentially represent a novel cell source for regenerative medicine of the liver. IMPACT AND IMPLICATIONS: Cell therapies for liver disease could represent an option to support liver function, but the identification of sustainable and viable cell sources is critical. Here, we describe a cell population with organoid formation capability and liver-specific regenerative potential in submucosal glands of the human duodenum. Duodenal submucosal gland cells are isolated from adult organs, do not require reprogramming, and could rescue hepatocellular damage in preclinical models of chronic, but not acute, liver injury. Duodenal submucosal gland cells could represent a potential candidate cell source for regenerative medicine of the liver, but the determination of cell dose and toxicity is needed before clinical testing in humans.
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Sistema Biliar , Hiperplasia Nodular Focal , Adulto , Humanos , Ratones , Animales , Ratones SCID , Regeneración Hepática , Hepatocitos , Hígado/lesiones , Diferenciación CelularRESUMEN
BACKGROUND: Thyroid hormones (TH)s are master regulators of mitochondrial activity and biogenesis. Nonthyroidal illness syndrome (NTIS) is generally considered an adaptative response to reduced energy that is secondary to critical illness, including COVID-19. COVID-19 has been associated with profound changes in the cell energy metabolism, especially in the cells of the immune system, with a central role played by the mitochondria, considered the power units of every cell. Infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) affects and alters mitochondrial functions, both to influence its intracellular survival and to evade host immunity. AIM OF THE STUDY: This study was undertaken to analyze the oxidative balance and mitochondrial respiration in COVID-19 patients with and without NTIS to elucidate the role that thyroid hormones (TH)s play in this context. METHODS: In our cohort of 54 COVID-19 patients, admitted to our University Hospital during the COVID-19 pandemic, we evaluated the generation of reactive oxygen species (ROS) by measuring the serum levels of derivatives of reactive oxygen metabolites (dROMs), and we analyzed the antioxidant capacity by measuring the serum biological antioxidant potential (BAP). We then analyzed the mitochondrial respiration in peripheral blood mononuclear cells (PBMC)s of 28 of our COVID-19 patients, using the seahorse instrument (Agilent). Results were correlated with the serum levels of THs and, in particular, of FT3. In addition, the role of T3 on bioelectrical impedance analysis (BIA) and mitochondrial respiration parameters was directly evaluated in two COVID-19 patients with NTIS, in which treatment with synthetic liothyronine (LT3) was given both in vivo and in vitro. RESULTS: In our COVID-19 patients with NTIS, the dROMs values were significantly lower and the BAP values were significantly higher. Consequently, the oxidative stress index (OSi), measured as BAP/dROMs ratio was reduced compared to that observed in COVID-19 patients without NTIS, indicating a protective role exerted by NTIS on oxidative stress. In our COVID-19 patients, the mitochondrial respiration, measured in PBMCs, was reduced compared to healthy controls. Those with NTIS showed a reduced maximal respiratory capacity and a reduced proton leak, compared to those with normal FT3 serum values. Such lowered mitochondrial respiratory capacity makes the cells more vulnerable to bioenergetic exhaustion. In a pilot study involving two COVID-19 patients with NTIS, we could reinforce our previous observation regarding the role of T3 in the maintenance of adequate peripheral hydroelectrolytic balance. In addition, in these two patients, we demonstrated that by treating their PBMCs with LT3, both in vitro and in vivo, all mitochondrial respiration parameters significantly increased. CONCLUSIONS: Our results regarding the reduction in the serum levels of the reactive oxygen species (ROS) of COVID-19 patients with NTIS support the hypothesis that NTIS could represent an adaptative response to severe COVID-19. However, beside this beneficial effect, we demonstrate that, in the presence of an acute reduction of FT3 serum levels, the mitochondrial respiration is greatly impaired, with a consequent establishment of a hypoenergetic state of the immune cells that may hamper their capacity to react to massive viral infection.
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Most cancer cells switch their metabolism from mitochondrial oxidative phosphorylation to aerobic glycolysis to generate ATP and precursors for the biosynthesis of key macromolecules. The aerobic conversion of pyruvate to lactate, coupled to oxidation of the nicotinamide cofactor, is a primary hallmark of cancer and is catalyzed by lactate dehydrogenase (LDH), a central effector of this pathological reprogrammed metabolism. Hence, inhibition of LDH is a potential new promising therapeutic approach for cancer. In the search for new LDH inhibitors, we carried out a structure-based virtual screening campaign. Here, we report the identification of a novel specific LDH inhibitor, the pyridazine derivative 18 (RS6212), that exhibits potent anticancer activity within the micromolar range in multiple cancer cell lines and synergizes with complex I inhibition in the suppression of tumor growth. Altogether, our data support the conclusion that compound 18 deserves to be further investigated as a starting point for the development of LDH inhibitors and for novel anticancer strategies based on the targeting of key metabolic steps.
