RESUMO
Extracellular matrix (ECM) is a major component of the tumor environment, promoting the establishment of a pro-invasive behavior. Such environment is supported by both tumor- and stromal-derived metabolites, particularly lactate. In prostate cancer (PCa), cancer-associated fibroblasts (CAFs) are major contributors of secreted lactate, able to impact on metabolic and transcriptional regulation in cancer cells. Here, we describe a mechanism by which CAF-secreted lactate promotes in PCa cells the expression of genes coding for the collagen family. Lactate-exploiting PCa cells rely on increased α-ketoglutarate (α-KG) which activates the α-KG-dependent collagen prolyl-4-hydroxylase (P4HA1) to support collagen hydroxylation. De novo synthetized collagen plays a signaling role by activating discoidin domain receptor 1 (DDR1), supporting stem-like and invasive features of PCa cells. Inhibition of lactate-induced collagen hydroxylation and DDR1 activation reduces the metastatic colonization of PCa cells. Overall, these results provide a new understanding of the link between collagen remodeling/signaling and the nutrient environment exploited by PCa.
Assuntos
Colágeno , Matriz Extracelular , Ácido Láctico , Metástase Neoplásica , Neoplasias da Próstata , Masculino , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Neoplasias da Próstata/genética , Humanos , Matriz Extracelular/metabolismo , Colágeno/metabolismo , Ácido Láctico/metabolismo , Linhagem Celular Tumoral , Fibroblastos Associados a Câncer/metabolismo , Fibroblastos Associados a Câncer/patologia , Transdução de Sinais , Regulação Neoplásica da Expressão Gênica , Animais , Camundongos , Microambiente Tumoral , Ácidos Cetoglutáricos/metabolismo , HidroxilaçãoRESUMO
Transcription of E-cadherin, a tumor suppressor that plays critical roles in cell adhesion and the epithelial-mesenchymal transition, is regulated by a promoter-associated non-coding RNA (paRNA). The sense-oriented paRNA (S-paRNA) includes a functional C/A single nucleotide polymorphism (SNP rs16260). The A-allele leads to decreased transcriptional activity and increased prostate cancer risk. The polymorphic site is known to affect binding of a microRNA-guided Argonaute 1 (AGO1) complex and recruitment of chromatin-modifying enzymes to silence the promoter. Yet the SNP is distant from the microRNA-AGO1 binding domain in both primary sequence and secondary structure, raising the question of how regulation occurs. Here we report the 3D NMR structure of the 104-nucleotide domain of the S-paRNA that encompasses the SNP and the microRNA-binding site. We show that the A to C change alters the locally dynamic and metastable structure of the S-paRNA, revealing how the single nucleotide mutation regulates the E-cadherin promoter through its effect on the non-coding RNA structure.
Assuntos
MicroRNAs , Polimorfismo de Nucleotídeo Único , Masculino , Humanos , Caderinas/genética , Caderinas/metabolismo , RNA não Traduzido/genética , MicroRNAs/genética , Nucleotídeos , Regulação Neoplásica da Expressão Gênica , Linhagem Celular TumoralRESUMO
MOTIVATION: Long non-coding RNAs (lncRNAs) have gained increasing relevance in epigenetic regulation and nuclear functional organization. High-throughput sequencing approaches have revealed frequent non-coding transcription in promoter-proximal regions. However, a comprehensive catalogue of promoter-associated RNAs (paRNAs) and an analysis of the possible interactions with neighboring genes and genomic regulatory elements are missing. RESULTS: Integrating data from multiple cell types and experimental platforms we identified thousands of paRNAs in the human genome. paRNAs are transcribed in both sense and antisense orientation, are mostly non-polyadenylated and retained in the cell nucleus. Transcriptional regulators, epigenetic effectors and activating chromatin marks are enriched in paRNA-positive promoters. Furthermore, paRNA-positive promoters exhibit chromatin signatures of both active promoters and enhancers. Promoters with paRNAs reside preferentially at chromatin loop boundaries, suggesting an involvement in