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1.
Nat Commun ; 15(1): 1393, 2024 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-38360927

RESUMO

Patients affected by neurofibromatosis type 1 (NF1) frequently show muscle weakness with unknown etiology. Here we show that, in mice, Neurofibromin 1 (Nf1) is not required in muscle fibers, but specifically in early postnatal myogenic progenitors (MPs), where Nf1 loss led to cell cycle exit and differentiation blockade, depleting the MP pool resulting in reduced myonuclear accretion as well as reduced muscle stem cell numbers. This was caused by precocious induction of stem cell quiescence coupled to metabolic reprogramming of MPs impinging on glycolytic shutdown, which was conserved in muscle fibers. We show that a Mek/Erk/NOS pathway hypersensitizes Nf1-deficient MPs to Notch signaling, consequently, early postnatal Notch pathway inhibition ameliorated premature quiescence, metabolic reprogramming and muscle growth. This reveals an unexpected role of Ras/Mek/Erk signaling supporting postnatal MP quiescence in concert with Notch signaling, which is controlled by Nf1 safeguarding coordinated muscle growth and muscle stem cell pool establishment. Furthermore, our data suggest transmission of metabolic reprogramming across cellular differentiation, affecting fiber metabolism and function in NF1.


Assuntos
Neurofibromatose 1 , Neurofibromina 1 , Camundongos , Humanos , Animais , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Neurofibromatose 1/genética , Neurofibromatose 1/metabolismo , Transdução de Sinais/fisiologia , Sistema de Sinalização das MAP Quinases , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo
2.
Mol Metab ; 80: 101876, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38216123

RESUMO

OBJECTIVE: NF1 is a tumor suppressor gene and its protein product, neurofibromin, is a negative regulator of the RAS pathway. NF1 is one of the top driver mutations in sporadic breast cancer such that 27 % of breast cancers exhibit damaging NF1 alterations. NF1 loss-of-function is a frequent event in the genomic evolution of estrogen receptor (ER)+ breast cancer metastasis and endocrine resistance. Individuals with Neurofibromatosis type 1 (NF) - a disorder caused by germline NF1 mutations - have an increased risk of dying from breast cancer [1-4]. NF-related breast cancers are associated with decreased overall survival compared to sporadic breast cancer. Despite numerous studies interrogating the role of RAS mutations in tumor metabolism, no study has comprehensively profiled the NF1-deficient breast cancer metabolome to define patterns of energetic and metabolic reprogramming. The goals of this investigation were (1) to define the role of NF1 deficiency in estrogen receptor-positive (ER+) breast cancer metabolic reprogramming and (2) to identify potential targeted pathway and metabolic inhibitor combination therapies for NF1-deficient ER + breast cancer. METHODS: We employed two ER+ NF1-deficient breast cancer models: (1) an NF1-deficient MCF7 breast cancer cell line to model sporadic breast cancer, and (2) three distinct, Nf1-deficient rat models to model NF-related breast cancer [1]. IncuCyte proliferation analysis was used to measure the effect of NF1 deficiency on cell proliferation and drug response. Protein quantity was assessed by Western Blot analysis. We then used RNAseq to investigate the transcriptional effect of NF1 deficiency on global and metabolism-related transcription. We measured cellular energetics using Agilent Seahorse XF-96 Glyco Stress Test and Mito Stress Test assays. We performed stable isotope labeling and measured [U-13C]-glucose and [U-13C]-glutamine metabolite incorporation and measured total metabolite pools using mass spectrometry. Lastly, we used a Bliss synergy model to investigate NF1-driven changes in targeted and metabolic inhibitor synergy. RESULTS: Our results revealed that NF1 deficiency enhanced cell proliferation, altered neurofibromin expression, and increased RAS and PI3K/AKT pathway signaling while constraining oxidative ATP production and restricting energetic flexibility. Neurofibromin deficiency also increased glutamine influx into TCA intermediates and dramatically increased lipid pools, especially triglycerides (TG). Lastly, NF1 deficiency alters the synergy between metabolic inhibitors and traditional targeted inhibitors. This includes increased synergy with inhibitors targeting glycolysis, glutamine metabolism, mitochondrial fatty acid transport, and TG synthesis. CONCLUSIONS: NF1 deficiency drives metabolic reprogramming in ER+ breast cancer. This reprogramming is characterized by oxidative ATP constraints, glutamine TCA influx, and lipid pool expansion, and these metabolic changes introduce novel metabolic-to-targeted inhibitor synergies.


