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Individuals with ultrarare disorders pose a structural challenge for healthcare systems since expert clinical knowledge is required to establish diagnoses. In TRANSLATE NAMSE, a 3-year prospective study, we evaluated a novel diagnostic concept based on multidisciplinary expertise in Germany. Here we present the systematic investigation of the phenotypic and molecular genetic data of 1,577 patients who had undergone exome sequencing and were partially analyzed with next-generation phenotyping approaches. Molecular genetic diagnoses were established in 32% of the patients totaling 370 distinct molecular genetic causes, most with prevalence below 1:50,000. During the diagnostic process, 34 novel and 23 candidate genotype-phenotype associations were identified, mainly in individuals with neurodevelopmental disorders. Sequencing data of the subcohort that consented to computer-assisted analysis of their facial images with GestaltMatcher could be prioritized more efficiently compared with approaches based solely on clinical features and molecular scores. Our study demonstrates the synergy of using next-generation sequencing and phenotyping for diagnosing ultrarare diseases in routine healthcare and discovering novel etiologies by multidisciplinary teams., (© 2024. The Author(s).)

Objective: To compare the incidence of right-sided versus left-sided, and unilateral versus bilateral, nonsyndromic clefting in the affected offspring of smoking and nonsmoking mothers., Design: Self-report data on periconceptual and first trimester smoking behavior were collected from 842 mothers of children with nonsyndromic orofacial clefting. Differences in the incidence of left- versus right-sided clefts, and of unilateral versus bilateral clefts, were analyzed between the children of smoking and nonsmoking mothers., Setting: Interviews and clinical examinations took place at 8 specialist centers in Germany., Patients and Participants: Children with nonsyndromic clefts were recruited during the course of surgical or orthodontic treatment, or within the context of the annual control consultation. Patients with cleft palate only or missing data were excluded. The final cohort comprised 842 patients (540 males and 302 females) with unilateral or bilateral clefts. The respective mothers were interviewed., Main Outcome Measure: Side and laterality of nonsyndromic clefts were the main outcome measures., Results: Children of smoking mothers more often had right-sided clefts than children of nonsmoking mothers (42% right-sided clefts in children of smoking mothers vs 31% of nonsmoking mothers). Children of smoking mothers more often had bilateral clefts than children of nonsmoking mothers (35% bilateral clefts in children of smoking mothers vs 29% of nonsmoking mothers). Sex-specific analyses confirmed substantially and statistically significant associations only for girls., Conclusions: The results suggest that maternal smoking is a sex-specific, exogenous determinant of laterality and side in nonsyndromic clefts.

Retinitis pigmentosa (RP) is an inherited degenerative disease causing severe retinal dystrophy and visual impairment mainly with onset in infancy or adolescence. Targeted next-generation sequencing (NGS) has become an efficient tool to encounter the enormous genetic heterogeneity of diverse retinal dystrophies, including RP. To identify disease-causing mutations in unselected, consecutive RP patients, we conducted Sanger sequencing of genes commonly involved in the suspected genetic RP subtype, followed by targeted large-panel NGS if no mutation was identified, or NGS as primary analysis. A high (70%) detection rate of disease-causing mutations was achieved in a large cohort of 116 unrelated patients. About half (48%) of the solved RP cases were explained by mutations in four genes: RPGR, EYS, PRPF31 and USH2A. Overall, 110 different mutations distributed across 30 different genes were detected, and 46 of these mutations were novel. A molecular diagnosis was achieved in the majority (82-100%) of patients if the family history was suggestive for a particular mode of inheritance, but only in 60% in cases of sporadic RP. The diagnostic potential of extensive molecular analysis in a routine setting is also illustrated by the identification of unexpected genotype-phenotype correlations for RP patients with mutations in CRX, CEP290, RPGRIP1, MFSD8. Furthermore, we identified numerous mutations in autosomal dominant (PRPF31, PRPH2, CRX) and X-linked (RPGR) RP genes in patients with sporadic RP. Variants in RP2 and RPGR were also found in female RP patients with apparently sporadic or dominant disease. In summary, this study demonstrates that massively parallel sequencing of all known retinal dystrophy genes is a valuable diagnostic approach for RP patients., Competing Interests: I have read the journal's policy and the authors of this manuscript have the following competing interests: Dres. Neuhaus, Lenzner, Zahnleiter, Betz, and Eisenberger are employees of Bioscientia, a publicly traded diagnostic company. Dr. Bolz has been employee of Bioscientia until 2016. There are no patents, products in development or marketed products to declare. This does not alter the authors' adherence to PLOS ONE policies on sharing data and materials. The other authors have declared that no competing interests exist.

Background: The genes Gremlin-1 (GREM1) and Noggin (NOG) are components of the bone morphogenetic protein 4 pathway, which has been implicated in craniofacial development. Both genes map to recently identified susceptibility loci (chromosomal region 15q13, 17q22) for nonsyndromic cleft lip with or without cleft palate (nsCL/P). The aim of the present study was to determine whether rare variants in either gene are implicated in nsCL/P etiology., Methods: The complete coding regions, untranslated regions, and splice sites of GREM1 and NOG were sequenced in 96 nsCL/P patients and 96 controls of Central European ethnicity. Three burden and four nonburden tests were performed. Statistically significant results were followed up in a second case-control sample (n = 96, respectively). For rare variants observed in cases, segregation analyses were performed., Results: In NOG, four rare sequence variants (minor allele frequency < 1%) were identified. Here, burden and nonburden analyses generated nonsignificant results. In GREM1, 33 variants were identified, 15 of which were rare. Of these, five were novel. Significant p-values were generated in three nonburden analyses. Segregation analyses revealed incomplete penetrance for all variants investigated., Conclusion: Our study did not provide support for NOG being the causal gene at 17q22. However, the observation of a significant excess of rare variants in GREM1 supports the hypothesis that this is the causal gene at chr. 15q13. Because no single causal variant was identified, future sequencing analyses of GREM1 should involve larger samples and the investigation of regulatory elements., (© 2014 Wiley Periodicals, Inc.)