anchor site recognition and chromatin looping. Importantly, these features are independent of the transcriptional state of neighboring genes. Thus, paRNAs may act as cis-regulatory modules with an impact on local recruitment of transcription factors, epigenetic state and chromatin loop organization. This study provides a comprehensive analysis of the promoter-proximal transcriptome and offers novel insights into the roles of paRNAs in epigenetic processes and human diseases. AVAILABILITY AND IMPLEMENTATION: Genomic coordinates of predicted paRNAs are available at https://figshare.com: https://doi.org/10.6084/m9.figshare.7392791.v1 and https://doi.org/10.6084/m9.figshare.4856630.v2. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Assuntos
RNA Longo não Codificante/genética , Transcriptoma , Cromatina , Epigênese Genética , Humanos , Regiões Promotoras GenéticasRESUMO
In addition to its canonical role in nuclear transcription, signal transducer and activator of transcription 3 (STAT3) is emerging as an important regulator of mitochondrial function. Here, we demonstrate that a novel inhibitor that binds with high affinity to the STAT3 SH2 domain triggers a complex cascade of events initiated by interference with mitochondrial STAT3 (mSTAT3). The mSTAT3-drug interaction leads to mitochondrial dysfunction, accumulation of proteotoxic STAT3 aggregates, and cell death. The cytotoxic effects depend directly on the drug's ability to interfere with mSTAT3 and mitochondrial function, as demonstrated by site-directed mutagenesis and use of STAT3 knockout and mitochondria-depleted cells. Importantly, the lethal consequences of mSTAT3 inhibition are enhanced by glucose starvation and by increased reliance of cancer cells and tumor-initiating cells on mitochondria, resulting in potent activity in cell cultures and tumor xenografts in mice. These findings can be exploited for eliciting synthetic lethality in metabolically stressed cancer cells using high-affinity STAT3 inhibitors. Thus, this study provides insights on the role of mSTAT3 in cancer cells and a conceptual framework for developing more effective cancer therapies.
Assuntos
Mitocôndrias/genética , Neoplasias/genética , Fator de Transcrição STAT3/genética , Mutações Sintéticas Letais/genética , Domínios de Homologia de src/genética , Animais , Morte Celular/genética , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos NusRESUMO
Transcription factors are proteins able to bind DNA and induce the transcription of specific genes. Consequently, they play a pivotal role in multiple cellular pathways and are frequently over-expressed or dysregulated in cancer. Here, we will focus on a specific "signal transducer and activator of transcription" (STAT3) factor that is involved in several pathologies, including cancer. For long time, the mechanism by which STAT3 exerts its cellular functions has been summarized by a three steps process: (1) Protein phosphorylation by specific kinases, (2) dimerization promoted by phosphorylation, (3) activation of gene expression by the phosphorylated dimer. Consequently, most of the inhibitors reported in literature aimed at blocking phosphorylation and dimerization. However, recent observations reopened the debate and the entire functional mechanism has been revisited stimulating the scientific community to pursue new inhibition strategies. In particular, the dimerization of the unphosphorylated species has been experimentally demonstrated and specific roles proposed also for these dimers. Despite difficulties in the expression and purification of the full length STAT3, structural biology investigations allowed the determination of atomistic structures of STAT3 dimers and several protein domains. Starting from this information, computational methods have been used both to improve the understanding of the STAT3 functional mechanism and to design new inhibitors to be used as anticancer drugs. In this review, we will focus on the contribution of structural biology to understand the roles of STAT3, to design new inhibitors and to suggest new strategies of pharmacological intervention.