Assuntos
Neurofibromatose 1 , Neurofibromina 1 , Animais , Ratos , Trifosfato de Adenosina/metabolismo , Glutamina/metabolismo , Lipídeos , Neurofibromatose 1/genética , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo
3.
Fam Cancer ; 23(1): 35-40, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38270845

RESUMO

Neurofibromatosis type 1 (NF1) is an autosomal dominant condition caused by neurofibromin haploinsufficiency due to pathogenic variants in the NF1 gene. Tumor predisposition has long been associated with NF1, and an increased breast cancer (BC) incidence and reduced survival have been reported in recent years for women with NF1. As breast density is another known independent risk factor for BC, this study aims to evaluate the variability of breast density in patients with NF1 compared to the general population. Mammograms from 98 NF1 women affected by NF1, and enrolled onto our monocentric BC screening program, were compared with those from 300 healthy subjects to verify differences in breast density. Mammograms were independently reviewed and scored by a radiologist and using a Computer-Aided Detection (CAD) software. The comparison of breast density between NF1 patients and controls was performed through Chi-squared test and with multivariable ordinal logistic models adjusted for age, body mass index (BMI), number of pregnancies, and menopausal status.breast density was influenced by BMI and menopausal status in both NF1 patients and healthy subjects. No difference in breast density was observed between NF1 patients and the healthy female population, even after considering the potential confounding factors.Although NF1 and a highly fibroglandular breast are known risk factors of BC, in this study, NF1 patients were shown to have comparable breast density to healthy subjects. The presence of pathogenic variants in the NF1 gene does not influence the breast density value.


Assuntos
Neoplasias da Mama , Neurofibromatose 1 , Humanos , Feminino , Neurofibromatose 1/diagnóstico por imagem , Neurofibromatose 1/genética , Neurofibromatose 1/complicações , Densidade da Mama , Estudos Retrospectivos , Neurofibromina 1/genética , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/genética , Neoplasias da Mama/epidemiologia
4.
Artigo em Chinês | MEDLINE | ID: mdl-38225833

RESUMO

Objective: To summarize the gene therapy strategies for neurofibromatosis type 1 (NF1) and related research progress. Methods: The recent literature on gene therapy for NF1 at home and abroad was reviewed. The structure and function of the NF1 gene and its mutations were analyzed, and the current status as well as future prospects of the transgenic therapy and gene editing strategies were summarized. Results: NF1 is an autosomal dominantly inherited tumor predisposition syndrome caused by mutations in the NF1 tumor suppressor gene, which impair the function of the neurofibromin and lead to the disease. It has complex clinical manifestations and is not yet curable. Gene therapy strategies for NF1 are still in the research and development stage. Existing studies on the transgenic therapy for NF1 have mainly focused on the construction and expression of the GTPase-activating protein-related domain in cells that lack of functional neurofibromin, confirming the feasibility of the transgenic therapy for NF1. Future research may focus on split adeno-associated virus (AAV) gene delivery, oversized AAV gene delivery, and the development of new vectors for targeted delivery of full-length NF1 cDNA. In addition, the gene editing tools of the new generation have great potential to treat monogenic genetic diseases such as NF1, but need to be further validated in terms of efficiency and safety. Conclusion: Gene therapy, including both the transgenic therapy and gene editing, is expected to become an important new therapeutic approach for NF1 patients.


Assuntos
Neurofibromatose 1 , Humanos , Neurofibromatose 1/genética , Neurofibromatose 1/terapia , Neurofibromatose 1/patologia , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Proteínas Ativadoras de GTPase , Mutação , Predisposição Genética para Doença , Terapia Genética
5.
Brain Res Bull ; 206: 110860, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38143008

RESUMO

Forkhead box A1 (FOXA1), a member of the forkhead family of transcription factors, plays a crucial role in the development of various organ systems and exhibits neuroprotective properties. This study aims to investigate the effect of FOXA1 on Parkinson's disease (PD) and unravel the underlying mechanism. Transcriptome analysis of PD was conducted using three GEO datasets to identify aberrantly expressed genes. A mouse model of PD was generated by injecting neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP), resulting in reduced FOXA1 expression. FOXA1 decline was also observed in 1-methyl-4-phenylpyridinium-treated SH-SY5Y cells. Artificial upregulation of FOXA1 improved motor abilities of mice according to rotarod and pole tests, and it mitigated tissue damage, cell loss, and neuronal damage in the mouse substantia nigra or in vitro. FOXA1 was found to bind to the neurofibromin 1 (NF1) promoter, thereby inducing its transcription and inactivating the mitogen-activated protein kinase (MAPK) signaling pathway. Further experimentation revealed that silencing NF1 in mice or SH-SY5Y cells counteracted the neuroprotective effects of FOXA1. In conclusion, this research suggests that FOXA1 activates NF1 transcription and inactivates the MAPK signaling pathway, ultimately ameliorating neuronal damage and motor disability in PD. The findings may offer novel ideas in the field of PD management.


