A unique case of two somatic APC mutations in an early onset cribriform-morular variant of papillary thyroid carcinoma and overview of the literature.

We report a case of a 22-year-old female patient who was diagnosed with a cribriform-morular variant of papillary thyroid carcinoma (CMV-PTC). While at early ages this thyroid cancer variant is highly suggestive for familial adenomatous polyposis (FAP), there was no family history of FAP. In the tumor biallelic, inactivating APC variants were identified. The patient tested negative for germline variants based on analysis of genomic DNA from peripheral blood leukocytes. Somatic mosaicism was excluded by subsequent deep sequencing of leukocyte and normal thyroid DNA using next generation sequencing (NGS). This report presents a rare sporadic case of CMV-PTC, and to the best of our knowledge the first featuring two somatic APC mutations underlying the disease, with an overview of CMV-PTC cases with detected APC and CTNNB1 pathogenic variants from the literature.

TP53 germline mutation testing in early-onset breast cancer: findings from a nationwide cohort.

Early-onset breast cancer may be due to Li-Fraumeni Syndrome (LFS). Current national and international guidelines recommend that TP53 genetic testing should be considered for women with breast cancer diagnosed before the age of 31 years. However, large studies investigating TP53 mutation prevalence in this population are scarce. We collected nationwide laboratory records for all young breast cancer patients tested for TP53 mutations in the Netherlands. Between 2005 and 2016, 370 women diagnosed with breast cancer younger than 30 years of age were tested for TP53 germline mutations, and eight (2.2%) were found to carry a (likely) pathogenic TP53 sequence variant. Among BRCA1/BRCA2 mutation negative women without a family history suggestive of LFS or a personal history of multiple LFS-related tumours, the TP53 mutation frequency was < 1% (2/233). Taking into consideration that TP53 mutation prevalence was comparable or even higher in some studies selecting patients with breast cancer onset at older ages or HER2-positive breast cancers, raises the question of whether a very early age of onset is an appropriate single TP53 genetic testing criterion.

Evaluation of current prediction models for Lynch syndrome: updating the PREMM5 model to identify PMS2 mutation carriers.

Until recently, no prediction models for Lynch syndrome (LS) had been validated for PMS2 mutation carriers. We aimed to evaluate MMRpredict and PREMM5 in a clinical cohort and for PMS2 mutation carriers specifically. In a retrospective, clinic-based cohort we calculated predictions for LS according to MMRpredict and PREMM5. The area under the operator receiving characteristic curve (AUC) was compared between MMRpredict and PREMM5 for LS patients in general and for different LS genes specifically. Of 734 index patients, 83 (11%) were diagnosed with LS; 23 MLH1, 17 MSH2, 31 MSH6 and 12 PMS2 mutation carriers. Both prediction models performed well for MLH1 and MSH2 (AUC 0.80 and 0.83 for PREMM5 and 0.79 for MMRpredict) and fair for MSH6 mutation carriers (0.69 for PREMM5 and 0.66 for MMRpredict). MMRpredict performed fair for PMS2 mutation carriers (AUC 0.72), while PREMM5 failed to discriminate PMS2 mutation carriers from non-mutation carriers (AUC 0.51). The only statistically significant difference between PMS2 mutation carriers and non-mutation carriers was proximal location of colorectal cancer (77 vs. 28%, p < 0.001). Adding location of colorectal cancer to PREMM5 considerably improved the models performance for PMS2 mutation carriers (AUC 0.77) and overall (AUC 0.81 vs. 0.72). We validated these results in an external cohort of 376 colorectal cancer patients, including 158 LS patients. MMRpredict and PREMM5 cannot adequately identify PMS2 mutation carriers. Adding location of colorectal cancer to PREMM5 may improve the performance of this model, which should be validated in larger cohorts.

Interdependence of clinical factors predicting cognition in children with tuberous sclerosis complex.

