Paul Critchley

A variant in the sonic hedgehog regulatory sequence (ZRS) is associated with triphalangeal thumb and deregulates expression in the developing limb

A locus for triphalangeal thumb, variably associated with pre-axial polydactyly, was previously identified in the zone of polarizing activity regulatory sequence (ZRS), a long range limb-specific enhancer of the Sonic Hedgehog (SHH) gene at human chromosome 7q36.3. Here, we demonstrate that a 295T>C variant in the human ZRS, previously thought to represent a neutral polymorphism, acts as a dominant allele with reduced penetrance. We found this variant in three independently ascertained probands from southern England with triphalangeal thumb, demonstrated significant linkage of the phenotype to the variant (LOD = 4.1), and identified a shared microsatellite haplotype around the ZRS, suggesting that the probands share a common ancestor. An individual homozygous for the 295C allele presented with isolated bilateral triphalangeal thumb resembling the heterozygous phenotype, suggesting that the variant is largely dominant to the wild-type allele. As a functional test of the pathogenicity of the 295C allele, we utilized a mutated ZRS construct to demonstrate that it can drive ectopic anterior expression of a reporter gene in the developing mouse forelimb. We conclude that the 295T>C variant is in fact pathogenic and, in southern England, appears to be the most common cause of triphalangeal thumb. Depending on the dispersal of the founding mutation, it may play a wider role in the aetiology of this disorder.

Genetic screening of 202 individuals with congenital limb malformations and requiring reconstructive surgery

Background: Congenital limb malformations (CLMs) are common and present to a variety of specialties, notably plastic and orthopaedic surgeons, and clinical geneticists. The authors aimed to characterise causative mutations in an unselected cohort of patients with CLMs requiring reconstructive surgery. Methods: 202 patients presenting with CLM were recruited. The authors obtained G-banded karyotypes and screened EN1, GLI3, HAND2, HOXD13, ROR2, SALL1, SALL4, ZRS of SHH, SPRY4, TBX5, TWIST1 and WNT7A for point mutations using denaturing high performance liquid chromatography (DHPLC) and direct sequencing. Multiplex ligation dependent probe amplification (MLPA) kits were developed and used to measure copy number in GLI3, HOXD13, ROR2, SALL1, SALL4, TBX5 and the ZRS of SHH. Results: Within the cohort, causative genetic alterations were identified in 23 patients (11%): mutations in GLI3 (n = 5), HOXD13 (n = 5), the ZRS of SHH (n = 4), and chromosome abnormalities (n = 4) were the most common lesions found. Clinical features that predicted the discovery of a genetic cause included a bilateral malformation, positive family history, and having increasing numbers of limbs affected (all p<0.01). Additionally, specific patterns of malformation predicted mutations in specific genes. Conclusions: Based on higher mutation prevalence the authors propose that GLI3, HOXD13 and the ZRS of SHH should be prioritised for introduction into molecular genetic testing programmes for CLM. The authors have developed simple criteria that can refine the selection of patients by surgeons for referral to clinical geneticists. The cohort also represents an excellent resource to test for mutations in novel candidate genes.

A 10-year review of benign and malignant peripheral nerve sheath tumors in a single center: clinical and radiographic features can help to differentiate benign from malignant lesions.

Background: Malignant peripheral nerve sheath tumors are rare, and their aggressive nature mandates treatment in specialist centers. In contrast, benign peripheral nerve sheath tumors are common and are treated by a variety of specialist surgeons, including plastic surgeons. The authors aimed to detect features in the clinical presentation of peripheral nerve sheath tumors that point toward a diagnosis of malignant peripheral nerve sheath tumor and therefore prompt referral to a specialist center. Methods: All histologically diagnosed primary peripheral nerve sheath tumors from January of 1995 to December of 2004 were identified from histopathology records. Notes were reviewed and analyzed with regard to symptoms, signs, radiology, electrophysiology, surgery, and pathology. Statistical comparisons used Fishers exact test and the Mann-Whitney test. Results: During the study period, 32 cases of malignant peripheral nerve sheath tumor in 30 patients were treated. Factors in the clinical evaluation that significantly predicted the presence of malignant peripheral nerve sheath tumor included site, large size, depth in relation to the deep fascia, short duration of symptoms, and pain. Magnetic resonance imaging and computed tomography were sensitive and specific ways of confirming the clinical diagnosis. Interestingly, schwannomata were harder to distinguish from malignant peripheral nerve sheath tumors both clinically and radiologically. Conclusions: The authors have reviewed their institutional experience of peripheral nerve sheath tumors over a 10-year period. Their results will help to focus clinical and radiologic investigation of patients presenting with these tumors.

Nonsense-mediated decay and the molecular pathogenesis of mutations in SALL1 and GLI3.

Mutations in SALL1 and GLI3 are responsible for human limb malformation syndromes. The molecular pathophysiology of these mutations is incompletely understood, and many conclusions have been drawn from studies performed in the mouse. We identified truncating mutations in SALL1 and GLI3 in patients with limb malformation and studied the contribution of nonsense‐mediated decay (NMD) to the expression of mutant mRNA in patient‐derived fibroblasts. Quantification of the relative proportions of mutant and wild‐type alleles was performed by pyrosequencing. In SALL1, a mutant allele causing Townes–Brocks syndrome was unexpectedly resistant to NMD, whereas a different mutation causing a much milder phenotype was susceptible to NMD. In GLI3, all three mutant alleles tested were susceptible to NMD. This work provides novel insights into the molecular pathophysiology of SALL1 and GLI3 mutations, extends the phenotypic spectrum of SALL1 mutations, and provides an example of a human mutation which does not follow the usual accepted positional rules governing mammalian NMD.