Assuntos
Pessoas com Deficiência , Transtornos Motores , Neuroblastoma , Fármacos Neuroprotetores , Doença de Parkinson , Animais , Humanos , Camundongos , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/farmacologia , Modelos Animais de Doenças , Neurônios Dopaminérgicos/metabolismo , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Fator 3-alfa Nuclear de Hepatócito/farmacologia , Sistema de Sinalização das MAP Quinases , Camundongos Endogâmicos C57BL , Transtornos Motores/tratamento farmacológico , Neuroblastoma/metabolismo , Neurofibromina 1/metabolismo , Neurofibromina 1/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Doença de Parkinson/metabolismo , Ativação Transcricional
6.
Dis Model Mech ; 16(12)2023 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-37990867

RESUMO

Neurofibromatosis type 1 (NF1) is an autosomal dominant condition caused by germline mutations in the neurofibromin 1 (NF1) gene. Children with NF1 are prone to the development of multiple nervous system abnormalities, including autism and brain tumors, which could reflect the effect of NF1 mutation on microglia function. Using heterozygous Nf1-mutant mice, we previously demonstrated that impaired purinergic signaling underlies deficits in microglia process extension and phagocytosis in situ. To determine whether these abnormalities are also observed in human microglia in the setting of NF1, we leveraged an engineered isogenic series of human induced pluripotent stem cells to generate human microglia-like (hiMGL) cells heterozygous for three different NF1 gene mutations found in patients with NF1. Whereas all NF1-mutant and isogenic control hiMGL cells expressed classical microglia markers and exhibited similar transcriptomes and cytokine/chemokine release profiles, only NF1-mutant hiMGL cells had defects in P2X receptor activation, phagocytosis and motility. Taken together, these findings indicate that heterozygous NF1 mutations impair a subset of the functional properties of human microglia, which could contribute to the neurological abnormalities seen in children with NF1.


Assuntos
Células-Tronco Pluripotentes Induzidas , Neurofibromatose 1 , Animais , Humanos , Camundongos , Genes da Neurofibromatose 1 , Microglia/patologia , Mutação/genética , Neurofibromatose 1/genética , Neurofibromina 1/genética
7.
Gynecol Oncol ; 178: 80-88, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37820398

RESUMO

OBJECTIVE: Inhibition of the MAPK pathway by MEK inhibitors (MEKi) is currently a therapeutic standard in several cancer types, including ovarian low-grade serous carcinoma (LGSC). A common MAPK pathway alteration in tubo-ovarian high-grade serous carcinoma (HGSC) is the genomic inactivation of neurofibromin 1 (NF1). The primary objectives of our study were to survey the prevalence of NF1 inactivation in the principal ovarian carcinoma histotype as well as to evaluate its associations with clinico-pathological parameters and key biomarkers including BRCA1/2 status in HGSC. METHODS: A recently commercialized NF1 antibody (clone NFC) was orthogonally validated on an automated immunohistochemistry (IHC) platform and IHC was performed on tissue microarrays containing 2140 ovarian carcinoma cases. Expression was interpreted as loss/inactivated (complete or subclonal) versus normal/retained. RESULTS: Loss of NF1 expression was detected in 250/1429 (17.4%) HGSC including 11% with subclonal loss. Survival of NF1-inactivated HGSC patients was intermediate between favorable BRCA1/2 mutated HGSC and unfavorable CCNE1 high-level amplified HGSC. NF1 inactivation was mutually exclusive with CCNE1 high-level amplifications, co-occurred with RB1 loss and occurred at similar frequencies in BRCA1/2 mutated versus wild-type HGSC. NF1 loss was found in 21/286 (7.3%) endometrioid carcinomas with a favorable prognostic association (p = 0.048), and in 4/64 (5.9%) LGSC, mutually exclusive with other driver events. CONCLUSIONS: NF1 inactivation occurs in a significant subset of BRCA1/2 wild-type HGSC and a subset of LGSC. While the functional effects of NF1 inactivation need to be further characterized, this signifies a potential therapeutic opportunity to explore targeting NF1 inactivation in these tumors.


Assuntos
Carcinoma Endometrioide , Cistadenocarcinoma Seroso , Neoplasias Ovarianas , Feminino , Humanos , Proteína BRCA1 , Neurofibromina 1/genética , Imuno-Histoquímica , Proteína BRCA2 , Neoplasias Ovarianas/patologia , Carcinoma Endometrioide/patologia , Cistadenocarcinoma Seroso/patologia , Carcinoma Epitelial do Ovário
8.
Clin Transl Med ; 13(9): e1427, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37743642

RESUMO

BACKGROUND: N6-methyladenosine (m6A) is an abundant reversible modification in eukaryotic mRNAs. Emerging evidences indicate that m6A modification plays a vital role in tumourigenesis. As a crucial reader of m6A, IGF2BP3 usually mediates the stabilisation of mRNAs via an m6A-dependent manner. But the underlying mechanism of IGF2BP3 in the tumourigenesis of triple-negative breast cancer (TNBC) is unclear. METHODS: TCGA cohorts were analysed for IGF2BP3 expression and IGF2BP3 promoter methylation levels in different breast cancer subtypes. Colony formation, flow cytometry assays and subcutaneous xenograft were performed to identify the phenotype of IGF2BP3 in TNBC. RNA/RNA immunoprecipitation (RIP)/methylated RNA immunoprecipitation (MeRIP) sequencing and luciferase assays were used to certify the target of IGF2BP3 in TNBC cells. RESULTS: IGF2BP3 was highly expressed in TNBC cell lines and tissues. TET3-mediated IGF2BP3 promoter hypomethylation led to the upregulation of IGF2BP3. Knocking down IGF2BP3 markedly reduced the proliferation of TNBC in vitro and in vivo. Intersection co-assays revealed that IGF2BP3 decreased neurofibromin 1 (NF1) stabilisation via an m6A-dependent manner. NF1 knockdown could rescue the phenotypes of IGF2BP3 knockdown cells partially. CONCLUSION: TET3-mediated IGF2BP3 accelerated the proliferation of TNBC by destabilising NF1 mRNA via an m6A-dependent manner. This suggests that IGF2BP3 could be a potential therapeutic target for TNBC.