Assuntos
Antineoplásicos/síntese química , DNA de Neoplasias/química , Regulação Neoplásica da Expressão Gênica , Neoplasias/tratamento farmacológico , Fator de Transcrição STAT3/antagonistas & inibidores , Antineoplásicos/farmacologia , Sítios de Ligação , DNA de Neoplasias/metabolismo , Desenho de Fármacos , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Fosforilação , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Multimerização Proteica , Fator de Transcrição STAT3/química , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Transdução de SinaisRESUMO
Cis-natural antisense transcripts (cis-NATs) are long noncoding RNAs transcribed from the opposite strand and overlapping coding and noncoding genes on the sense strand. cis-NATs are widely present in the human genome and can be involved in multiple mechanisms of gene regulation. Here, we describe the presence of cis-NATs in the 3' distal region of the c-MYC locus and investigate their impact on transcriptional regulation of this key oncogene in human cancers. We found that cis-NATs are produced as consequence of the activation of cryptic transcription initiation sites in the 3' distal region downstream of the c-MYC 3'UTR. The process is tightly regulated and leads to the formation of two main transcripts, NAT6531 and NAT6558, which differ in their ability to fold into stem-loop secondary structures. NAT6531 acts as a substrate for DICER and as a source of small RNAs capable of modulating c-MYC transcription. This complex system, based on the interplay between cis-NATs and NAT-derived small RNAs, may represent an important layer of epigenetic regulation of the expression of c-MYC and other genes in human cells.
Assuntos
Regulação da Expressão Gênica , Genes myc , RNA Antissenso/genética , Transcrição Gênica , Regiões 3' não Traduzidas , Acetilação , Linhagem Celular Tumoral , Clonagem Molecular , Epigênese Genética , Loci Gênicos , Histonas/metabolismo , Humanos , Modelos Biológicos , Conformação de Ácido Nucleico , RNA não Traduzido/genética , Ribonuclease III/metabolismo , Sítio de Iniciação de TranscriçãoRESUMO
MicroRNAs (miRNAs) originate from stem-loop-containing precursors (pre-miRNAs, pri-miRNAs) and mature by means of the Drosha and Dicer endonucleases and their associated factors. The let-7 miRNAs have prominent roles in developmental differentiation and in regulating cell proliferation. In cancer, the tumor suppressor function of let-7 is abrogated by overexpression of Lin28, one of several RNA-binding proteins that regulate let-7 biogenesis by interacting with conserved motifs in let-7 precursors close to the Dicer cleavage site. Using in vitro assays, we have identified a binding site for short modified oligoribonucleotides ('looptomirs') overlapping that of Lin28 in pre-let-7a-2. These looptomirs selectively antagonize the docking of Lin28, but still permit processing of pre-let-7a-2 by Dicer. Looptomirs restored synthesis of mature let-7 and inhibited growth and clonogenic potential in Lin28 overexpressing hepatocarcinoma cells, thereby demonstrating a promising new means to rescue defective miRNA biogenesis in Lin28-dependent cancers.
Assuntos
MicroRNAs/metabolismo , Neoplasias/genética , Oligorribonucleotídeos/metabolismo , Processamento Pós-Transcricional do RNA , Proteínas de Ligação a RNA/antagonistas & inibidores , Sítios de Ligação , Linhagem Celular Tumoral , Proliferação de Células , Células HEK293 , Humanos , MicroRNAs/química , Neoplasias/enzimologia , Neoplasias/patologia , Oligorribonucleotídeos/química , Precursores de RNA/química , Precursores de RNA/metabolismo , Proteínas de Ligação a RNA/metabolismo , Ribonuclease III/metabolismoRESUMO
Mutations of the K-Ras gene occur in over 90 % of pancreatic carcinomas, and to date, no targeted therapies exist for this genetically defined subset of cancers. STAT3 plays a critical role in KRAS-driven pancreatic tumorigenesis, suggesting its potential as a therapeutic target in this cancer. Therefore, finding novel and potential drugs to inhibit oncogenic K-Ras is a major challenge in cancer therapy. In an attempt to develop novel anti-KRAS mutant chemotherapeutics, we isolated three novel triterpenoids from Amoora rohituka stem and their chemical structures were characterized by extensive 1H-NMR, 13C-NMR, Mass, IR spectroscopic studies and chemical transformations. Aphanin (3 alpha-angeloyloxyolean-12-en-28-oic acid) is one of the isolated novel triterpenoid compounds. We found aphanin exhibited antiproliferative effects, caused G0-G1 cell cycle arrest, inhibits K-Ras G12D mutant activity by decreased STAT3, p-STAT3, Akt, p-Akt, cyclin D1 and c-Myc expressions, and induced apoptosis in pancreatic cancer HPAF-II (ΔKRAS G12D ) cells. The apoptosis proceeded through depletion of GSH with a concomitant increase in the reactive oxygen species production. The results of our study have important implications for the development of aphanin as potential novel agent for the treatment of K-Ras mutant pancreatic cancer, and STAT3-cMyc-cyclinD1 axis may serve as an important predictive biomarker for the therapeutic efficacy.