Cognitive development in patients with tuberous sclerosis complex is highly variable. Predictors in the infant years would be valuable to counsel parents and to support development. The aim of this study was to confirm factors that have been reported to be independently correlated with cognitive development. 102 patients included in this study were treated at the ENCORE-TSC expertise center of the Erasmus Medical Center-Sophia Children's Hospital. Data from the first 24 months of life were used, including details on epilepsy, motor development and mutation status. Outcome was defined as cognitive development (intellectual equivalent, IE) as measured using tests appropriate to the patients age and cognitive abilities (median age at testing 8.2 years, IQR 4.7-12.0). Univariable and multivariable regression analyses were used. In a univariable analysis, predictors of lower IE were: the presence of infantile spasms (ß = -18.3, p = 0.000), a larger number of antiepileptic drugs used (ß = -6.3, p = 0.000), vigabatrin not used as first drug (ß = -14.6, p = 0.020), corticosteroid treatment (ß = -33.2, p = 0.005), and a later age at which the child could walk independently (ß = -2.1, p = 0.000). An older age at seizure onset predicted higher IE (ß = 1.7, p = 0.000). In a multivariable analysis, only age at seizure onset was significantly correlated to IE (ß = 1.2, p = 0.005), contributing to 28% of the variation in IE. In our cohort, age at seizure onset was the only variable that independently predicted IE. Factors predicting cognitive development could aid parents and physicians in finding the appropriate support and schooling for these patients.

The growth response to GH treatment is greater in patients with SHOX enhancer deletions compared to SHOX defects.

OBJECTIVE: Short stature caused by point mutations or deletions of the short stature homeobox (SHOX) gene (SHOX haploinsufficiency (SHI)) is a registered indication for GH treatment. Patients with a SHOX enhancer deletion (SED) have a similar phenotype, but their response to GH is unknown. It is uncertain if duplications of SHOX or its enhancer (SDUP) cause short stature. This study aimed to describe the clinical characteristics and growth response to GH treatment in patients with aberrations of SHOX and its enhancers. DESIGN: In this retrospective multi-center study (2002-March 2014) clinical information was available from 130 patients (72 SHI, 44 SED, and 14 SDUP) of whom 52 patients were treated with GH. We evaluated height, sitting height (SH), arm span, dysmorphic features and indicators of the growth response to GH (delta height SDS, height velocity, and index of responsiveness). RESULTS: Patients with SEDs showed similar HtSDS to patients with SHI (-2.3 and -2.6, respectively, P=0.2), but they were less disproportionate (SH/height ratio SDS 2.0 vs 3.1 (P<0.01) and extremities/trunk ratio 2.57 vs 2.43 (P=0.03)). The 1st year growth response to GH treatment was significantly greater in prepubertal patients with SEDs than SHI. None of the patients with an SDUP was disproportionate and SDUP cosegregated poorly with short stature; their growth response to GH treatment (n=3) was similar to the other groups. CONCLUSIONS: Patients with SEDs are equally short, but less disproportionate than patients with SHI, and show a greater response to GH.

A clinical and genetic overview of 18 years neurofibromatosis type 1 molecular diagnostics in the Netherlands.

NF1 mutations are the underlying cause of neurofibromatosis type 1 (NF1), a neuro-cardio-facio-cutaneous syndrome (NCFC). Because of the clinical overlap between NCFCs, genetic analysis of NF1 is necessary to confirm a clinical diagnosis NF1. This report describes the clinical and genetic findings of 18 years of NF1 molecular diagnostics in the Netherlands. A pathogenic mutation was found in 59.3% (1178/1985) of the index patients, mostly de novo (73.8%). The majority of the index patients (64.3%) fulfilled the National Institute of Health NF1 criteria, a pathogenic mutation was found in 80.9% of these patients. Seventy-four percent of the index patients with an NF1 pathogenic mutation and not fulfilling the NF1 criteria is <12 years, in agreement with the fact that some NF1 symptoms appear after puberty. Genotype-phenotype correlations were studied for 527 index patients. NF1 patients with a type 1 microdeletion have a sixfold higher risk of special education vs NF1 patients with an intragenic mutation. No evidently milder NF1 phenotype for patients with a missense mutation was observed. Forty-six prenatal analyses were performed in 28 (2.4%) families, of which 29 (63%) showed heterozygosity for the familial pathogenic mutation. This indicates that there is a need for prenatal NF1 testing.

Phenotypes of craniofrontonasal syndrome in patients with a pathogenic mutation in EFNB1.

