Apoptotic and proliferative markers in chordomas: a study of 26 tumors

Few studies have examined the role of cell proliferation and apoptotic markers in chordomas. This study retrospectively reviews the clinicopathologic features of 26 chordomas and examines MIB-1, p53, bcl-2, and cyclin D1 immunoreactivity in these neoplasms. Patients ranged in age from 34 to 78 years (mean, 60.7 years) and included 14 females. The most common presentations included lower back pain (N = 15) and headaches (N = 10). Sixteen tumors arose in the lumbosacral region and 10 in the clivus. Initial surgery included biopsy (N = 17), subtotal resection (N = 4), and gross total resection (N = 5). The single highest mitosis count per 10 high power fields ranged from 0 to 6 (mean, 1). Marked nuclear pleomorphism was identified in seven tumors. Marked hypercellularity was seen in two tumors. Focal necrosis was identified in seven tumors. MIB-1 labeling indices (LI) in 22 tumors ranged from 0 to 3.8 (mean, 0.5). Cyclin D1 LI ranged from 0 to 82.4 (mean, 35.6). Seven tumors had positive p53 immunostaining and three demonstrated focal positive staining with bcl-2 antibody. Five tumors locally recurred; two patients developed metastatic disease. Thirteen patients received adjuvant chemotherapy and/or radiation therapy. At last known follow-up, seven patients died with tumor (12 to 132 months follow-up). Five additional patients died, two without tumor at 36 and 72 months follow-up and three patients in whom the tumor status at death was not known. Seven patients were alive with evidence of tumor (1 to 120 months) and five patients were alive without evidence of tumor (12 to 84 months). Clinical follow-up was not available in one patient. In conclusion, the low MIB-1 LIs and the lack of p53 and bcl-2 staining is in keeping with the low-grade nature of most chordomas. High cyclin D1 LIs may be reflective of a tendency to accumulate cyclin D1 protein; however, there appears to be a block in the effect of cyclin D1 on cell proliferation in these tumors. Cyclin D1, MIB-1, p53, and bcl-2 immunostaining does not appear to improve one's ability to predict behavior versus routine light microscopy.     

Subependymomas: clinicopathologic study of 14 tumors, including comparative MIB-1 immunohistochemical analysis with other ependymal neoplasms.

BACKGROUND: Subependymomas are uncommonly encountered ependymal tumors, which are important to distinguish from ordinary ependymomas because of their generally better prognosis. OBJECTIVE: To review the clinicopathologic features and MIB-1 labeling indices (marker of cell proliferation) of 14 subependymomas. DESIGN: Retrospective review of 14 subependymomas encountered in a tertiary care setting. RESULTS: Fourteen ependymomas presenting in 8 men and 6 women between the ages of 18 and 78 years (mean, 53.6 years) comprise the study group. The most common clinical presentations included ataxia (n = 4), dizziness/vertigo (n = 3), nausea/vomiting (n = 3), headaches (n = 3), and incidental finding at autopsy (n = 2). Tumor locations included fourth ventricle (n = 7), lateral ventricle (n = 4), third ventricle (n = 2), and thoracic spinal cord (n = 1). Eight patients underwent gross total resection, and 4 had subtotal resection. Tumors were characterized by clustering of cell nuclei arranged against a fibrillary background. Focal cystic degeneration was seen in 10 tumors, hemosiderin deposition in 8 tumors, sclerotic vessels in 8 tumors, calcifications in 5 tumors, and focal nuclear pleomorphism in 2 tumors. Mitotic figures, vascular endothelial proliferation, and necrosis were not seen in any of these tumors. Cell proliferation marker MIB-1 labeling indices (percentage of positive staining tumor cells) ranged from 0 to 1.4 (mean, 0.3). In comparison, 13 myxopapillary ependymomas had labeling indices ranging from 0 to 5.5 (mean, 1.1). Thirty-nine low-grade ependymomas had MIB-1 labeling indices of 0.1 to 5.4 (mean, 1.1). Fourteen anaplastic/malignant ependymomas had MIB-1 labeling indices ranging from 0.4 to 34.0 (mean, 12.8). One subependymoma was treated with radiation therapy. Six patients were alive with no evidence of tumor at a mean follow-up of 94.4 months. Two patients were alive with residual tumor (follow-up of 4 and 53 months). Two patients died with tumor at 0.67 and 43.4 months. One patient was lost to follow-up, 1 is a recent case, and 2 were incidental findings at autopsy. None of the patients developed tumor recurrence. CONCLUSIONS: Subependymomas are generally low-grade lesions, as evidenced by their benign clinical course and low MIB-1 labeling indices. Compared with other ependymal tumors, subependymomas have the lowest rate of cell proliferation as evidenced by MIB-1 immunostaining.     