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L-Lactato Deshidrogenasa , Neoplasias , Línea Celular , Inhibidores Enzimáticos/farmacología , Glucólisis , Humanos , L-Lactato Deshidrogenasa/metabolismo , Ácido Láctico , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Fosforilación OxidativaRESUMEN
Biguanides are a family of antidiabetic drugs with documented anticancer properties in preclinical and clinical settings. Despite intensive investigation, how they exert their therapeutic effects is still debated. Many studies support the hypothesis that biguanides inhibit mitochondrial complex I, inducing energy stress and activating compensatory responses mediated by energy sensors. However, a major concern related to this "complex" model is that the therapeutic concentrations of biguanides found in the blood and tissues are much lower than the doses required to inhibit complex I, suggesting the involvement of additional mechanisms. This comprehensive review illustrates the current knowledge of pharmacokinetics, receptors, sensors, intracellular alterations, and the mechanism of action of biguanides in diabetes and cancer. The conditions of usage and variables affecting the response to these drugs, the effect on the immune system and microbiota, as well as the results from the most relevant clinical trials in cancer are also discussed.
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Neuroblastoma (NB) is a common malignant solid tumor in children and accounts for 15% of childhood cancer mortality. Amplification of the N-Myc oncogene is a well-established poor prognostic marker in NB patients and strongly correlates with higher tumor aggression and resistance to treatment. New therapies for patients with N-Myc-amplified NB need to be developed. After treating NB cells with BSAO/SPM, the detection of apoptosis was determined after annexin V-FITC labeling and DNA staining with propidium iodide. The mitochondrial membrane potential activity was checked, labeling cells with the probe JC-1 dye. We analyzed, by real-time RT-PCR, the transcript of genes involved in the apoptotic process, to determine possible down- or upregulation of mRNAs after the treatment on SJNKP and the N-Myc-amplified IMR5 cell lines with BSAO/SPM. The experiments were carried out considering the proapoptotic genes Tp53 and caspase-3. After treatment with BSAO/SPM, both cell lines displayed increased mRNA levels for all these proapoptotic genes. Western blotting analysis with PARP and caspase-3 antibody support that BSAO/SPM treatment induces high levels of apoptosis in cells. The major conclusion is that BSAO/SPM treatment leads to antiproliferative and cytotoxic activity of both NB cell lines, associated with activation of apoptosis.
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Amina Oxidasa (conteniendo Cobre)/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Apoptosis/efectos de los fármacos , Caspasa 3/metabolismo , MicroARNs/metabolismo , Proteína Proto-Oncogénica N-Myc/metabolismo , Neuroblastoma/tratamiento farmacológico , Espermina/farmacología , Proteína p53 Supresora de Tumor/metabolismo , Animales , Caspasa 3/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , MicroARNs/genética , Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/enzimología , Neuroblastoma/genética , Ratas Wistar , Transducción de Señal , Espermina/metabolismo , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Eukaryotic Translation Initiation Factor 5A (EIF5A) is a translation factor regulated by hypusination, a unique posttranslational modification catalyzed by deoxyhypusine synthetase (DHPS) and deoxyhypusine hydroxylase (DOHH) starting from the polyamine spermidine. Emerging data are showing that hypusinated EIF5A regulates key cellular processes such as autophagy, senescence, polyamine homeostasis, energy metabolism, and plays a role in cancer. However, the effects of EIF5A inhibition in preclinical cancer models, the mechanism of action, and specific translational targets are still poorly understood. We show here that hypusinated EIF5A promotes growth of colorectal cancer (CRC) cells by directly regulating MYC biosynthesis at specific pausing motifs. Inhibition of EIF5A hypusination with the DHPS inhibitor GC7 or through lentiviral-mediated knockdown of DHPS or EIF5A reduces the growth of various CRC cells. Multiplex gene expression analysis reveals that inhibition of hypusination impairs the expression of transcripts regulated by MYC, suggesting the involvement of this oncogene in the observed effect. Indeed, we demonstrate that EIF5A regulates MYC elongation without affecting its mRNA content or protein stability, by alleviating ribosome stalling at five distinct pausing motifs in MYC CDS. Of note, we show that blockade of the hypusination axis elicits a remarkable growth inhibitory effect in preclinical models of CRC and significantly reduces the size of polyps in APCMin/+ mice, a model of human familial adenomatous polyposis (FAP). Together, these data illustrate an unprecedented mechanism, whereby the tumor-promoting properties of hypusinated EIF5A are linked to its ability to regulate MYC elongation and provide a rationale for the use of DHPS/EIF5A inhibitors in CRC therapy.
