TGFBR2 Mutations Research Articles

Cholangiocarcinoma Heterogeneity Revealed by Multigene Mutational Profiling: Clinical and Prognostic Relevance in Surgically Resected Patients.

Cholangiocarcinoma can be classified in intrahepatic cholangiocarcinoma (ICC) and perihilar cholangiocarcinoma (PCC). Moreover, PCC includes two different forms: extrahepatic (EH) PCC, which arises from the perihilar EH large ducts, and intrahepatic (IH) PCC, in which a significant liver mass invades the perihilar bile ducts. In this study, we investigated the molecular profile and molecular prognostic factors in EH-PCC, IH-PCC, and ICC submitted to curative surgery. Ninety-one patients with cholangiocarcinoma (38 EH-PCC, 18 IH-PCC, and 35 ICC), who underwent curative surgery in a single tertiary hepatobiliary surgery referral center were assessed for mutational status in 56 cancer-related genes. The most frequently mutated genes in EH-PCC were KRAS (47.4%), TP53 (23.7%) and ARID1A (15.8%); in IH-PCC were KRAS (22.2%), PBRM1 (16.7%), and PIK3CA (16.7%); and in ICC were IDH1 (17.1%), NRAS (17.1%), and BAP1 (14.3%). The presence of mutations in ALK, IDH1, and TP53 genes was significantly associated with poor prognosis in patients with EH-PCC (p<0.001, p=0.043, and p=0.019, respectively). Mutation of the TP53 gene was significantly associated with poor prognosis in patients with IH-PCC (p=0.049). The presence of mutations in ARID1A, PIK3C2G, STK11, TGFBR2, and TP53 genes was significantly associated with poor prognosis in patients with ICC (p=0.012, p=0.030, p=0.030, p=0.011, and p=0.011, respectively). Mutational gene profiling identified different gene mutations in EH-PCC, IH-PCC, and ICC. Moreover, our study reported specific prognostic genes that can identify patients with poor prognosis after curative surgery who may benefit from traditional or target adjuvant treatments.

Genetic heterogeneity in synchronous colorectal cancers impacts genotyping approaches and therapeutic strategies.

Synchronous colorectal carcinomas (sCRC) are clinically challenging neoplasms. Although the epidemiological characteristics are quite well established, their biological basis is still poorly understood. Hence, we performed comprehensive molecular profiling of 23 sCRC cases comprising 50 synchronous primary tumors, 5 metastases, and corresponding normal tissue by targeted deep sequencing of 30 CRC-related genes, microsatellite analysis and analysis for methylated MLH1. We identified a striking inter- and intratumoral genetic heterogeneity of sCRC. Twenty (87%) cases showed genetic heterogeneity leaving only three cases with tumors that had an identical genetic make-up. Intertumoral heterogeneity was frequently observed for clinically actionable genes, including KRAS. Specifically, 44% of the cases harbored tumors of which at least one was KRAS mutated and the other KRAS wildtype. Moreover, 48% of the cases had at least double, sometimes even triple or quadruple mutations in KRAS, APC, TP53, PIK3CA, and TGFBR2, most of them being subclonal events. Lastly, we detected four cases (17%) with microsatellite instable (MSI) tumors with one case harboring one MSI- and a distinct microsatellite stable carcinoma. Our data demonstrate a striking genetic heterogeneity not only between different sCRC of a single case but also within a single tumor. These results contribute to the biological understanding of sCRC and directly impact genotyping strategies and oncological decision making. Testing one tumor or a single metastasis may not suffice in the sCRC setting as clinically relevant and tumor-specific genetic information may be left undetected compromising optimal oncological therapy. © 2015 Wiley Periodicals, Inc.