Assuntos
Meliaceae/química , Mutação , Neoplasias Pancreáticas/tratamento farmacológico , Proteínas Proto-Oncogênicas p21(ras)/genética , Fator de Transcrição STAT3/fisiologia , Triterpenos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Glutationa/metabolismo , Humanos , Neoplasias Pancreáticas/genética , Fosforilação , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-myc/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Neoplasias PancreáticasRESUMO
Signal transducer and activator of transcription factors (STATs) are proteins that can translocate into the nucleus, bind DNA, and activate gene transcription. STAT proteins play a crucial role in cell proliferation, apoptosis, and differentiation. The prevalent view is that STAT proteins are able to form dimers and bind DNA only upon phosphorylation of specific tyrosine residues in the transactivation domain. However, this paradigm has been questioned recently by the observation of dimers of unphosphorylated STATs (USTATs) by X-ray, Förster resonance energy transfer, and site-directed mutagenesis. A more complex picture of the dimerization process and of the role of the dimers is, thus, emerging. Here we present an integrated modeling study of STAT3, a member of the STAT family of utmost importance in cancer development and therapy, in which we combine available experimental data with several computational methodologies such as homology modeling, protein-protein docking, and molecular dynamics to build reliable atomistic models of USTAT3 dimers. The models generated with the integrative approach presented here were then validated by performing computational alanine scanning for all the residues in the protein-protein interface. These results confirmed the experimental observation of the importance of some of these residues (in particular Leu78 and Asp19) in the USTAT3 dimerization process. Given the growing importance of USTAT3 dimers in several cellular pathways, our models provide an important tool for studying the effects of pathological mutations at the molecular and/or atomistic level, and in the rational design of new inhibitors of dimerization.
Assuntos
Modelos Moleculares , Multimerização Proteica , Fator de Transcrição STAT3/química , Fator de Transcrição STAT3/genética , Sequência de Aminoácidos , Animais , Camundongos , Dados de Sequência Molecular , Fosforilação/fisiologia , Multimerização Proteica/fisiologia , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Fator de Transcrição STAT3/metabolismoRESUMO
Castration-resistant prostate cancer (CRPC) is a frequently occurring disease with adverse clinical outcomes and limited therapeutic options. Here, we identify methionine adenosyltransferase 2a (MAT2A) as a critical driver of the androgen-indifferent state in ERG fusion-positive CRPC. MAT2A is upregulated in CRPC and cooperates with ERG in promoting cell plasticity, stemness and tumorigenesis. RNA, ATAC and ChIP-sequencing coupled with histone post-translational modification analysis by mass spectrometry show that MAT2A broadly impacts the transcriptional and epigenetic landscape. MAT2A enhances H3K4me2 at multiple genomic sites, promoting the expression of pro-tumorigenic non-canonical AR target genes. Genetic and pharmacological inhibition of MAT2A reverses the transcriptional and epigenetic remodeling in CRPC models and improves the response to AR and EZH2 inhibitors. These data reveal a role of MAT2A in epigenetic reprogramming and provide a proof of concept for testing MAT2A inhibitors in CRPC patients to improve clinical responses and prevent treatment resistance.
