Andrew E. Rosenberg

CDKN2A Gene Deletions and Loss of p16 Expression Occur in Osteosarcomas That Lack RB Alterations

Osteosarcomas often suffer mutations of the RB (retinoblastoma) gene, with resultant inactivation of the pRb protein. pRb is one component in a cell-cycle control pathway that includes the p16 (encoded by the CDKN2A gene) and cyclin-dependent kinase 4 (cdk4, encoded by the CDK4 gene) proteins. We therefore sought to determine whether the CDKN2A and CDK4 genes were altered in those osteosarcomas that lacked RB inactivation. Twenty-one osteosarcomas (2 low-grade and 19 high-grade) were evaluated for homozygous deletion of the CDKN2A gene, CDK4 amplification, and allelic loss of the RB gene, as well as for expression of p16 and pRb proteins. Five high-grade osteosarcomas showed loss of p16 expression; four of these had homozygous CDKN2A deletions, and the fifth had a probable deletion obscured by numerous nonneoplastic, p16-immunopositive multinucleated giant cells. Thus, p16 immunohistochemistry may provide a sensitive means for assessing CDKN2A status. Twelve tumors (including the two low-grade osteosarcomas) were immunopositive for pRb, and nine tumors were immunonegative for pRb. Of the five cases with CDKN2A/p16 alterations, none had allelic loss of the RB gene and all expressed pRb, suggesting that each of these tumors had an intact RB gene. None of the tumors showed CDK4 amplification. No alterations were detected in the two low-grade osteosarcomas. This study suggests that CDKN2A is a tumor suppressor inactivated in osteosarcomas that lack RB mutations and that the p16-pRb cell-cycle control pathway is deregulated in a large number of high-grade osteosarcomas.

Most osteomalacia-associated mesenchymal tumors are a single histopathologic entity: an analysis of 32 cases and a comprehensive review of the literature.

Oncogenic osteomalacia (OO) is a rare paraneoplastic syndrome of osteomalacia due to phosphate wasting. The phosphaturic mesenchymal tumor (mixed connective tissue variant) (PMTMCT) is an extremely rare, distinctive tumor that is frequently associated with OO. Despite its association with OO, many PMTMCTs go unrecognized because they are erroneously diagnosed as other mesenchymal tumors. Expression of fibroblast growth factor-23 (FGF-23), a recently described protein putatively implicated in renal tubular phosphate loss, has been shown in a small number of mesenchymal tumors with known OO. The clinicopathological features of 32 mesenchymal tumors either with known OO (29) or with features suggestive of PMTMCT (3) were studied. Immunohistochemistry for cytokeratin, S-100, actin, desmin, CD34, and FGF-23 was performed. The patients (13 male, 19 female) ranged from 9 to 80 years in age (median 53 years). A long history of OO was common. The cases had been originally diagnosed as PMTMCT (15), hemangiopericytoma (HPC) (3), osteosarcoma (3), giant cell tumor (2), and other (9). The tumors occurred in a variety of soft tissue (21) and bone sites (11) and ranged from 1.7 to 14 cm. Twenty-four cases were classic PMTMCT with low cellularity, myxoid change, bland spindled cells, distinctive “grungy” calcified matrix, fat, HPC-like vessels, microcysts, hemorrhage, osteoclasts, and an incomplete rim of membranous ossification. Four of these benign-appearing PMTMCTs contained osteoid-like matrix. Three other PMTMCTs were hypercellular and cytologically atypical and were considered malignant. The 3 cases without known OO were histologically identical to the typical PMTMCT. Four cases did not resemble PMTMCT: 2 sinonasal HPC, 1 conventional HPC, and 1 sclerosing osteosarcoma. Three cases expressed actin; all other markers were negative. Expression of FGF-23 was seen in 17 of 21 cases by immunohistochemistry and in 2 of 2 cases by RT-PCR. Follow-up (25 cases, 6-348 months) indicated the following: 21 alive with no evidence of disease and with normal serum chemistry, 4 alive with disease (1 malignant PMTMCT with lung metastases). We conclude that most cases of mesenchymal tumor-associated OO, both in the present series and in the reported literature, are due to PMTMCT. Improved recognition of their histologic spectrum, including the presence of bone or osteoid-like matrix in otherwise typical cases and the existence of malignant forms, should allow distinction from other mesenchymal tumors. Recognition of PMTMCT is critical, as complete resection cures intractable OO. Immunohistochemistry and RT-PCR for FGF-23 confirm the role of this protein in PMTMCT-associated OO.

