Oliveira, André M.

ObjectiveThe relationship between inflammation, obesity, and adverse metabolic conditions is associated with adipose tissue macrophages (ATM). This study compared the measurements of human ATM using flow cytometry, immunohistochemistry (IHC), and real‐time polymerase chain reaction (RT‐PCR) of ATM markers.MethodsA new software program (AMCounter) was evaluated to help measure ATM using IHC, and this was compared to flow cytometry and RT‐PCR.ResultsIHC had good intraindividual reproducibility for total (CD68), proinflammatory (CD14), and anti‐inflammatory (CD206) ATM. The AMCounter improved interreader agreement and was more time efficient. Flow cytometry had acceptable intraindividual reproducibility for the percentage of CD68+ cells that were CD14+ or CD206+, but not for ATMs per gram of tissue. ATMs per gram of tissue was much greater using IHC than flow cytometry. The flow cytometry and IHC measures of ATM from the same biopsies were not correlated. There were statistically significant correlations between RT‐PCR CD68 and IHC CD68, CD14, and CD206 ATMs per 100 adipocytes. Also of interest were statistically significant correlations between RT‐PCR CD68 and IHC CD68, CD14, and adipose flow cytometry measures of CD68+, CD68+/CD14+, and CD68+/CD206+ ATMs per gram of tissue.ConclusionsThe AMCounter software helps provide reproducible and efficient measures of IHC ATMs. Flow cytometry, IHC, and RT‐PCR measures of adipose inflammation provide somewhat different information.

Pulmonary vein stenosis (PVS) is a luminal narrowing of extrapulmonary pulmonary veins. In pediatric patients, it arises following repair of congenital heart disease, particularly anomalous pulmonary venous return; in lung disease, especially prematurity; and rarely in isolation. The etiology is unknown and the course often fatal without lung transplantation. We hypothesized that systematic clinicopathologic review of pediatric PVS could provide further pathogenic insight. We included patients who underwent first resection of pulmonary venous tissue for symptomatic PVS at our pediatric referral center from 1995 to 2014. Clinical records and hematoxylin and eosin slides were reviewed. Subsets were immunostained for smooth muscle actin, Ki-67, β-catenin, estrogen receptor, and other markers and analyzed forUSP6gene rearrangement. A total of 97 patients (57% male; median age: 6 mo) were identified. Overall, 59 (61%) had prior congenital heart disease repair, 35 involving pulmonary vein manipulation. Samples included 213 separate anatomic sites (median: 2/patient). Histologically, all showed sparsely cellular intimal expansion composed of haphazardly arranged fibroblasts with slender nuclei in myxoid matrix. This tissue merged with underlying collagen. Most samples had a variably continuous sheath of cardiomyocytes. Ancillary tests supported a reactive fibroblastic proliferation; in particular, fibroblasts showed cytoplasmic β-catenin localization, no estrogen receptor expression, and noUSP6rearrangement. At last follow-up (mean: 2.3 y), 46% of patients had died of disease. Pediatric PVS uniformly consists of a paucicellular fibrointimal proliferation, irrespective of clinical scenario. It may be best conceived of as a form of reactive hyperplasia. As with other forms of vascular remodeling, trauma (iatrogenic or occult) is likely an inciting factor. A comprehensive understanding of the surgical pathology of PVS may further inform therapeutic strategies in this lethal disease.

PURPOSE: The p27kip1 protein (p27) is a cyclin-dependent kinase inhibitor that has been shown to be an independent prognostic factor in a variety of human neoplasms. Low expression of p27 tends to occur in more aggressive neoplasms. The role of p27 as an independent prognostic factor in the spectrum of myxoid and round-cell liposarcomas has not been examined. MATERIALS AND METHODS: Forty-seven cases of myxoid and round-cell liposarcomas were examined. Clinicopathologic features and immunohistochemical expression of p27 and Ki-67 antigen were studied in all cases. Survival analysis was performed using the log-rank test and the Cox multivariate regression model. RESULTS: The male:female ratio was 1.4:1, and the mean age at diagnosis was 45 years. The tumors were located in the lower extremities (94%) and retroperitoneum (6%). The median tumor size was 13.5 cm. The median follow-up was 6.3 years, and the overall 5- and 10-year survival rates were 76% and 67%, respectively. Low expression of p27 was identified in 34 cases (72%) and correlated with decreased metastasis-free (P = .026) and overall survival (P = .008). In a multivariate analysis, only round-cell differentiation and low expression of p27 independently predicted decreased metastasis-free and overall survival. CONCLUSION: p27 expression predicts the clinical behavior of myxoid and round-cell liposarcomas, even in neoplasms with few or no round-cell differentiation.

