RNF213 Mutation Research Articles

Loss of mitochondrial ClpP, Lonp1, and Tfam triggers transcriptional induction of Rnf213, a susceptibility factor for moyamoya disease

Human RNF213, which encodes the protein mysterin, is a known susceptibility gene for moyamoya disease (MMD), a cerebrovascular condition with occlusive lesions and compensatory angiogenesis. Mysterin mutations, together with exposure to environmental trigger factors, lead to an elevated stroke risk since childhood. Mysterin is induced during cell stress, to function as cytosolic AAA+ ATPase and ubiquitylation enzyme. Little knowledge exists, in which context mysterin is needed. Here, we found that genetic ablation of several mitochondrial matrix factors, such as the peptidase ClpP, the transcription factor Tfam, as well as the peptidase and AAA+ ATPase Lonp1, potently induces Rnf213 transcript expression in various organs, in parallel with other components of the innate immune system. Mostly in mouse fibroblasts and human endothelial cells, the Rnf213 levels showed prominent upregulation upon Poly(I:C)-triggered TLR3-mediated responses to dsRNA toxicity, as well as upon interferon gamma treatment. Only partial suppression of Rnf213 induction was achieved by C16 as an antagonist of PKR (dsRNA-dependent protein kinase). Since dysfunctional mitochondria were recently reported to release immune-stimulatory dsRNA into the cytosol, our results suggest that mysterin becomes relevant when mitochondrial dysfunction or infections have triggered RNA-dependent inflammation. Thus, MMD has similarities with vasculopathies that involve altered nucleotide processing, such as Aicardi-Goutières syndrome or systemic lupus erythematosus. Furthermore, in MMD, the low penetrance of RNF213 mutations might be modified by dysfunctions in mitochondria or the TLR3 pathway.

Mutations of RNF213 are responsible for sporadic cerebral cavernous malformation and lead to a mulberry-like cluster in zebrafish

Although familial forms of cerebral cavernous malformation are mainly attributed to three CCM genes (KRIT1, CCM2 and PDCD10), no mutation is identified in sporadic cerebral cavernous malformation cases with a unique lesion, indicating additional genes for sporadic cerebral cavernous malformation. To screen the candidate genes, we conducted whole exome sequencing in 31 sporadic cerebral cavernous malformation patients and 32 healthy controls, and identified 5 affected individuals carrying 6 heterozygous deleterious mutations in RNF213 but no RNF213 mutation in healthy individuals. To further confirm RNF213 was associated with cerebral cavernous malformation, we generated rnf213a homozygous knockout zebrafish and found mutation of rnf213a in zebrafish led to a mulberry-like cluster of disordered-flow vascular channels which was reminiscent of human cerebral cavernous malformation. In addition, we revealed kbtbd7 and anxa6 were significantly downregulated due to rnf213a mutation through transcriptomic sequencing and RT-qPCR analysis. Based on the mulberry-like phenotype partly rescued by mRNA of kbtbd7 as well as anxa6, we suggested that rnf213a promoted mulberry-like cluster via downregulation of kbtbd7 and anxa6. Altogether, we firstly demonstrate RNF213is a novel candidate gene for sporadic cerebral cavernous malformation and the mutation of rnf213a is responsible for the mulberry-like cluster in zebrafish.

Reversible Cerebral Angiopathy after Viral Infection in a Pediatric Patient with Genetic Variant of RNF213

Ring finger protein (RNF) 213 is known as a susceptibility gene for moyamoya disease (MMD), which is characterized by bilateral carotid folk stenosis. Cerebral angiopathy after viral infection has been known to present angiographical appearance resembling MMD, however its pathogenesis and genetic background are not well known. We report a case of reversible cerebral angiopathy after viral infection in a pediatric patient with genetic variant of RNF213 mutation. The patient had developed a severe headache after hand, foot, and mouth disease. Magnetic resonance imaging and magnetic resonance angiography (MRA) performed 2-3 weeks after disease onset revealed bilateral carotid folk stenosis and an old cerebral infarction in the left putamen. The patient's headache spontaneously resolved and the follow-up MRA showed a complete spontaneous resolution of the arterial stenosis after 9 months. We were able to determine genetic predisposition to angiopathy by identifying the RNF213 c.14576G>A (rs112735431, p.R4859K) mutation. Based on the present case, we hypothesize that an RNF213 variant might play an important role for the onset of postviral cerebral angiopathy.