Chondrosarcoma of Bone: An Assessment of Outcome*

BACKGROUND The data on 227 patients who had been managed for a chondrosarcoma at one institution were reviewed to determine the nature of the lesions, the predictors of outcome, and whether there were any ways to change the treatment approaches to improve the results. METHODS The patients were followed for a mean duration of six years (range, three to twenty-five years). The mean age of the patients was forty-seven years (range, nine to eighty-four years). The most prevalent sites of the tumors were the femur (seventy-eight), the pelvis (fifty-one), and the humerus (thirty-nine). The tumors were divided into two groups according to histological grade. Eighty-six tumors (sixteen atypical enchondromas and seventy grade-1 chondrosarcomas) that were locally destructive but were associated with a low likelihood of metastasis were considered to be low-grade. The remaining 141 lesions, which were locally destructive, potentially metastatic, and capable of causing death, were thought to be high-grade. One hundred and three of these 141 lesions were grade 2, and thirty-eight were grade 3 (eighteen of the thirty-eight were grade 3 only, and twenty were both grade 3 and dedifferentiated). Two hundred and twenty-four patients were managed with resection and a limb-sparing procedure; the remaining three patients had an amputation. Postoperative adjuvant radiation was used for fifty-six patients; chemotherapy, for thirty-five; and both radiation and chemotherapy, for nineteen. Flow cytometric patterns were analyzed for 105 patients. RESULTS The patients who had a high-grade tumor were older than those who had a low-grade tumor (mean age [and standard deviation], 50+/-17.0 years compared with 40+/-15.9 years; p < 0.001). Pathological fracture, metastasis, local recurrence, and death were more prevalent in the group that had a high-grade lesion (p < 0.001). Predictors of metastasis and death in that group of patients included local recurrence, a pelvic location of the tumor, a tumor that was more than 100 cubic centimeters in size, a ploidic abnormality (aneuploidy coupled with a high mean DNA index), a histological grade of 3, and a dedifferentiated type of tumor (p < 0.001). CONCLUSIONS Although the data are suggestive, with the numbers available for study we could not detect a significant difference in the rates of pulmonary metastasis and death between the patients who had a grade-3 lesion and those who had a grade-3 lesion that was also dedifferentiated. However, the interval between diagnosis and death was 32+/-22.8 months for the patients who had a grade-3 lesion compared with 5+/-3.7 months for those who had a grade-3 lesion that was also dedifferentiated (p < 0.001). Overall, patients who had had a resection with wide margins (margins extending outside the reactive zone) had a longer duration of survival than did those who had had a so-called marginal resection (margins extending outside the tumor but within the reactive zone) or an intralesional resection (margins within the lesion) (p < 0.04). Adjunctive chemotherapy or radiation, or both (which, it must be noted, was used, without a protocol, in a relatively small number of patients), after an intralesional resection, for recurrent disease, or for distant metastasis did not appear to alter the outcome.

