Elsahy–Waters syndrome is caused by biallelic mutations in CDH11

Elsahy–Waters syndrome (EWS), also known as branchial–skeletal–genital syndrome, is a distinct dysmorphology syndrome characterized by facial asymmetry, broad forehead, marked hypertelorism with proptosis, short and broad nose, midface hypoplasia, intellectual disability, and hypospadias. We have recently published a homozygous potential loss of function variant in CDH11 in a boy with a striking resemblance to EWS. More recently, another homozygous truncating variant in CDH11 was reported in two siblings with suspected EWS. Here, we describe in detail the clinical phenotype of the original CDH11‐related patient with EWS as well as a previously unreported EWS‐affected girl who was also found to have a novel homozygous truncating variant in CDH11, which confirms that EWS is caused by biallelic CDH11 loss of function mutations. Clinical features in the four CDH11 mutation‐positive individuals confirm the established core phenotype of EWS. Additionally, we identify upper eyelid coloboma as a new, though infrequent clinical feature. The pathomechanism underlying EWS remains unclear, although the limited phenotypic data on the Cdh11^?/? mouse suggest that this is a potentially helpful model to explore the craniofacial and brain development in EWS‐affected individuals.

     

Androgen depletion up‐regulates cadherin‐11 expression in prostate cancer

Men with castration‐resistant prostate cancer (PCa) frequently develop metastasis in bone. The reason for this association is unclear. We have previously shown that cadherin‐11 (also known as OB‐cadherin), a homophilic cell adhesion molecule that mediates osteoblast adhesion, plays a role in the metastasis of PCa to bone. Here, we report that androgen‐deprivation therapy up‐regulates cadherin‐11 expression in PCa. In human PCa specimens, immunohistochemical staining showed that 22/26 (85%) primary PCa tumours from men with castration‐resistant PCa expressed cadherin‐11. In contrast, only 7/50 (14%) androgen‐dependent PCa tumours expressed cadherin‐11. In the MDA–PCa‐2b xenograft animal model, cadherin‐11 was expressed in the recurrent tumours following castration. In the PCa cell lines, there is an inverse correlation between expression of cadherin‐11 and androgen receptor (AR), and cadherin‐11 is expressed in very low levels or not expressed in AR‐positive cell lines, including LNCaP, C4‐2B4 and VCaP cells. We showed that AR likely regulates cadherin‐11 expression in PCa through an indirect mechanism. Although re‐expression of AR in the AR‐negative PC3 cells led to the inhibition of cadherin‐11 expression, depletion of androgen from the culture medium or down‐regulation of AR by RNA interference in the C4‐2B4 cells or VCaP cells only produced a modest increase of cadherin‐11 expression. Promoter analysis indicated that cadherin‐11 promoter does not contain a typical AR‐binding element, and AR elicits a modest inhibition of cadherin‐11 promoter activity, suggesting that AR does not regulate cadherin‐11 expression directly. Together, these results suggest that androgen deprivation up‐regulates cadherin‐11 expression in prostate cancer, and this may contribute to the metastasis of PCa to bone. Our study suggests that therapeutic strategies that block cadherin‐11 expression or function may be considered when applying androgen‐ablation therapy. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Epigenetic silencing in non‐neoplastic epithelia identifies E‐cadherin (CDH1) as a target for chemoprevention of lobular neoplasia

Invasive lobular carcinoma (ILC) of the breast is believed to develop from in situ lesions, atypical lobular hyperplasia (ALH), and lobular carcinoma in situ (LCIS). Down‐regulation of the cell–cell adhesion protein E‐cadherin is a defining feature of lobular breast cancer (LBC) and already occurs in ALH and LCIS. Apart from mutational mechanisms, epigenetic silencing of the E‐cadherin gene (CDH1) is thought to be involved in E‐cadherin down‐regulation and has been observed at a high frequency in ILC. Whether CDH1 promoter methylation is already present in in situ lesions and thus contributes to the initiation of LBC is not established. We thus examined microdissected archived tissue from 20 LBCs by methylation‐specific PCR to determine the CDH1 methylation status of lobular lesions. Nineteen of the 20 LBCs had a hypermethylated CDH1 promoter, including 13/14 ILCs and 13/13 ALHs or LCIS. Bisulphite sequencing indicated that methylation was complete within the investigated promoter fragment. Intriguingly, CDH1 methylation was likewise present in 8/8 adjacent non‐neoplastic epithelia, but not in 6/6 mammary epithelia from healthy subjects. E‐cadherin protein and mRNA were down‐regulated in in situ lesions relative to adjacent epithelia. Together, these results indicate that CDH1 promoter methylation occurs in LBC prior to E‐cadherin down‐regulation and neoplastic formation. We thus propose that epigenetic silencing represents the first of the two hits required to silence both CDH1 alleles for LBC to develop. Because promoter methylation is in principle reversible, our findings suggest that chemoprevention of LBC by epigenetic drugs should be feasible. Furthermore, the presence of CDH1 methylation in pre‐neoplastic epithelia suggests the existence of mammary regions with increased disease susceptibility, providing an explanation for the often multifocal presentation of LBC. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

