RESUMO
The aberrant expression of protein arginine methyltransferase 5 (PRMT5) has been associated with multiple cancers. Using the proteolysis targeting chimera technology, we discovered a first-in-class PRMT5 degrader 15 (MS4322). Here, we report the design, synthesis, and characterization of compound 15 and two structurally similar controls 17 (MS4370) and 21 (MS4369), with impaired binding to the von Hippel-Lindau E3 ligase and PRMT5, respectively. Compound 15, but not 17 and 21, effectively reduced the PRMT5 protein level in MCF-7 cells. Our mechanism studies indicate that compound 15 degraded PRMT5 in an E3 ligase- and proteasome-dependent manner. Compound 15 also effectively reduced the PRMT5 protein level and inhibited growth in multiple cancer cell lines. Moreover, compound 15 was highly selective for PRMT5 in a global proteomic study and exhibited good plasma exposure in mice. Collectively, compound 15 and its two controls 17 and 21 are valuable chemical tools for exploring the PRMT5 functions in health and disease.
Assuntos
Dipeptídeos/farmacologia , Inibidores Enzimáticos/farmacologia , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Dipeptídeos/síntese química , Dipeptídeos/farmacocinética , Desenho de Fármacos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacocinética , Humanos , Ligantes , Masculino , Camundongos , Estrutura Molecular , Proteína-Arginina N-Metiltransferases/metabolismo , Proteólise/efeitos dos fármacos , Relação Estrutura-Atividade , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismoRESUMO
Transcription factor (TF)-based reprogramming of somatic tissues holds great promise for regenerative medicine. Previously, we demonstrated that the TFs GATA2, GFI1B, and FOS convert mouse and human fibroblasts to hemogenic endothelial-like precursors that generate hematopoietic stem progenitor (HSPC)-like cells over time. This conversion is lacking in robustness both in yield and biological function. Herein, we show that inclusion of GFI1 to the reprogramming cocktail significantly expands the HSPC-like population. AFT024 coculture imparts functional potential to these cells and allows quantification of stem cell frequency. Altogether, we demonstrate an improved human hemogenic induction protocol that could provide a valuable human in vitro model of hematopoiesis for disease modeling and a platform for cell-based therapeutics. DATABASE: Gene expression data are available in the Gene Expression Omnibus (GEO) database under the accession number GSE130361.
Assuntos
Diferenciação Celular/genética , Reprogramação Celular/genética , Hemangioblastos/citologia , Células-Tronco Hematopoéticas/citologia , Animais , Técnicas de Cocultura/métodos , Fibroblastos/citologia , Fibroblastos/metabolismo , Fator de Transcrição GATA2/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Hemangioblastos/metabolismo , Hematopoese/genética , Células-Tronco Hematopoéticas/metabolismo , Humanos , Camundongos , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genéticaRESUMO
Osteosarcoma (OS), the most common primary bone tumor, is highly metastatic with high chemotherapeutic resistance and poor survival rates. Using induced pluripotent stem cells (iPSCs) generated from Li-Fraumeni syndrome (LFS) patients, we investigate an oncogenic role of secreted frizzled-related protein 2 (SFRP2) in p53 mutation-associated OS development. Interestingly, we find that high SFRP2 expression in OS patient samples correlates with poor survival. Systems-level analyses identified that expression of SFRP2 increases during LFS OS development and can induce angiogenesis. Ectopic SFRP2 overexpression in normal osteoblast precursors is sufficient to suppress normal osteoblast differentiation and to promote OS phenotypes through induction of oncogenic molecules such as FOXM1 and CYR61 in a ß-catenin-independent manner. Conversely, inhibition of SFRP2, FOXM1, or CYR61 represses the tumorigenic potential. In summary, these findings demonstrate the oncogenic role of SFRP2 in the development of p53 mutation-associated OS and that inhibition of SFRP2 is a potential therapeutic strategy.
Assuntos
Neoplasias Ósseas/genética , Carcinogênese/genética , Síndrome de Li-Fraumeni/patologia , Proteínas de Membrana/genética , Osteossarcoma/genética , Proteína Supressora de Tumor p53/genética , Animais , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Proteína Rica em Cisteína 61/antagonistas & inibidores , Proteína Rica em Cisteína 61/genética , Proteína Forkhead Box M1/antagonistas & inibidores , Proteína Forkhead Box M1/genética , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Síndrome de Li-Fraumeni/genética , Masculino , Proteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Nus , Neovascularização Patológica/genética , Osteoblastos/citologia , Osteossarcoma/patologiaRESUMO
The advent of induced pluripotent stem cells (iPSCs) together with recent advances in genome editing, microphysiological systems, tissue engineering and xenograft models present new opportunities for the investigation of hematological diseases and cancer in a patient-specific context. Here we review the progress in the field and discuss the advantages, limitations, and challenges of iPSC-based malignancy modeling. We will also discuss the use of iPSCs and its derivatives as cellular sources for drug target identification, drug development and evaluation of pharmacological responses.
Assuntos
Doenças Hematológicas/genética , Doenças Hematológicas/patologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Neoplasias/genética , Neoplasias/patologia , Animais , Reprogramação Celular , Modelos Animais de Doenças , Desenvolvimento de Medicamentos , Edição de Genes , Xenoenxertos , Humanos , Engenharia TecidualRESUMO
In the present study, we investigated the effect of melatonin on the preimplantation development of porcine parthenogenetic and somatic cell nuclear transfer (SCNT) embryos. Parthenogenetic embryos were cultured in mNCSU-23 supplemented with various concentrations of melatonin for 7 days. The results revealed that 100 pM was the optimal concentration, which resulted in significantly increased cleavage and blastocyst formation rates. Additionally, 100 pM melatonin provided the highest increase in total cell number of blastocysts. Therefore, the subsequent experiments were performed with 100 pM melatonin. ROS level in 2-8 cell stage embryos in the presence or absence of melatonin was evaluated. Embryos cultured with melatonin showed significantly decreased ROS. Blastocysts cultured with melatonin for 7 days were analyzed by the TUNEL assay. It was observed that melatonin not only increased (P < 0.05) the total cell number but also decreased (P < 0.05) the rate of apoptotic nuclei. Blastocysts cultured with melatonin were assessed for the expression of apoptosis-related genes Bcl-xl and Bax, and of pluripotency marker gene Oct-4 by real-time quantitative PCR. Analysis of data showed that the expression of Bcl-xl was higher (1.7-fold) compared to the control while the expression of Bax was significantly decreased relative to the control (0.7-fold) (P < 0.05). Moreover, the expression of Oct-4 was 1.7-fold higher than the control. These results indicated that melatonin had beneficial effects on the development of porcine parthenogenetic embryos. Based on the findings of parthenogenetic embryos, we investigated the effect of melatonin on the development of porcine SCNT embryos. The results also demonstrated increased cleavage and blastocyst formation rates, and the total cell numbers in blastocysts were significantly higher when the embryos were cultured with melatonin. Therefore, these data suggested that melatonin may have important implications for improving porcine preimplantation SCNT embryo development.