Clinicopathologic study of forty-four histologically pure supratentorial oligodendrogliomas

Few studies in recent years have specifically focused on pure oligodendroglial neoplasms. We retrospectively reviewed the clinicopathologic features of 44 patients with supratentorial oligodendroglioma diagnosed over a 19-year period (1974 to 1993). The study group consisted of 44 patients (age range, 8 to 69 years; median, 42 years), including 31 males. Thirty-one initially resected tumors (70%) were low grade and 13 (30%) were high grade (anaplastic). Using the St Anne-Mayo criteria for astrocytic tumors, 19 tumors (43%) were grade 2, 17 (39%) were grade 3, and 8 were (18%) grade 4. Histologic features of the tumors at initial resection included prominent nucleoli (N = 18, 41%), vascular proliferation (N = 9, 20%), necrosis (N = 6, 14%), and microcystic degeneration (N = 23, 52%). Nuclear atypia was graded as mild in 22 tumors (50%), moderate in 18 (41%), and marked in four (9%). The highest mitosis counts ranged from 0 to 10 mitotic figures (MF)/10 high-power fields (HPF) (mean, 2.4). Twelve patients (27%) had four or more MF/10 HPF. Initial surgery included gross total resection in 10 patients, subtotal resection in 16 patients, and biopsy in 14 patients. Thirty-one patients received adjuvant radiotherapy and 15 received chemotherapy. MIB-1 labeling indices ranged from 0 to 42.3 (median, 1.2 [low grade tumor median, 0.5; anaplastic tumor median, 6.2]). p53 immunostaining was observed in 18 of 43 stained tumors (41%). Overall, 5- and 10-year survival rates were 71% and 63%, respectively. The entire group had a median follow-up of 5.2 years. Age greater than 45 years (P = .02), mitosis counts of > or =4 MF/10 HPF (P = .0004), and MIB-1 labeling indices <2 (P = .03) were independent predictors of survival (Kaplan-Meier analysis). MIB-1 labeling indices <2 (P = .0009) was an independent predictor of disease-free survival. Low cell density (P = .04) and low histologic grade (P = .03) show trends with regard to being associated with longer survival. In conclusion, older patients (>45 years) or patients with tumors with an increased rate of cell proliferation generally have a worse prognosis. Although tumors of high histologic grade generally have a worse survival, the correlation was not statistically significant.     

Expression of cyclin A and topoisomerase IIalpha of oligodendrogliomas is correlated with tumour grade,MIB-1 labelling index and survival

AIMS: This study was designed to investigate immunoexpression of cyclin A and D1, and topoisomerase IIalpha in oligodendrogliomas and to evaluate the correlation with MIB-1 (Ki67), tumour grade, and survival of the patients. METHODS AND RESULTS: Forty cases of oligodendrogliomas (20 high- and 20 low-grade) were studied immunohistochemically with the above-mentioned monoclonal antibodies. RESULTS: Normal brain tissues included in tumour sections did not express any of cyclin A, MIB-1 and topoisomerase IIalpha except cyclin D1, which was shown in perineuronal and interfascicular normal oligodendroglial cells. In low-grade and high-grade oligodendrogliomas, the mean cyclin A labelling index (LI) was 1.18 +/- 0.98% versus 4.65 +/- 1.99%, respectively; the mean topoisomerase IIalpha LI was 1.32 +/- 1.04% versus 6.63 +/- 4.31%, respectively; and the mean MIB-1 LI was 1.69 +/- 1.55% versus 9.46 +/- 4.66%, respectively. Interestingly, cyclin D1 was not expressed in any oligodendrogliomas. Both cyclin A and topoisomerase IIalpha LI showed a significant positive correlation with MIB-1 LI and histological grade of oligodendrogliomas (P < 0.01) and an inverse correlation with overall survival (P < 0.01). Univariate analysis showed that cyclin A and topoisomerase IIalpha LIs with a cut-off point at 3% were a significant prognostic factor (P: cyclin A = 0.0040, topoisomerase IIalpha = 0.0033). CONCLUSION: Cyclin A and topoisomerase IIalpha expression are closely correlated with anaplastic oligodendrogliomas and worse clinical outcomes. Cyclin D1 seems not to be involved in the tumorigenesis of oligodendrogliomas.     