αE‐catenin is a candidate tumor suppressor for the development of E‐cadherin‐expressing lobular‐type breast cancer

Although mutational inactivation of E‐cadherin (CDH1) is the main driver of invasive lobular breast cancer (ILC), approximately 10–15% of all ILCs retain membrane‐localized E‐cadherin despite the presence of an apparent non‐cohesive and invasive lobular growth pattern. Given that ILC is dependent on constitutive actomyosin contraction for tumor development and progression, we used a combination of cell systems and in vivo experiments to investigate the consequences of α‐catenin (CTNNA1) loss in the regulation of anchorage independence of non‐invasive breast carcinoma. We found that inactivating somatic CTNNA1 mutations in human breast cancer correlated with lobular and mixed ducto‐lobular phenotypes. Further, inducible loss of α‐catenin in mouse and human E‐cadherin‐expressing breast cancer cells led to atypical localization of E‐cadherin, a rounded cell morphology, and anoikis resistance. Pharmacological inhibition experiments subsequently revealed that, similar to E‐cadherin‐mutant ILC, anoikis resistance induced by α‐catenin loss was dependent on Rho/Rock‐dependent actomyosin contractility. Finally, using a transplantation‐based conditional mouse model, we demonstrate that inducible inactivation of α‐catenin instigates acquisition of lobular features and invasive behavior. We therefore suggest that α‐catenin represents a bona fide tumor suppressor for the development of lobular‐type breast cancer and as such provides an alternative event to E‐cadherin inactivation, adherens junction (AJ) dysfunction, and subsequent constitutive actomyosin contraction. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.     

Zebrafish cadherin-11 participates in retinal differentiation and retinotectal axon projection during visual system development

Background: Cadherins orchestrate tissue morphogenesis by controlling cell adhesion, migration and differentiation. Various cadherin family members are expressed in the retina and other neural tissues during embryogenesis, regulating development of these tissues. Cadherin‐11 (Cdh11) is expressed in mesenchymal, bone, epithelial, neural and other tissues, and this cadherin was shown to control cell migration and differentiation in neural crest, tumor and bone cells. Our previous studies characterized Cdh11 expression and function in zebrafish. Results: Here, we report effects of Cdh11 loss‐of‐function on visual system development using morpholino oligonucleotide knockdown methods. Cdh11 is expressed in the retina and lens during retinal differentiation. Cdh11 loss‐of‐function produced defects in retinal differentiation and lens development. Cdh11 loss‐of‐function also reduced retinotectal axon projection and organization, consistent with known Cdh11 function in cell migration. Conclusion: Cdh11 expression in the developing visual system and Cdh11 loss‐of‐function phenotype illustrates the critical role for differential cadherin activity in visual system differentiation and organization. Developmental Dynamics 241:442–454, 2012. © 2012 Wiley Periodicals, Inc.     

A prenatally diagnosed case of Donnai‐Barrow syndrome: Highlighting the importance of whole exome sequencing in cases of consanguinity

Donnai‐Barrow syndrome (DBS) is an autosomal recessive disorder characterized by typical craniofacial features, vision and hearing loss, intellectual disability, agenesis of the corpus callosum (ACC), congenital diaphragmatic hernia (CDH), and omphalocele. This condition is associated with loss‐of‐function mutations in the LRP2 gene. Few cases have been described in the literature. In our case, CDH and ACC were prenatally diagnosed by ultrasound, and the fetus was the product of a first‐degree union. Single‐nucleotide polymorphism‐microarray showed large regions of homozygosity. Whole exome sequencing (WES) was performed and revealed a homozygous frameshift pathogenic variant in LRP2 (c.6978dupG). Here, we present a case of DBS, which diagnosed prenatally via WES in a fetus with CDH and ACC.     

