Antisense oligonucleotides targeting exon 11 are able to partially rescue the NF2-related schwannomatosis phenotype in vitro.

NF2-related schwannomatosis (NF2-related SWN) is an autosomal dominant condition caused by loss of function variants in the NF2 gene, which codes for the protein Merlin and is characterized by the development of multiple tumors of the nervous system. The clinical presentation of the disease is variable and related to the type of the inherited germline variant. Here, we tested if phosphorodiamidate morpholino oligomers (PMOs) could be used to correct the splice signaling caused by variants at ±13 within the intron-exon boundary region and showed that the PMOs designed for these variants do not constitute a therapeutic approach. Furthermore, we evaluated the use of PMOs to decrease the severity of the effects of NF2 truncating variants with the aim of generating milder hypomorphic isoforms in vitro through the induction of the in-frame deletion of the exon-carrying variant. We were able to specifically induce the skipping of exons 4, 8, and 11 maintaining the NF2 gene reading frame at cDNA level. Only the skipping of exon 11 produced a hypomorphic Merlin (Merlin-e11), which is able to partially rescue the observed phenotype in primary fibroblast cultures from NF2-related SWN patients, being encouraging for the treatment of patients harboring truncating variants located in exon 11.

Neurofibromatosis type 1 families with first-degree relatives harbouring distinct NF1 pathogenic variants. Genetic counselling and familial diagnosis: what should be offered?

Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder caused by pathogenic variants in NF1 Recently, NF1 testing has been included as a clinical criterion for NF1 diagnosis. Additionally, preconception genetic counselling in patients with NF1 focuses on a 50% risk of transmitting the familial variant as the risk of having a sporadic NF1 is considered the same as the general population. METHODS: 829 individuals, 583 NF1 sporadic cases and 246 patients with NF1 with documented family history, underwent genetic testing for NF1. Genotyping and segregation analysis of NF1 familial variants was determined by microsatellite analysis and NF1 sequencing. RESULTS: The mutational analysis of NF1 in 154 families with two or more affected cases studied showed the co-occurrence of two different NF1 germline pathogenic variants in four families. The estimated mutation rate in those families was 3.89×10-3, 20 times higher than the NF1 mutation rate (~2×10-4) (p=0.0008). Furthermore, the co-occurrence of two different NF1 germline pathogenic variants in these families was 1:39, 60 times the frequency of sporadic NF1 (1:2500) (p=0.003). In all cases, the de novo NF1 pathogenic variant was present in a descendant of an affected male. In two cases, variants were detected in the inherited paternal wild-type allele. CONCLUSIONS: Our results, together with previous cases reported, suggest that the offspring of male patients with NF1 could have an increased risk of experiencing de novo NF1 pathogenic variants. This observation, if confirmed in additional cohorts, could have relevant implications for NF1 genetic counselling, family planning and NF1 genetic testing.

Revisiting the UK Genetic Severity Score for NF2: a proposal for the addition of a functional genetic component.

BACKGROUND: Neurofibromatosis type 2 (NF2) is an autosomal dominant disorder characterised by the development of multiple schwannomas, especially on vestibular nerves, and meningiomas. The UK NF2 Genetic Severity Score (GSS) is useful to predict the progression of the disease from germline NF2 pathogenic variants, which allows the clinical follow-up and the genetic counselling offered to affected families to be optimised. METHODS: 52 Spanish patients were classified using the GSS, and patients' clinical severity was measured and compared between GSS groups. The GSS was reviewed with the addition of phenotype quantification, genetic variant classification and functional assays of Merlin and its downstream pathways. Principal component analysis and regression models were used to evaluate the differences between severity and the effect of NF2 germline variants. RESULTS: The GSS was validated in the Spanish NF2 cohort. However, for 25% of mosaic patients and patients harbouring variants associated with mild and moderate phenotypes, it did not perform as well for predicting clinical outcomes as it did for pathogenic variants associated with severe phenotypes. We studied the possibility of modifying the mutation classification in the GSS by adding the impact of pathogenic variants on the function of Merlin in 27 cases. This revision helped to reduce variability within NF2 mutation classes and moderately enhanced the correlation between patient phenotype and the different prognosis parameters analysed (R2=0.38 vs R2=0.32, p>0001). CONCLUSIONS: We validated the UK NF2 GSS in a Spanish NF2 cohort, despite the significant phenotypic variability identified within it. The revision of the GSS, named Functional Genetic Severity Score, could add value for the classification of mosaic patients and patients showing mild and moderate phenotypes once it has been validated in other cohorts.

Mechanical Reinforced Terminolateral Ileo-Transverse Anastomosis: An Option After Right Hemicolectomy-A 452 Patients Study.

INTRODUCTION: Bowel reconstruction techniques after right hemicolectomy has currently been objective of review, due to the high rate of anastomotic leak. The aim of this study is to analyse our results of the mechanical reinforced terminolateral ileo-transverse anastomosis. MATERIALS AND METHODS: A prospective and descriptive study of a consecutive series of right colonic cancer cases that underwent right hemicolectomy. Mechanical reinforced terminolateral ileo-transverse anastomosis technique was carried out in all patients. Demographics, emergency or elective surgery, surgical management, postoperative complications, rate of anastomotic leak, need for surgical procedure after complication, average stay, and mortality were analysed. RESULTS: A total of 452 patients underwent surgery between 2010 and 2017. Of those, 40.6% were female and 59.4% were male. The average age and body mass index (BMI) was 72±11.3 years old, and 26±7.1, respectively. Elective surgery was carried out in 405 (89.6%) patients. Laparoscopic approach was used in 250 patients (61.7%) and 6% needed conversion. Only 41 patients (10.6%) had major complications (Clavien-Dindo III-IV). The rate of postoperative paralytic ileus reach was up to 13.9%. Reintervention was needed in five patients (1.1%) due to anastomotic leak and three (0.7%) of them from the elective surgery subgroup. There were 10 patients (2.2%) with postoperative gastrointestinal bleeding. The average stay was 8.2±2.8 days and late postoperative mortality in the first 30 days was 2%. CONCLUSIONS: Mechanical reinforced terminolateral ileo-colic anastomosis is a safe technique with a low anastomotic leak rate. Although our results using this approach seem promising, postoperative paralytic ileus is still a high-rate complication.