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BioRxiv : the Preprint Server For... Mar 2024The mechanisms driving late relapse in uveal melanoma (UM) patients remains a medical mystery and major challenge. Clinically it is inferred that UM disseminated cancer...
UNLABELLED
The mechanisms driving late relapse in uveal melanoma (UM) patients remains a medical mystery and major challenge. Clinically it is inferred that UM disseminated cancer cells (DCCs) persist asymptomatic for years-to-decades mainly in the liver before they manifest as symptomatic metastasis. Here we reveal using Gαq/11 /BAP human uveal melanoma models and human UM metastatic samples, that the neural crest lineage commitment nuclear receptor NR2F1 is a key regulator of spontaneous UM DCC dormancy in the liver. Using a quiescence reporter, RNA-seq and multiplex imaging we revealed that rare dormant UM DCCs upregulate NR2F1 expression and genes related to neural crest programs while repressing gene related to cell cycle progression. Gain and loss of function assays showed that NR2F1 silences YAP1/TEAD1 transcription downstream of Gαq/11 signaling and that NR2F1 expression can also be repressed by YAP1. YAP1 expression is repressed by NR2F1 binding to its promoter and changing the histone H3 tail activation marks to repress YAP1 transcription. CRISPR KO of NR2F1 led dormant UM DCCs to awaken and initiate relentless liver metastatic growth. Cut&Run and bulk RNA sequencing further confirmed that NR2F1 epigenetically stimulates neuron axon guidance and neural lineage programs, and it globally represses gene expression linked to G-protein signaling to drive dormancy. Pharmacological inhibition of Gαq/11 signaling resulted in NR2F1 upregulation and robust UM growth arrest, which was also achieved using a novel NR2F1 agonist. Our work sheds light on the molecular underpinnings of UM dormancy revealing that transcriptional programs driven by NR2F1 epigenetically short-circuit Gαq/11 signaling to its downstream target YAP1.
HIGHLIGHTS
Quiescent solitary uveal melanoma (UM) DCCs in the liver up- and down-regulate neural crest and cell cycle progression programs, respectively.NR2F1 drives solitary UM DCC dormancy by antagonizing the Gαq/11-YAP1 pathway; small molecule Gαq/11 inhibition restores NR2F1 expression and quiescence. NR2F1 short-circuits oncogenic YAP1 and G-protein signaling a chromatin remodeling program. Loss of function of NR2F1 in dormant UM DCCs leads to aggressive liver metastasis.
PubMed: 38496663
DOI: 10.1101/2024.03.05.583565 -
JPMA. the Journal of the Pakistan... Feb 2024To identify the mutation in codon 201 of the deleted in colorectal cancer gene in colorectal cancer, and to correlate that mutation to the histopathological grading of...
OBJECTIVE
To identify the mutation in codon 201 of the deleted in colorectal cancer gene in colorectal cancer, and to correlate that mutation to the histopathological grading of colorectal cancer.
METHODS
The cross-sectional study was conducted from February 2019 to February 2021 after approval from the ethics review board of the Dow University of Health Sciences, Karachi, and comprised biopsy-proven colorectal cancer patients regardless of age and gender. After histopathological reporting, formalin-fixed paraffin-embedded tissue blocks of colorectal cancer were used for deoxyribonucleic acid extraction, followed by polymerase chain reaction optimisation and deoxyribonucleic acid Sanger sequencing for mutational analysis. Data was analysed using SPSS 25.
RESULTS
Of the 100 biopsy specimens assessed, 45(45%) were selected. Of them, 13(29%) samples failed to show any band on gel electrophoresis. The remaining 32(71%) samples were used for Sanger sequencing. Of these, 1(3%) sample did not sequence, while 31(97%) showed sequencing. All the sequenced samples identified a mutation in codon 201 of exon 3 in the deleted in colorectal cancer gene; 30(97%) showed homozygosity, and 1(3%) showed heterozygosity. No significant association of point mutation was noted with various demographic and clinicopathological parameters (p>0.05).
CONCLUSION
The deleted in colorectal cancer gene's missense mutation in codon 201 was frequently observed in colorectal cancer patients.