Abstract 16575: Arterial Tortuosity is Present in Multiple Genetic Conditions Associated With Aortopathy, a Study From the National Registry of Genetically Triggered Thoracic Aortic Aneurysms and Cardiovascular Conditions (GenTAC)

Background: Arterial tortuosity is emerging as an imaging marker of adverse events in aortopathy, but quantification has focused on patients with Marfan and Loeys-Dietz syndromes. Our objective was to characterize vertebral artery tortuosity among a variety of aortopathy phenotypes and genotypes. Methods: We performed a retrospective analysis of patients ≤50 years old with at least one of the following: aortic dilation or dissection, bicuspid aortic valve (BAV), or a genotype associated with aortopathy. Patients included were either enrolled in GenTAC or were cared for at our institution, and had undergone a magnetic resonance or computed tomography angiogram that included the vertebral arteries. The disease-causing gene mutation was available for a subset of the patients. The vertebral artery tortuosity index (VTI) was utilized to quantify tortuosity, and was compared among phenotypes and genotypes. Increased tortuosity was defined as ≥2SD above the mean VTI of previously reported controls undergoing evaluation to exclude arrhythmogenic right ventricular dysplasia (mean 4.5±2.5, cutoff=10.5). Results: We included 456 patients, of median age 29 years (IQR 15-40 years), and 58% male. The figure shows VTI by phenotype and genotype. VTI was higher in all phenotypic groups than controls (p ≤0.01 for all). All genotypes (for which n&gt;1) had increased VTI compared to controls (p≤0.01 for all). Patients with TGFBR2 mutations had significantly greater VTI (median 59, IQR 20-85) than those with FBN1 mutations (median 25, IQR 13-51, p=0.01); there was no difference in VTI between TGFBR1 (median 19, IQR 12-64) and FBN1 mutations (p=0.79). There was no overall correlation between age and VTI; when evaluated by diagnosis, there was a weak correlation in MFS (Spearman's rho=0.213, p=0.01). Conclusions: Increased arterial tortuosity as measured by VTI is present in many phenotypes and genotypes associated with aortopathy, and is not limited to those with mutations in TGFBR1/2.

Arterial tortuosity in genetic arteriopathies.

Arterial tortuosity is emerging as a common feature in genetically mediated thoracic aortic disease that may be prognostic. This review will summarize recent literature on arterial tortuosity in the setting of genetic arteriopathies. Although arterial tortuosity has been primarily described in Loeys-Dietz syndrome due to TGFBR1 and TGFBR2 mutations and in arterial tortuosity syndrome due to SLC210A mutations, recent studies that use quantitative measures of tortuosity suggest that tortuosity is present in many other genetic conditions associated with aortic dilation and dissection. The mechanisms of the development of tortuosity in these disorders are not fully understood, but are founded in the concept that there is abnormal, pathologic arterial lengthening in a fixed space, resulting in more tortuous vessels. Further studies suggest that patients with increased arterial tortuosity are at increased risk of adverse cardiovascular events, including aortic surgery, aortic dissection, and death. Arterial tortuosity is commonly present in genetically mediated aortic disease. Given the suboptimal performance of aortic dimension alone in predicting aortic dissection, quantification of tortuosity may augment the current algorithms for determining risk in patients with aortic disease.

Clonal origins and parallel evolution of regionally synchronous colorectal adenoma and carcinoma.

Although the colorectal adenoma-to-carcinoma sequence represents a classical cancer progression model, the evolution of the mutational landscape underlying this model is not fully understood. In this study, we analyzed eight synchronous pairs of colorectal high-grade adenomas and carcinomas, four microsatellite-unstable (MSU) and four-stable (MSS) pairs, using whole-exome sequencing. In the MSU adenoma-carcinoma pairs, we observed no subclonal mutations in adenomas that became fixed in paired carcinomas, suggesting a 'parallel' evolution of synchronous adenoma-to-carcinoma, rather than a 'stepwise' evolution. The abundance of indel (in MSU and MSS pairs) and microsatellite instability (in MSU pairs) was noted in the later adenoma- or carcinoma-specific mutations, indicating that the mutational processes and functional constraints operative in early and late colorectal carcinogenesis are different. All MSU cases exhibited clonal, truncating mutations in ACVR2A, TGFBR2, and DNA mismatch repair genes, but none were present in APC or KRAS. In three MSS pairs, both APC and KRAS mutations were identified as both early and clonal events, often accompanying clonal copy number changes. An MSS case uniquely exhibited clonal ERBB2 amplification, followed by APC and TP53 mutations as carcinoma-specific events. Along with the previously unrecognized clonal origins of synchronous colorectal adenoma-carcinoma pairs, our study revealed that the preferred sequence of mutational events during colorectal carcinogenesis can be context-dependent.