Chondrosarcoma of the base of the skull : A clinicopathologic study of 200 cases with emphasis on its distinction from chordoma

Conventional chondrosarcoma (CSA) of the skull base is an uncommon neoplasm that can resemble chordoma, and indeed it is misdiagnosed frequently as such. This has important clinical implications, because when treated with similar aggressive treatment strategies, CSA has a much better prognosis than chordoma. In an effort to identify those morphologic and immunohistochemical features that help to identify conventional skull base CSA correctly and to understand its prognosis better, particularly compared with chordoma, when treated with surgery and proton beam irradiation, the authors performed a clinicopathologic analysis of 200 CSAs. The patients ranged in age from 10 to 79 years (mean, 39 years), 87 patients were male and 113 patients were female, and most presented with symptoms related to the central nervous system. Approximately 6% of the tumors arose in the sphenoethmoid complex, 28% originated in the clivus, and 66% developed in the temperooccipital junction. Histologically, 15 tumors (7.5%) were classified as hyaline CSA, 59 (29.5%) as myxoid CSA, and 126 (63%) as mixed hyaline and myxoid CSA. A total of 101 (50.5%) tumors were grade 1, 57 (28.5%) had areas of grades 1 and 2, and 42 (21%) were pure grade 2 neoplasms. The vast majority of patients originated from referring hospitals, and the diagnosis was changed prospectively at our institution to CSA from chordoma in 74 patients (37%). Of the tumors studied immunohistochemically, 96 of 97 (98.9%) stained for S-100 protein, 0 of 97 (0%) stained for keratin, and faint staining for epithelial membrane antigen was seen in 7 of 88 tumors (7.95%). All patients underwent high-dose postoperative fractionated precision conformal radiation therapy with a dose that ranged from 64.2 to 79.6 Cobalt-Gray-equivalents (median, 72.1 Cobalt-Gray-equivalents, given in 38 fractions. The 200 patients had a median follow-up of 63 months (range, 2.1 mos - 18.5 yrs). Tumor control was defined as lack of progression by clinical and radiographic assessment. Based on this definition, there were three local recurrences, and two of these patients died of tumor-related complications. The 5- and 10-year local control rates were 99% and 98% respectively, and the 5- and 10-year disease-specific survival rates were both 99%. In contrast to CSA, the 5- and 10-year survival rates of chordoma have been reported to be approximately 51 % and 35% respectively, and in our institution intensive treatment has resulted in 5- and 10-year progression-free survival rates of 70% and 45% respectively. CSA of the skull base can be distinguished reliably from chordoma, and this distinction is important because skull base CSA has an excellent prognosis when treated with surgery and proton beam irradiation, whereas chordomas have a substantially poorer clinical course despite similar aggressive management.

EWSR1‐POU5F1 fusion in soft tissue myoepithelial tumors. A molecular analysis of sixty‐six cases, including soft tissue, bone, and visceral lesions, showing common involvement of the EWSR1 gene

The diagnosis of myoepithelial (ME) tumors outside salivary glands remains challenging, especially in unusual clinical presentations, such as bone or visceral locations. A few reports have indicated EWSR1 gene rearrangement in soft tissue ME tumors, and, in one case each, the fusion partner was identified as either PBX1 or ZNF444. However, larger studies to investigate whether these genetic abnormalities are recurrent or restricted to tumors in soft tissue locations are lacking. Sixty‐six ME tumors mainly from soft tissue (71%), but also from skin, bone, and visceral locations, characterized by classic morphological features and supporting immunoprofile were studied. Gene rearrangements in EWSR1, FUS, PBX1, and ZNF444 were investigated by fluorescence in situ hybridization. EWSR1 gene rearrangement was detected in 45% of the cases. A EWSR1‐POU5F1 fusion was identified in a pediatric soft tissue tumor by 3′Rapid Amplification of cDNA Euds (RACE) and subsequently confirmed in four additional soft tissue tumors in children and young adults. An EWSR1‐PBX1 fusion was seen in five cases, whereas EWSR1‐ZNF444 and FUS gene rearrangement was noted in one pulmonary tumor each. In conclusion, EWSR1 gene rearrangement is a common event in ME tumors arising outside salivary glands, irrespective of anatomical location. EWSR1‐negative tumors were more often benign, superficially located, and showed ductal differentiation, suggesting the possibility of genetically distinct groups. A subset of soft tissue ME tumors with clear cell morphology harbor an EWSR1‐POU5F1 fusion, which can be used as a molecular diagnostic test in difficult cases. These findings do not support a pathogenetic relationship between soft tissue ME tumors and their salivary gland counterparts.