Assuntos
Neurofibromina 1 , Estabilidade de RNA , Proteínas de Ligação a RNA , Neoplasias de Mama Triplo Negativas , Humanos , Carcinogênese , Transformação Celular Neoplásica , Neurofibromina 1/genética , RNA , Neoplasias de Mama Triplo Negativas/genética , Proteínas de Ligação a RNA/genética
9.
Exp Dermatol ; 32(11): 2012-2022, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37724850

RESUMO

The formation of hypertrophic scars and keloids is strongly associated with mechanical stimulation, and myofibroblasts are known to play a major role in abnormal scar formation. Wounds in patients with neurofibromatosis type 1 (NF1) become inconspicuous and lack the tendency to form abnormal scars. We hypothesized that there would be a unique response to mechanical stimulation and subsequent scar formation in NF1. To test this hypothesis, we investigated the molecular mechanisms of differentiation into myofibroblasts in NF1-derived fibroblasts and neurofibromin-depleted fibroblasts and examined actin dynamics, which is involved in fibroblast differentiation, with a focus on the pathway linking LIMK2/cofilin to actin dynamics. In normal fibroblasts, expression of α-smooth muscle actin (α-SMA), a marker of myofibroblasts, significantly increased after mechanical stimulation, whereas in NF1-derived and neurofibromin-depleted fibroblasts, α-SMA expression did not change. Phosphorylation of cofilin and subsequent actin polymerization did not increase in NF1-derived and neurofibromin-depleted fibroblasts after mechanical stimulation. Finally, in normal fibroblasts treated with Jasplakinolide, an actin stabilizer, α-SMA expression did not change after mechanical stimulation. Therefore, when neurofibromin was dysfunctional or depleted, subsequent actin polymerization did not occur in response to mechanical stimulation, which may have led to the unchanged expression of α-SMA. We believe this molecular pathway can be a potential therapeutic target for the treatment of abnormal scars.


Assuntos
Cicatriz Hipertrófica , Neurofibromatose 1 , Humanos , Miofibroblastos/metabolismo , Actinas/metabolismo , Neurofibromina 1/metabolismo , Fibroblastos/metabolismo , Cicatriz Hipertrófica/metabolismo , Neurofibromatose 1/patologia , Fatores de Despolimerização de Actina/metabolismo , Diferenciação Celular , Células Cultivadas , Fator de Crescimento Transformador beta1/metabolismo
10.
Expert Opin Investig Drugs ; 32(10): 941-957, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37747491

RESUMO

INTRODUCTION: NF1 is a tumor suppressor gene encoding neurofibromin, an inhibitor of the RAS/MAPK and PI3K-AKT-mTOR signaling pathways. NF1 germline pathogenic variants cause the tumor predisposition syndrome neurofibromatosis type 1. Targeted therapies (MEK inhibitors) have been approved for benign nerve sheath tumors in neurofibromatosis type 1 patients. NF1 somatic alterations are present in ~5% of all human sporadic cancers. In melanomas, acute myeloid leukemias and lung adenocarcinomas, the NF1 somatic alteration frequency is higher (~15%). However, to date, the therapeutic impact of NF1 somatic alterations is poorly investigated. AREAS COVERED: This review presents a comprehensive overview of targeted therapies and immunotherapies currently developed and evaluated in vitro and in vivo for NF1-altered cancer treatment. A PubMed database literature review was performed to select relevant original articles. Active clinical trials were researched in ClinicalTrials.gov database in August 2022. TCGA and HGMD® databases were consulted. EXPERT OPINION: This review highlights the need to better understand the molecular mechanisms of NF1-altered tumors and the development of innovative strategies to effectively target NF1-loss in human cancers. One of the current major challenges in cancer management is the targeting of tumor suppressor genes such as NF1 gene. Currently, most studies are focusing on inhibitors of the RAS/MAPK and PI3K-AKT-mTOR pathways and immunotherapies.


Assuntos
Neurofibromatose 1 , Humanos , Neurofibromatose 1/genética , Neurofibromatose 1/metabolismo , Neurofibromatose 1/patologia , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Genes da Neurofibromatose 1 , Proteínas Proto-Oncogênicas c-akt , Medicina de Precisão , Fosfatidilinositol 3-Quinases/genética , Serina-Treonina Quinases TOR/metabolismo
11.
Kidney Blood Press Res ; 48(1): 568-577, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37562365