Abstract 2301: The generation of colorectal cancer mouse model based on microsatellite instability and the identification of transforming growth factor-beta signal target

Abstract Background &amp; Aims: The transforming growth factor-beta (TGF-beta) signal is a tumor-suppressor pathway that is commonly inactivated in about 90% of microsatellite instability (MSI) colorectal cancer (CRC). However, there was little evidence what gene is regulated by TGF-beta signal in the multistep progression sequence of CRC. The first aim of the present study was to generate a mouse model that is null for Tgfbr2 and Apc in the colon epithelium and forms tumors in the colon. The second aim was to analyze the tumors that arose in the mice model for the purpose to identify the gene regulated by TGF-beta signal. Method &amp; Result: Previously we have described the generation of the ‘CDX2P-G19Cre;Apcflox/flox mice’ (called Apc KO mice) which is randomly null for Apc in the colonic epithelium. By mating Tgfbr2flox/flox mice with Apcflox/flox and CDX2P9.5-G19Cre mice, we have finally generated a mouse model ‘CDX2P-G19Cre;Apcflox/flox;Tgfbr2flox/flox mice’ (called Apc+Tgfbr2 KO mice) which is null for Apc and Tgfbr2. In these model, the tumors with well differentiated adenocarcinoma arose mainly in proximal colon and most of mice died at 4 weeks age due to tumor bleeding. Therefore the mice were harvested at 3 weeks age to evaluate the development of colon tumors. Total RNAs of only cancerous tissue areas were extracted from frozen samples by the laser capture microdissection method. We compared gene expression profiles of these mice's tumors (n = 3, respectively) with Mouse Exon 1.0 ST Array (Affymetrix). Gene X expression of Apc+Tgfbr2 KO mice tumors was most highly upregurated by 9.25-fold compared with Apc KO mice (p = 0.045). The array data was validated by quantitative PCR. For human CRC samples, mutations of repetitive mononucleotide tracts in the coding regions of TGFBR2 were identified by direct sequencing. By immunohistochemical analysis, the expression of X was classified according to the percentage of stained cancer cells. The expression was considered to be ‘positive’ if ≥30% of cancer cells were stained. An analysis demonstrated that 11 (100%) of 11 mutated TGFBR2 cases were positive for X, whereas 10 (66.7%) of 15 wild type TGFBR2 cases were positive (P = 0.033), indicating that high expression of X was correlated with TGFBR2 mutation in human CRCs samples. Additionally, the cell proliferation assay revealed that silencing of X led to a significant reduction in CRC cell proliferation. Conversely, forced expression of X enhanced CRC cell proliferation in vitro. Conclusion: We have generated an in vivo model system that Apc and Tgfbr2 were inactivated only in the colonic epithelium and tumors with well differentiated adenocarcinoma arose mainly in proximal colon. The analysis of this model revealed that Gene X is regulated by a TGF-beta signal and likely promotes cell proliferation in CRC. Citation Format: Masashi Miguchi, Takao Hinoi, Manabu Shimomura, Tomohiro Adachi, Yasufumi Saito, Hiroaki Niitsu, Masatoshi Kochi, Yusuke Sotomaru, Hideaki Ijichi, Tsuneo Ikenoue, Kunitoshi Shigeyasu, Kohji Tanakaya, Kazuhiro Sentani, Naohide Oue, Wataru Yasui, Hideki Ohdan. The generation of colorectal cancer mouse model based on microsatellite instability and the identification of transforming growth factor-beta signal target. [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 2301. doi:10.1158/1538-7445.AM2015-2301

Transforming Growth Factor β Signaling in Colorectal Cancer Cells With Microsatellite Instability Despite Biallelic Mutations in TGFBR2