Assuntos
Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Metionina Adenosiltransferase , Neoplasias de Próstata Resistentes à Castração , Regulador Transcricional ERG , Masculino , Humanos , Regulador Transcricional ERG/genética , Regulador Transcricional ERG/metabolismo , Metionina Adenosiltransferase/genética , Metionina Adenosiltransferase/metabolismo , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Epigênese Genética/efeitos dos fármacos , Animais , Androgênios/metabolismo , Epigenoma , Camundongos , Histonas/metabolismo , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidoresRESUMO
Small interfering RNAs (siRNAs) directed to gene promoters can silence genes at the transcriptional level. siRNA-directed transcriptional silencing (RdTS) was first described in plants and yeasts and more recently in mammalian cells. RdTS has been associated with the induction of epigenetic changes and the formation of complexes containing RNA interference and chromatin-remodelling factors. Here, we show that a promoter-targeted siRNA inhibits transcription of the c-myc gene. Transcriptional silencing of c-myc did not involve changes of known epigenetic marks. Instead, the c-myc promoter-targeted siRNA interfered with transcription initiation blocking the assembly of the pre-initiation complex. Transcriptional interference depended on Argonaute 2 and a noncoding promoter-associated RNA initiated upstream and overlapping the transcription start site. Silencing of c-myc led to growth arrest, reduced clonogenic potential and senescence of c-myc over-expressing prostate cancer cells with minimal effect on normal cells. RNA-directed transcriptional interference may be a natural mechanism of transcriptional control and siRNAs targeting noncoding RNAs participating in this regulatory pathway could be valuable tools to control expression of deregulated genes in human diseases.
Assuntos
Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas c-myc/genética , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transcrição Gênica , Proteínas Argonautas , Linhagem Celular Tumoral , Proliferação de Células , Senescência Celular , Fator de Iniciação 2 em Eucariotos/metabolismo , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/metabolismo , RNA não Traduzido/genética , Ensaio Tumoral de Célula-TroncoRESUMO
Prostate cancer (PCa) is the most common malignancy and the second leading cause of cancer-related death amongst men in the United States. Neuroendocrine prostate cancer (NEPC) can either arise de novo or emerge as a consequence of therapy. De novo NEPC is rare, with an incidence of <2% of all PCa cases. In contrast, treatment-induced NEPC is frequent with >20% of patients with metastatic castration-resistant prostate cancer (CRPC) reported to progress to neuroendocrine (NE) differentiation. The emergence of treatment-induced NEPC is linked to the increased therapeutic pressure, due to the broad application of androgen deprivation therapy (ADT) for PCa management and the development of novel more potent androgen receptor (AR) pathway inhibitors. NEPC is a high-grade tumor type characterized by aggressive phenotype and clinical behavior. Patients affected by NEPC frequently develop visceral metastases and have a poor prognosis. The molecular mechanisms underlying the development and progression of NEPC are still poorly understood. Transcriptional and epigenetic reprogramming appears to be involved in NE progression. In this review, we aim to provide a comprehensive view of the available models for NEPC detailing their strengths and limitations. Moreover, we describe novel approaches to expand the repertoire of preclinical models to better study, prevent, or reverse NEPC. The integration of multiple preclinical models along with molecular and omics approaches will provide important insights to understand disease progression and to devise novel therapeutic strategies for the management of NEPC in the near future. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Generation of organoids starting from the prostate gland of a GEMM or a human PDX Basic Protocol 2: Ex vivo tumor sphere formation.
Assuntos
Neoplasias da Próstata , Masculino , Humanos , Neoplasias da Próstata/genética , Neoplasias da Próstata/terapia , Neoplasias da Próstata/metabolismo , Antagonistas de Androgênios/uso terapêutico , Próstata/metabolismo , Próstata/patologia , Antagonistas de Receptores de Andrógenos/uso terapêuticoRESUMO
Elucidating the mechanisms by which immune cells become dysfunctional in tumors is critical to developing next-generation immunotherapies. We profiled proteomes of cancer tissue as well as monocyte/macrophages, CD4+ and CD8+ T cells, and NK cells isolated from tumors, liver, and blood of 48 patients with hepatocellular carcinoma. We found that tumor macrophages induce the sphingosine-1-phospate-degrading enzyme SGPL1, which dampened their inflammatory phenotype and anti-tumor function in vivo. We further discovered that the signaling scaffold protein AFAP1L2, typically only found in activated NK cells, is also upregulated in chronically stimulated CD8+ T cells in tumors. Ablation of AFAP1L2 in CD8+ T cells increased their viability upon repeated stimulation and enhanced their anti-tumor activity synergistically with PD-L1 blockade in mouse models. Our data reveal new targets for immunotherapy and provide a resource on immune cell proteomes in liver cancer.