Topics: Humans; Genes, DCC; Colorectal Neoplasms; Cross-Sectional Studies; Mutation; Codon; DNA; DCC Receptor
PubMed: 38419228
DOI: 10.47391/JPMA.9158 -
Nucleic Acids Research May 2024Acetylation of lysine 16 of histone H4 (H4K16ac) stands out among the histone modifications, because it decompacts the chromatin fiber. The metazoan acetyltransferase...
Acetylation of lysine 16 of histone H4 (H4K16ac) stands out among the histone modifications, because it decompacts the chromatin fiber. The metazoan acetyltransferase MOF (KAT8) regulates transcription through H4K16 acetylation. Antibody-based studies had yielded inconclusive results about the selectivity of MOF to acetylate the H4 N-terminus. We used targeted mass spectrometry to examine the activity of MOF in the male-specific lethal core (4-MSL) complex on nucleosome array substrates. This complex is part of the Dosage Compensation Complex (DCC) that activates X-chromosomal genes in male Drosophila. During short reaction times, MOF acetylated H4K16 efficiently and with excellent selectivity. Upon longer incubation, the enzyme progressively acetylated lysines 12, 8 and 5, leading to a mixture of oligo-acetylated H4. Mathematical modeling suggests that MOF recognizes and acetylates H4K16 with high selectivity, but remains substrate-bound and continues to acetylate more N-terminal H4 lysines in a processive manner. The 4-MSL complex lacks non-coding roX RNA, a critical component of the DCC. Remarkably, addition of RNA to the reaction non-specifically suppressed H4 oligo-acetylation in favor of specific H4K16 acetylation. Because RNA destabilizes the MSL-nucleosome interaction in vitro we speculate that RNA accelerates enzyme-substrate turn-over in vivo, thus limiting the processivity of MOF, thereby increasing specific H4K16 acetylation.
Topics: Animals; Male; Acetylation; Drosophila melanogaster; Drosophila Proteins; Histone Acetyltransferases; Histones; Lysine; Nuclear Proteins; Nucleosomes; Substrate Specificity; Transcription Factors; Histone Code
PubMed: 38407474
DOI: 10.1093/nar/gkae123 -
Journal of Clinical Medicine Feb 2024: Pathogenic variants in the deleted in colorectal cancer gene (DCC), encoding the Netrin-1 receptor, may lead to mirror movements (MMs) associated with...
: Pathogenic variants in the deleted in colorectal cancer gene (DCC), encoding the Netrin-1 receptor, may lead to mirror movements (MMs) associated with agenesis/dysgenesis of the corpus callosum (ACC) and cognitive and/or neuropsychiatric issues. The clinical phenotype is related to the biological function of DCC in the corpus callosum and corticospinal tract development as Netrin-1 is implicated in the guidance of developing axons toward the midline. We report on a child with a novel inherited, monoallelic, pathogenic variant in the DCC gene. : Standardized measures and clinical scales were used to assess psychomotor development, communication and social skills, emotional and behavioural difficulties. MMs were measured via the Woods and Teuber classification. Exome sequencing was performed on affected and healthy family members. : The patient's clinical presentation during infancy consisted of paroxysmal dystonic posturing when asleep, mimicking nocturnal leg cramps. A brain magnetic resonance imaging (MRI) showed complete ACC. He developed typical upper limb MMs during childhood and a progressively evolving neuro-phenotype with global development delay and behavioural problems. We found an intrafamilial clinical variability associated with DCC mutations: the proband's father and uncle shared the same DCC variant, with a milder clinical phenotype. The atypical early clinical presentation of the present patient expands the clinical spectrum associated with DCC variants, especially those in the paediatric age. : This study underlines the importance of in-depth genetic investigations in young children with ACC and highlights the need for further detailed analyses of early motor symptoms in infants with DCC mutations.
PubMed: 38398422
DOI: 10.3390/jcm13041109 -
Circulation. Genomic and Precision... Apr 2024Less than 40% of patients with dilated cardiomyopathy (DCM) have a pathogenic/likely pathogenic genetic variant identified. has been linked to congenital heart defects;...
BACKGROUND
Less than 40% of patients with dilated cardiomyopathy (DCM) have a pathogenic/likely pathogenic genetic variant identified. has been linked to congenital heart defects; although an association with left ventricular noncompaction (LVNC) and DCM has been proposed, it is still considered a gene with limited evidence for these phenotypes. This study sought to investigate the association between the truncating variant () and DCM/LVNC.