Cyclin D1 expression in astrocytomas is associated with cell proliferation activity and patient prognosis

An important positive regulator of the cell cycle, cyclin D1, is often amplified and overexpressed in malignancies. Cyclin D1 aberrations were analysed in grade II–IV astrocytomas by fluorescence in situ hybridization (FISH), mRNA in situ hybridization and immunohistochemistry. Proliferation activity was determined by Ki-67^MIB-1 immunolabelling and mitotic counting. High cyclin D1 expression was observed in grade IV astrocytomas (grades II–III versus grade IV; mRNA expression: p < 0·001; immunoexpression: p = 0·013), and correlated with poor patient survival (p < 0·001, n = 46). Upregulated cyclin D1 expression was also closely associated with poor patient prognosis in grade II–III astrocytomas (p < 0·001, n = 30). Cyclin D1 gene was not found to be amplified (n = 7). Cell proliferation activity was significantly increased in tumours exhibiting high cyclin D1 mRNA levels (Ki-67^MIB-1: p < 0·001; mitotic count: p < 0·001) and high cyclin D1 protein expression (Ki-67^MIB-1: p = 0·002; mitotic count: p = 0·012). These results indicate that increased production of cyclin D1 is closely associated with high cell proliferation activity and aggressive behaviour in diffusely infiltrating astrocytomas. Copyright © 1999 John Wiley & Sons, Ltd.     

MIB-1 labeling indices in benign, aggressive, and malignant meningiomas: A study of 90 tumors

Predicting tumor behavior in meningiomas based on histology alone has been problematic. This study retrospectively compares histology and MIB-1 (cell proliferation marker) labeling indices (LI) in benign, aggressive, and malignant meningiomas. Six histological features, including mitoses, necrosis, loss of pattern, hypervascularity/ hemosiderin deposition, prominent nucleoli, and nuclear pleomorphism, were compared in 90 meningiomas (Fisher's exact test). Tumors with two or more of the above features were designated as aggressive meningiomas. Malignant meningiomas were characterized by brain invasion or metastasis. The MIB-1 Us (% positive tumor cell nuclei) were compared between the three groups (Kruskal-Wallis test, Wilcoxon two-sample test). Of the benign meningiomas (n = 37; mean age, 54 years), 41% had one of the six histological features, with nuclear pleomorphism (n = 10) being the most frequent. The aggressive tumors (n = 29; mean age, 61 years) were characterized by nuclear pleomorphism (n = 28), mitoses (n = 20), necrosis (n = 16), loss of pattern (n = 16), prominent nucleoli (n = 6), and hypervascularity/hemosiderin deposition (n = 5). Malignant tumors (n = 24; mean age, 59 years) were characterized by nuclear pleomorphism (n = 22), mitoses (n = 21), loss of pattern (n = 21), necrosis (n = 21), nucleoli (n = 17), and hypervascularity/hemosiderin deposition (n = 3). Significant differences were found between the aggressive and malignant groups with regard to loss of pattern, necrosis, and nucleoli (P = .0043, .011, and .00029, respectively). Mean MIB-1 LIs for the benign, aggressive, and malignant groups were 1.0% (range, 0 to 5.5%), 5.5% (range, 0.1 to 32.5%), and 12.0% (range, 0.3 to 32.5%), respectively. Differences in the mean MIB-1 LI between groups were statistically significant, with P values of <.0001 (benign v aggressive) and .0012 (aggressive v malignant). Mean MIB-1 LIs for recurrent versus nonrecurrent tumors were 7.1% (range, 0 to 32.5%) versus 3.8% (range, 0 to 20.9%) (P = .32). The mean MIB-1 LI for patients who were alive with or without tumor was 6.2% (range, 0 to 32.5%) versus a mean MIB-1 LI of 14.2% (range, 2.8% to 32.5%) for patients who died of or with tumor (P = .0013). In conclusion, (1) There is a statistically significant difference in the increasing MIB-1 LI means between benign, aggressive, and malignant meningiomas and between patients who were alive versus those who died; (2) there is some overlap in MIB-1 LI ranges between groups, which warrants caution in interpreting an individual MIB-1 LI in a given tumor.     

Proliferative Activity in Pancreatic Endocrine Tumors: Association with Function,Metastases, and Survival