Associations Between Loss of ARID1A Expression and Clinicopathologic and Genetic Variables in T1 Early Colorectal Cancer.

To evaluate the relationships between adenine-thymine-rich interactive domain 1A (ARID1A) expression and the clinicopathologic features in T1 colorectal cancer (CRC) and to investigate whether the presence of ARID1A protein is related to genetic changes. We retrospectively studied 219 surgically resected T1 CRCs. ARID1A expression was assessed by immunohistochemical methods, and the correlation between ARID1A expression and clinicopathologic features was evaluated. The relationship between ARID1A expression and 409 cancer-related gene mutations was also evaluated using next-generation sequencing (NGS). Immunohistochemical staining indicated negative ARID1A expression in 4.6%. Loss of ARID1A expression was significantly associated with younger age, lymphatic invasion, and lymph node metastasis (LNM). NGS showed that PKHD1, RNF213, and MSH6 mutations were more frequent in ARID1A-negative tumors, whereas KRAS mutations were more common in ARID1A-positive tumors. In T1 CRC, negative ARID1A expression was correlated with early onset, lymphatic invasion, and LNM. Mutations in some cancer-related genes were possibly related with ARID1A expression.

Abstract 4024: Phase 1/2a LYM1002 study of ibrutinib (ibr) + nivolumab (nivo): Exome and gene expression profiling (GEP) analyses by histology and responder status

Abstract A phase 1/2a study (LYM1002 [EudraCT 2014-005191-28]) of ibr (560 mg once daily) + nivo (3 mg/kg on a 14-day cycle) demonstrated acceptable safety and promising efficacy vs single-agent ibr in Richter’s transformation (RT), follicular lymphoma (FL), and diffuse large B-cell lymphoma (DLBCL). We examined potential biomarkers of treatment (tx) response using archived biopsy samples. GEP was used for DLBCL subtyping and to assess proportions of 22 distinct immune cells. Exome data were generated from 72 formalin-fixed paraffin-embedded samples, and sequencing analysis was used to identify mutations in genes of interest and assess somatic mutation burden. The correlations of immune cell proportions and gene variants were evaluated by investigator-assessed responses in each histology and by ongoing responses in DLBCL patients (pts; progression-free survival [PFS] > 24 months, n = 7 vs not, n = 20). In pts with available GEP and response data, overall response rates were 29.6% (8/27) for DLBCL, 43.5% (10/23) for FL and 81.3% (13/16) for RT. Proportions of CD8 and follicular helper T cells, M1 macrophages, and resting dendritic cells were higher in DLBCL responders vs nonresponders, while the proportion of regulatory T cells was decreased. These subsets did not differ by response in FL although an increase of CD4 memory resting T-cells was noted in responders. The trend toward increased follicular helper T cell and resting dendritic cell proportions in DLBCL pts was also associated with longer survival (PFS > 24 months) vs not. Gene variant data and responder status were available for 26 pts with DLBCL, 26 with FL, and 17 with RT. Comparison between responders and nonresponders showed that DLBCL pts with RNF213 (4/10 [40.0%] vs 1/16 [6.2%]) and KLHL14 (3/10 [30.0%] vs 0/16) mutations were more likely to respond to ibr + nivo. Conversely, nonresponders were associated with variants in EBF1, ADAMTS20, AKAP9, SOCS1, TP53, and genes in BCR pathways such as TNFRSF14, MYD88, and NFKB1B. BCL2 mutation in FL (9/12 [75.0%] vs 4/14 [28.6%]) and ROS1 mutation in RT (0/13 vs 2/4 [50.0%]) were associated with response; both are involved in the NF-kB pathway. In DLBCL, the most frequent gene mutations were RNF213, NBPF1, and BCL2 in pts who had PFS > 24 months (3/7 [42.9%] each), and KMT2D (8/20 [40.0%]) and CSMD3 (8/20 [40.0%]) in pts who did not. Somatic mutation burden was lower in responders vs nonresponders, especially in germinal center B-cell-DLBCL, and in DLBCL pts with PFS > 24 months vs not. In conclusion, we report gene variations among DLBCL, FL, and RT pts associated with response or durable PFS with ibr + nivo. While ibr inhibits Bruton’s tyrosine kinase-dependent pathways, we identify alternative gene pathway variants that may impact tx outcomes. Immune cell infiltration into the microenvironment relates to differential tx response with this immune combination and is histology dependent. Citation Format: Brendan Hodkinson, Michael Schaffer, Joshua Brody, Wojciech Jurczak, Cecilia Carpio, Dina Ben-Yehuda, Irit Avivi, Rao Saleem, Muhit Özcan, John Alvarez, Rob Ceulemans, Nele Fourneau, Sriram Balasubramanian, Anas Younes. Phase 1/2a LYM1002 study of ibrutinib (ibr) + nivolumab (nivo): Exome and gene expression profiling (GEP) analyses by histology and responder status [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4024.