METHODS
was sequenced by next-generation sequencing in 7463 unrelated probands with a diagnosis of DCM or LVNC, 22 773 probands of an internal comparison group (hypertrophic cardiomyopathy, channelopathies, or aortic diseases), and 124 098 external controls (individuals from the gnomAD database). Enrichment of in DCM/LVNC was calculated, cosegregation was determined in selected families, and clinical characteristics and outcomes were analyzed in carriers.
RESULTS
was enriched in DCM/LVNC (24/7463; 0.32%) compared with internal (1/22 773; 0.004%) and external comparison groups (4/124 098; 0.003%), with odds ratios of 73.23 (95% CI, 9.90-541.45; <0.0001) and 99.76 (95% CI, 34.60-287.62; <0.0001), respectively. was cosegregated with DCM/LVNC phenotype in 21 families for a combined logarythm of the odds score of 4.53 (strong linkage). Among 57 individuals with (49.1% men; mean age, 35.9±20.8 years), 41 (71.9%) exhibited DCM/LVNC, of whom 14 (34.1%) had also congenital heart defects. After a median follow-up of 6.9 (95% CI, 25-75:3.6-14.5) years, 9.7% of patients with DCM/LVNC had end-stage heart failure events and 4.8% experienced malignant ventricular arrhythmias.
CONCLUSIONS
is associated with DCM/LVNC; congenital heart defect is also present in around one-third of cases. -associated DCM/LVNC is characterized by a nonaggressive phenotype, with a low incidence of major cardiovascular events. should be considered a definitive gene for DCM and LVNC and routinely included in genetic testing panels for these phenotypes.
Topics: Male; Humans; Adolescent; Young Adult; Adult; Middle Aged; Female; Cardiomyopathy, Dilated; Heart Defects, Congenital; Arrhythmias, Cardiac; Phenotype; T-Box Domain Proteins
PubMed: 38353104
DOI: 10.1161/CIRCGEN.123.004404 -
Scientific Reports Jan 2024Among the most prevalent neurodevelopmental disorders, Autism Spectrum Disorder (ASD) is highly diverse showing a broad phenotypic spectrum. ASD also couples with a...
Among the most prevalent neurodevelopmental disorders, Autism Spectrum Disorder (ASD) is highly diverse showing a broad phenotypic spectrum. ASD also couples with a broad range of mutations, both de novo and inherited. In this study, we used a proprietary SNP genotyping chip to analyze the genomic DNA of 250 Vietnamese children diagnosed with ASD. Our Single Nucleotide Polymorphism (SNP) genotyping chip directly targets more than 800 thousand SNPs in the genome. Our primary focus was to identify pathogenic/likely pathogenic mutations that are potentially linked to more severe symptoms of autism. We identified and validated 23 pathogenic/likely pathogenic mutations in this initial study. The data shows that these mutations were detected in several cases spanning multiple biological pathways. Among the confirmed SNPs, mutations were identified in genes previously known to be strongly associated with ASD such as SLCO1B1, ACADSB, TCF4, HCP5, MOCOS, SRD5A2, MCCC2, DCC, and PRKN while several other mutations are known to associate with autistic traits or other neurodevelopmental disorders. Some mutations were found in multiple patients and some patients carried multiple pathogenic/likely pathogenic mutations. These findings contribute to the identification of potential targets for therapeutic solutions in what is considered a genetically heterogeneous neurodevelopmental disorder.