Most colorectal cancer (CRC) cells with high levels of microsatellite instability (MSI-H) accumulate mutations at a microsatellite sequence in the gene encoding transforming growth factor β receptor II (TGFBR2). TGFβ signaling therefore is believed to be defective in these tumors, although CRC cells with TGFBR2 mutations have been reported to remain sensitive to TGFβ. We investigated how TGFβ signaling might continue in MSI-H CRC cells. We sequenced the 10-adenines microsatellite sequence in the TGFBR2 gene of 32 MSI-H colon cancer tissues and 6 cell lines (HCT116, LS180, LS411N, RKO, SW48, and SW837). Activation of TGFβ signaling was detected by SMAD2 phosphorylation and through use of a TGFβ-responsive reporter construct in all CRC cell lines. Transcripts of TGFBR2 were knocked-down in CRC cells using short hairpin RNA. Full-length and mutant forms of TGFBR2 were expressed in LS411N cells, which do not respond to TGFβ, and their activities were measured. SMAD2 was phosphorylated in most MSI-H CRC tissues (strong detection in 44% and weak detection in 34% of MSI-H tumors). Phosphorylation of SMAD2 in MSI-H cells required TGFBR2—even the form encoding a frameshift mutation. Transcription and translation of TGFBR2 with a 1-nucleotide deletion at its microsatellite sequence still produced a full-length TGFBR2 protein. However, protein expression required preservation of the TGFBR2 microsatellite sequence; cells in which this sequence was replaced with a synonymous nonmicrosatellite sequence did not produce functional TGFBR2 protein. TGFβ signaling remains active in some MSI-H CRC cells despite the presence of frameshift mutations in the TGFBR2 gene because the mutated gene still expresses a functional protein. Strategies to reactivate TGFβ signaling in colorectal tumors might not be warranted, and the functional effects of mutations at other regions of microsatellite instability should be evaluated.

Gene panel sequencing in heritable thoracic aortic disorders and related entities - results of comprehensive testing in a cohort of 264 patients.

BackgroundHeritable Thoracic Aortic Disorders (H-TAD) may present clinically as part of a syndromic entity or as an isolated (nonsyndromic) manifestation. About one dozen genes are now available for clinical molecular testing. Targeted single gene testing is hampered by significant clinical overlap between syndromic H-TAD entities and the absence of discriminating features in isolated cases. Therefore panel testing of multiple genes has now emerged as the preferred approach. So far, no data on mutation detection rate with this technique have been reported.MethodsWe performed Next Generation Sequencing (NGS) based screening of the seven currently most prevalent H-TAD-associated genes (FBN1, TGFBR1/2, TGFB2, SMAD3, ACTA2 and COL3A1) on 264 samples from unrelated probands referred for H-TAD and related entities. Patients fulfilling the criteria for Marfan syndrome (MFS) were only included if targeted FBN1 sequencing and MLPA analysis were negative.ResultsA mutation was identified in 34 patients (13%): 12 FBN1, one TGFBR1, two TGFBR2, three TGFB2, nine SMAD3, four ACTA2 and three COL3A1 mutations. We found mutations in FBN1 (N = 3), TGFBR2 (N = 1) and COL3A1 (N = 2) in patients without characteristic clinical features of syndromal H-TAD. Six TAD patients harboring a mutation in SMAD3 and one TAD patient with a TGFB2 mutation fulfilled the diagnostic criteria for MFS.ConclusionNGS based H-TAD panel testing efficiently reveals a mutation in 13% of patients. Our observations emphasize the clinical overlap between patients harboring mutations in syndromic and nonsyndromic H-TAD related genes as well as within syndromic H-TAD entities, justifying a widespread application of this technique.Electronic supplementary materialThe online version of this article (doi:10.1186/s13023-014-0221-6) contains supplementary material, which is available to authorized users.

Abstract 18498: Description of Aortic Root Growth and Outcomes in a Cohort of Pediatric Patients with Loeys-Dietz Syndrome

Background: Loeys Dietz Syndrome (LDS) is associated with rapid aortic dilation and aortic dissection, but data on children with LDS are limited. The goal of this study is to describe aortic root growth and outcomes in children with LDS. Methods: Patients with LDS were identified from an institutional database. Data regarding genetic mutation, medications, aortic root dimensions by transthoracic echocardiography (TTE), aortic dissection and surgical intervention were collected. For those with &gt;2 TTEs 1 year apart, rate of change in z-score was calculated using linear regression. TTEs performed after aortic surgery were excluded. We examined if variables were associated with rate of aortic root growth. Results: Of 16 patients, 8 were female. Five had a TGFBR1 and 9 had a TGFBR2 mutation; 2 patients did not have genetic data available. Median aortic root Z-score at diagnosis was 3.5 (range 0.5-25.4). Fifteen patients were on medication (2 beta-blocker (BB), 5 angiotensin receptor blocker (ARB), 5 BB+ARB, and 3 with past use of both). Four patients underwent prophylactic root replacement at ages 3.3, 6.7, 8.7, and 9.4 years at root dimensions 3.2, 6.5, 4.0, and 4.1 cm respectively. One patient had a Type A dissection at age 15y after prior root replacement and underwent repeat surgery. Another underwent heart transplant at 6.9 years old after prior root replacement. Ten patients had serial TTE data. Median change in aortic root diameter and Z-score was 0.11cm/year and 0.1/year respectively. Mean change in z-score per year for those on on BB was -0.1± (range -1.2 to 0.7), ARB 0.5 (range 0.1 to 1.1) and both 0.0 (range -0.2 to 0.2, p=NS). No variables studied were associated with faster aortic growth. Conclusions: Degree of aortic root dilation and rate of aortic root growth is highly variable in children with LDS, although factors associated more aggressive disease are unclear. The high proportion of patients with adverse outcomes including aortic dissection and surgery is concerning.