RESUMO

INTRODUCTION: Neurofibromatosis type 1 (NF-1) is caused by mutations in the NF1 gene that encodes neurofibromin, a negative regulator of RAS proto-oncogene. Approximately one-third of the reported pathogenic mutations in NF1 are splicing mutations, but most consequences are unclear. The objective of this study was to identify the pathogenicity of splicing mutation in a Chinese family with NF-1 and determine the effects of the pre-mRNA splicing mutation by in vitro functional analysis. METHODS: Next-generation sequencing was used to screen candidate mutations. We performed a minigene splicing assay to determine the effect of the splicing mutation on NF1 expression, and three-dimensional structure models of neurofibromin were generated using SWISS-MODEL and PROCHECK methods, respectively. RESULTS: A pathogenic splicing mutation c.479+1G>C in NF1 was found in the proband characterized by childhood-onset refractory hypertension. In vitro analysis demonstrated that c.479+1G>C mutation caused the skipping of exon 4, leading to a glutamine-to-valine substitution at position 97 in neurofibromin and an open reading frame shift terminating at codon 108. Protein modeling showed that several major domains were missing in the truncated neurofibromin protein. CONCLUSION: The splicing mutation c.479+1G>C identified in a Chinese patient with NF-1 and childhood-onset refractory hypertension caused the skipping of exon 4 and a truncated protein. Our findings offer new evidence for the molecular diagnosis of NF-1.


Assuntos
Hipertensão , Neurofibromatose 1 , Criança , Humanos , Genes da Neurofibromatose 1 , Hipertensão/genética , Mutação , Neurofibromatose 1/genética , Neurofibromatose 1/diagnóstico , Neurofibromina 1/genética
12.
Cancer Res Commun ; 3(7): 1366-1377, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37501682

RESUMO

NF1 is a key tumor suppressor that represses both RAS and estrogen receptor-α (ER) signaling in breast cancer. Blocking both pathways by fulvestrant (F), a selective ER degrader, together with binimetinib (B), a MEK inhibitor, promotes tumor regression in NF1-depleted ER+ models. We aimed to establish approaches to determine how NF1 protein levels impact B+F treatment response to improve our ability to identify B+F sensitive tumors. We examined a panel of ER+ patient-derived xenograft (PDX) models by DNA and mRNA sequencing and found that more than half of these models carried an NF1 shallow deletion and generally have low mRNA levels. Consistent with RAS and ER activation, RET and MEK levels in NF1-depleted tumors were elevated when profiled by mass spectrometry (MS) after kinase inhibitor bead pulldown. MS showed that NF1 can also directly and selectively bind to palbociclib-conjugated beads, aiding quantification. An IHC assay was also established to measure NF1, but the MS-based approach was more quantitative. Combined IHC and MS analysis defined a threshold of NF1 protein loss in ER+ breast PDX, below which tumors regressed upon treatment with B+F. These results suggest that we now have a MS-verified NF1 IHC assay that can be used for patient selection as a complement to somatic genomic analysis. Significance: A major challenge for targeting the consequence of tumor suppressor disruption is the accurate assessment of protein functional inactivation. NF1 can repress both RAS and ER signaling, and a ComboMATCH trial is underway to treat the patients with binimetinib and fulvestrant. Herein we report a MS-verified NF1 IHC assay that can determine a threshold for NF1 loss to predict treatment response. These approaches may be used to identify and expand the eligible patient population.


Assuntos
Neoplasias da Mama , Proteogenômica , Humanos , Feminino , Neoplasias da Mama/tratamento farmacológico , Neurofibromina 1/genética , Fulvestranto/farmacologia , Receptores de Estrogênio/genética , Inibidores de Proteínas Quinases/farmacologia , Fatores de Transcrição NFI , RNA Mensageiro , Quinases de Proteína Quinase Ativadas por Mitógeno
13.
Physiol Genomics ; 55(10): 415-426, 2023 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-37519249

RESUMO

Congenital heart disease is the most frequent congenital disorder, affecting a significant number of live births. Gaining insights into its genetic etiology could lead to a deeper understanding of this condition. Although the Nf1 gene has been identified as a potential causative gene, its role in congenital heart disease has not been thoroughly clarified. We searched and summarized evidence from cohort-based and experimental studies on the issue of Nf1 and heart development in congenital heart diseases from various databases. Available evidence demonstrates a correlation between Nf1 and congenital heart diseases, mainly pulmonary valvar stenosis. The mechanism underlying this correlation may involve dysregulation of epithelial-mesenchymal transition (EMT). The Nf1 gene affects the EMT process via multiple pathways, including directly regulating the expression of EMT-related transcription factors and indirectly regulating the EMT process by regulating the MAPK pathway. This narrative review provides a comprehensive account of the Nf1 involvement in heart development and congenital cardiovascular diseases in terms of epidemiology and potential mechanisms. RAS signaling may contribute to congenital heart disease independently or in cooperation with other signaling pathways. Efficient management of both NF1 and cardiovascular disease patients would benefit from further research into these issues.