Topics: Child; Humans; Autism Spectrum Disorder; Polymorphism, Single Nucleotide; Genotype; Vietnam; Genetic Predisposition to Disease; Mutation; Liver-Specific Organic Anion Transporter 1; Sulfurtransferases; Membrane Proteins; 3-Oxo-5-alpha-Steroid 4-Dehydrogenase
PubMed: 38287090
DOI: 10.1038/s41598-024-52777-y -
Translational Pediatrics Dec 2023Gene editing of the porcine genome has enabled the production of pigs that do not express the three known carbohydrate antigens that are associated with hyperacute... (Review)
Review
Gene editing of the porcine genome has enabled the production of pigs that do not express the three known carbohydrate antigens that are associated with hyperacute rejection of a pig organ xenotransplant. In addition, it is now possible to insert a variety of human transgenes to protect against the human immune response, e.g., to protect from complement and coagulation activation. As a result, cardiac xenotransplantation of the gene-edited porcine heart is progressing towards clinical application. Many hope that it will definitively address the disparity between organ supply and demand. The role of cardiac xenotransplantation in pediatric care remains controversial but we believe there is an infant patient population with complex congenital heart disease (CHD) (not optimally managed by conventional surgical approaches) that is ideally suited to initial clinical application of this new technology. The most efficacious start would be to initiate clinical use as a short-term bridge to allotransplantation, particularly in infants with single ventricle pathology and significant risk factors for first stage Norwood palliation. Infants with end-stage heart failure after first stage palliation would represent a second target population. Infants experience unacceptably high mortality and morbidity when placed on mechanical circulatory support as a bridge to allotransplant. Effectively bridging these vulnerable populations could promote acceptance of cardiac xenotransplantation, allowing indications and use to expand, e.g., by (I) bridging patients with failed second and third stage single ventricle disease, or (II) with complex biventricular CHD, or (III) those with a restrictive or dilated cardiomyopathy. Finally, there is a reasonable expectation that the immunologic privilege of infants will allow porcine heart xenotransplantation to be destination therapy for some patients. In summary, heart allotransplantation in infants offers superior outcomes when compared to three-stage single ventricle palliation, but there is a continual shortage of deceased human donor organs. We should pursue research towards the application of xenotransplantation in patients with single ventricle pathology, in whom the results of staged palliation are likely to be suboptimal. There are many remaining issues to be resolved before cardiac xenotransplantation enters regular pediatric clinical use, but experience in this field is progressing rapidly.
PubMed: 38197100
DOI: 10.21037/tp-22-664 -
Scientific Reports Jan 2024Lewis lung carcinoma (LLC), as a widely used preclinical cancer model, has still not been genetically and genomically characterized. Here, we performed a whole-exome...
Lewis lung carcinoma (LLC), as a widely used preclinical cancer model, has still not been genetically and genomically characterized. Here, we performed a whole-exome sequencing analysis on the LLC cell line to elucidate its molecular characteristics and etiologies. Our data showed that LLC originated from a male mouse belonging to C57BL/6L (a transitional strain between C57BL/6J and C57BL/6N) and contains substantial somatic SNV and InDel mutations (> 20,000). Extensive regional mutation clusters are present in its genome, which were caused mainly by the mutational processes underlying the SBS1, SBS5, SBS15, SBS17a, and SBS21 signatures during frequent structural rearrangements. Thirty three deleterious mutations are present in 30 cancer genes including Kras, Nras, Trp53, Dcc, and Cacna1d. Cdkn2a and Cdkn2b are biallelically deleted from the genome. Five pathways (RTK/RAS, p53, cell cycle, TGFB, and Hippo) are oncogenically deregulated or affected. The major mutational processes in LLC include chromosomal instability, exposure to metabolic mutagens, spontaneous 5-methylcytosine deamination, defective DNA mismatch repair, and reactive oxygen species. Our data also suggest that LLC is a lung cancer similar to human lung adenocarcinoma. This study lays a molecular basis for the more targeted application of LLC in preclinical research.
Topics: Male; Humans; Mice; Animals; Adenocarcinoma; Carcinoma, Lewis Lung; Exome Sequencing; Mice, Inbred C57BL; Mutation; Lung Neoplasms; Proto-Oncogene Proteins p21(ras)
PubMed: 38167599
DOI: 10.1038/s41598-023-50703-2 -
Circulation Research Jan 2024Single-nucleotide polymorphisms linked with the rs1474868 T allele ( [mitofusin-2] T/T) in the human mitochondrial fusion protein gene are associated with reduced...
BACKGROUND
Single-nucleotide polymorphisms linked with the rs1474868 T allele ( [mitofusin-2] T/T) in the human mitochondrial fusion protein gene are associated with reduced platelet RNA expression and platelet counts. This study investigates the impact of MFN2 on megakaryocyte and platelet biology.