G.P.148: FHL1 mutations are causing familial aortic and other arterial aneurysms with scapuloperoneal myopathy

Familial aortic and other arterial aneurysms have been associated with Marfan syndrome, usually related to mutation in FBN1, and mutations in TGFBR1, TGFBR2, ACTA2, MYH11, and SMAD3. FHL1 mutations have been associated with various disorders that include reducing body myopathy (RBM), Emery–Dreifuss-like muscular dystrophy, isolated hypertrophic cardiomyopathy, and some overlapping conditions. So far no aortic diseases have been reported in these patients. Here we report a German family with a scapuloperoneal myopathy and aortic and other arterial aneurysms in two patients. Patient 1 had an aneurysm of the sinus of Valsalva and of the carotid artery, and mitral and tricuspidal valve prolapse. He received artery stenting of the carotid artery and aortic aneurysm repair. Main clinical neuromuscular features in this index patient were elevated CK levels, rigid spine, and Achilles tendon contractures. The muscle biopsy did not reveal protein aggregation. Patient 2 is also diagnosed with an aneurysm of the sinus of Valsalva and cardiac arrhythmia, but did not present any neuromuscular symptoms. Of note is that both patients presented a hallux abducto valgus deformity. We identified a novel mutation in the FHL1 gene (IVS6+1, c688+1: G>A) in four family members. The mutations were hemizygote in the two affected male patients and heterozygote in two unaffected female family members. These phenotypes provide evidence that mutations in FHL1 not only lead to neuromuscular symptoms, but also can be associated with aortic and other arterial aneurysms. This data suggest that all individuals with pathogenic FHL1 mutations should be assessed for aortic disease. Further studies are necessary to elucidate the factors responsible for the different phenotypes associated with these mutations in these patients.

Identification and Surgical Repair of Familial Thoracic Aortic Aneurysm and Dissection Caused by TGFBR1 Mutation

This study aimed at exploring the causative gene and summarizing the clinical characteristics in a Chinese thoracic aortic aneurysm and dissection (TAAD) family. Family members were examined for features of syndromic genetic diseases by clinician and geneticist. Genomic DNA was extracted from 2 distantly related members with definite TAAD for exome sequencing. A pathogenic mutation (rs111426349, c.1459C >T) of transforming growth factor β receptor 1 (TGFBR1) was confirmed, which result in the amino acid substitution p.R487W. Fourteen TGFBR1 mutation carriers were detected among 39 tested members in this family. The average age at diagnosis of aortic root dilatation or aneurysm was 23.2 ± 12.6 years (range 3-37 years). Early onset of aortic root dilatation was significant in this family without reported phenotypes. The David procedure was performed prophylactically in 3 carriers of this family. Familial TAAD caused by TGFBR1 mutation (c.1459C >T) was confirmed in a large Chinese Han ethnic family using exome sequencing. Aggressively prophylactic David procedure may be not necessary at a smaller aortic size in familial TAAD patients with TGFBR1 mutation and further observation is warranted.