Although the majority of giant cell tumors (GCTs) of the bone occur in adult patients, occasionally they arise in the pediatric population. In this setting they may be mistaken for tumors more commonly seen in this age group, including osteosarcoma, aneurysmal bone cyst, and chondroblastoma. All cases of primary GCT of the bone arising in patients 18 years and below were retrieved from our institutional archives and examined with emphasis on the evaluation of various morphologic patterns. Clinical/radiologic records were reviewed when available. Analysis for H3F3A/H3F3B mutations was performed in a subset of cases. Sixty-three (of 710) patients treated at our institution for GCT were 18 years of age and below. The following morphologic patterns were identified: fibrosis (31 cases, 49%), reactive-appearing bone (26, 41%), cystic change (7, 11%), foamy histiocytes (6, 10%), secondary aneurysmal bone cyst (3, 5%), and cartilage (2, 3%). Infarct-like necrosis was present in 17 tumors (27%), and the mitotic rate ranged from 0 to 35 mitoses/10 high-power fields (median 5 mitoses/10 high-power field). Follow-up information (n=55; 6 mo to 69.6 y; median, 11.6 y) showed 21 patients with local recurrence (38%) and 2 patients with lung metastasis (4%). Polymerase chain reaction with sequencing showed that 5 of 5 tested cases harbored H3F3A mutations. In summary, GCT arising in the pediatric population is rare, representing 9% of GCTs seen at our institution. The morphologic spectrum of these tumors is broad and similar to that seen in patients above 18 years of age. It is important to recognize that matrix formation may be observed in GCT, including reactive-appearing bone and cartilage, as well as areas of fibrosis mimicking osteoid production, to avoid misclassification as osteosarcoma or other giant cell–rich lesions common in children.

AimsBiphenotypic sinonasal sarcoma (SNS) is a locally aggressive tumour that occurs in the sinonasal region. PAX3–MAML3 has recently been identified as a recurrent fusion gene event in this entity; however, a subset of tumours harbour alternative PAX3 rearrangement without the involvement of MAML3. In this study we sought to characterize the molecular profile of a large series of cases, with a special emphasis on tumours with alternative fusions.Methods and resultsForty‐four examples of SNS were screened by fluorescence in‐situ hybridization and reverse transcription polymerase chain reaction to better characterize its molecular profile and identify potential novel fusion genes. Twenty‐four were positive for PAX3–MAML3 (55%), 15 showed rearrangements of PAX3 without MAML3 involvement (34%), one showed rearrangement of MAML3 without PAX3 involvement, and four were negative for the involvement of either gene (9%). Among 15 cases with PAX3 involvement only, three were found to harbour PAX3–FOXO1. Two of these cases arose in the nasal cavities of female patients (aged 31 and 47 years), and one showed bilateral involvement of the nasal cavities of a 35‐year‐old male. A fourth case involved the skull base of a 47‐year‐old male, and was positive for PAX3–NCOA1. Patients with fusion‐negative tumours were slightly older.ConclusionMore than half of the SNSs in this series were positive for PAX3–MAML3. However, a subset of tumours may harbour alternative PAX3 fusion genes or show no involvement of PAX3. Except for a possible weak association between age and molecular profile, the overall morphological and immunophenotypic features of all cases seem to be similar. Because of the rarity of these tumours, the impact of the molecular profile on the clinical course of these tumours remains to be determined.

Abstract Bone and soft tissue tumors (BSTT) are relatively poorly understood, hampering the development of effective therapies. Here we report a critical effector pathway for the ubiquitin-specific protease 6 (USP6)/TRE17 oncogene, which is overexpressed upon chromosome translocation in various human tumors, including aneurysmal bone cyst (ABC) and the related benign lesion nodular fasciitis. Ectopic expression of USP6 is known to drive formation of tumors, which recapitulate key features of ABC and nodular fasciitis; however, the identity of USP6′s relevant substrates has been obscure. Here we report that the Jak1–STAT3 signaling pathway serves as an essential effector of USP6 in BSTT formation. We found that USP6 directly deubiquitinated Jak1, leading to its stabilization and activation of STAT3. The tumorigenic potential of USP6 was attenuated significantly by CRISPR-mediated deletion of Jak1 or STAT3, or by administration of a Jak family inhibitor. Analysis of primary clinical samples of nodular fasciitis confirmed the activation of a Jak1–STAT3 gene signature in vivo. Together, our studies highlight Jak1 as the first identified substrate for USP6, and they offer a mechanistic rationale for the clinical investigation of Jak and STAT3 inhibitors as therapeutics for the treatment of bone and soft tissue tumors along with other neoplasms driven by USP6 overexpression. Cancer Res; 76(18); 5337–47. ©2016 AACR.

A cancer-promoting histone protein Mutations in the chromatin protein histone H3 are found in a number of pediatric cancers. The lysine-36–to–methionine (K36M) “oncohistone” mutation is seen in almost all chondroblastomas. Fang et al. show that the K36M mutant histones inhibit the normal methylation of this same residue in wild-type H3 histones. They do so by interfering with the enzymes that normally methylate this residue. The altered chromatin methylation patterns alter the expression of known cancer-related genes and impart cancer-related characteristics to the chondrocyte cells. Science , this issue p. 1344

Significance Ubiquitin-specific protease 6 (USP6) is a deubiquitylase that is overexpressed by chromosome translocation in two human neoplasms, aneurysmal bone cyst and nodular fasciitis. The relevant substrates of this ubiquitin-specific protease are not clear. Here, we identify the Wnt receptor Frizzled (Fzd) as a key target of the USP6 oncogene. Increased expression of USP6 increases the membrane abundance of Fzd, and hence increases cellular sensitivity to Wnts. USP6 opposes the activity of the ubiquitin ligase and tumor suppressor ring finger protein 43 (RNF43). This study identifies a new mechanism for pathological Wnt pathway activation in human disease and suggests a new approach to regulate Wnt activity therapeutically.