RESUMO
A series of novel σ1 receptor ligands with a 4-(2-aminoethyl)piperidine scaffold was prepared and biologically evaluated. The underlying concept of our project was the improvement of the lipophilic ligand efficiency of previously synthesized potent σ1 ligands. The key steps of the synthesis comprise the conjugate addition of phenylboronic acid at dihydropyridin-4(1H)-ones 7, homologation of the ketones 8 and introduction of diverse amino moieties and piperidine N-substituents. 1-Methylpiperidines showed particular high σ1 receptor affinity and selectivity over the σ2 subtype, whilst piperidines with a proton, a tosyl moiety or an ethyl moiety exhibited considerably lower σ1 affinity. Molecular dynamics simulations with per-residue binding free energy deconvolution demonstrated that different interactions of the basic piperidine-N-atom and its substituents (or the cyclohexane ring) with the lipophilic binding pocket consisting of Leu105, Thr181, Leu182, Ala185, Leu186, Thr202 and Tyr206 are responsible for the different σ1 receptor affinities. Recorded logD7.4 and calculated clogP values of 4a and 18a indicate low lipophilicity and thus high lipophilic ligand efficiency. Piperidine 4a inhibited the growth of human non-small cell lung cancer cells A427 to a similar extent as the σ1 antagonist haloperidol. 1-Methylpiperidines 20a, 21a and 22a showed stronger antiproliferative effects on androgen negative human prostate cancer cells DU145 than the σ1 ligands NE100 and S1RA.
Assuntos
Antineoplásicos , Piperidinas , Receptores sigma , Antineoplásicos/química , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas , Linhagem Celular Tumoral , Humanos , Ligantes , Neoplasias Pulmonares , Masculino , Piperidinas/química , Piperidinas/farmacologia , Neoplasias da Próstata , Receptores sigma/metabolismo , Relação Estrutura-AtividadeRESUMO
Lin28 RNA-binding proteins are stem-cell factors that play key roles in development. Lin28 suppresses the biogenesis of let-7 microRNAs and regulates mRNA translation. Notably, let-7 inhibits Lin28, establishing a double-negative feedback loop. The Lin28/let-7 axis resides at the interface of metabolic reprogramming and oncogenesis and is therefore a potential target for several diseases. In this study, we use compound-C1632, a drug-like Lin28 inhibitor, and show that the Lin28/let-7 axis regulates the balance between ketogenesis and lipogenesis in liver cells. Hence, Lin28 inhibition activates synthesis and secretion of ketone bodies whilst suppressing lipogenesis. This occurs at least partly via let-7-mediated inhibition of nuclear receptor co-repressor 1, which releases ketogenesis gene expression mediated by peroxisome proliferator-activated receptor-alpha. In this way, small-molecule Lin28 inhibition protects against lipid accumulation in multiple cellular and male mouse models of hepatic steatosis. Overall, this study highlights Lin28 inhibitors as candidates for the treatment of hepatic disorders of abnormal lipid deposition.
