ABSTRACT
BACKGROUND: Deletion of 1p is associated with poor prognosis in neuroblastoma, however selected 1p-intact patients still experience poor outcomes. Since mutations of 1p genes may mimic the deleterious effects of chromosomal loss, we studied the incidence, spectrum and effects of mutational variants in 1p-intact neuroblastoma. METHODS: We characterized the 1p status of 325 neuroblastoma patients, and correlated the mutational status of 1p tumor suppressors and neuroblastoma candidate genes with survival outcomes among 100 1p-intact cases, then performed functional validation of selected novel variants of 1p36 genes identified from our patient cohort. RESULTS: Among patients with adverse disease characteristics, those who additionally had 1p deletion had significantly worse overall survival. Among 100 tumor-normal pairs sequenced, somatic mutations of 1p tumor suppressors KIF1Bß and CHD5 were most frequent (2%) after ALK and ATRX (8%), and BARD1 (3%). Mutations of neuroblastoma candidate genes were associated with other synchronous mutations and concurrent 11q deletion (P = 0.045). In total, 24 of 38 variants identified were novel and predicted to be deleterious or pathogenic. Functional validation identified novel KIF1Bß I1355M variant as a gain-of-function mutation with increased expression and tumor suppressive activity, correlating with indolent clinical behavior; another novel variant CHD5 E43Q was a loss-of-function mutation with decreased expression and increased long-term cell viability, corresponding with aggressive disease characteristics. CONCLUSIONS: Our study showed that chromosome 1 gene mutations occurred frequently in 1p-intact neuroblastoma, but may not consistently abrogate the function of bonafide 1p tumor suppressors. These findings may augment the evolving model of compounding contributions of 1p gene aberrations toward tumor suppressor inactivation in neuroblastoma.
Subject(s)
Genes, Tumor Suppressor , Neuroblastoma , Chromosome Aberrations , Chromosome Deletion , Chromosomes, Human, Pair 1/genetics , Cohort Studies , DNA Helicases/genetics , Humans , Mutation , Nerve Tissue Proteins/genetics , Neuroblastoma/genetics , Neuroblastoma/pathologyABSTRACT
Ex vivo evaluation of personalized models can facilitate individualized treatment selection for patients, and advance the discovery of novel therapeutic options. However, for embryonal malignancies, representative primary cultures have been difficult to establish. We developed patient-derived cell cultures (PDCs) from chemo-naïve and post-treatment neuroblastoma tumors in a consistent and efficient manner, and characterized their in vitro growth dynamics, histomorphology, gene expression, and functional chemo-response. From 34 neuroblastoma tumors, 22 engrafted in vitro to generate 31 individual PDC lines, with higher engraftment seen with metastatic tumors. PDCs displayed characteristic immunohistochemical staining patterns of PHOX2B, TH, and GD2 synthase. Concordance of MYCN amplification, 1p and 11q deletion between PDCs and patient tumors was 83.3%, 72.7%, and 80.0% respectively. PDCs displayed a predominantly mesenchymal-type gene expression signature and showed upregulation of pro-angiogenic factors that were similarly enriched in culture medium and paired patient serum samples. When tested with standard-of-care cytotoxics at human Cmax -equivalent concentrations, MYCN-amplified and non-MYCN-amplified PDCs showed a differential response to cyclophosphamide and topotecan, which mirrored the corresponding patients' responses, and correlated with gene signatures of chemosensitivity. In this translational proof-of-concept study, early-phase neuroblastoma PDCs enriched for the mesenchymal cell subpopulation recapitulated the individual molecular and phenotypic profile of patient tumors, and highlighted their potential as a platform for individualized ex vivo drug-response testing.