METHODS
Mice with megakaryocyte/platelet deletion of ( [ conditional knockout]) were generated using Pf4-Cre crossed with floxed mice. Human megakaryocytes were generated from cord blood and platelets isolated from healthy subjects genotyped for rs1474868. Ex vivo approaches assessed mitochondrial morphology, function, and platelet activation responses. In vivo measurements included endogenous/transfused platelet life span, tail bleed time, transient middle cerebral artery occlusion, and pulmonary vascular permeability/hemorrhage following lipopolysaccharide-induced acute lung injury.
RESULTS
Mitochondria was more fragmented in megakaryocytes derived from mice and from human cord blood with T/T genotype compared with control megakaryocytes. Human resting platelets of T/T genotype had reduced MFN2 protein, diminished mitochondrial membrane potential, and an increased rate of phosphatidylserine exposure during ex vivo culture. Platelet counts and platelet life span were reduced in mice accompanied by an increased rate of phosphatidylserine exposure in resting platelets, especially aged platelets, during ex vivo culture. also decreased platelet mitochondrial membrane potential (basal) and activated mitochondrial oxygen consumption rate, reactive oxygen species generation, calcium flux, platelet-neutrophil aggregate formation, and phosphatidylserine exposure following dual agonist activation. Ultimately, mice showed prolonged tail bleed times, decreased ischemic stroke infarct size after cerebral ischemia-reperfusion, and exacerbated pulmonary inflammatory hemorrhage following lipopolysaccharide-induced acute lung injury. Analysis of SNPs in the iSPAAR study (Identification of SNPs Predisposing to Altered ALI Risk) identified a significant association between and 28-day mortality in patients with acute respiratory distress syndrome.
CONCLUSIONS
Mfn2 preserves mitochondrial phenotypes in megakaryocytes and platelets and influences platelet life span, function, and outcomes of stroke and lung injury.
Topics: Aged; Animals; Humans; Mice; Acute Lung Injury; Blood Platelets; Hemorrhage; Lipopolysaccharides; Mitochondria; Phosphatidylserines
PubMed: 38156445
DOI: 10.1161/CIRCRESAHA.123.322914 -
PloS One 2023The aim of this study was to compare the ability of demineralized (DMB) and decellularized (DCC) bovine bone granules to support bone regeneration in rat calvaria...
The aim of this study was to compare the ability of demineralized (DMB) and decellularized (DCC) bovine bone granules to support bone regeneration in rat calvaria critical-size defects. DMB and DCC were prepared using a previously published method. The granule size used ranged between 500 and 750 μm. A total of forty-eight Sprague-Dawley rats were divided into two groups (n = 24). A pair of 5 mm diameter defects were created on the calvaria of the rats in the right and left parietal bone in both groups. Group A animals received DMB granules and Group B received DCC granules in the right parietal defect side while the left parietal untreated defect acted as sham surgery for both groups. Four animals per group were euthanized in a CO2 chamber at day 7, 14 and 21 post-surgery and the calvaria implantation site biopsy harvested was subjected to osteogenic gene expression analysis. Another four animals per group were euthanized at days 15, 30 and 60 post surgery and the calvaria implantation site biopsy harvested was subjected to histological, immunohistochemistry, RAMAN spectroscopy and Micro-CT analysis at the mentioned time points. Statistical analysis was conducted using t-tests and ANOVA. Histomorphometry showed significantly higher new bone formation in the DCC sites (p<0.05) compared to DMB. Both DMB and DCC implantation sites showed distinct staining for osteocalcin and osteopontin proteins compared to their respective sham sites. By day 21 after implantation, DCC sites demonstrated significantly elevated mRNA levels of osteonectin (p<0.001), osteopontin (p<0.001), osteocalcin (p<0.0001), ALP (p<0.01), and BMP-2 (p<0.001) compared to DMB. However, VEGF expression showed no significant differences at this time point between the two groups. Micro-CT analysis also showed enhanced defect closure and higher bone density in DCC implanted sites while RAMAN spectra demonstrated increased abundance of collagen and bone minerals, especially, PO43- ions than DMB. In conclusion, both DMB and DCC granules demonstrated favorable osteogenic potential in critical-sized defects, with DCC exhibited superior osteoconductive, osteoinductive and osteogenesis properties.
Topics: Rats; Animals; Cattle; Osteogenesis; Rats, Sprague-Dawley; Osteopontin; Osteocalcin; Skull; Bone Regeneration; Minerals
PubMed: 38127838
DOI: 10.1371/journal.pone.0294291