Craniofrontonasal syndrome (CFNS) is an X-linked developmental malformation, caused by mutations in the EFNB1 gene, which have only been described since 2004. A genotype-phenotype correlation seems not to be present. As it is of major importance to adequately counsel patients with EFNB1 mutations and their parents, and to improve diagnosis of new patients, more information about the phenotypic features is needed. This study included 23 patients (2 male, 21 female) with confirmed EFNB1 mutations. All patients underwent a thorough physical examination and photographs were taken. If available, radiological images were also consulted. Hypertelorism, longitudinal ridging and/or splitting of nails, a (mild) webbed neck and a clinodactyly of one or more toes were the only consistent features observed in all patients. Frequently observed phenotypic features were bifid tip of the nose (91%), columellar indentation (91%) and low implantation of breasts (90%). In comparison with anthropometric data of facial proportions, patients with CFNS had a significantly different face in multiple respects. An overview of all phenotypic features is shown. Patients with EFNB1 mutations have a clear phenotype. This study will facilitate genetic counseling of parents and patients, and contribute to the diagnostic and screening process of patients with suspected CFNS.

Distress in partners of individuals diagnosed with or at high risk of developing tumors due to rare hereditary cancer syndromes.

OBJECTIVE: Li Fraumeni syndrome (LFS) and Von Hippel-Lindau disease (VHL) are two rare hereditary tumor syndromes, characterized by a high risk of developing multiple tumors at various sites and ages for which preventive and treatment options are limited. For partners, it may be difficult to deal with the on-going threat of tumors in both their spouse and children. Therefore, this study aims to evaluate the prevalence of and factors associated with psychological distress among partners of individuals with or at high risk of LFS or VHL. METHODS: As part of a nationwide, cross-sectional study, partners of individuals diagnosed with or at high risk of LFS or VHL were invited to complete a self-report questionnaire assessing distress, worries, and health-related quality of life. RESULTS: Fifty-five (58%) of those high-risk individuals with a partner consented to having their partner approached for the study. In total, 50 partners (91%) completed the questionnaire, of whom 28% reported clinically relevant levels of syndrome-related distress. Levels of distress and worries of the partners and their high-risk spouse were significantly correlated. Younger age and a lack of social support were also associated significantly with heightened levels of distress and worries. The majority of partners (76%) believed that professional psychosocial support should be routinely offered to them. CONCLUSIONS: Approximately one-quarter of the partners exhibit clinically relevant levels of distress that warrant psychological support. The distress levels of the 'patient' could potentially be used to identify partners at risk of developing clinically relevant levels of distress.

Pitfalls in molecular analysis for mismatch repair deficiency in a family with biallelic pms2 germline mutations.

Heterozygous germline mutations in the mismatch repair (MMR) genes MLH1, MSH2, MSH6 and PMS2 cause Lynch syndrome. Biallelic mutations in the MMR genes are associated with a childhood cancer syndrome [constitutional mismatch repair deficiency (CMMR-D)]. This is predominantly characterized by hematological malignancies and tumors of the bowel and brain, often associated with signs of neurofibromatosis type 1 (NF1). Diagnostic strategies for selection of patients for MMR gene analysis include analysis of microsatellite instability (MSI) and immunohistochemical (IHC) analysis of MMR proteins in tumor tissue. We report the clinical characterization and molecular analyses of tumor specimens from a family with biallelic PMS2 germline mutations. This illustrates the pitfalls of present molecular screening strategies. Tumor tissues of five family members were analyzed for MSI and IHC. MSI was observed in only one of the analyzed tissues. However, IHC analysis of brain tumor tissue of the index patient and his sister showed absence of PMS2 expression, and germline mutation analyses showed biallelic mutations in PMS2: p.Ser46IIe and p.Pro246fs. The same heterozygous mutations were confirmed in the father and mother, respectively. These data support the conclusion that in case of a clinical phenotype of CMMR-D, it is advisable to routinely combine MSI analysis with IHC analysis for the expression of MMR proteins. With inconclusive or conflicting results, germline mutation analysis of the MMR genes should be considered after thorough counselling of the patients and/or their relatives.

Psychosocial impact of Von Hippel-Lindau disease: levels and sources of distress.

