Identification and characterization of NF1 and non-NF1 congenital pseudarthrosis of the tibia based on germline NF1 variants: genetic and clinical analysis of 75 patients

Zhu, Guoqiang; Zheng, Yu; Liu, Yaoxi; An, Yan; Hu, Zhengmao; Yang, Yongjia; Xiang, Shiting; Li, Liping; Chen, Weijian; Yu, Ping; Zhong, Nanbert; Mei, Haibo

doi:10.1186/s13023-019-1196-0

Cited by 28 publications

(27 citation statements)

References 55 publications

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“…The double inactivation of the NF-1 gene with subsequent clonal growth was suggested as a possible pathophysiologic mechanism at least in some of the CPT patients. However, these data need still to be confirmed in larger studies [26,27].…”

Section: Etiologymentioning

confidence: 94%

Management of spinal deformities and tibial pseudarthrosis in children with neurofibromatosis type 1 (NF-1)

et al. 2020

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Summary of background data The skeletal system is affected in up to 60% of patients with neurofibromatosis type 1. The most commonly observed entities are spinal deformities and tibial dysplasia. Early recognition of radiologic osseous dystrophy signs is of utmost importance because worsening of the deformities without treatment is commonly observed and surgical intervention is often necessary. Due to the relative rarity and the heterogenic presentation of the disease, evidence regarding the best surgical strategy is still lacking. Purpose To report our experience with the treatment of skeletal manifestations in pediatric patients with (neurofibromatosis type 1) NF-1 and to present the results with our treatment protocols. Materials and methods This is a retrospective, single expert center study on children with spinal deformities and tibial dysplasia associated with NF-1 treated between 2006 and 2020 in a tertiary referral institution. Results Spinal deformity: Thirty-three patients (n = 33) were included. Mean age at index surgery was 9.8 years. In 30 patients (91%), the deformity was localized in the thoracic and/or lumbar spine, and in 3 patients (9%), there was isolated involvement of the cervical spine. Eleven patients (33%) received definitive spinal fusion as an index procedure and 22 (67%) were treated by means of “growth-preserving” spinal surgery. Halo-gravity traction before index surgery was applied in 11 patients (33%). Progression of deformity was stopped in all patients and a mean curve correction of 60% (range 23–98%) was achieved. Mechanical problems with instrumentation requiring revision surgery were observed in 55% of the patients treated by growth-preserving techniques and in none of the patients treated by definitive fusion. One patient (3%) developed a late incomplete paraplegia due to a progressive kyphotic deformity. Tibial dysplasia: The study group comprised of 14 patients. In 5 of them (36%) pathological fractures were present on initial presentation. In the remaining 9 patients (64%), anterior tibial bowing without fracture was observed initially. Four of them (n = 4, 28%) subsequently developed a pathologic fracture despite brace treatment. Surgical treatment was indicated in 89% of the children with pathological fractures. This involved resection of the pseudarthrosis, autologous bone grafting, and intramedullary nailing combined with external fixation in some of the cases. In 50% of the patients, bone morphogenic protein was used “off-label” in order to promote union. Healing of the pseudarthrosis was achieved in all of the cases and occurred between 5 to 13 months after the index surgical intervention. Four of the patients treated surgically needed more than one surgical intervention in order to achieve union; one patient had a re-fracture. All patients had a good functional result at last follow-up. Conclusion Early surgical intervention is recommended for the treatment dystrophic spinal deformity in children with NF-1. Good and sustainable curve correction without relevant thoracic growth inhibition can be achieved with growth-preserving techniques alone or in combination with short spinal fusion at the apex of the curve. Preoperative halo-gravity traction is a safe and very effective tool for the correction of severe and rigid deformity in order to avoid neurologic injury. Fracture union in tibial dysplasia with satisfactory functional results can be obtained in over 80% of the children by means of surgical resection of the pseudarthrosis, intramedullary nailing, and bone grafting. Wearing a brace until skeletal maturity is achieved is mandatory in order to minimize the risk of re-fracture.