Assuntos
Doenças Cardiovasculares , Cardiopatias Congênitas , Neurofibromatose 1 , Humanos , Neurofibromatose 1/genética , Neurofibromatose 1/metabolismo , Genes da Neurofibromatose 1 , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Coração , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/epidemiologia , Doenças Cardiovasculares/genética
14.
Cell Death Dis ; 14(6): 373, 2023 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-37355626

RESUMO

Phosphodiesterase 4D interacting protein (PDE4DIP) is a centrosome/Golgi protein associated with cyclic nucleotide phosphodiesterases. PDE4DIP is commonly mutated in human cancers, and its alteration in mice leads to a predisposition to intestinal cancer. However, the biological function of PDE4DIP in human cancer remains obscure. Here, we report for the first time the oncogenic role of PDE4DIP in colorectal cancer (CRC) growth and adaptive MEK inhibitor (MEKi) resistance. We show that the expression of PDE4DIP is upregulated in CRC tissues and associated with the clinical characteristics and poor prognosis of CRC patients. Knockdown of PDE4DIP impairs the growth of KRAS-mutant CRC cells by inhibiting the core RAS signaling pathway. PDE4DIP plays an essential role in the full activation of oncogenic RAS/ERK signaling by suppressing the expression of the RAS GTPase-activating protein (RasGAP) neurofibromin (NF1). Mechanistically, PDE4DIP promotes the recruitment of PLCγ/PKCε to the Golgi apparatus, leading to constitutive activation of PKCε, which triggers the degradation of NF1. Upregulation of PDE4DIP results in adaptive MEKi resistance in KRAS-mutant CRC by reactivating the RAS/ERK pathway. Our work reveals a novel functional link between PDE4DIP and NF1/RAS signal transduction and suggests that targeting PDE4DIP is a promising therapeutic strategy for KRAS-mutant CRC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Neoplasias Colorretais , Proteínas do Citoesqueleto , Neurofibromina 1 , Humanos , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Resistencia a Medicamentos Antineoplásicos/genética , Mutação , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo
15.
Br J Haematol ; 202(2): 328-343, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37144690

RESUMO

Juvenile myelomonocytic leukaemia (JMML) is an aggressive paediatric leukaemia characterized by mutations in five canonical RAS pathway genes, including the NF1 gene. JMML is driven by germline NF1 gene mutations, with additional somatic aberrations resulting in the NF1 biallelic inactivation, leading to disease progression. Germline mutations in the NF1 gene alone primarily cause benign neurofibromatosis type 1 (NF1) tumours rather than malignant JMML, yet the underlying mechanism remains unclear. Here, we demonstrate that with reduced NF1 gene dose, immune cells are promoted in anti-tumour immune response. Comparing the biological properties of JMML and NF1 patients, we found that not only JMML but also NF1 patients driven by NF1 mutations could increase monocytes generation. But monocytes cannot further malignant development in NF1 patients. Utilizing haematopoietic and macrophage differentiation from iPSCs, we revealed that NF1 mutations or knockout (KO) recapitulated the classical haematopoietic pathological features of JMML with reduced NF1 gene dose. NF1 mutations or KO promoted the proliferation and immune function of NK cells and iMacs derived from iPSCs. Moreover, NF1-mutated iNKs had a high capacity to kill NF1-KO iMacs. NF1-mutated or KO iNKs administration delayed leukaemia progression in a xenograft animal model. Our findings demonstrate that germline NF1 mutations alone cannot directly drive JMML development and suggest a potential cell immunotherapy for JMML patients.


Assuntos
Leucemia Mielomonocítica Juvenil , Neurofibromatose 1 , Animais , Humanos , Leucemia Mielomonocítica Juvenil/genética , Leucemia Mielomonocítica Juvenil/terapia , Leucemia Mielomonocítica Juvenil/metabolismo , Neurofibromina 1/genética , Genes da Neurofibromatose 1 , Mutação em Linhagem Germinativa , Neurofibromatose 1/genética , Neurofibromatose 1/terapia , Mutação , Imunidade , Células Germinativas/metabolismo , Células Germinativas/patologia
16.
J Biol Chem ; 299(6): 104789, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-37149146

RESUMO

Sprouty-related EVH-1 domain-containing (SPRED) proteins are a family of proteins that negatively regulate the RAS-Mitogen-Activated Protein Kinase (MAPK) pathway, which is involved in the regulation of the mitogenic response and cell proliferation. However, the mechanism by which these proteins affect RAS-MAPK signaling has not been elucidated. Patients with mutations in SPRED give rise to unique disease phenotypes; thus, we hypothesized that distinct interactions across SPRED proteins may account for alternative nodes of regulation. To characterize the SPRED interactome and evaluate how members of the SPRED family function through unique binding partners, we performed affinity purification mass spectrometry. We identified 90-kDa ribosomal S6 kinase 2 (RSK2) as a specific interactor of SPRED2 but not SPRED1 or SPRED3. We identified that the N-terminal kinase domain of RSK2 mediates the interaction between amino acids 123 to 201 of SPRED2. Using X-ray crystallography, we determined the structure of the SPRED2-RSK2 complex and identified the SPRED2 motif, F145A, as critical for interaction. We found that the formation of this interaction is regulated by MAPK signaling events. We also find that this interaction between SPRED2 and RSK2 has functional consequences, whereby the knockdown of SPRED2 resulted in increased phosphorylation of RSK substrates, YB1 and CREB. Furthermore, SPRED2 knockdown hindered phospho-RSK membrane and nuclear subcellular localization. We report that disruption of the SPRED2-RSK complex has effects on RAS-MAPK signaling dynamics. Our analysis reveals that members of the SPRED family have unique protein binding partners and describes the molecular and functional determinants of SPRED2-RSK2 complex dynamics.