Vascular tortuosity of the internal carotid artery is related to the RNF213 c.14429G\u2009>\u2009A variant in moyamoya disease

Recent studies have implicated RNF213 mutations in the pathogenesis of moyamoya disease (MMD). However, the underlying mechanism of disease development is not fully elucidated. Nonetheless, a possible relationship between vascular morphology and hemodynamics related with MMD has been proposed. Here, we aimed to investigate the relationship between a variant of RNF213 and the morphology of the internal carotid artery (ICA). We enrolled bilateral MMD patients who had undergone genetic testing for RNF213. Patients were divided into mutant and wild-type groups. Six anatomy-specific three-dimensional coordinates were collected using magnetic-resonance angiography. From these, five vectors between two adjacent points and four angles between two adjacent vectors were calculated. The tortuosity was defined as the ratio between the actual and the linear length of the ICAs. Among 58 patients, 44 and 14 belonged to the mutant and wild-type groups, respectively. The tortuosity of ICAs was significantly lower in the mutant group (p = 0.010). The change in blood flow direction was more prominent in the wild-type group (p = 0.002). The tortuosity was significantly lower in MMD patients than normal controls (p < 0.001). Our results indicate that RNF213 could play a role in the lower tortuosity observed in patients with RNF213 mutation.

Near-Comprehensive Resequencing of Cancer-Associated Genes in Surgically Resected Metastatic Liver Tumors of Gastric Cancer

Liver metastasis is a major cause of death in patients with gastric cancer. The molecular alterations in clinically resected liver metastases of gastric cancer were evaluated to identify candidate biomarkers and therapeutic targets. Seventy-four patients, including 37 with liver metastasis who underwent gastrectomy and hepatectomy for gastric cancer and 37 without liver metastasis who underwent gastrectomy for gastric cancer, were studied. Next-generation resequencing was performed for 412 cancer-associated genes in metastatic and/or primary tumors from 30 patients and somatic mutations in TP53, LRP1B, PIK3CA, ADAMTS20, PAX7, FN1, FOXO3, WRN, PTEN, ETV4, and RNF213 were found in metastatic tumors. TP53 mutations were studied by Sanger sequencing in the remaining patients; the number of patients with TP53 mutations in metastatic tumors was significantly higher among those with liver metastasis (86.5%, 32/37) versus those without liver metastasis (40.5%; 15/37; P<0.0001). TP53 mutations in metastatic liver tumors and corresponding primary tumors were identical in 96.9% (31/32), including some patients with heterogeneous primary tumor components. Immunohistochemical analyses showed aberrant p53 expression in tumors with TP53 mutations. In silico functional evaluations indicated functional loss of missense-mutated TP53. Thus, the p53 pathway may facilitate the development of biomarkers and therapeutic approaches to treat gastric cancer metastases to the liver.

Posterior circulation involvement and collateral flow pattern in moyamoya disease with the RNF213 polymorphism.