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Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Lisina/análogos & derivados , Factores de Iniciación de Péptidos/metabolismo , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteínas de Unión al ARN/metabolismo , Poliposis Adenomatosa del Colon/genética , Poliposis Adenomatosa del Colon/patología , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Animales , Línea Celular Tumoral , Proliferación Celular/genética , Neoplasias Colorrectales/genética , Regulación hacia Abajo , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Lisina/metabolismo , Ratones Desnudos , Sistemas de Lectura Abierta/genética , Oxidorreductasas actuantes sobre Donantes de Grupo CH-NH/antagonistas & inhibidores , Oxidorreductasas actuantes sobre Donantes de Grupo CH-NH/metabolismo , Factores de Iniciación de Péptidos/química , Péptidos/metabolismo , Poliaminas/metabolismo , Biosíntesis de Proteínas , Proteínas de Unión al ARN/química , Factor 5A Eucariótico de Iniciación de TraducciónRESUMEN
Unfolded protein response (UPR) is a conserved adaptive response that tries to restore protein homeostasis after endoplasmic reticulum (ER) stress. Recent studies highlighted the role of UPR in acute leukemias and UPR targeting has been suggested as a therapeutic approach. Aberrant Notch signaling is a common feature of T-cell acute lymphoblastic leukemia (T-ALL), as downregulation of Notch activity negatively affects T-ALL cell survival, leading to the employment of Notch inhibitors in T-ALL therapy. Here we demonstrate that Notch3 is able to sustain UPR in T-ALL cells, as Notch3 silencing favored a Bip-dependent IRE1α inactivation under ER stress conditions, leading to increased apoptosis via upregulation of the ER stress cell death mediator CHOP. By using Juglone, a naturally occurring naphthoquinone acting as an anticancer agent, to decrease Notch3 expression and induce ER stress, we observed an increased ER stress-associated apoptosis. Altogether our results suggest that Notch3 inhibition may prevent leukemia cells from engaging a functional UPR needed to compensate the Juglone-mediated ER proteotoxic stress. Notably, in vivo administration of Juglone to human T-ALL xenotransplant models significantly reduced tumor growth, finally fostering the exploitation of Juglone-dependent Notch3 inhibition to perturb the ER stress/UPR signaling in Notch3-dependent T-ALL subsets.
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The Wnt/ß-catenin pathway is often found deregulated in cancer. The aberrant accumulation of ß-catenin in the cell nucleus results in the development of various malignancies. Specific drugs against this signaling pathway for clinical treatments have not been approved yet. Herein we report inhibitors of ß-catenin signaling of potential therapeutic value as anticancer agents. Ethyl 4-((4-(trifluoromethyl)phenyl)sulfonamido)benzoate (compound 14) inhibits the effect on Wnt reporter with an IC50 value of 7.0â µM, significantly reduces c-MYC levels, inhibits HCT116 colon cancer cell growth (IC50 20.2â µM), does not violate Lipinski and Veber rules, and shows predicted Caco-2 and MDCK cell permeability Papp >500â nm s-1 . Compound 14 seems to have potential for the development of new anticancer therapies.
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Antineoplásicos/farmacología , Proteínas Proto-Oncogénicas c-myc/antagonistas & inhibidores , Sulfonamidas/farmacología , beta Catenina/antagonistas & inhibidores , Antineoplásicos/síntesis química , Antineoplásicos/química , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Estructura Molecular , Proteínas Proto-Oncogénicas c-myc/metabolismo , Sulfonamidas/síntesis química , Sulfonamidas/química , Vía de Señalización Wnt/efectos de los fármacos , beta Catenina/metabolismoRESUMEN
The antidiabetic drug phenformin displays potent anticancer activity in different tumors, but its mechanism of action remains elusive. Using Shh medulloblastoma as model, we show here that at clinically relevant concentrations, phenformin elicits a significant therapeutic effect through a redox-dependent but complex I-independent mechanism. Phenformin inhibits mitochondrial glycerophosphate dehydrogenase (mGPD), a component of the glycerophosphate shuttle, and causes elevations of intracellular NADH content. Inhibition of mGPD mimics phenformin action and promotes an association between corepressor CtBP2 and Gli1, thereby inhibiting Hh transcriptional output and tumor growth. Because ablation of CtBP2 abrogates the therapeutic effect of phenformin in mice, these data illustrate a biguanide-mediated redox/corepressor interplay, which may represent a relevant target for tumor therapy.