Assuntos
Proteínas Quinases Ativadas por Mitógeno , Proteínas Repressoras , Proteínas Quinases S6 Ribossômicas 90-kDa , Transdução de Sinais , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosforilação , Proteínas Quinases S6 Ribossômicas 90-kDa/química , Proteínas Quinases S6 Ribossômicas 90-kDa/genética , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Transdução de Sinais/genética , Humanos , Linhagem Celular , Domínios Proteicos , Proteínas Repressoras/química , Proteínas Repressoras/metabolismo , Técnicas de Silenciamento de Genes , Transporte Proteico/genética , Ligação Proteica , Estrutura Terciária de Proteína , Modelos Moleculares , Neurofibromina 1/metabolismo
17.
J Transl Med ; 21(1): 306, 2023 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-37147639

RESUMO

BACKGROUND: Gallbladder cancer (GBC) is the most prevalent and invasive biliary tract malignancy. As a GTPase-activating protein, Neurofibromin 1 (NF1) is a tumor suppressor that negatively regulates the RAS signaling pathway, and its abnormality leads to neurofibromatosis type 1 (NF-1) disease. However, the role of NF1 playing in GBC and the underlying molecular mechanism has not been defined yet. METHODS: A combination of NOZ and EH-GB1 cell lines as well as nude mice, were utilized in this study. mRNA expression and protein levels of NF1 and YAP1 were evaluated by quantitative real-time PCR (qRT-PCR), western blot (WB), and immunohistochemistry (IHC). In vitro and in vivo assays were performed to explore the biological effects of NF1 in NOZ and EH-GB1 cells via siRNA or lv-shRNA mediated knockdown. Direct interaction between NF1 and YAP1 was detected by confocal microscopy and co-immunoprecipitation (Co-IP), and further confirmed by GST pull-down assay and isothermal titration calorimetry assay (ITC). The stability of proteins was measured by western blot (WB) in the presence of cycloheximide. RESULTS: This study showed that a higher level of NF1 and YAP1 was found in GBC samples than in normal tissues and associated with worse prognoses. The NF1 knockdown impaired the proliferation and migration of NOZ in vivo and in vitro by downregulating YAP1 expression. Moreover, NF1 co-localized with YAP1 in NOZ and EH-GB1 cells, and the WW domains of YAP1 specifically recognized the PPQY motif of NF1. The structural modeling also indicated the hydrophobic interactions between YAP1 and NF1. On the other hand, YAP1 knockdown also impaired the proliferation of NOZ in vitro, phenocopying the effects of NF1 knockdown. Overexpression of YAP1 can partially rescue the impaired proliferation in NF1 stably knockdown cells. In mechanism, NF1 interacted with YAP1 and increased the stability of YAP1 by preventing ubiquitination. CONCLUSIONS: Our findings discovered a novel oncogenic function of NF1 by directly interacting with YAP1 protein and stabilizing YAP1 to protect it from proteasome degradation in NOZ cells. NF1 may serve as a potential therapeutic target in GBC.


Assuntos
Neoplasias da Vesícula Biliar , Neurofibromina 1 , Proteínas de Sinalização YAP , Animais , Camundongos , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias da Vesícula Biliar/genética , Regulação Neoplásica da Expressão Gênica , Camundongos Nus , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , RNA Interferente Pequeno , Transdução de Sinais , Humanos , Proteínas de Sinalização YAP/genética , Proteínas de Sinalização YAP/metabolismo
18.
Eur J Hum Genet ; 31(8): 931-938, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37217626

RESUMO

Spinal neurofibromatosis (SNF) is a form of neurofibromatosis type 1 (NF1) characterized by bilateral neurofibromas involving all spinal roots. The pathogenic mechanisms determining the SNF form are currently unknown. To verify the presence of genetic variants possibly related to SNF or classic NF1, we studied 106 sporadic NF1 and 75 SNF patients using an NGS panel of 286 genes encoding RAS pathway effectors and neurofibromin interactors and evaluated the expression of syndecans (SDC1, SDC2, SDC3, SDC4), the NF1 3' tertile interactors, by quantitative real-time PCR. We previously identified 75 and 106 NF1 variants in SNF and NF1 cohorts, respectively. The analysis of the distribution of pathogenic NF1 variants in the three NF1 tertiles showed a significantly higher prevalence of NF1 3' tertile mutations in SNF than in the NF1 cohort. We hypothesized a potential pathogenic significance of the 3' tertile NF1 variants in SNF. The analysis of syndecan expression on PBMCs RNAs from 16 SNF, 16 classic NF1 patients and 16 healthy controls showed that the expression levels of SDC2 and SDC3 were higher in SNF and NF1 patients than in controls; moreover, SDC2, SDC3 and SDC4 were significantly over expressed in patients mutated in the 3' tertile compared to controls. Two different mutational NF1 spectra seem to characterize SNF and classic NF1, suggesting a pathogenic role of NF1 3' tertile and its interactors, syndecans, in SNF. Our study, providing new insights on a possible role of neurofibromin C-terminal in SNF, could address effective personalized patient management and treatments.