Von Hippel-Lindau disease (VHL) is a hereditary tumor susceptibility syndrome, characterized by an increased risk of developing multiple benign and malignant tumors at various sites and ages with limited preventive options. This study evaluates the prevalence of distress among VHL family members and factors associated significantly with such distress. Forty-eight families with a VHL mutation were identified via the nine family cancer clinics in the Netherlands. In total, 171 family members (carriers, 50% at-risk, non-carriers) were approached, of whom 123 (72%) completed a self-report questionnaire. Approximately 40% of the VHL family members reported clinically relevant levels of distress, approaching 50% among the carriers and, possibly even more striking, 36% among the non-carriers. Having lost a first degree relative due to VHL during adolescence (OR 11.2; 95% CI 1.4-86.9) was related significantly to heightened levels of distress. Approximately, only one-third of those who reported heightened levels of distress had received professional psychosocial support. A substantial percentage of family members experience clinically relevant levels of distress. We would recommend the introduction of a procedure for screening for distress in this vulnerable population. Special attention should be paid to those individuals who have lost a close relative due to VHL during adolescence.

Inflammatory processes in cortical tubers and subependymal giant cell tumors of tuberous sclerosis complex.

Cortical tubers and subependymal giant cell tumors (SGCT) are two major cerebral lesions associated with tuberous sclerosis complex (TSC). In the present study, we investigated immunocytochemically the inflammatory cell components and the induction of two major pro-inflammatory pathways (the interleukin (IL)-1beta and complement pathways) in tubers and SGCT resected from TSC patients. All lesions were characterized by the prominent presence of microglial cells expressing class II-antigens (HLA-DR) and, to a lesser extent, the presence of CD68-positive macrophages. We also observed perivascular and parenchymal T lymphocytes (CD3(+)) with a predominance of CD8(+) T-cytotoxic/suppressor lymphoid cells. Activated microglia and reactive astrocytes expressed IL-1beta and its signaling receptor IL-1RI, as well as components of the complement cascade, such as C1q, C3c and C3d. Albumin extravasation, with uptake in astrocytes, was observed in both tubers and SGCT, suggesting that alterations in blood brain barrier permeability are associated with inflammation in TSC-associated lesions. Our findings demonstrate a persistent and complex activation of inflammatory pathways in cortical tubers and SGCT.

Frequency of Von Hippel-Lindau germline mutations in classic and non-classic Von Hippel-Lindau disease identified by DNA sequencing, Southern blot analysis and multiplex ligation-dependent probe amplification.

The current clinical diagnosis of Von Hippel-Lindau (VHL) disease demands at least one specific [corrected] VHL manifestation in a patient with familial VHL disease, or, in a [corrected] sporadic patient, at least two or more hemangioblastomas or a single hemangioblastoma in combination with a typical visceral lesion. To evaluate this definition, we studied the frequency of germline VHL mutation in three patients groups: (i) multi-organ involvement (classic VHL), (ii) limited VHL manifestations meeting criteria (non-classic VHL) and (iii) patients with VHL-associated tumors not meeting current diagnostic VHL criteria. In addition, we validated multiplex ligation-dependent probe amplification (MLPA) as a rapid and reliable quantitative method for the identification of germline VHL deletions. The frequency of germline VHL mutations was very high in classic VHL cases with multi-organ involvement (95%), lower in non-classic cases that meet current diagnostic criteria but have limited VHL manifestations or single-organ involvement (24%) and low (3.3%), but tangible in cases not meeting current diagnostic VHL criteria. The detection of germline VHL mutations in patients or families with limited VHL manifestations, or single-organ involvement is relevant for follow-up of probands and early identification of at-risk relatives.

[Tuberous sclerosis complex type 1 (TSC1): diagnostic significance of the cutaneous mini-lesions in a familial presentation]. / Complejo esclerosis tuberosa tipo 1 (CET1): importancia diagnóstica de las minilesiones cutáneas en los casos de presentación familiar.