Recurrence of pleomorphic adenoma of the parotid gland—predictive value of cadherin‐11 and fascin

The predictive value of cadherin‐11, tenascin, fascin, and mucin‐1 as markers for the likelihood of recurrence in pleomorphic adenoma of the parotid gland was examined. In this retrospective study we analysed 20 tumours from16 patients by immunohistochemistry. Staining intensities were measured using a semiquantitative scoring approach; localisation (tumour centre vs border) as well as clinical data were analysed and correlated with follow‐up. Cadherin‐11 was increased in recurrent tumours. However, no changes of fascin, tenascin or mucin‐1 were observed. Cadherin‐11 and fascin were increased in primary tumours of patients with later recurrence, with fascin upregulation restricted to the tumour border. In conclusion, cadherin‐11 and fascin should be further analysed for their value as markers for later recurrence in pleomorphic adenoma. Our observations might reflect dysregulation of cellular pathways contributing to cellular dissemination, which might potentially result in later recurrence.     

Three new cases of ataxia‐telangiectasia‐like disorder: No impairment of the ATM pathway,but S‐phase checkpoint defect

Ataxia‐telangiectasia‐like disorder (ATLD) is a rare genomic instability syndrome caused by biallelic variants of MRE11 (meiotic recombination 11) characterized by progressive cerebellar ataxia and typical karyotype abnormalities. These symptoms are common to those of ataxia‐telangiectasia, which is consistent with the key role of MRE11 in ataxia‐telangiectasia mutated (ATM) activation after DNA double‐strand breaks. Three unrelated French patients were referred with ataxia. Only one had typical karyotype abnormalities. Unreported biallelic MRE11 variants were found in these three cases. Interestingly, one variant (c.424G>A) was present in two cases and haplotype analysis strongly suggested a French founder variant. Variants c.544G>A and c.314+4_314+7del lead to splice defects. The level of MRE11 in lymphoblastoid cell lines was consistently and dramatically reduced. Functional consequences were evaluated on activation of the ATM pathway via phosphorylation of ATM targets (KAP1 and CHK2), but no consistent defect was observed. However, an S‐phase checkpoint activation defect after camptothecin was observed in these patients with ATLD. In conclusion, we report the first three French ATLD patients and a French founder variant, and propose an S‐phase checkpoint activation study to evaluate the pathogenicity of MRE11 variants.     

Comprehensive genetic and functional characterization of IPH‐926: a novel CDH1‐null tumour cell line from human lobular breast cancer

Infiltrating lobular breast cancer (ILBC) is a clinically and biologically distinct tumour entity defined by a characteristic linear cord invasion pattern and inactivation of the CDH1 tumour suppressor gene encoding for E‐cadherin. ILBCs also lack β‐catenin expression and show aberrant cytoplasmic localization of the E‐cadherin binding protein p120‐catenin. The lack of a well‐characterized ILBC cell line has hampered the functional characterization of ILBC cells in vitro. We report the establishment of a permanent ILBC cell line, named IPH‐926, which was derived from a patient with metastatic ILBC. The DNA fingerprint of IPH‐926 verified genetic identity with the patient and had no match among the human cell line collections of several international biological resource banks. IPH‐926 expressed various epithelial cell markers but lacked expression of E‐cadherin due to a previously unreported, homozygous CDH1 241ins4 frameshift mutation. Detection of the same CDH1 241ins4 mutation in archival tumour tissue of the corresponding primary ILBC proved the clonal origin of IPH‐926 from this particular tumour. IPH‐926 also lacked β‐catenin expression and showed aberrant cytoplasmic localization of p120‐catenin. Array‐CGH analysis of IPH‐926 revealed a profile of genomic imbalances that included many distinct alterations previously observed in primary ILBCs. Spectral karyotyping of IPH‐926 showed a hyperdiploid chromosome complement and numerous clonal, structural aberrations. IPH‐926 cells were anti‐cancer drug‐resistant, clonogenic in soft agar, and tumourigenic in SCID mice. In xenograft tumours, IPH‐926 cells recapitulated the linear cord invasion pattern that defines ILBCs. In summary, IPH‐926 significantly extends the biological spectrum of the established breast cancer cell lines and will facilitate functional analyses of genuine human ILBC cells in vitro and in vivo. Copyright © 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Protein‐elongating mutations in MYH11 are implicated in a dominantly inherited smooth muscle dysmotility syndrome with severe esophageal,gastric, and intestinal disease