Endocrine tumors of the pancreas are slow-growing lesions, yet one-third to one-half will metastasize. It is generally accepted that histopathologic features do not reliably predict metastatic potential or outcome. We investigated whether proliferative activity, as determined by MIB-1 labeling, correlated with tumor type, metastasis, or patient survival. Formalin-fixed sections of pancreatic endocrine tumors were immunohistochemically stained for the MIB-1 antibody against Ki-67 using the avidin-biotin complex technique. Labeling index (LI) was determined by counting 1000 consecutive tumor cells in an area of greatest staining intensity at ×400 and expressed as a percentage. The study group included 37 patients, including 10 gastrinomas, 9 insulinomas, 4 glucagonomas, 2 VIPomas, and 12 nonfunctioning tumors. Twenty-one patients had metastases, primarily to regional lymph nodes and the liver. Five patients had MEN I. MIB-1 LI was significantly greater in the nonfunctioning tumors (mean 20.9%) than in the functioning tumors (mean 5.1%) (p = 0.01). LI for functional tumors (insulinomas 6.4%, glucagonoma 4.4%, gastrinomas 3.2%, VIPomas 3.2%) were similar to each other. MIB-1 was significantly higher in those tumors that metastasized (mean 15.6%) compared to those that did not (mean 3.1%), (p = 0.04). All tumors with MIB-1 LI≥10% developed metastases. Logistic regression showed that MIB-1 was a significant predictor of metastases (p = 0.003) after adjusting for functional status. MIB-1 LI also correlated with outcome in that those patients with MIB-1 LI ≥10% had a mean survival of 19 mo compared to 72 mo for those with levels <10% (p = 0.0001). Results of the proportional hazards model showed that MIB-1 remained a significant (p = 0.03) and independent predictor of survival times after adjustment for tumor size and functional status. Higher MIB-1 LI values, were significantly associated with shorter survival times. In conclusion, MIB-1 LI appears to be a useful indicator of metastatic potential and is predictive of outcome in PET.     

Microscopic thrombi in anaplastic astrocytoma predict worse survival?

The purpose of this study is to determine whether anaplastic astrocytoma patients with intratumoral vascular thrombi have a worse survival than anaplastic astrocytoma patients without thrombi. A retrospective review of 101 patients (60 males; mean age, 53.3 years) with anaplastic astrocytoma (World Health Organization grade III) was conducted. Thrombi were counted relative to the number of involved blood vessels in the initially resected tumor (69 biopsies, 32 subtotal resections) and were correlated with survival and development of postoperative deep venous thrombosis (DVT). Of tumors with thrombi (n = 17), the percentage of blood vessels with thrombi ranged from 1.5% to 20% (mean, 5.6%). Of these patients, 16 died of tumor (mean survival, 15.4 months), and 1 patient was alive with tumor at 180 months. Eighty-four patients with anaplastic astrocytoma had no intravascular tumor thrombi; 75 of these patients died of tumor (mean survival, 26.5 months), 4 patients were alive, and 5 patients were lost to follow-up. Evidence of DVT was found in 2 (18.2%) of 11 tested patients with thrombi vs 10 (18.5%) of 54 patients without thrombi. Patients with microscopic intratumoral thrombi (17% of anaplastic astrocytoma) had a worse survival compared with patients without thrombi; the difference did not reach statistical significance. There was no correlation between the presence of intratumoral thrombi and the development of DVT.     

Apoptotic activity and bcl-2 immunoreactivity in meningiomas. Association with grade and outcome

We retrospectively evaluated 90 meningiomas for bcl-2 expression, apoptosis counts (per 10 high-power fields [HPF]), MIB-1 labeling indices (LI), and mitosis counts (per 10 HPF). Characteristics were as follows: 37 low-grade (benign) meningiomas: mean apoptosis count, 1.2; MIB-1 LI, 1.0; mitosis count, 0.1; and bcl-2 positivity, 40%; 29 atypical meningiomas: apoptosis count, 3.3; MIB-1 LI, 5.5; mitosis count, 2.2; and bcl-2 positivity, 62%; 24 malignant meningiomas: apoptosis count, 6.5; MIB-1 LI, 12.0; mitosis count, 6.0; and bcl-2 positivity, 67%. By univariate analysis, MIB-1 LI, apoptosis and mitosis counts, and tumor grade were associated significantly with death due to tumor; by multivariate analysis, only mitosis count was independently associated with death due to tumor. We compared similar data for 27 patients with nonrecurrent tumors and 32 patients with recurrent meningiomas. Histologic sections from the initially resected tumor and from the most recent recurrence were reviewed. Only the apoptosis count was significantly higher by univariate analysis in the initial resection specimens from tumors that ultimately recurred vs nonrecurrent tumors. Expression of bcl-2, MIB-1 LI, and mitosis count did not correlate with recurrence. By multivariate analysis, only extent of surgical resection was associated significantly with tumor recurrence. Although bcl-2 immunostaining was not associated statistically with outcome, bcl-2 positivity was more common in atypical and malignant meningiomas than in low-grade tumors.     