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Antineoplásicos/uso terapéutico , Proteínas Co-Represoras/efectos de los fármacos , Proteínas Hedgehog/efectos de los fármacos , Hipoglucemiantes/uso terapéutico , Neoplasias/tratamiento farmacológico , Fenformina/uso terapéutico , Animales , Antineoplásicos/farmacología , Humanos , Hipoglucemiantes/farmacología , Ratones , Fenformina/farmacologíaRESUMEN
Medulloblastoma (MB) is the most common malignant childhood brain tumor. About 30% of all MBs belong to the I molecular subgroup, characterized by constitutive activation of the Sonic Hedgehog (Hh) pathway. The Hh pathway is involved in several fundamental processes during embryogenesis and in adult life and its deregulation may lead to cerebellar tumorigenesis. Indeed, Hh activity must be maintained via a complex network of activating and repressor signals. One of these repressor signals is KCASH2, belonging to the KCASH family of protein, which acts as negative regulators of the Hedgehog signaling pathway during cerebellar development and differentiation. KCASH2 leads HDAC1 to degradation, allowing hyperacetylation and inhibition of transcriptional activity of Gli1, the main effector of the Hh pathway. In turn, the KCASH2 loss leads to persistent Hh activity and eventually tumorigenesis. In order to better characterize the physiologic role and modulation mechanisms of KCASH2, we have searched through a proteomic approach for new KCASH2 interactors, identifying Potassium Channel Tetramerization Domain Containing 15 (KCTD15). KCTD15 is able to directly interact with KCASH2, through its BTB/POZ domain. This interaction leads to increase KCASH2 stability which implies a reduction of the Hh pathway activity and a reduction of Hh-dependent MB cells proliferation. Here we report the identification of KCTD15 as a novel player in the complex network of regulatory proteins, which modulate Hh pathway, this could be a promising new target for therapeutic approach against MB.
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The Hedgehog (Hh) pathway is essential for embryonic development and tissue homeostasis. Aberrant Hh signaling may occur in a wide range of human cancers, such as medulloblastoma, the most common brain malignancy in childhood. Here, we identify endoplasmic reticulum aminopeptidase 1 (ERAP1), a key regulator of innate and adaptive antitumor immune responses, as a previously unknown player in the Hh signaling pathway. We demonstrate that ERAP1 binds the deubiquitylase enzyme USP47, displaces the USP47-associated ßTrCP, the substrate-receptor subunit of the SCFßTrCP ubiquitin ligase, and promotes ßTrCP degradation. These events result in the modulation of Gli transcription factors, the final effectors of the Hh pathway, and the enhancement of Hh activity. Remarkably, genetic or pharmacological inhibition of ERAP1 suppresses Hh-dependent tumor growth in vitro and in vivo. Our findings unveil an unexpected role for ERAP1 in cancer and indicate ERAP1 as a promising therapeutic target for Hh-driven tumors.
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Aminopeptidasas/fisiología , Antígenos de Histocompatibilidad Menor/fisiología , Proteasas Ubiquitina-Específicas/metabolismo , Proteínas con Repetición de beta-Transducina/metabolismo , Aminopeptidasas/genética , Aminopeptidasas/metabolismo , Animales , Carcinogénesis/genética , Proteínas Hedgehog/metabolismo , Ratones , Antígenos de Histocompatibilidad Menor/genética , Antígenos de Histocompatibilidad Menor/metabolismo , Células 3T3 NIH , Estabilidad Proteica , Proteolisis , Transducción de SeñalRESUMEN
Targeted approaches aiming at modulating NHERF1 activity, rather than its overall expression, would be preferred to preserve the normal functions of this versatile protein. We focused our attention on the NHERF1/PDZ1 domain that governs its membrane recruitment/displacement through a transient phosphorylation switch. We herein report the design and synthesis of novel NHERF1 PDZ1 domain inhibitors. These compounds have potential therapeutic value when used in combination with antagonists of ß-catenin to augment apoptotic death of colorectal cancer cells refractory to currently available Wnt/ß-catenin-targeted agents.