Gastrointestinal motility disorders include a spectrum of mild to severe clinical phenotypes that are caused by smooth muscle dysfunction. We investigated the genetic etiology of severe esophageal, gastric, and colonic dysmotility in two unrelated families with autosomal dominant disease presentation. Using exome sequencing, we identified a 2 base pair insertion at the end of the myosin heavy chain 11 (MYH11) gene in all affected members of Family 1 [NM_001040113:c.5819_5820insCA(p.Gln1941Asnfs*91)] and a 1 base pair deletion at the same genetic locus in Proband 2 [NM_001040113:c.5819del(p.Pro1940Hisfs*91)]. Both variants are predicted to result in a similarly elongated protein product. Heterozygous dominant negative MYH11 pathogenic variants have been associated with thoracic aortic aneurysm and dissection while biallelic null alleles have been associated with megacystis microcolon intestinal hypoperistalsis syndrome. This report highlights heterozygous protein‐elongating MYH11 variants affecting the SM2 isoforms of MYH11 as a cause for severe gastrointestinal dysmotility, and we hypothesize that the mechanistic pathogenesis of this disease, dominant hypercontractile loss‐of‐function, is distinct from those implicated in other diseases involving MYH11 dysfunction.     

GM‐CSF and IL‐4 are not involved in IVIG‐mediated amelioration of ITP in mice: a role for IL‐11 cannot be ruled out

Previously, we have reported that interleukin (IL)‐4, granulocyte–macrophage colony‐stimulating factor (GM‐CSF), and IL‐11, but not IL‐33, are up‐regulated in two strains of mice with immune thrombocytopenia (ITP) that are responsive to intravenous immunoglobulin (IVIg) treatment. Previously, IL‐4 was ruled out in the mechanism of IVIg; however, other publications have suggested this cytokine as a major player in the mechanism of IVIg action. Thus, we sought to further investigate a role for IL‐4 and, in addition, GM‐CSF and IL‐11 in the mechanism of action of IVIg using a murine model of ITP. A passive platelet antibody model was used to generate ITP in IL‐4 receptor knock‐out (IL‐4R^–/–), IL‐11 receptor knock‐out (IL‐11Rα^–/–) and GM‐CSF knock‐out (Csf2^–/–) mice. We also used a neutralizing antibody to IL‐11 and recombinant human IL‐11 (rhIL‐11) in addition to depleting basophils in vivo to study the effect of IVIg to ameliorate ITP. Our results showed that basophils, IL‐4 and GM‐CSF were unimportant in both ITP induction and its amelioration by IVIg. The role of IL‐11 in these processes was less clear. Even though IL‐11Rα^–/– mice with ITP responded to IVIg similarly to wild‐type (WT) mice, treatment of ITP WT mice with rhIL‐11 instead of IVIg showed an increase in platelet numbers and WT mice administered anti‐IL‐11 showed a significant reduction in the ability of IVIg to ameliorate the ITP. Our findings indicate that neither IL‐4, basophils or GM‐CSF have roles in IVIg amelioration of ITP; however, a role for IL‐11 requires further study.     

Large deletions and splicing‐site mutations in the STK11 gene in Peutz‐Jeghers Chilean families

Peutz‐Jeghers syndrome (PJS) is an autosomal dominant disorder characterized by mucocutaneous melanocytic macules, gastrointestinal hamartomatous polyposis and an increased risk of various neoplasms. Germline mutations in the serine/threonine kinase 11 (STK11) gene have been identified as a cause for PJS. The aim of this study was to characterize the genotype of Chilean PJS patients. Mutation screening of 13 patients from eight PJS families was performed using a single strand conformation polymorphism analysis, DNA sequencing and multiplex ligation‐dependent probe amplification assay. The breakpoints of the genomic rearrangements were assessed by a long‐range polymerase chain reaction and sequencing. The results revealed the existence of seven different pathogenic mutations in STK11 gene in seven unrelated families, including three point mutations and four large genomic deletions. Three of these point mutations (43%, 3/7) may be considered as novel. Our results showed that a germline mutation is present in STK11 in 88% of probands fulfilling the diagnostic criteria of PJS. In this study, the combination of two different experimental approaches in the screening of the STK11 in PJS, led to a higher percentage of mutation detection.