Clinicopathological and immunohistochemical features of three pilomyxoid astrocytomas: Comparative study with 11 pilocytic astrocytomas

Pilomyxoid astrocytoma, first described by Tihan et al ., was recently included as an established variant of pilocytic astrocytoma in the World Health Organization classification of CNS tumors. Histologically, it much resembles pilocytic astrocytoma, but monomorphic myxoid tumor of pilocytic cells with prominent angiocentric growth pattern without Rosenthal fibers or eosinophilic granular bodies is characteristic of pilomyxoid astrocytoma. Pilomyxoid astrocytoma is thought to be more aggressive with more frequent local recurrence as well as cerebrospinal spread. The authors recently encountered a case of pilomyxoid astrocytoma, therefore the purpose of the present study was undertake a retrospective review of pilocytic astrocytomas previously diagnosed during the past 10 years. Consequently, two of them were found to have histological features suggestive of pilomyxoid astrocytoma and both involved multiple recurrence, suggesting aggressive behavior in comparison to pilocytic astrocytoma. Therefore, knowledge of this entity is essential to surgical pathologists and clinicians for patient management.     

Cyclin alterations in giant cell tumor of bone.

Cyclins play an important role in regulating the passage of dividing cells through critical checkpoints in the cell cycle. Because alterations of several cyclins, especially cyclin D1, have been implicated in the development of many human neoplasms, we examined 32 cases of giant cell tumor of long bones for cyclin D1 gene amplification and protein overexpression using differential polymerase chain reaction and immunohistochemistry, respectively. In addition, the expression of cyclin D3, cyclin B1, and the proliferation-associated antigen Ki-67 (MIB-1) was assessed immunohistochemically. Low-level cyclin D1 gene amplification was detected in 61% of giant cell tumor cases. All tumors showed cyclin D1, cyclin D3, cyclin B1, and Ki-67 (MIB-1) staining; however, the distribution was very characteristic. Cyclin D1 protein expression was seen predominantly in the nuclei of the giant cells, with occasional mononuclear cells staining. There was no correlation between cyclin D1 gene amplification and protein overexpression. Cyclin D3 staining showed a similar distribution, with 88% of cases showing protein overexpression. Cyclin D1 and/or D3 staining in the giant cells was never associated with staining for either cyclin B1 or Ki-67 (MIB-1), as the expression of the latter two proteins was restricted to the mononuclear cells. Cyclin B1 overexpression was seen in 44% of cases. Ki-67 (MIB-1) staining was present in all cases, and between 10 to 50% of the mononuclear cells were positive. These results suggest that alterations in cyclin D1 and/or D3 might play a role in the pathogenesis of giant cell tumor of bone.     

Expression of cyclin D1 and c-Ki-ras gene product in human epithelial ovarian tumors

The expression of cyclin Dl gene product in human ovarian tumors was studied. We found that cyclin D1 is expressed at high levels in several ovarian cancer cell lines. Immunohistochemical study also showed that a significant proportion of primary ovarian tumor tissues overexpressed cyclin D1 gene product. Clear nuclear staining of cyclin Dl protein was detected in 28% of the cases. We also characterized the expression of c-Ki-ras gene product in ovarian cancer cell lines and tumor tissues. Amplification or overexpression of this protooncogene has been reported in ovarian tumors from Taiwan. These results show that c-Ki-ras is strongly expressed in PA-1 and NIH: OVCAR-3 cells in which cyclin D1 also expressed at high levels. Specific cytoplasmic staining of c-Ki-ras protein was detected in 11 tumors (52%). Statistical analyses show a strong positive correlation between cyclin D1 and c-Ki-ras immunoexpression. Thus, these data support the ideas that cyclin D1 may be involved in the pathogenesis of ovarian cancer, and coactivation of cyclin D1 and c-Ki-ras gene expression may represent one of the major pathways that lead to the development of ovarian cancer in Taiwan.     

Expression of KIT Receptor Tyrosine Kinase in Endothelial Cells of Juvenile Brain Tumors

KIT receptor tyrosine kinase is expressed in tumor endothelial cells of adult glioblastomas, but its expression in pediatric brain tumor endothelial cells is unknown. We assessed expression of KIT, phosphorylated KIT, stem cell factor (SCF) and vascular endothelial growth factor receptor-2 (VEGFR-2) in 35 juvenile pilocytic astrocytomas and 49 other pediatric brain tumors using immunohistochemistry, and KIT messenger RNA (mRNA) using in situ hybridization. KIT and phospho-KIT were moderately or strongly expressed in tumor endothelia of 37% and 35% of pilocytic astrocytomas, respectively, whereas marked SCF and VEGFR-2 expression was uncommon. KIT mRNA was detected in tumor endothelial cells. Tumor endothelial cell KIT expression was strongly (P < 0.01) associated with endothelial cell phospho-KIT and SCF expression, and with tumor KIT (P = 0.0011) and VEGFR-2 expression (P = 0.022). KIT and phospho-KIT were present in endothelia of other pediatric brain tumors, notably ependymomas. Endothelial cell KIT expression was associated with a young age at diagnosis of pilocytic astrocytoma or ependymoma, and it was occasionally present in histologically normal tissue of the fetus and children. We conclude that KIT is commonly present in endothelial cells of juvenile brain tumors and thus may play a role in angiogenesis in these neoplasms.     