Background: The prognosis of patients with advanced oropharyngeal carcinoma (OPSCC) is generally poor. The aim of this study is to investigate the different therapeutic approaches and identify prognostic factors associated with a worse outcome for patients treated for T4a OPSCC, in order to improve treatment selection for the individual., Methods: A retrospective study was conducted on 426 patients with T4a OPC treated between 1980 and 2010. Eleven prognostic factors including treatment modality, lymph node staging, and p16 status as a surrogate marker for human papillomavirus (HPV) infection were analyzed., Results: Univariate analysis showed a significant difference in DSS between N0 and N+ (57.1% versus 26.9%, P < 0.001), primary surgical and primary nonsurgical treatment (52.7% versus 31.4%, P < 0.001), and perinodal invasion (51.7% versus 19.9%, P = 0.011). P16-negative patients tended towards a worse DSS than p16-positive patients (40.2% versus 64.6%, P = 0.126) but responded better to primary surgery than to nonsurgical treatment (71.4% versus 34.0%, P = 0.113). Multivariate analysis identified the N category as an independent prognostic factor for survival., Conclusion: The survival of p16-negative patients was worse than p16-positive patients, although they seem to respond better to primary surgery. The strongest independent prognostic factor for T4a carcinomas proved to be the presence of lymph node metastases.

The MSH2 c.388&#95;389del mutation has occasionally been described in Lynch families worldwide. At the Portuguese Oncology Institute in Porto, Portugal, we have identified 16 seemingly unrelated families with this germline mutation. To evaluate if this alteration is a founder or a recurrent mutation we performed haplotype analysis in the 16 Portuguese index cases and 55 relatives, as well as in four index cases and 13 relatives reported from Germany, Scotland, England, and Argentina. In the Portuguese families we observed a shared haplotype of approximately 10 Mb and all were originated from the north of Portugal. These results suggest that this alteration is a founder mutation in Portugal with a relatively recent origin. In the reported families outside Portugal with this mutation different haplotype backgrounds were observed, supporting the hypothesis that it occurred de novo on multiple occasions. We also conclude that the high proportion of families with the MSH2 c.388&#95;389del mutation indicates that screening for this alteration as a first step may be cost-effective in the genetic testing of Lynch syndrome suspects of Portuguese ancestry, especially those originating from the north of Portugal., (© 2012 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Many genome-wide association studies (GWAS) have signals with unknown etiology. This paper addresses the question-is such an association signal caused by rare or common variants that lead to increased disease risk? For a genomic region implicated by a GWAS, we use single nucleotide polymorphism (SNP) data in a case-control setting to predict how many common or rare variants there are, using a Bayesian analysis. Our objective is to compute posterior probabilities for configurations of rare and/or common variants. We use an extension of coalescent trees--the ancestral recombination graphs--to model the genealogical history of the samples based on marker data. As we expect SNPs to be in linkage disequilibrium with common disease variants, we can expect the trees to reflect the type of variants. To demonstrate the application, we apply our method to candidate gene sequencing data from a German case-control study on nonsyndromic cleft lip with or without cleft palate., (© 2012 Wiley Periodicals, Inc.)

Ten non-synonymous single nucleotide polymorphisms (nsSNPs), which were recently associated with colorectal cancer risk in a comprehensive, array based study (AKAP9 M463I, DKK3 G335R, AMPD1 Q12X, LIPC L356F, PSMB9 V32I, THBS1 N700S, CA6 S90G, ASCC3 C1995S, DHX36 S416C and CPA4 G303C) were re-evaluated in the present study based on 626 German familial non-HNPCC colorectal cancer patients and 736 healthy controls. No associations of any of the 10 nsSNPs with colorectal cancer could be replicated. The combined analyses indicated that further research based on additional independent samples is required.

Purpose: Lynch syndrome is linked to germline mutations in mismatch repair genes. We analyzed the genotype-phenotype correlations in the largest cohort so far reported., Patients and Methods: Following standard algorithms, we identified 281 of 574 unrelated families with deleterious germline mutations in MLH1 (n = 124) or MSH2 (n = 157). A total of 988 patients with 1,381 cancers were included in this analysis., Results: We identified 181 and 259 individuals with proven or obligatory and 254 and 294 with assumed MLH1 and MSH2 mutations, respectively. Age at diagnosis was younger both in regard to first cancer (40 v 43 years; P < .009) and to first colorectal cancer (CRC; 41 v 44 years; P = .004) in MLH1 (n = 435) versus MSH2 (n = 553) mutation carriers. In both groups, rectal cancers were remarkably frequent, and the time span between first and second CRC was smaller if the first primary occurred left sided. Gastric cancer was the third most frequent malignancy occurring without a similarly affected relative in most cases. All prostate cancers occurred in MSH2 mutation carriers., Conclusion: The proportion of rectal cancers and shorter time span to metachronous cancers indicates the need for a defined treatment strategy for primary rectal cancers in hereditary nonpolyposis colorectal cancer patients. Male MLH1 mutation carriers require earlier colonoscopy beginning at age 20 years. We propose regular gastric surveillance starting at age 35 years, regardless of the familial occurrence of this cancer. The association of prostate cancer with MSH2 mutations should be taken into consideration both for clinical and genetic counseling practice.