Assuntos
Neurofibromatoses , Neurofibromatose 1 , Humanos , Neurofibromatose 1/genética , Neurofibromina 1/genética , Mutação , Sindecanas/genética , Genes da Neurofibromatose 1
19.
Biol Sex Differ ; 14(1): 24, 2023 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-37101298

RESUMO

BACKGROUND: Neurofibromatosis type 1 (NF1) is an inherited neurocutaneous disorder associated with neurodevelopmental disorders including autism spectrum disorder (ASD). This condition has been associated with an increase of gamma-aminobutyric acid (GABA) neurotransmission and, consequently, an excitation/inhibition imbalance associated with autistic-like behavior in both human and animal models. Here, we explored the influence of biological sex in the GABAergic system and behavioral alterations induced by the Nf1+/- mutation in a murine model. METHODS: Juvenile male and female Nf1+/- mice and their wild-type (WT) littermates were used. Hippocampus size was assessed by conventional toluidine blue staining and structural magnetic resonance imaging (MRI). Hippocampal GABA and glutamate levels were determined by magnetic resonance spectroscopy (MRS), which was complemented by western blot for the GABA(A) receptor. Behavioral evaluation of on anxiety, memory, social communication, and repetitive behavior was performed. RESULTS: We found that juvenile female Nf1+/- mice exhibited increased hippocampal GABA levels. Moreover, mutant female displays a more prominent anxious-like behavior together with better memory performance and social behavior. On the other hand, juvenile Nf1+/- male mice showed increased hippocampal volume and thickness, with a decrease in GABA(A) receptor levels. We observed that mutant males had higher tendency for repetitive behavior. CONCLUSIONS: Our results suggested a sexually dimorphic impact of Nf1+/- mutation in hippocampal neurochemistry, and autistic-like behaviors. For the first time, we identified a "camouflaging"-type behavior in females of an animal model of ASD, which masked their autistic traits. Accordingly, like observed in human disorder, in this animal model of ASD, females show larger anxiety levels but better executive functions and production of normative social patterns, together with an imbalance of inhibition/excitation ratio. Contrary, males have more externalizing disorders, such as hyperactivity and repetitive behaviors, with memory deficits. The ability of females to camouflage their autistic traits creates a phenotypic evaluation challenge that mimics the diagnosis difficulty observed in humans. Thus, we propose the study of the Nf1+/- mouse model to better understand the sexual dimorphisms of ASD phenotypes and to create better diagnostic tools.


Assuntos
Transtorno do Espectro Autista , Neurofibromatose 1 , Animais , Feminino , Humanos , Masculino , Camundongos , Transtorno do Espectro Autista/genética , Transtorno do Espectro Autista/complicações , Transtorno do Espectro Autista/diagnóstico , Ácido gama-Aminobutírico , Neurofibromatose 1/genética , Neurofibromatose 1/complicações , Receptores de GABA-A , Caracteres Sexuais , Neurofibromina 1/genética , Neurofibromina 1/metabolismo
20.
BMC Med Genomics ; 16(1): 85, 2023 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-37095468

RESUMO

Neurofibromatosis type 1 (NF1) presents an autosomal dominant, haploinsufficient, and multisystemic disorder with patches of skin café-au-lait spots, lisch nodules in the iris, even tumors in the peripheral nervous system or fibromatous skin. In this study, a Chinese young woman who suffered from NF1 disease with first-trimester spontaneous abortion was recruited. Analysis for whole exome sequencing (WES), Sanger sequencing, short tandem repeat (STR), and co-segregation was carried out. As results, a novel, heterozygous, de novo pathogenic variant (c.4963delA:p.Thr1656Glnfs*42) of the NF1 gene in the proband was identified. This pathogenic variant of the NF1 gene produced a truncated protein that lost more than one-third of the NF1 protein at the C-terminus including half of the CRAL-TRIO lipid-binding domain and nuclear localization signal (NLS), thus leading to pathogenicity (ACMG criteria: PVS1 + PM2 + PM2). Analysis for NF1 conservation in species revealed high conservation in different species. Analysis of NF1 mRNA levels in different human tissues showed low tissue specificity, which may affect multiple organs presenting other symptoms or phenotypes. Moreover, prenatal NF1 gene diagnosis showed both alleles as wild types. Thus, this NF1 novel variant probably underlays the NF1 pathogenesis in this pedigree, which would help for the diagnosis, genetic counseling, and clinical management of this disorder.


Assuntos
Neurofibromatose 1 , Feminino , Humanos , Manchas Café com Leite/diagnóstico , Manchas Café com Leite/genética , População do Leste Asiático , Genes da Neurofibromatose 1 , Neurofibromatose 1/diagnóstico , Neurofibromatose 1/genética , Neurofibromatose 1/patologia , Neurofibromina 1/genética
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