MIB1 and p53 immunoreactivity in protoplasmic astrocytomas

Pure protoplasmic astmqtomas am a group of rarely encountered low grade astfocytic neoplasms. Relatively few studies have speclfically examined this subset of tumors. A series of 18 protoplasmic astrocytomas in 14 males and four females (age range 2.5–52, mean 22 years) were studied in order to examine MlB1 (a marker of cell proliferation) and p53 (a tumor suppressor gene) immunoreactivity. All patients presented with seizures (mean duration 94 months) and three with headaches also. Elght tumors were located in the temporal lobe and six in the frontal lobe. All tumors were characterized by a proliferation of astrocytes with round nuclear contours arranged against a microcystic background. Only rare focl of mild vascular proliferation (3 tumors), rare mitotic figures (1 tumor), and mild nuclear atypia (3 tumors) were observed. Most tumors were primarily cortical in location. Necrosls was not seen in any of the tumors. MiB1 indices (number of MiB1 positive tumor cells/1000 tumor cells evaluated x100) ranged from 0 to 4.3 (mean 0.7); in five tumors, no MiB1 staining was observed. p53 immunoreactivity was noted in 5 of 18 tumors (28%). Five patients received adjuvant radiation therapy and one adjuvant chemotherapy. At last known follow-up, 11 patients are alive with no evidence of residual tumor (mean 70 months), six patients are alive with evidence of residual tumor (mean 58 months), and one patlent died of complications unrelated to the tumor (36 months) postoperatively. Based on these findings, the conclusions presented are as follows: (i) MlBl indices are generally low in these tumors, corroboratlng the clinical impredon of a slow growing neoplasm; and (ii) p53 immunoreactivity is obsewed in a minority of protoplasmic astrocytomas.     

Clinicopathologic study of retinoblastoma including MIB-1, p53, and CD99 immunohistochemistry.

Retinoblastoma is the most common intraocular tumor of childhood and has served as a model for the understanding or tumorigenesis. This study retrospectively examines the clinicopathologic features of 19 retinoblastomas and defines the MIB-1 (cell proliferation marker), p53 (tumor suppression gene), and CD99 (HBA71 or MIC2 antibody) immunoreactivity in 10 selected cases. Nineteen patients (11 boys), ranging in age from 6 to 47 months (mean, 20 months), were included for study. Clinical presentations included: leukocoria (n = 12), strabismus (n = 6), apparent decreased visual acuity (n = 5), and proptosis (n = 1). Five patients had bilateral tumors and one neoplasm arose in a patient with a known family history of retinoblastoma. All tumors were histologically characterized by a proliferation of small cells with high nuclear-to-cytoplasmic ratios. Commonly encountered histologic features included necrosis (n = 17, 89%), calcification (n = 16, 84%), fleurettes (n = 14, 74%), and Flexner-Wintersteiner rosettes (n = 11, 58%). Retinal involvement was noted in 18 tumors (95%) and optic nerve invasion in six cases (32%). The surgical optic nerve margin was positive in one case. Mitosis counts were evaluable in 18 cases and ranged from 1 to 42 mitotic figures/10 high power field (mean, 13 mitotic figures/10 high power field). Ten tumors were evaluated with MIB-1, p53, and CD99 antibodies by paraffin immunohistochemistry. MIB-1 labeling indices ranged from 31.4 to 77.1 (mean, 49.4). p53 immunostaining was observed in six tumors; less than 10% of tumor cells were noted to be p53 positive in each case. CD99 positivity was demonstrable focally in three tumors. Adjuvant chemotherapy and/or radiation therapy was administered in six patients. Tumor recurrence was not observed in any of the patients with a mean follow-up of 8.9 years. Only one patient died (20 years after enucleation) because of metastatic osteosarcoma. In conclusion: (1) Fleurettes and Flexner-Wintersteiner rosettes are variable findings in retinoblastoma. (2) Retinoblastomas are characterized by marked cell proliferation as evidenced by generally high mitosis counts and extremely high MIB-1 labeling indices, but this does not appear to adversely impact on prognosis. (3) Unlike peripheral primitive neuroectodermal tumors, most retinoblastomas do not stain positively with antibody to CD99. (4) Limited p53 immunostaining was present in 60% of tumors studied. (5) Enucleation with negative optic nerve margin is potentially curative in patients with retinoblastoma.     