We present a family with tuberous sclerosis complex type 1 (TSC1). The family is formed by six patients. Two sibs, female and male, both patients having minor signs of the disorder. The woman have two daughters, one of them with severe clinical and radiological features of TSC1, and the other who only has a small cutaneous acromic spot. The man has two sons, one with severe features of TSC1, and the other with cutaneous mini-lesions. DNA study of the two cousins, woman and man, with cutaneous mini-lesions, showed a mutation in the chromosome 9 (1749 del GA) (TSC1 ex 15) in both patients. No pathogenic mutation in the TSC2 gene was detected in the two cousins.

Hyperechogenic fetal bowel: counseling difficulties.

The detection of echodense fetal bowel on ultrasound examination in the second trimester of pregnancy justifies invasive procedures such as amniocentesis to detect an underlying cause. We present a case in which initial tests identified only one mutation in the cystic fibrosis transmembrane regulator (CFTR)-gene of the fetus, the family history being negative for CF. Strongly reduced intestinal enzyme activities suggested intestinal obstruction and further increased the estimated risk for CF. After the 24th gestational week, a second mutation was found, confirming cystic fibrosis in this child. Problems in counseling in this particular case are discussed.

Large deletion at the TSC1 locus in a family with tuberous sclerosis complex.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterized by seizures, mental retardation and the development of hamartomas in a variety of organs and tissues. The disease is caused by mutations in either the TSC1 gene on chromosome 9q34, or the TSC2 gene on chromosome 16p13.3. Here we describe a deletion encompassing the TSC1 gene and two neighboring transcripts on chromosome 9q34 in six affected individuals from a family with TSC. To our knowledge, this is the first report of such a large deletion at the TSC1 locus and indicates that screening for similar mutations at the TSC1 locus is warranted in individuals with TSC.

Ipsilateral breast tumour recurrence in hereditary breast cancer following breast-conserving therapy.

The overall rate of an ipsilateral breast tumour recurrence (IBTR) after breast-conserving therapy (BCT) ranges from 1% to 2% per year. Risk factors include young age but data on the impact of BRCA1/2 mutations or a definite positive family history for breast cancer are scarce. We investigated IBTR after BCT in patients with hereditary breast cancer (HBC). Through our family cancer clinic we identified 87 HBC patients, including 26 BRCA1/2 carriers, who underwent BCT between 1980 and 1995 (cases). They were compared to 174 patients with sporadic breast cancer (controls) also treated with BCT, matched for age and year of diagnosis. Median follow up was 6.1 years for the cases and 6.0 years for controls. Patient and tumour characteristics were similar in both groups. An IBTR was observed in 19 (21.8%) hereditary and 21 (12.1%) sporadic patients. In the hereditary patients more recurrences occurred elsewhere in the breast (21% versus 9.5%), suggestive of new primaries. Overall, the actuarial IBTR rate was similar at 2 years, but higher in hereditary as compared to sporadic patients at 5 years (14% versus 7%) and at 10 years (30% versus 16%) (P=0.05). Post-relapse and overall survival was not different between hereditary and sporadic cases. Hereditary breast cancer was therefore associated with a higher frequency of early (2-5 years) and late (>5 years) local recurrences following BCT. These data suggest an indication for long-term follow up in HBC and should be taken into account when additional 'risk-reducing' surgery after primary BCT is eventually considered.

Frequent loss of the AXIN1 locus but absence of AXIN1 gene mutations in adenocarcinomas of the gastro-oesophageal junction with nuclear beta-catenin expression.