INDIVIDUALIZING MANAGEMENT OF AGGRESSIVE FIBROMATOSES

PURPOSE To examine prognostic indicators in aggressive fibromatoses that may be used to optimize case-specific management strategy. METHODS AND MATERIALS One hundred and seven fibromatoses presenting between 1971 and 1992 were analyzed. The following treatment modalities were utilized: (a) surgery alone for 51 tumors; (b) radiation alone for 15 tumors; and (c) radiation and surgery (combined modality) for 41 tumors. Outcome analysis was based on 5-year actuarial local control rates. RESULTS Control rates among surgery, radiation therapy, and combined modality groups were 69%, 93%, and 72%. Multivariate analysis identified age < 18 years, recurrent disease, positive surgical margins, and treatment with surgery alone as predictors for failure. Patients treated with surgery alone had control rates of 50% (3 of 6) for gross residual, 56% for microscopically positive margins, and 77% for negative margins. Radiation and surgery resulted in rates of 59% for gross residual, 78% for microscopically positive margins, and 100% (6 of 6) for negative margins. For recurrent vs. primary tumors, control was achieved in 48% vs. 77%, 90% vs. 100% (5 of 5), and 67% vs. 79% in the Surgery, Radiation, and Combined modality Groups, respectively. Patients presenting with multiple disease sites tended to have aggressive disease. A radiation dose-control relation to > 60 Gy was seen in patients with unresected or gross residual disease. Of the patients, 23 with disease involving the plantar region had a control rate of 62%, with significantly worse outcomes in children. CONCLUSIONS These results are consistent with those found in the relevant literature. They support primary resection with negative margins when feasible. Radiation is a highly effective alternative in situations where surgery would result in major functional or cosmetic defects. When negative surgical margins are not achieved in recurrent tumors, radiation is recommended. Perioperative radiation should be considered in other high-risk groups (recurrent disease, positive margins, and plantar tumors in young patients). Doses of 60-65 Gy for gross disease and 50-60 Gy for microscopic residual are recommended. Observation may be considered for primary tumors with disease remaining in situ when they are located such that progression would not cause significant morbidity. Although plantar lesions in children may represent a group at high risk for recurrence or aggressive behavior, the greater potential for radiation-induced morbidity in this group must also temper its use. Given the inconsistent nature and treatment response of this tumor, it is fundamental that treatment recommendations should be made based on the risk:benefit analysis for the individual patient, dependent on tumor characteristics and location, as well as patient characteristics and preferences.

Treatment of the patient with stage M0 soft tissue sarcoma.

During the period 1971 to 1985, 220 patients with soft tissue sarcoma of the extremities, torso, and head-neck region were managed by radiation and resectional surgery at the Massachusetts General Hospital (MGH). Actuarial 5-year local control and disease-free survival rates were 86% and 70%, respectively. The success rate improved during this time period. Namely, the local control rates for 1971 to 1975, 1976 to 1980, and 1981 to 1985 were 81%, 81%, and 94%, respectively. For the same time periods, the 5-year disease-free survival rates were 64%, 70%, and 76%. One hundred thirty-one patients were treated with postoperative radiation, and 89 with preoperative radiation. In the most recent 5-year period, the local control rates were 91% and 97% for the two groups (number of patients being 50 and 57 in the post- and preoperative groups, respectively). Treatment by preoperative radiation appears to have a major advantage for patients with very large sarcomas, ie, greater than 15 cm in maximum dimension. None of our patients with local control of grade 1 sarcoma have developed distant metastasis (DM). In contrast, among patients with grade 2 or 3 sarcomas, there is a relentless and progressive increase in the frequency of DM with size of the primary lesion, namely, 6% at less than or equal to 2.5 cm, congruent to 60% at 15 to 20 cm, and congruent to 80% at greater than 20 cm.