Subject(s)
Homeodomain Proteins/genetics , N-Myc Proto-Oncogene Protein/genetics , Neuroblastoma/drug therapy , Transcription Factors/genetics , Tyrosine 3-Monooxygenase/genetics , Animals , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cyclophosphamide/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Heterografts , Humans , Mice , N-Acetylgalactosaminyltransferases/genetics , Neuroblastoma/genetics , Neuroblastoma/pathology , Precision Medicine , Topotecan/pharmacology , Transcriptome/geneticsABSTRACT
BACKGROUND: Primary paediatric epidural sarcomas are extremely rare. Overall, there remains a paucity of knowledge in paediatric epidural sarcomas owing to the infrequent number of cases. The Archer FusionPlex Sarcoma Kit (ArcherDX, Inc) is a next-generation sequencing assay that has been reported to be a useful technique to detect recurrent fusion in sarcomas. We report the molecular exploration of 3 primary paediatric epidural sarcomas-one in the cranium (mesenchymal chondrosarcoma) and 2 in the spine (mesenchymal chondrosarcoma and Ewing sarcoma respectively). CASE PRESENTATION: This is a study approved by the hospital ethics board. Clinico-pathological information from 3 consenting patients with primary epidural sarcomas was collected. These selected tumours are interrogated via Archer FusionPlex Sarcoma Kit (ArcherDX, Inc) for genomic aberrations. Results were validated with RT-PCR and Sanger sequencing. All findings are corroborated and discussed in concordance with current literature. Our findings show 2 variants of the HEY1-NCOA2 gene fusion: HEY1 (exon 4)-NCOA2 (exon 13) and HEY1 (exon 4)-NCOA2 (exon 14), in both mesenchymal chondrosarcoma patients. Next, the Ewing sarcoma tumour is found to have EWSR1 (exon 10)-FLI1 (exon 8) translocation based on NGS. This result is not detected via conventional fluorescence in situ testing. CONCLUSIONS: This is a molecularly-centered study based on 3 unique primary paediatric epidural sarcomas. Our findings to add to the growing body of literature for these exceptionally rare and malignant neoplasms. The authors advocate global collaborative efforts and in-depth studies for targeted therapy to benefit affected children.
Subject(s)
Epidural Neoplasms/diagnosis , Sarcoma/diagnosis , Age Factors , Biomarkers, Tumor , Biopsy , Child , Chondrosarcoma, Mesenchymal/diagnosis , Chondrosarcoma, Mesenchymal/genetics , DNA Mutational Analysis , Epidural Neoplasms/genetics , Female , Humans , Magnetic Resonance Imaging , Sarcoma/genetics , Sarcoma, Ewing/diagnosis , Sarcoma, Ewing/genetics , Symptom AssessmentABSTRACT
AIMS: Clear cell sarcoma of the kidney (CCSK) is a rare paediatric renal malignant tumour. The majority of CCSKs have internal tandem duplications (ITDs) of the BCOR gene, whereas a minority have the YWHAE-NUTM2 gene fusion. A third 'double-negative' (DN) category comprises CCSKs with neither BCOR ITDs nor YWHAE-NUTM2 fusion. The aim of this study was to characterise 11 histologically diagnosed CCSKs immunohistochemically (with CCND1, BCOR and CCNB3 stains) and genetically. METHODS AND RESULTS: By next-generation sequencing, 10 cases (90.9%) had BCOR exon 15 ITDs, with positive BCOR immunoreactivity being found in four (36%) or eight (72%) cases, depending on the antibody clone. By reverse transcription polymerase chain reaction, none had the YWHAE-NUTM2 fusion. The DN case had a BCOR-CCNB3 fusion and strong nuclear CCNB3 and BCOR immunoreactivity. Quantitative polymerase chain reaction showed markedly elevated BCOR expression in this case, whereas BCOR ITD cases had lower levels of elevated BCOR expression. CONCLUSIONS: The majority of the CCSKs in our cohort had BCOR ITDs, and none had the YWHAE-NUTM2 fusion. We verified the strong, diffuse cyclin D1 (CCND1) immunoreactivity in CCSKs described in recent reports. BCOR immunoreactivity was not consistently positive in all CCSKs with BCOR ITDs, and therefore cannot be used as a diagnostic immunohistochemical stain to identify BCOR ITD cases. The DN case was a BCOR-CCNB3 fusion sarcoma. BCOR-CCNB3 sarcoma is typically a primary bone sarcoma affecting male adolescents, and this is the first report of it presenting in a kidney of a young child as a CCSK. The full spectrum of DN CCSKs awaits more comprehensive characterisation.