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Section: Etiologymentioning

confidence: 94%

Management of spinal deformities and tibial pseudarthrosis in children with neurofibromatosis type 1 (NF-1)

et al. 2020

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“…In our series, we included seven exonic variants: two synonym substitutions that, as expected, were shown not to affect splicing and 5 missense variants/inframe deletions. Except for the c.1466A > G change, for all the other exonic variants, no splicing alterations were observed with the minigene assay: in three cases (c.278G > A, c.3112A > G, c.7250_7252delACT) population data, segregation analysis, previous findings [ 23 , 48 , 49 ] and in silico tools allowed to classify them as LP or P. In accordance with RNA studies performed by our group and others [ 35 ], these findings indicate that the c.1466A > G p.(Cys489Tyr) acts through two distinct mechanisms: in addition to the deleterious effect on splicing, a significant proportion of residual mutant protein is still produced, whose activity should be investigated by further functional studies.…”

Section: Discussionmentioning

confidence: 75%

“…Data from RNA classical analysis performed in our lab and by other groups [ 23 , 34 , 35 , 45 , 46 , 48 ] were available for 10 out of the 29 NF1 variants of our series ( Table 2 and Table 3 ). Remarkably, in four cases minigene assay revealed a more complex splicing profile compared to cDNA analysis, owing to the presence of multiple aberrant transcripts.…”

Section: Discussionmentioning

confidence: 99%

Hybrid Minigene Assay: An Efficient Tool to Characterize mRNA Splicing Profiles of NF1 Variants

Morbidoni

Baschiera

Forzan

et al. 2021

Cancers

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Neurofibromatosis type 1 (NF1) is caused by heterozygous loss of function mutations in the NF1 gene. Although patients are diagnosed according to clinical criteria and few genotype-phenotype correlations are known, molecular analysis remains important. NF1 displays allelic heterogeneity, with a high proportion of variants affecting splicing, including deep intronic alleles and changes outside the canonical splice sites, making validation problematic. Next Generation Sequencing (NGS) technologies integrated with multiplex ligation-dependent probe amplification (MLPA) have largely overcome RNA-based techniques but do not detect splicing defects. A rapid minigene-based system was set up to test the effects of NF1 variants on splicing. We investigated 29 intronic and exonic NF1 variants identified in patients during the diagnostic process. The minigene assay showed the coexistence of multiple mechanisms of splicing alterations for seven variants. A leaky effect on splicing was documented in one de novo substitution detected in a sporadic patient with a specific phenotype without neurofibromas. Our splicing assay proved to be a reliable and fast method to validate novel NF1 variants potentially affecting splicing and to detect hypomorphic effects that might have phenotypic consequences, avoiding the requirement of patient’s RNA.

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“…DNA were fragmented and exomes were captured with the Agilent SureSelect Human All Exon Kit V5 (Agilent, United States). DNA was sequenced with 200bp reads by Illumina HiSeq2000 platform (Illumina, United States) as previously described (Zhu et al, 2019). Data were mapped to the Genome Reference Consortium Human genome build 37 (GRCh37) and only the variants locating in the coding sequence or splice site regions would be retained.…”

Section: Whole-exome Sequencingmentioning

confidence: 99%

Case Report: A Synonymous Mutation in NF1 Located at the Non-canonical Splicing Site Leading to Exon 45 Skipping

Jin

Yan

et al. 2021

Front. Genet.

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Synonymous mutations are generally considered non-pathogenic because it did not alter the amino acids of the encoded protein. Publications of the associations between synonymous mutations and abnormal splicing have increased recently, however, not much observations available described the synonymous mutations at the non-canonical splicing sites leading to abnormal splicing. In this pedigree, the proband was diagnosed Neurofibromatosis type I due to the presence of typical cafe’ au lait macules and pectus carinatum. Whole-exome sequencing identified a synonymous mutation c.6795C > T (p.N2265N) of the NF1 gene which was located at the non-canonical splicing sites. Reverse transcription polymerase chain reaction followed by Sanger sequencing was carried out, and the skipping of exon 45 was observed. Therefore, the pathogenicity of the synonymous mutation c.6795C > T was confirmed. Our finding expanded the spectrum of pathogenic mutations in Neurofibromatosis type I and provided information for genetic counseling.

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Identification and characterization of NF1 and non-NF1 congenital pseudarthrosis of the tibia based on germline NF1 variants: genetic and clinical analysis of 75 patients

Cited by 28 publications

References 55 publications

Management of spinal deformities and tibial pseudarthrosis in children with neurofibromatosis type 1 (NF-1)

Management of spinal deformities and tibial pseudarthrosis in children with neurofibromatosis type 1 (NF-1)

Hybrid Minigene Assay: An Efficient Tool to Characterize mRNA Splicing Profiles of NF1 Variants

Case Report: A Synonymous Mutation in NF1 Located at the Non-canonical Splicing Site Leading to Exon 45 Skipping

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