Assuntos
MicroRNAs , Hepatopatia Gordurosa não Alcoólica , Animais , Camundongos , Masculino , MicroRNAs/genética , MicroRNAs/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/genética , Homeostase , LipídeosRESUMO
Lactate is an abundant oncometabolite in the tumor environment. In prostate cancer, cancer-associated fibroblasts (CAF) are major contributors of secreted lactate, which can be taken up by cancer cells to sustain mitochondrial metabolism. However, how lactate impacts transcriptional regulation in tumors has yet to be fully elucidated. Here, we describe a mechanism by which CAF-secreted lactate is able to increase the expression of genes involved in lipid metabolism in prostate cancer cells. This regulation enhanced intracellular lipid accumulation in lipid droplets (LD) and provided acetyl moieties for histone acetylation, establishing a regulatory loop between metabolites and epigenetic modification. Inhibition of this loop by targeting the bromodomain and extraterminal protein family of histone acetylation readers suppressed the expression of perilipin 2 (PLIN2), a crucial component of LDs, disrupting lactate-dependent lipid metabolic rewiring. Inhibition of this CAF-induced metabolic-epigenetic regulatory loop in vivo reduced growth and metastasis of prostate cancer cells, demonstrating its translational relevance as a therapeutic target in prostate cancer. Clinically, PLIN2 expression was elevated in tumors with a higher Gleason grade and in castration-resistant prostate cancer compared with primary prostate cancer. Overall, these findings show that lactate has both a metabolic and an epigenetic role in promoting prostate cancer progression. SIGNIFICANCE: This work shows that stromal-derived lactate induces accumulation of lipid droplets, stimulates epigenetic rewiring, and fosters metastatic potential in prostate cancer.
Assuntos
Metabolismo dos Lipídeos , Neoplasias da Próstata , Epigênese Genética , Humanos , Ácido Láctico/metabolismo , Metabolismo dos Lipídeos/genética , Masculino , Próstata/patologia , Neoplasias da Próstata/patologiaRESUMO
State-of-the-art small interfering RNA (siRNA) therapeutics such as givosiran and fitusiran are constructed from three variable components: a fully-modified RNA core that conveys metabolic stability, a targeting moiety that mediates target-cell uptake, and a linker. This structural complexity poses challenges for metabolite characterization and risk assessment after long-term patient exposure. In this study, we show that basic phosphorothioate modification of a siRNA targeting the oncoprotein Lin28B provides a useful increase in metabolic stability, without greatly compromising potency. We found that its stability in vitro matched that of nanoparticle-free patisiran in serum and surpassed it in liver tritosome extracts, although it did not reach the stability of the fitusiran siRNA core structure. Liver and kidney were the main sites of accumulation after its subcutaneous administration in mice. Despite the lack of a delivery agent-free antitumor effect, we anticipate our study to be a starting point to develop alternative siRNA scaffolds that can be degraded into naturally-occurring metabolites and help alleviate the aforementioned challenges. Furthermore, Lin28B is a promising target for cancers, and the development of such simplified siRNA analogs, possibly together with novel targeting units, holds potential.
Assuntos
Nanopartículas , Neoplasias , Animais , Inativação Gênica , Humanos , Camundongos , RNA Interferente Pequeno/genéticaRESUMO
1,3-Dioxanes 1 and cyclohexanes 2 bearing a phenyl ring and an aminoethyl moiety in 1,3-relationship to each other represent highly potent σ1 receptor antagonists. In order to increase the chemical stability of the acetalic 1,3-dioxanes 1 and the polarity of the cyclohexanes 2, tetrahydropyran derivatives 3 equipped with the same substituents were designed, synthesized and pharmacologically evaluated. The key step of the synthesis was a lipase-catalyzed enantioselective acetylation of the alcohol (R)-5 leading finally to enantiomerically pure test compounds 3a-g. With respect to σ1 receptor affinity and selectivity over a broad range of related (σ2, PCP binding site) and further targets, the enantiomeric benzylamines 3a and cyclohexylmethylamines 3b represent the most promising drug candidates of this series. However, the eudismic ratio for σ1 binding is only in the range of 2.5-3.3. Classical molecular dynamics (MD) simulations confirmed the same binding pose for both the tetrahydropyran 3 and cyclohexane derivatives 2 at the σ1 receptor, according to which: i) the protonated amino moiety of (2S,6R)-3a engages the same key polar interactions with Glu172 (ionic) and Phe107 (π-cation), ii) the lipophilic parts of (2S,6R)-3a are hosted in three hydrophobic regions of the σ1 receptor, and iii) the O-atom of the tetrahydropyran derivatives 3 does not show a relevant interaction with the σ1 receptor. Further in silico evidences obtained by the application of free energy perturbation and steered MD techniques fully supported the experimentally observed difference in receptor/ligand affinities. Tetrahydropyrans 3 require a lower dissociative force peak than cyclohexane analogs 2. Enantiomeric benzylamines 3a and cyclohexylmethylamines 3b were able to inhibit the growth of the androgen negative human prostate cancer cell line DU145. The cyclohexylmethylamine (2S,6R)-3b showed the highest σ1 affinity (Ki(σ1) = 0.95 nM) and the highest analgesic activity in vivo (67%).