Up to 60% of gastro-oesophageal junction (GEJ) adenocarcinomas show nuclear beta-catenin expression, pointing to activated T-cell factor (TCF)/beta-catenin-driven gene transcription. We demonstrate in five human GEJ adenocarcinoma cell lines that nuclear beta-catenin expression indeed correlates with enhanced TCF-mediated transcription of a reporter gene. In several tumour types, TCF/beta-catenin activation is caused by mutations in either adenomatous polyposis coli (APC), beta-catenin exon 3, AXIN1, AXIN2 or beta-transducin repeat-containing protein (beta-TrCP). In GEJ adenocarcinomas, very few APC and beta-catenin mutations have been found. Therefore, the mechanism of Wnt pathway activation remains unclear. In the present study, we did not find AXIN1 gene mutations in 17 GEJ tumours with nuclear beta-catenin expression (without beta-catenin exon 3 mutations). Six intragenic single nucleotide polymorphisms (SNPs) were identified. One of these, the AXIN1 gene T1942C SNP, has a frequency of 21% but is only very recently described despite numerous AXIN1 gene mutational studies. We provide evidence why this SNP was missed in single strand conformation polymorphism analyses. The AXIN1 gene G2063A variation was previously described as a gene mutation but we demonstrate that this is a polymorphism. With these six SNPs loss of heterozygosity (LOH) was found in 11 of 15 (73%) informative tumours. To investigate a possible AXIN1 gene dosage effect in GEJ tumours expressing nuclear beta-catenin, AXIN1 locus LOH was determined in 20 tumours expressing membranous and no nuclear beta-catenin. LOH was found in 10 of 13 (77%) informative cases. AXIN1 protein immunohistochemistry revealed cytoplasmic expression in all tumours irrespective of the presence of AXIN1 locus LOH. These data indicate that nuclear beta-catenin expression is indicative for activated Wnt signalling and that neither AXIN1 gene mutations nor AXIN1 locus LOH are involved in Wnt pathway activation in GEJ adenocarcinomas.

[From gene to disease; craniosynostosis syndromes due to FGFR2-mutation]. / Van gen naar ziekte; craniosynostosesyndromen door FGFR2-mutaties.

One of the genes involved in craniosynostosis syndromes is the fibroblast growth factor receptor 2 (FGFR2) gene, a tyrosine kinase receptor gene. Upon ligand binding the FGFR2 receptors dimerise, and this is followed by activation of the intracellular tyrosine kinase domains. This initiates a cascade of signals that influence cell division and differentiation. FGFR2 mutations have been found in the Apert, Crouzon and Pfeiffer craniosynostosis syndromes. Most mutations are gain of function mutations, inducing ligand-independent receptor activation or altered ligand binding. With the exception of Apert syndrome, there is no clear genotype-phenotype correlation. Many different mutations have been found in Pfeiffer and Crouzon syndrome, but all of the mutations occur in the same extracellular region of the receptor. Identical mutations have been found in Pfeiffer and Crouzon syndrome. So within one family, both Crouzon and Pfeiffer syndrome may occur. Mutations in other FGFR-genes have also been found in craniosynostosis syndromes.

TSC2 missense mutations inhibit tuberin phosphorylation and prevent formation of the tuberin-hamartin complex.

Tuberous sclerosis (TSC) is an autosomal dominant disorder characterized by a broad phenotypic spectrum that includes seizures, mental retardation, renal dysfunction and dermatological abnormalities. Inactivating mutations to either of the TSC1 and TSC2 tumour suppressor genes are responsible for the disease. TSC1 and TSC2 encode two large novel proteins called hamartin and tuberin, respectively. Hamartin and tuberin interact directly with each other and it has been reported that tuberin may act as a chaperone, preventing hamartin self-aggregation and maintaining the tuberin-hamartin complex in a soluble form. In this study, the ability of tuberin to act as a chaperone for hamartin was used to investigate the tuberin-hamartin interaction in more detail. A domain within tuberin necessary for the chaperone function was identified, and the effects of TSC2 missense mutations on the tuberin-hamartin interaction were investigated to allow specific residues within the central domain of tuberin that are important for the interaction with hamartin to be pin-pointed. In addition, the results confirm that phosphorylation may play an important role in the formation of the tuberin-hamartin complex. Although mutations that prevent tuberin tyrosine phosphorylation also inhibit tuberin-hamartin binding and the chaperone function, our results indicate that only hamartin is phosphorylated in the tuberin-hamartin complex.

[From gene to disease; TSC1 and TSC2 genes and tuberous sclerosis complex]. / Van gen naar ziekte; de TSC1- en TSC2-genen en tubereuze-sclerosiscomplex.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterised by the presence of multiple hamartomas in different parts of the body: the skin, central nervous system, retina, heart, and kidneys. The diagnosis is based on clinical criteria. Screening for TSC must include investigation of skin, CT-scan of the brain and retinal examination. Mutations in TSC patients are present in either the TSC1 or the TSC2 gene. Due to the complexity of the genes and the observation that almost each family has a unique mutation, DNA analysis is not suitable for fast diagnosis of the index patient, but may be used for testing relatives at risk.