Subject(s)
Cyclin B/genetics , Kidney Neoplasms/genetics , Oncogene Proteins, Fusion/genetics , Proto-Oncogene Proteins/genetics , Repressor Proteins/genetics , Sarcoma, Clear Cell/genetics , Child , Child, Preschool , Exons , Female , Humans , MaleABSTRACT
PURPOSE: Posterior fossa syndrome (PFS) is a serious complication faced by neurosurgeons and their patients, especially in paediatric medulloblastoma patients. The uncertain aetiology of PFS, myriad of cited risk factors and therapeutic challenges make this phenomenon an elusive entity. The primary objective of this study was to identify associative factors related to the development of PFS in medulloblastoma patient post-tumour resection. METHODS: This is a retrospective study based at a single institution. Patient data and all related information were collected from the hospital records, in accordance to a list of possible risk factors associated with PFS. These included pre-operative tumour volume, hydrocephalus, age, gender, extent of resection, metastasis, ventriculoperitoneal shunt insertion, post-operative meningitis and radiological changes in MRI. Additional variables included molecular and histological subtypes of each patient's medulloblastoma tumour. Statistical analysis was employed to determine evidence of each variable's significance in PFS permanence. RESULTS: A total of 19 patients with appropriately complete data was identified. Initial univariate analysis did not show any statistical significance. However, multivariate analysis for MRI-specific changes reported bilateral DWI restricted diffusion changes involving both right and left sides of the surgical cavity was of statistical significance for PFS permanence. CONCLUSION: The authors performed a clinical study that evaluated possible risk factors for permanent PFS in paediatric medulloblastoma patients. Analysis of collated results found that post-operative DWI restriction in bilateral regions within the surgical cavity demonstrated statistical significance as a predictor of PFS permanence-a novel finding in the current literature.
Subject(s)
Cerebellar Neoplasms/surgery , Cranial Fossa, Posterior/diagnostic imaging , Cranial Fossa, Posterior/pathology , Diffusion Magnetic Resonance Imaging , Medulloblastoma/surgery , Neurosurgical Procedures/adverse effects , Postoperative Complications/diagnostic imaging , Adolescent , Child , Child, Preschool , Databases, Bibliographic/statistics & numerical data , Female , Glasgow Coma Scale , Humans , Male , Multivariate Analysis , Postoperative Period , Retrospective StudiesSubject(s)
Gene Fusion , Proteins/genetics , Proto-Oncogene Proteins c-ret/genetics , Soft Tissue Neoplasms/genetics , Thigh/pathology , Antigens, CD34/genetics , Antigens, CD34/metabolism , Child, Preschool , Humans , Male , Proteins/metabolism , Proto-Oncogene Proteins c-ret/metabolism , S100 Proteins/genetics , S100 Proteins/metabolism , Soft Tissue Neoplasms/metabolism , Soft Tissue Neoplasms/pathologySubject(s)
Brain Edema/etiology , Brain Neoplasms/complications , Cerebral Peduncle/pathology , Germinoma/complications , Brain Neoplasms/diagnostic imaging , Brain Neoplasms/pathology , Brain Neoplasms/surgery , Child , Germinoma/diagnostic imaging , Germinoma/pathology , Humans , Ki-67 Antigen/metabolism , Magnetic Resonance Imaging , Magnetic Resonance Spectroscopy , Male , Neurosurgery/methods , Octamer Transcription Factor-3/metabolism , Proteins/metabolism , Proto-Oncogene Proteins c-kit/metabolismSubject(s)
Arnold-Chiari Malformation/complications , Medulloblastoma/complications , Syringomyelia/complications , Arnold-Chiari Malformation/diagnostic imaging , Brain/diagnostic imaging , Brain/metabolism , Child , Humans , Image Processing, Computer-Assisted , Magnetic Resonance Imaging , Male , Medulloblastoma/diagnostic imaging , Spinal Cord/diagnostic imaging , Spinal Cord/metabolism , Synaptophysin/metabolism , Syringomyelia/diagnostic imagingABSTRACT