Pathologic features, proliferative activity, and cyclin D1 expression in Hurthle cell neoplasms of the thyroid.

Making a histologic distinction between Hurthle cell adenomas and carcinomas sometimes may be difficult. We analyzed a series of Hurthle cell lesions to determine whether specific histologic features and expression of Ki67 and cyclin D1 could be useful in distinguishing Hurthle cell adenomas from carcinomas. Formalin-fixed, paraffin-embedded tissues from 128 Hurthle cell neoplasms, including 59 adenomas; 55 carcinomas; and 14 tumors classified as neoplasms of uncertain malignant behavior (UMB), which had equivocal capsular invasion but no vascular invasion, were analyzed for expression of Ki67 and cyclin D1 by immunostaining. The distribution of immunoreactivity for Ki67 with antibody MIB-1 was analyzed by quantifying the percentage of positive nuclei that was expressed as the labeling index. None of the patients with adenomas or UMB tumors developed recurrent or metastatic disease after a mean follow-up of 7.8 and 7.9 years, respectively. Of the 55 patients with Hurthle cell carcinoma, 19 were associated with metastatic disease, 13 of whom died with disease. No patient with a Hurthle cell carcinoma without vascular invasion developed metastatic disease. The mean tumor size for Hurthle cell carcinomas (4.8 cm) was significantly larger than that of Hurthle cell adenomas (3.1 cm) or UMB tumors (3.7 cm). No patient with a Hurthle cell tumor smaller than 3.5 cm developed metastatic disease, even when vascular invasion was present. The Ki67 labeling index in Hurthle cell carcinomas (10.0 +/- 1.2) was 3-fold higher than in Hurthle cell adenomas (3.2 +/- 0.3). The Ki67 labeling index in the UMB group was 5.0 +/- 0.7. Cyclin D1 showed diffuse nuclear staining in 1 of the 59 (1.7%) Hurthle cell adenomas, in 10 of the 55 (18%) Hurthle cell carcinomas, and in none of the UMB tumors. In summary, analyses of the cell cycle proteins Ki67 and cyclin D1 in Hurthle cell thyroid neoplasms indicate that these markers may assist in distinguishing some Hurthle cell carcinomas from adenomas. Among the Hurthle cell carcinomas, large tumor size and vascular invasion are associated with clinically aggressive tumors. Our study also suggests that Hurthle cell neoplasms with only equivocal capsular invasion and no vascular invasion should behave in a benign manner.     

Interobserver variability in determining MIB-1 labeling indices in oligodendrogliomas

Several studies have shown that MIB-1 labeling indices correlate well with tumor grade and prognosis in a variety of tumor types. Several factors are responsible for some degree of variability in the determination of labeling indices. Interobserver variability is one of the factors often cited as responsible for this variability. A slide from each of 30 oligodendrogliomas, stained with MIB-1 antibody, was distributed to six pathologists. The same set of slides was reviewed by each individual. Each pathologist was instructed to determine a MIB-1 labeling index by evaluating 1,000 tumor cell nuclei from the area of the slide with the most staining. The labeling index record reflected a percentage of positive-staining tumor cells. Interobserver agreement was compared. MIB-1 labeling indices ranged from 0 to 45.7. Overall agreement was good (> or =0.75) with a concordance coefficient of 0.832 (confidence interval, 0.700 to 0.909). Variability was greater among tumors with higher labeling indices as compared with tumors with labeling indices closer to 0. The overall agreement of MIB-1 labeling indices, while not perfect, was good. The generally minor variability among observers may be related to differences in the area of the slide evaluated and in differing lower thresholds for interpreting positivity. Further improvement of concordance may theoretically be attainable by further training and discussion among observers.     

Overexpression of vascular adhesion protein‐1 is associated with poor prognosis of astrocytomas

Vascular adhesion protein‐1 (VAP‐1) is one of the endothelial adhesion molecules that is believed to play a role in tumor progression and metastasis, supporting cancer cell extravasation. Very few studies have been performed on analyzing the contribution of VAP‐1 in brain tumor. Astrocytomas are the most common type of brain tumors, which are classified by World Health Organization (WHO) into four grades according to the degree of malignancy. This study was designed to investigate VAP‐1 expression level in different astrocytoma grades and its correlation with clinicopathological features as well as prognosis of astrocytoma patients. Eighty‐seven patients with different grades of astrocytoma (WHO Grade I–Grade IV) were enrolled in this study. The expression of VAP‐1 was assayed by immunohistochemistry. The correlation between VAP‐1 expression and clinicopathological features was evaluated by Chi‐square test, and overall survival was analyzed by Kaplan–Meier method. Cox regression analysis was applied to analyze the independent influence of each parameter on overall survival. The expression level of VAP‐1 was significantly higher in diffuse astrocytoma than those of pilocytic astrocytoma (p < 0.0001). In the subgroup analysis, upregulated VAP‐1 expression was frequently found in older age patients (≥50 years). The VAP‐1 expression was found to be significantly correlated with the overall survival (p = 0.0002). There was a statistical correlation between VAP‐1^high tumors in diffuse astrocytoma and VAP‐1^low tumors in pilocytic astrocytoma (p < 0.0001). Multivariate Cox analysis indicated VAP‐1 was an independent predictive marker for poorer prognosis (p = 0.0036). Therefore, VAP‐1 could be a promising prognostic biomarker in astrocytoma.     