Whole exome sequencing for the identification of a new mutation in TGFB2 involved in a familial case of non-syndromic aortic disease

BackgroundNon-syndromic aortic disease (NSAD) is a frequently asymptomatic but potentially lethal disease characterised by familial cases of thoracic aortic aneurysms and dissections. This monogenic but genetically heterogeneous condition is primarily inherited as an autosomal dominant disorder with low penetrance and variable expression. Mutations in ACTA2, TGFBR1, TGFBR2, MYH11, SMAD3, MYLK, and FBN1 genes have been described but still, there are many unresolved familial cases. MethodsThe whole exome of two distantly related and affected members of a Spanish family with multiple cases of NSAD was analysed through 5500 SOLiD™ System for the identification of shared and putative pathogenic variants. ResultsA new mutation termed c.C1042T:p.R348C (NM_001135599.2) was identified in TGFB2, a gene located in an evolutionary highly conserved region (Chr1: 218,519,577-218,617,961) that has been recently connected to this disease. The analysis of other family members using capillary sequencing confirmed cosegregation of the mutation with the disease and its incomplete penetrance. ConclusionsThe repeated implication of TGFB2 in the development of thoracic aortic aneurysms and dissections suggests that this gene should be considered during genetic diagnosis of this disease. An accurate diagnosis of affected individuals and additional family members at risk allows for a personalised and more efficient gene-based follow-up and treatment. Finally, the reiterative presence of common musculoskeletal and craniofacial additional features in patients with TGFB2 mutations suggests the existence of a new yet undefined connective tissue syndrome responsible for not only aortic dilation, but also for the other extracardiac alterations present in the affected patients.

Immune perturbation in patients with TGFbeta pathway defects (LYM7P.729)

Abstract Intro: Mutations in TGFbR1, TGFbR2 and SMAD3 are associated with familial thoracic aortic aneurysms and aortic dissections (TAAD). These patients offer an opportunity to study their immune development. Methods: PBMC from TAAD (n=9) and controls (CT, n=8) were analyzed by FACS. Th1 (IFNg) and Th17 (IL17A) were determined with intracellular cytokine staining. Foxp3+ Tregs were detected with anti-Foxp3. CD19+ were analyzed for naive (IgD+CD27-), unswitched (IgD+CD27+) and switched memory (IgD-CD27+). Plasmacytoid (CD303+pDC) and myeloid (CD1c+mDC) were defined within lineage-1 negative population. Results: %CD3-CD16+NK, CD3+CD16+NKT, CD4+, CD8+ and CD4+CD45RA+ in TAAD were similar to HC. Average %CD19+ (20.8vs7.3, p=0.006) and naive B cells (81.3vs66.6, p=0.004) were higher in TAADvsCT. The unswitched were similar but the switched B cells were lower (8.6vs15.5, p=0.01). While the %Tregs were similar, there was a remarkable reduction (1/2-3 folds, p&amp;lt;0.05) in Foxp3 concentration based on median fluorescence intensity of Foxp3 in TAAD. There was a significant reduction of %Th17 (0.14vs0.61, p=0.01), while the Th1 were similar. %pDC (9.4vs24.1, p=0.009) and %mDC (11.4vs17.9, p=0.01) were also lower in TAAD. Conclusions: These results demonstrate the involvement of TGFbeta signaling in B cells, DCs, Th17 and Treg development. Further studies and monitoring of the clinical effects of these immunological perturbations are needed to appreciate the impact of their underlying disease.

FKBP14‐related Ehlers‐Danlos syndrome: Expansion of the phenotype to include vascular complications

Biallelic mutations in FKBP14 cause a recessive form of Ehlers-Danlos syndrome (EDS) characterized by progressive kyphoscoliosis, myopathy, and hearing loss. To date, four children and one adult with this condition have been reported. We recently identified a 42-year-old man with severe kyphoscoliosis, restrictive/obstructive lung disease, short stature, mild hearing loss, decreased muscle mass, and a dissection of the celiac artery at age 41. He also had complete occlusion of the superior mesenteric artery with compensatory flow through an enlarged and tortuous inferior mesenteric artery. He was homozygous for a previously identified FKBP14 mutation, c.362dupC, p.(Glu122Argfs*7). He had no mutations in COL3A1, ACTA2, TGFBR1, TGFBR2, or SMAD3. The FKBP14 mutations in our patient occurred on the same haplotype as others with this same mutation. Although one family member in a previous report was thought to have early vascular complications, it could not be confirmed that she had biallelic mutations in FKBP14. This report expands the phenotype of FKBP14-related EDS to include risk for vascular complications and also raises the question of whether the shared haplotype represents a risk allele or founder mutation.