USP6 and CDH11 Oncogenes Identify the Neoplastic Cell in Primary Aneurysmal Bone Cysts and Are Absent in So-Called Secondary Aneurysmal Bone Cysts

Aneurysmal bone cyst (ABC) is a locally recurrent bone lesion that has been regarded as a reactive process. Recently, a neoplastic basis in primary ABC was evidenced by demonstration of clonal chromosome band 17p13 translocations that place the USP6 (TRE2 or TRE17) oncogene under the regulatory influence of the highly active CDH11 promoter. Herein, we report CDH11 and/or USP6 rearrangements in 36 of 52 primary ABCs (69%), of which 10 had CDH11-USP6 fusion, 23 had variant USP6 rearrangements without CDH11 rearrangement, and three had variant CDH11 rearrangements without USP6 rearrangement. USP6 and CDH11 rearrangements were restricted to spindle cells in the ABC and were not found in multinucleated giant cells, inflammatory cells, endothelial cells, or osteoblasts. CDH11 and USP6 rearrangements did not correlate with recurrence-free survival, or with other clinicopathological features. CDH11 and USP6 rearrangements were not found in any of 17 secondary ABC associated with giant cell tumor, chondroblastoma, osteoblastoma, and fibrous dysplasia. These findings demonstrate that primary ABC are mesenchymal neoplasms exhibiting USP6 and/or CDH11 oncogenic rearrangements. By contrast, secondary ABC lack CDH11 and USP6 rearrangements, and although morphological mimics of primary ABC, appear to represent a non-specific morphological pattern of a diverse group of non-ABC neoplasms.

Cytokeratin expression in epithelioid vascular neoplasms

Seven epithelioid and eight non-epithelioid vascular tumors were studied by the avidin-biotin-peroxidase method for the presence of endothelial- and epithelial-associated markers, using Ulex europaeus agglutinin-1 (UEA-1) lectin, and antibodies directed against factor VIII-related antigen, (FVIII-RA), vimentin, keratin, carcinoembryonic antigen, and epithelial membrane antigen. The cases included four epithelioid hemangiomas, two epithelioid hemangioendotheliomas (EHE), one epithelioid angiosarcoma (EAS), four common non-epithelioid capillary hemangiomas, and four non-epithelioid angiosarcomas. Staining for FVIII-RA, UEA-1, and vimentin were observed in all cases. The EAS showed staining for keratin in formalin-fixed, paraffin-embedded sections and in frozen sections. Staining for keratin was also observed in frozen sections of one EHE. Both keratin-positive vascular tumors were confirmed with electron microscopy. Carcinoembryonic antigen and epithelial membrane antigen stains were negative in all cases. Our results show that the epithelioid vascular tumors EHE and EAS, in addition to staining for the endothelial markers and vimentin, may also express the epithelial marker keratin. This is important since these tumors may closely resemble carcinomas by routine light microscopy. This study further underscores the importance of using a broad panel of immunohistochemical markers in the diagnostic workup of soft-tissue neoplasms.

Solitary fibrous tumor of the orbit

We report three cases of an orbital soft tissue lesion that fulfills the histologic, immunohistochemical, and electron microscopic criteria for solitary fibrous tumor, an entity previously described as a pleural tumor, but recently reported to occur in other locations. All three patients presented with proptosis. Two of the patients were cured by simple excision, and one patient had two recurrences, the last recurrence incompletely excised. The findings indicate that solitary fibrous tumor can occur in the orbit and, like solitary fibrous tumors of other anatomic sites, may behave in a nonaggressive or occasionally, locally aggressive fashion, with as yet no metastatic potential demonstrated in orbital lesions.