Increased cyclin D3 expression significantly correlates with p27 nuclear positivity in gastrointestinal stromal tumors

Gastrointestinal stromal tumors, the most common mesenchymal tumors of the gastrointestinal tract, are characterized by strong expression of c-Kit protein. Recently, it has been shown that gastrointestinal stromal tumors may also contain alterations of genes involved in the regulation of cell cycle. In this study, we evaluate the prevalence and clinical significance of cyclin D1 and D3, Ki-67, p27, and retinoblastoma protein expression in a group of 50 human gastrointestinal stromal tumors selected from the files of the Moffitt Cancer Center. Tissue sections from each case were subjected to immunostaining using the avidin-biotin complex method. Cyclin D1 nuclear positivity was detected in 21 of 50 (42%) and cyclin D3 in 24 of 50 (48%) cases. p27 high immunoreactivity and negative or decreased retinoblastoma protein expression were identified in 33 of 50 (66%) gastrointestinal stromal tumors. In 19 of 50 (38%) tumors, Ki-67 had high labeling index. Direct correlation was observed between cyclin D3 and p27 expression (P < .0001), and between cyclin D1 and retinoblastoma protein (P = .03). Coexpression of cyclin D3 and p27 was demonstrated by immunofluorescence. The p27 protein expression inversely correlated with tumor size (P = .004), but was not correlated with tumor grade (P = .12). Ki-67 directly correlated with both tumor size (P = .03) and tumor grade (P = .008). We report a direct correlation between cyclin D3 and p27 expression in gastrointestinal stromal tumors. Additional alterations in cyclin D1, Ki-67, and retinoblastoma protein expression indicate a disregulated cell cycle in these tumors.     

Biology and grading of pleomorphic xanthoastrocytoma—what have we learned about it?

Pleomorphic xanthoastrocytoma (PXA) is a rare astrocytoma predominantly affecting children and young adults. We performed comprehensive genomic characterization on a cohort of 67 patients with histologically defined PXA (n = 53, 79%) or anaplastic PXA (A‐PXA, n = 14, 21%), including copy number analysis (ThermoFisher Oncoscan, n = 67), methylation profiling (Illumina EPIC array, n = 43) and targeted next generation sequencing (n = 32). The most frequent alterations were CDKN2A/B deletion (n = 63; 94%) and BRAF p.V600E (n = 51, 76.1%). In 7 BRAF p.V600 wild‐type cases, alternative driver alterations were identified involving BRAF, RAF1 and NF1. Downstream phosphorylation of ERK kinase was uniformly present. Additional pathogenic alterations were rare, with TERT, ATRX and TP53 mutations identified in a small number of tumors, predominantly A‐PXA. Methylation‐based classification of 46 cases utilizing a comprehensive reference tumor allowed assignment to the PXA methylation class in 40 cases. A minority grouped with the methylation classes of ganglioglioma or pilocytic astrocytoma (n = 2), anaplastic pilocytic astrocytoma (n = 2) or control tissues (n = 2). In 9 cases, tissue was available from matched primary and recurrent tumors, including 8 with anaplastic transformation. At recurrence, two tumors acquired TERT promoter mutations and the majority demonstrated additional non‐recurrent copy number alterations. Methylation class was preserved at recurrence. For 62 patients (92.5%), clinical follow‐up data were available (median follow‐up, 5.4 years). Overall survival was significantly different between PXA and A‐PXA (5‐year OS 80.8% vs. 47.6%; P = 0.0009) but not progression‐free survival (5‐year PFS 59.9% vs. 39.8%; P = 0.05). WHO grade remained a strong predictor of overall survival when limited to 38 cases defined as PXA by methylation‐based classification. Our data confirm the importance of WHO grading in histologically and epigenetically defined PXA. Methylation‐based classification may be helpful in cases with ambiguous morphology, but is largely confirmatory in PXA with well‐defined morphology.