Quality of Life and Surgical Outcomes After Soft-Tissue Reconstruction of Complex Oncologic Defects of the Spine and Sacrum.

BACKGROUND Tumor-related spinal surgery has been revolutionized by recent advances in spinal stabilization, modern neuroimaging, and perioperative intensive medicine. This study examines clinical outcomes and factors associated with complications following reconstruction of complex oncologic defects of the spine and sacrum, in an attempt to increase preoperative recognition of high-risk patients with diminished wound-healing capacity and to optimize clinical outcomes in this cohort. METHODS We performed a retrospective analysis of fifty-five consecutive patients who underwent soft-tissue reconstruction with or without osseous stabilization of defects following spinal or sacral tumor resection at a quaternary referral center over a twelve-year period. Surgical outcomes included the prevalence of postoperative complications and success of wound closure at the latest follow-up. Health-related quality-of-life outcomes were assessed using the EORTC QLQ-C30 (European Organization for Research and Treatment of Cancer 30-Item Core Quality of Life Questionnaire) and SF-36 (Short Form-36) questionnaires. RESULTS The mean age of the cohort was 46.7 years (range, eighteen to seventy-one years), with a male preponderance (3:1). Soft-tissue reconstructions (n = 70 flaps) were performed in the fifty-five patients. Overall, 36.3% of patients had wound complications. There was a twofold higher wound complication rate after delayed (60%) compared with immediate (29%) reconstruction (p = 0.03). Patients undergoing delayed reconstruction reported significantly lower SF-36 and EORTC QLQ-C30 scores. CONCLUSIONS Orthoplastic management of spinal tumors should involve a strategy for preoperative recognition of patients at risk of compromised wound-healing. Prophylactic soft-tissue reconstruction can achieve stable definitive wound closure and potentially avoid the need for secondary procedures in appropriately selected patients. LEVEL OF EVIDENCE Prognostic Level IV. See Instructions for Authors for a complete description of levels of evidence.

A misdiagnosed burn: necrotising fasciitis in an elderly patient

A 91-year-old man was admitted having been found unresponsive in his bed. He was previously well, but had sustained a laceration over the dorsum of his forearm after a recent fall. Initial appearances were innocuous, and suggestive of a superficial wound infection. A review of the wound by the plastic surgeons led to the diagnosis of a full-thickness burn, despite a lack of supporting evidence in the history. No clinician who had initially assessed the patient was involved in his ongoing care. Rapid clinical deterioration ensued, with sepsis and deteriorating mental state. Necrosis spread rapidly up his arm, and the diagnosis of necrotising fasciitis was made. The patient was urgently taken to theatre for circumferential excision of the necrotic tissue. He was subsequently managed on the intensive care unit where he made a promising initial recovery, but later died from pneumonia in the ward.

Retrospective audit of 957 consecutive 18F-FDG PET–CT scans compared to CT and MRI in 493 patients with different histological subtypes of bone and soft tissue sarcoma

BackgroundThe use of 18F-FDG PET–CT (PET–CT) is widespread in many cancer types compared to sarcoma. We report a large retrospective audit of PET–CT in bone and soft tissue sarcoma with varied grade in a single multi-disciplinary centre. We also sought to answer three questions. Firstly, the correlation between sarcoma sub-type and grade with 18FDG SUVmax, secondly, the practical uses of PET–CT in the clinical setting of staging (during initial diagnosis), restaging (new baseline prior to definitive intervention) and treatment response. Finally, we also attempted to evaluate the potential additional benefit of PET–CT over concurrent conventional CT and MRI.MethodsA total of 957 consecutive PET–CT scans were performed in a single supra-regional centre in 493 sarcoma patients (excluding GIST) between 2007 and 2014. We compared, PET–CT SUVmax values in relation to histology and FNCCC grading. We compared PET–CT findings relative to concurrent conventional imaging (MRI and CT) in staging, restaging and treatment responses.ResultsHigh-grade (II/III) bone and soft tissue sarcoma correlated with high SUVmax, especially undifferentiated pleomorphic sarcoma, leiomyosarcoma, translocation induced sarcomas (Ewing, synovial, alveolar rhabdomyosarcoma), de-differentiated liposarcoma and osteosarcoma. Lower SUVmax values were observed in sarcomas of low histological grade (grade I), and in rare subtypes of intermediate grade soft tissue sarcoma (e.g. alveolar soft part sarcoma and solitary fibrous tumour). SUVmax variation was noted in malignant peripheral nerve sheath tumours, compared to the histologically benign plexiform neurofibroma, whereas PET–CT could clearly differentiate low from high-grade chondrosarcoma. We identified added utility of PET–CT in addition to MRI and CT in high-grade sarcoma of bone and soft tissues. An estimated 21% overall potential benefit was observed for PET–CT over CT/MRI, and in particular, in ‘upstaging’ of high-grade disease (from M0 to M1) where an additional 12% of cases were deemed M1 following PET–CT.ConclusionsPET–CT in high-grade bone and soft tissue sarcoma can add significant benefit to routine CT/MRI staging. Further prospective and multi-centre evaluation of PET–CT is warranted to determine the actual predictive value and cost-effectiveness of PET–CT in directing clinical management of clinically complex and heterogeneous high-grade sarcomas.