Assuntos
Analgésicos/síntese química , Antineoplásicos/síntese química , Piranos/química , Receptores sigma/metabolismo , Analgésicos/metabolismo , Analgésicos/uso terapêutico , Animais , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Sítios de Ligação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Humanos , Hiperalgesia/tratamento farmacológico , Ligantes , Camundongos , Simulação de Dinâmica Molecular , Ligação Proteica , Piranos/metabolismo , Receptores sigma/química , Estereoisomerismo , Relação Estrutura-Atividade , Termodinâmica , Receptor Sigma-1RESUMO
BACKGROUND: Men with African ancestry are more likely to develop aggressive prostate cancer (PCa) and to die from this disease. The study of PCa in the South African population represents an opportunity for biomedical research due to the high prevalence of aggressive PCa. While inflammation is known to play a significant role in PCa progression, its association with tumor stage in populations of African descent has not been explored in detail. Identification of new metabolic biomarkers of inflammation may improve diagnosis of patients with aggressive PCa. METHODS: Plasma samples were profiled from 41 South African men with PCa using nuclear magnetic resonance (NMR) spectroscopy. A total of 41 features, including metabolites, lipid classes, total protein, and the inflammatory NMR markers, GlycA, and GlycB, were quantified from each NMR spectrum. The Bruker's B.I.-LISA protocols were used to characterize 114 parameters related to the lipoproteins. The unsupervised KODAMA method was used to stratify the patients of our cohort based on their metabolic profile. RESULTS: We found that the plasma of patients with very high risk, aggressive PCa and high level of C-reactive protein have a peculiar metabolic phenotype (metabotype) characterized by extremely high levels of GlycA and GlycB. The inflammatory processes linked to the higher level of GlycA and GlycB are characterized by a deep change of the plasma metabolome that may be used to improve the stratification of patients with PCa. We also identified a not previously known relationship between high values of VLDL and low level of GlycB in a different metabotype of patients characterized by lower-risk PCa. CONCLUSIONS: For the first time, a portrait of the metabolic changes in African men with PCa has been delineated indicating a strong association between inflammation and metabolic profiles. Our findings indicate how the metabolic profile could be used to identify those patients with high level of inflammation, characterized by aggressive PCa and short life expectancy. Integrating a metabolomic analysis as a tool for patient stratification could be important for opening the door to the development of new therapies. Further investigations are needed to understand the prevalence of an inflammatory metabotype in patients with aggressive PCa.
RESUMO
The multi-kinase inhibitor sorafenib is a primary treatment modality for advanced-stage hepatocellular carcinoma (HCC). However, the therapeutic benefits are short-lived due to innate and acquired resistance. Here, we examined how HCC cells respond to sorafenib and adapt to continuous and prolonged exposure to the drug. Sorafenib-adapted HCC cells show a profound reprogramming of mitochondria function and marked activation of genes required for mitochondrial protein translation and biogenesis. Mitochondrial ribosomal proteins and components of translation and import machinery are increased in sorafenib-resistant cells and sorafenib-refractory HCC patients show similar alterations. Sorafenib-adapted cells also exhibited increased serine 727 phosphorylated (pSer727) STAT3, the prevalent form in mitochondria, suggesting that STAT3 might be an actionable target to counteract resistance. Consistently, a small-molecule STAT3 inhibitor reduces pSer727, reverts mitochondrial alterations, and enhances the response to sorafenib in resistant cells. These results sustain the importance of mitochondria plasticity in response to sorafenib and identify a clinically actionable strategy for improving the treatment efficacy in HCC patients.