Pediatric mesotheliomas are rare and their pathogenesis remains undefined. In this study, we report 5 cases of malignant mesothelioma in children, characterized by fusions involving the anaplastic lymphoma kinase (ALK) gene. Four cases occurred in females involving the abdominal cavity and were characterized by a pure epithelioid morphology. The fifth arose in the tunica vaginalis of a 15-year-old male and displayed a biphasic epithelioid-sarcomatoid phenotype. All cases demonstrated the classic morphologic and immunohistochemical features of malignant mesothelioma, including tubulopapillary architecture and cuboidal epithelioid cells with eosinophilic cytoplasm and uniform nuclei with vesicular chromatin. Immunohistochemically, all cases showed labeling for ALK, cytokeratins, WT1, and calretinin, while lacking expression of adenocarcinoma immunomarkers. Four cases demonstrated weak-moderate labeling for PAX8 protein, which resulted in diagnostic challenges with primary peritoneal serous carcinoma. The ALK genetic abnormalities were investigated by a combination of molecular methods. Archer FusionPlex was performed in 2 cases, showing fusions between ALK with either STRN or TPM1 genes, resulting in a transcript that retained the ALK kinase domain. One case was further studied by DNA targeted sequencing, but no additional genetic alterations were observed. In 1 case, cytogenetic analysis showed the presence of a t(2;15)(p23;q22) and fluorescence in situ hybridization confirmed the ALK gene break-apart. In the remaining 2 cases, ALK gene rearrangements were demonstrated by fluorescence in situ hybridization. Unlike adult mesotheliomas, which are tightly linked to asbestos exposure, often show loss of BAP1 expression and have complex karyotypes, ALK-rearranged mesothelioma appears to be similar to other fusion-positive mesotheliomas, such as those harboring EWSR1/FUS-ATF1 fusions, sharing significant morphologic overlap, occurring in young patients and displaying a simple, translocation-driven genetic profile.
Subject(s)
Abdominal Neoplasms/genetics , Anaplastic Lymphoma Kinase/genetics , Biomarkers, Tumor/genetics , Gene Fusion , Gene Rearrangement , Mesothelioma/genetics , Testicular Neoplasms/genetics , Abdominal Neoplasms/enzymology , Abdominal Neoplasms/pathology , Adolescent , Biomarkers, Tumor/analysis , Child , Female , Genetic Predisposition to Disease , Humans , Immunohistochemistry , Male , Mesothelioma/enzymology , Mesothelioma/pathology , Molecular Diagnostic Techniques , Testicular Neoplasms/enzymology , Testicular Neoplasms/pathologyABSTRACT
Importance: Three-dimensionally printed nasopharyngeal swabs (3DP swabs) have been used to mitigate swab shortages during the coronavirus disease 2019 (COVID-19) pandemic. Clinical validation for diagnostic accuracy and consistency, as well as patient acceptability, is crucial to evaluate the swab's performance. Objective: To determine the accuracy and acceptability of the 3DP swab for identifying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Design, Setting, and Participants: A diagnostic study was conducted from May to July 2020 at 2 tertiary care centers in Singapore with different reference swabs (FLOQSwab [COPAN Diagnostics] or Dacron swab [Deltalab]) and swab processing techniques (wet or dry) to evaluate the performance of the 3DP swab compared with traditional, standard-of-care nasopharyngeal swabs used in health care institutions. The participants were patients with COVID-19 in the first 2 weeks of illness and controls with acute respiratory illness with negative test results for SARS-CoV-2. Paired nasopharyngeal swabs were obtained from the same nostril and tested for SARS-CoV-2 by reverse-transcriptase polymerase chain reaction. The sequence of swabs was randomized based on odd and even participant numbers. Main Outcomes and Measures: Primary outcome measures were overall agreement (OA), positive percentage agreement (PPA), and negative percentage agreement of the 3DP swab compared with reference swabs. Secondary outcome measures were the correlation of cycle threshold (Ct) values of both swabs. Results: The mean (SD) age of participants was 45.4 (13.1) years, and most participants were men (87 of 89 [97.8%]), in keeping with the epidemiology of the COVID-19 pandemic in Singapore. A total of 79 patients with COVID-19 and 10 controls were recruited. Among the patients with COVID-19, the overall agreement and PPA of the 3DP swab was 91.1% and 93.5%, respectively, compared with reference swabs. The PPA was 100% for patients with COVID-19 who were tested within the first week of illness. All controls tested negative. The reverse-transcriptase polymerase chain reaction Ct values for the ORF1ab and E-gene targets showed a strong correlation (intraclass correlations coefficient, 0.869-0.920) between the 3DP and reference swab on independent testing at each institution despite differences in sample processing. Discordant results for both gene targets were observed only at high Ct values. Conclusions and Relevance: In this diagnostic study of 79 patients with COVID-19 and 10 controls, the 3DP swab performed accurately and consistently across health care institutions and could help mitigate strained resources in the escalating COVID-19 pandemic.