In silico analysis of the potential effects of the disease-associated point mutations on the kinase activity of TGFBR1 and TGFBR2 receptors

Annick Barre, Jean-Philippe Borges, Stephanie Caze-Subra, Camille Gironde, Pierre Rouge Universite de Toulouse; UPS; UMR 152 Pharma-Dev; Universite Toulouse 3, Faculte des Sciences Pharmaceutiques, F-31062 Toulouse cedex 09, France Institut de Recherche pour le Developpement (IRD); UMR 152 Pharma-Dev; F-31062 Toulouse cedex 09, France TGFBR1 and TGFBR2 mutations associated with Marfan syndrome-related diseases have been recently suspected to play a key role in triggering the immune changes responsible for allergic diseases. In this respect, patients suffering from Marfan and Loeys-Dietz syndromes exhibited higher rates of (food) allergies, compared to other allergic patients. A possible common mechanism underlying both disorders should consist of the altered ability of TGBR1 and TGFBR2 receptors to properly respond to the recognition of the TGFβ signal, thus preventing the TGFβ−signaling pathway from operating correctly. To check such a hypothesis, we performed an in silico analysis to predict the effects of point mutations in TGFBR1 and TGFBR2 receptors associated to the Marfan and Loeys-Dietz syndromes, on the kniase activity of the serine-kinase domain of both receptors. The most frequently observed disease-related point mutations in TGFBR1 and TGFBR2 receptors essentially occur at hot spots that are located in the surrounding vicinity of the catalytic loop of the kinase domain. They are susceptible to modify the local conformation of the catalytic loop and thus to alter the phosphorylative activity of the kinase domain for the corresponding substrates, e.g. the SMAD proteins. Additionally, another point mutation associated to the Loeys-Dietz disease should modify the local conformation close to the P glycin-rich loop of the kinase domain, involved in the anchorage of an ATP molecule which serves as a phosphate-group donor necessary for the phosphorylation process of the substrate. This point mutation should also prevent a correct operation for the kinase domain of the TGFβ receptors. Although they remain speculative, our modeling results bring some credit to the hypothesis that selected point mutations occurring at hot spots in both TGFBR1 and TGFBR2 kinase domains, consist of a possible driven mechanism in promoting allergic disorders via an alteration of the phosphorylation capacity of the kinase domain of the TGFβ receptors. Obviously, these predictions need to be substanciated by experimental results.

Familial Thoracic Aortic Aneurysm with Dissection Presenting as Flash Pulmonary Edema in a 26-Year-Old Man

We are reporting a case of familial thoracic aortic aneurysm and dissection in a 26-year-old man with no significant past medical history and a family history of dissecting aortic aneurysm in his mother at the age of 40. The patient presented with cough, shortness of breath, and chest pain. Chest X-ray showed bilateral pulmonary infiltrates. CT scan of the chest showed a dissection of the ascending aorta. The patient underwent aortic dissection repair and three months later he returned to our hospital with new complaints of back pain. CT angiography showed a new aortic dissection extending from the left carotid artery through the bifurcation and into the iliac arteries. The patient underwent replacement of the aortic root, ascending aorta, total aortic arch, and aortic valve. The patient recovered well postoperatively. Genetic studies of the patient and his children revealed no mutations in ACTA2, TGFBR1, TGFBR2, TGFB2, MYH11, MYLK, SMAD3, or FBN1. This case report focuses on a patient with familial TAAD and discusses the associated genetic loci and available screening methods. It is important to recognize potential cases of familial TAAD and understand the available screening methods since early diagnosis allows appropriate management of risk factors and treatment when necessary.