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Hedgehog (Hh) signaling is a critical developmental regulator and its aberrant activation,due to somatic or germline mutations of genes encoding pathway components, causes Basal CellCarcinoma (BCC) and medulloblastoma (MB). A growing effort has been devoted at theidentification of druggable vulnerabilities of the Hedgehog signaling, leading to the identificationof various compounds with variable efficacy and/or safety. Emerging evidence shows that anaberrant polyamine metabolism is a hallmark of Hh-dependent tumors and that itspharmacological inhibition elicits relevant therapeutic effects in clinical or preclinical models ofBCC and MB. We discuss here the current knowledge of polyamine metabolism, its role in cancerand the available targeting strategies. We review the literature about the connection betweenpolyamines and the Hedgehog signaling, and the potential therapeutic benefit of targetingpolyamine metabolism in two malignancies where Hh pathways play a well-established role: BCCand MB.
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Carcinoma Basocelular/metabolismo , Carcinoma Basocelular/terapia , Proteínas Hedgehog/metabolismo , Meduloblastoma/metabolismo , Meduloblastoma/terapia , Terapia Molecular Dirigida , Poliaminas/metabolismo , Animales , Carcinogénesis/metabolismo , HumanosRESUMEN
The aberrant activation of hedgehog (HH) signaling is a leading cause of the development of medulloblastoma, a pediatric tumor of the cerebellum. The FDAapproved HH inhibitor, Vismodegib, which targets the transmembrane transducer SMO, has shown limited efficacy in patients with medulloblastoma, due to compensatory mechanisms that maintain an active HHGLI signaling status. Thus, the identification of novel actionable mechanisms, directly affecting the activity of the HHregulated GLI transcription factors is an important goal for these malignancies. In this study, using gene expression and reporter assays, combined with biochemical and cellular analyses, we demonstrate that mitogenactivated kinase kinase kinase 1 (MEKK1), the most upstream kinase of the mitogenactivated protein kinase (MAPK) phosphorylation modules, suppresses HH signaling by associating and phosphorylating GLI1, the most potent HHregulated transcription factor. Phosphorylation occurred at multiple residues in the Cterminal region of GLI1 and was followed by an increased association with the cytoplasmic proteins 1433. Of note, the enforced expression of MEKK1 or the exposure of medulloblastoma cells to the MEKK1 activator, Nocodazole, resulted in a marked inhibitory effect on GLI1 activity and tumor cell proliferation and viability. Taken together, the results of this study shed light on a novel regulatory mechanism of HH signaling, with potentially relevant implications in cancer therapy.
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Proteínas Hedgehog/genética , Quinasa 1 de Quinasa de Quinasa MAP/genética , Meduloblastoma/genética , Proteína con Dedos de Zinc GLI1/genética , Anilidas/administración & dosificación , Animales , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Meduloblastoma/tratamiento farmacológico , Meduloblastoma/patología , Ratones , Células 3T3 NIH , Fosforilación/efectos de los fármacos , Piridinas/administración & dosificación , ARN Mensajero/genética , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/genéticaRESUMEN
SHH Medulloblastoma (SHH-MB) is a pediatric brain tumor characterized by an inappropriate activation of the developmental Hedgehog (Hh) signaling. SHH-MB patients treated with the FDA-approved vismodegib, an Hh inhibitor that targets the transmembrane activator Smoothened (Smo), have shown the rapid development of drug resistance and tumor relapse due to novel Smo mutations. Moreover, a subset of patients did not respond to vismodegib because mutations were localized downstream of Smo. Thus, targeting downstream Hh components is now considered a preferable approach. We show here that selective inhibition of the downstream Hh effectors HDAC1 and HDAC2 robustly counteracts SHH-MB growth in mouse models. These two deacetylases are upregulated in tumor and their knockdown inhibits Hh signaling and decreases tumor growth. We demonstrate that mocetinostat (MGCD0103), a selective HDAC1/HDAC2 inhibitor, is a potent Hh inhibitor and that its effect is linked to Gli1 acetylation at K518. Of note, we demonstrate that administration of mocetinostat to mouse models of SHH-MB drastically reduces tumor growth, by reducing proliferation and increasing apoptosis of tumor cells and prolongs mouse survival rate. Collectively, these data demonstrate the preclinical efficacy of targeting the downstream HDAC1/2-Gli1 acetylation in the treatment of SHH-MB.