Subject(s)
COVID-19 Nucleic Acid Testing/instrumentation , COVID-19/diagnosis , Nasopharynx/virology , Printing, Three-Dimensional , Adult , Equipment Design , Humans , Middle Aged , Pandemics , SARS-CoV-2ABSTRACT
Congenital myeloproliferative disorders and transient leukemic disorders have been described in the perinatal period in infants with trisomy 21 (Down syndrome). We report a novel case of a neonate with trisomy 21 with GATA1-mutated congenital myeloproliferative disorder complicated by placental fetal thrombotic vasculopathy featuring chorionic vessel leukemic thrombi, fetal circulation vascular injuries, and large aggregates of avascular villi. These thrombotic and vasculopathic changes within the placenta are likely a reflection of the hypercoagulable state caused by the myeloproliferative disorder. Placental fetal thrombotic vasculopathy is associated with adverse outcomes for the infant, and should be documented during formal pathological examination of the placenta.
Subject(s)
Down Syndrome/complications , GATA1 Transcription Factor/genetics , Myeloproliferative Disorders/congenital , Thrombosis/congenital , Vascular Diseases/congenital , Adult , Down Syndrome/pathology , Female , Humans , Infant , Myeloproliferative Disorders/complications , Myeloproliferative Disorders/genetics , Myeloproliferative Disorders/pathology , Pregnancy , Pregnancy Trimester, Third , Thrombosis/chemically induced , Thrombosis/genetics , Thrombosis/pathology , Vascular Diseases/genetics , Vascular Diseases/pathologyABSTRACT
Klinefelter syndrome is a clinical syndrome with a distinct 47, XXY karyotype. Patients are characterized by a tall eunuchoid stature, small testes, hypergonotrophic hypogonadism, gynecomastia, learning difficulties and infertility. These patients have also been found to have raised estrogen levels. We report a 16 year old boy with Klinefelter syndrome presenting to our institution with gross hematuria. Cystoscopy and biopsy revealed the diagnosis of pseudomembranous trigonitis. Immunohistochemical stains showed an increase in estrogen and progesterone receptors in the trigone area but not in the rest of the bladder. In view of the patient's mildly raised estrogen levels and the histological findings, we postulate that estrogen is the driver of the development of pseudomembranous trigonitis. This is the first reported case of pseudomembranous trigonitis seen in association with Klinefelter syndrome, and also the first case of pseudomembranous trigonitis occurring within the male adolescent age group.
Subject(s)
Cystitis/pathology , Klinefelter Syndrome/complications , Urinary Bladder/pathology , Cystitis/complications , Humans , Immunohistochemistry , Male , Metaplasia/pathology , Young AdultABSTRACT
Fetal thrombotic vasculopathy (FTV) is a thrombo-occlusive disorder of the placenta that has been reported in association with perinatal conditions such as cardiac abnormalities, neurological injury, and perinatal liver disease. These complications are related to fetal circulation vascular compromise. We herein report a previously undocumented association of congenital intestinal atresia and placental FTV. Vascular occlusion of the fetal mesenteric vessels has been hypothesized to result in congenital intestinal atresia. Our report provides support for this vascular hypothesis and illustrates the value of formal pathological examination of the placenta in explaining this occurrence of congenital intestinal atresia.