Moyamoya disease is a chronic cerebrovascular disorder characterized by progressive stenosis of the circle of Willis with a compensatory collateral vessel network. Recent studies have identified the ring finger protein 213 gene (RNF213) as the unique susceptibility gene for moyamoya disease. The purpose of this study was to compare clinical features of moyamoya disease, especially angiographic findings, between patients with and without the RNF213 mutation. Blood samples from 35 patients with moyamoya disease were obtained between May 2016 and May 2017. Information on age at the time of diagnosis, sex, and initial symptom were obtained via retrospective chart review. Angiographic records were evaluated. RNF213 variants were detected in the 28 of 35 patients (80%), including all pediatric patients (100%) and 18 of 25 adult patients (72%) in our cohort. Leptomeningeal collateral flow from posterior to anterior circulation was more frequent in the RNF213-negative group than in the RNF213-positive group (100% versus 38.9%; p = 0.020). Posterior cerebral arterial territorial involvement was more frequently observed in RNF213-positive patients than in RNF213-negative patients (50% versus 0%; p = 0.027). RNF213 may play a significant role in the development of collateral anastomoses.

Mutational landscapes of tongue carcinoma reveal recurrent mutations in genes of therapeutic and prognostic relevance.

BackgroundCarcinoma of the oral tongue (OTSCC) is the most common malignancy of the oral cavity, characterized by frequent recurrence and poor survival. The last three decades has witnessed a change in the OTSCC epidemiological profile, with increasing incidence in younger patients, females and never-smokers. Here, we sought to characterize the OTSCC genomic landscape and to determine factors that may delineate the genetic basis of this disease, inform prognosis and identify targets for therapeutic intervention.MethodsSeventy-eight cases were subjected to whole-exome (n = 18) and targeted deep sequencing (n = 60).ResultsWhile the most common mutation was in TP53, the OTSCC genetic landscape differed from previously described cohorts of patients with head and neck tumors: OTSCCs demonstrated frequent mutations in DST and RNF213, while alterations in CDKN2A and NOTCH1 were significantly less frequent. Despite a lack of previously reported NOTCH1 mutations, integrated analysis showed enrichments of alterations affecting Notch signaling in OTSCC. Importantly, these Notch pathway alterations were prognostic on multivariate analyses. A high proportion of OTSCCs also presented with alterations in drug targetable and chromatin remodeling genes. Patients harboring mutations in actionable pathways were more likely to succumb from recurrent disease compared with those who did not, suggesting that the former should be considered for treatment with targeted compounds in future trials.ConclusionsOur study defines the Asian OTSCC mutational landscape, highlighting the key role of Notch signaling in oral tongue tumorigenesis. We also observed somatic mutations in multiple therapeutically relevant genes, which may represent candidate drug targets in this highly lethal tumor type.Electronic supplementary materialThe online version of this article (doi:10.1186/s13073-015-0219-2) contains supplementary material, which is available to authorized users.

Abstract 3874: Mutational landscapes of oral tongue squamous cell carcinoma reveal recurrent mutations in genes of therapeutic and prognostic relevance

Abstract Carcinoma of the oral tongue (OTSCC) is the most common malignancy of the oral cavity, characterized by frequent recurrence and poor survival. The last three decades has witnessed a change in the OTSCC epidemiological profile, with increasing incidence in younger patients, females and never-smokers. This study aims to characterize the OTSCC genomic landscape and to determine factors that may delineate the genetic basis of this disease, inform prognosis and identify targets for therapeutic intervention. Seventy-eight Asian OTSCC cases were subjected to whole-exome and targeted deep sequencing. While the most common mutation seen was in TP53, the OTSCC genetic landscape differed from previously described cohorts of patients with head and neck squamous cell cancers: OTSCCs demonstrated frequent mutations in DST, FSIP2 and RNF213 while alterations in CDKN2A and NOTCH1 were significantly less frequent than previously reported. Despite a lack of previously-known NOTCH1 mutations, integrated analysis showed enrichments of alterations affecting Notch signaling, more commonly seen in women. Importantly, Notch pathway alterations were prognostic on uni- and multivariate analyses. Furthermore, a high proportion of OTSCCs presented with alterations in drug targetable genes and chromatin remodeling genes. Mutations in these groups were also prognostic, where patients harboring mutations in these actionable pathways more likely to succumb from recurrent disease compared to those who did not, suggesting that these patients should indeed be considered for treatment with targeted compounds in future trial designs. In conclusion, these findings define the mutational landscape of OTSCC, highlight the key role of the Notch signaling pathway in oral tongue tumorigenesis and uncover somatic mutations in therapeutically relevant genes which may represent candidate drug targets. Citation Format: Andre L. Vettore, Kalpana Ramnarayanan, Choon Kiat Ong, Hui Sun Leong, Weng Khong Lim, Ioana Cutcutache, John R. Mcpherson, Shenli Zhang, Thankshayeni Skanthakumar, Daniel SW Tan, Bin Tean Teh, Steve Rozen, Patrick Tan, N Gopalakrishna Iyer. Mutational landscapes of oral tongue squamous cell carcinoma reveal recurrent mutations in genes of therapeutic and prognostic relevance. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3874. doi:10.1158/1538-7445.AM2015-3874