Angiotensin II–dependent TGF-β signaling contributes to Loeys-Dietz syndrome vascular pathogenesis

Loeys-Dietz syndrome (LDS) is a connective tissue disorder that is characterized by a high risk for aneurysm and dissection throughout the arterial tree and phenotypically resembles Marfan syndrome. LDS is caused by heterozygous missense mutations in either TGF-β receptor gene (TGFBR1 or TGFBR2), which are predicted to result in diminished TGF-β signaling; however, aortic surgical samples from patients show evidence of paradoxically increased TGF-β signaling. We generated 2 knockin mouse strains with LDS mutations in either Tgfbr1 or Tgfbr2 and a transgenic mouse overexpressing mutant Tgfbr2. Knockin and transgenic mice, but not haploinsufficient animals, recapitulated the LDS phenotype. While heterozygous mutant cells had diminished signaling in response to exogenous TGF-β in vitro, they maintained normal levels of Smad2 phosphorylation under steady-state culture conditions, suggesting a chronic compensation. Analysis of TGF-β signaling in the aortic wall in vivo revealed progressive upregulation of Smad2 phosphorylation and TGF-β target gene output, which paralleled worsening of aneurysm pathology and coincided with upregulation of TGF-β1 ligand expression. Importantly, suppression of Smad2 phosphorylation and TGF-β1 expression correlated with the therapeutic efficacy of the angiotensin II type 1 receptor antagonist losartan. Together, these data suggest that increased TGF-β signaling contributes to postnatal aneurysm progression in LDS.

Dural ectasia in Loeys–Dietz syndrome: comprehensive study of 30 patients with a TGFBR1 or TGFBR2 mutation

The purpose of this study was to assess the frequency, severity, and clinical associations of dural ectasia (DE) in Loeys-Dietz syndrome (LDS). Database analysis of three German metropolitan regions identified 30 patients with LDS and TGFBR1 mutation in 6 and a TGFBR2 mutation in 24 individuals (17 men; mean age: 31 ± 19 years), as well as 60 age and sex-matched control patients with Marfan syndrome carrying a FBN1 mutation. DE was present in 22 patients with LDS (73%), and it related to skeletal score points (p = 0.008), non-skeletal score points (p < 0.001), and to the presence of ≥7 systemic score points (p = 0.010). Similarly, the severity of DE was related to body height (p = 0.010) and non-skeletal score points (p = 0.004). Frequency (p = 0.131) and severity of DE (p = 0.567) was similar in LDS and Marfan syndrome. DE is a manifestation of LDS that occurs with similar frequency and severity as in Marfan syndrome. Severity of DE may serve as a marker of the overall connective tissue disease severity. LDS may be considered in patients with DE.

Severe eczema and Hyper-IgE in Loeys–Dietz-syndrome — Contribution to new findings of immune dysregulation in connective tissue disorders

Loeys-Dietz syndrome (LDS) is a connective tissue disorder caused by monoallelic mutations in TGFBR1 and TGFBR2, which encode for subunits of the transforming growth factor beta (TGFβ) receptor. Affected patients are identified by vascular aneurysms with tortuosity and distinct morphological presentations similar to Marfan syndrome; however, an additional predisposition towards asthma and allergy has recently been found. We describe two patients with a novel missense mutation in TGFBR1 presenting with highly elevated levels of IgE and severe eczema similar to autosomal-dominant Hyper-IgE syndrome (HIES). Mild allergic manifestations with normal up to moderately increased IgE were observed in 3 out of 6 additional LDS patients. A comparison of this cohort with 4 HIES patients illustrates the significant overlap of both syndromes including eczema and elevated IgE as well as skeletal and connective tissue manifestations.

Screening of TGFBR1, TGFBR2, and FLNA in familial mitral valve prolapse

So far only mutations in the filamin A gene (FLNA) have been identified as causing familial mitral valve prolapse (MVP). Previous studies have linked dysregulation of the transforming growth factor beta (TGF-β) cytokine family to MVP. We investigated whether mutations in the TGF-β receptors genes type I (TGFBR1) and II (TGFBR2) underlie isolated familial MVP cases. Eight families with isolated familial MVP were evaluated clinically and genetically. Ventricular arrhythmias were present in five of the eight families and sudden cardiac death occurred in six patients. Tissue obtained during mitral valve surgery or autopsy was available for histological examination in six cases; all demonstrated myxomatous degeneration. A previously described FLNA missense mutation (p.G288R) was identified in one large family, but no mutations were discovered in TGFBR1 or TGFBR2. An FLNA missense mutation was identified in one family but we found no TGFBR1 or TGFBR2 mutations. Our results suggest that TGFBR1 and TGFBR2 mutations do not play a major role in isolated myxomatous valve dystrophy. Screening for FLNA mutations is recommended in familial myxomatous valvular dystrophy, particularly if X-linked inheritance is suspected.