Mutation genotypes of RNF213 gene from moyamoya patients in Taiwan

Moyamoya disease (MMD) is a disorder characterized by stenosis of bilateral internal carotid arteries with compensatory angiogenesis of the perforating blood vessels. Familial transmission in MMD is common. Recently, mutations in human RNF213 and ACTA2 genes were identified to be responsible for MMD. The present study was to determine whether Taiwanese MMD patients carried mutations in these two genes. Of the 36 MMD patients, eleven was found to have RNF213 mutations. Direct genetic sequencing identified four different RNF213 mutations in the 11 patients from 8 families: five with a p.R4810K, one with p.A1622V, one with p.V3933M, and the other one with p.R4131C. The latter three represent novel missense mutations. No mutation in ACTA2 gene was identified. Clinically, cerebral infarction was common in patients with an RNF213 mutation (9/11). In addition, four mutant patients had developmental delay (4/11) and two had mental dysfunction (2/11). The magnetic resonance angiography of asymptomatic mutant carriers demonstrated high incidence of multiple stenosis of intracranial vessels (3/6, 50%). Since 30.6% (11/36) of Taiwanese moyamoya patients carry an RNF213 mutation and intracranial arterial stenosis was found in half of the asymptomatic mutant carriers, it is suggested that the RNF213 mutation should form part of the diagnostic workup for MMD in clinical practice.

Molecular Analysis of RNF213 Gene for Moyamoya Disease in the Chinese Han Population

BackgroundMoyamoya disease (MMD) is an uncommon cerebrovascular disorder characterized by progressive occlusion of the internal carotid artery causing cerebral ischemia and hemorrhage. Genetic factors in the etiology and pathogenesis of MMD are being increasingly recognized. Previous studies have shown that the RNF213 gene was related to MMD susceptibility in the Japanese population. However, there is no large scale study of the association between this gene and MMD in the Chinese Han population. Thus we designed this case-control study to validate the R4810K mutation and to define the further spectrum of RNF213 mutations in Han Chinese.Methodology/Principal FindingsGenotyping of the R4810K mutation in the RNF213 gene was performed in 170 MMD cases and 507 controls from a Chinese Han population. The R4810K mutation was identified in 22 of 170 MMD cases (13%), including 21 heterozygotes and a single familial homozygote. Two of the 507 controls (0.4%) were heterozygous R4810K carriers. The R4810K mutation greatly increased the risk for MMD (OR = 36.7, 95% CI: 8.6∼156.6, P = 6.1 E-15). The allele frequency of R4810K was significantly different between patients with ischemia and hemorrhage (OR = 5.4, 95% CI: 1.8∼16.1, P = 0.001). Genomic sequencing covering RNF213 exon 40 to exon 68 also identified eight other non-R4810K variants; P4007R, Q4367L, A4399T, T4586P, L4631V, E4950D, A5021V and M5136I. Among them A4399T polymorphism was found in 28/170 cases (16.5%) and 45/507 controls (8.9%) and was associated with MMD (OR = 2.0, 95% CI: 1.2∼3.3, P = 0.004), especially with hemorrhage (OR = 2.8, 95% CI: 1.2∼6.5, P = 0.014).Conclusions RNF213 mutations are associated with MMD susceptibility in Han Chinese. The ischemic type MMD is particularly related to the R4810K mutation. However, A4399T is also a susceptible variant for MMD, primarily associated with hemorrhage. Identification of novel variants in the RNF213 gene further highlights the genetic heterogeneity of MMD.