Masato Takano

9p21 deletion in the diagnosis of malignant mesothelioma, using fluorescence in situ hybridization analysis.

Homozygous deletion of 9p21, the locus harboring the p16 gene, has been reported as one of the most common genetic alterations in malignant mesotheliomas (MMs). Previous studies showed that this alteration might be useful for differentiating benign from malignant mesothelial tumors in cytology and surgical specimens. Although the diagnostic utility of 9p21 homozygous deletion by fluorescence in situ hybridization (FISH) analysis has been reported only recently, it has not been well demonstrated. The purpose of this study is to evaluate the diagnostic utility of 9p21 homozygous deletion assessed by FISH in mesothelial neoplasm and hyperplasia of Japanese patients using paraffin‐embedded tissue. Simultaneously, p16 protein immunoexpression was explored as a potential diagnostic aid. FISH analysis demonstrated 9p21 deletion in 35 of 40 cases with MM (88%) (P < 0.001). In contrast, no cases of adenomatoid tumor, benign mesothelial multicystic tumor, reactive mesothelial hyperplasia or pleuritis showed 9p21 deletion (P < 0.005). 9p21 homozygous deletion correlated well with p16 protein expression in the tumor cells. Our study suggests that 9p21 homozygous deletion assessed by FISH on paraffin‐embedded tissue may be very useful for differentiating MM from reactive mesothelial proliferation.

Genomic gains and losses in malignant mesothelioma demonstrated by FISH analysis of paraffin-embedded tissues

Aims Malignant mesothelioma (MM) results from the accumulation of a number of acquired genetic events at the onset. In MM, the most frequent changes were losses in 9p21, 1p36, 14q32 and 22q12, and gains in 5p, 7p and 8q24 by comparative genomic hybridisation analysis. Although the diagnostic utility of 9p21 homozygous deletion by fluorescence in situ hybridisation (FISH) analysis in MM has been reported recently, alterations of other genes have not been examined to any great extent. This study analysed the frequency of various genomic gains and losses in MM using FISH analysis. Materials and methods The authors performed a FISH analysis using paraffin-embedded tissues from 42 cases of MM. Results Chromosomal losses in MM were found at 9p21 (83%), 1p36 (43%), 14q32 (43%) and 22q12 (38%), whereas gains were found at 5p15 (48%), 7p12 (38%) and 8q24 (45%). There were no cases of adenomatoid tumour, benign mesothelial multicystic tumour, reactive mesothelial hyperplasia or pleuritis showing any gains or losses. At least one genomic abnormality was identified in all cases of MM. Among various histological subtypes, the chromosomal abnormality tended to be more common in cases showing sarcomatous elements (biphasic or pure sarcomatoid) than in cases showing an epithelioid histology. Conclusions The authors found various genomic gains and losses in MM by FISH analysis. The frequency of each genomic gain or loss examined in MM by FISH analysis almost agreed with the comparative genomic hybridisation technique in previous studies. This study suggests that genomic evaluation by FISH analysis might be helpful in distinguishing MM from benign mesothelial proliferation.

Angiomyolipoma of the liver: a reappraisal of morphological features and delineation of new characteristic histological features from the clinicopathological findings of 55 tumours in 47 patients.

Nonomura A, Enomoto Y, Takeda M, Takano M, Morita K & Kasai T 
(2012) Histopathology 61, 863–880

Pharmacokinetics and Antitumor Efficacy of Chemoembolization Using 40 µm Irinotecan-Loaded Microspheres in a Rabbit Liver Tumor Model

PURPOSE To evaluate the pharmacokinetics and antitumor efficacy of 40 μm irinotecan-loaded drug-eluting microspheres (Embozene TANDEM Microspheres; CeloNova BioSciences, Inc, San Antonio, Texas) (TANDEM-IRI). MATERIALS AND METHODS The following three groups included eight VX2 rabbits each: group 1, full-loaded (50 mg irinotecan/1 mL TANDEM)/high-dose injection (1 mg irinotecan/kg); group 2, full-loaded (50 mg irinotecan/1 mL TANDEM)/low-dose injection (0.5 mg irinotecan/kg); and group 3, half-loaded (25 mg irinotecan/1 mL TANDEM)/low-dose injection (0.5 mg irinotecan/kg). Irinotecan and SN-38 in the plasma and tumors were measured within 72 hours. Histologic examinations were conducted on days 1, 3, and 7. RESULTS Serum irinotecan levels remained near the maximum concentration for 180 minutes after transarterial chemoembolization; in group 1, levels were 351.4 ng/mL at 30 minutes, 329.0 ng/mL at 60 minutes, and 333.5 ng/mL at 180 minutes. The area under the curve for 0-24 hours of irinotecan in group 1 was approximately two times higher than the same value in groups 2 and 3. High irinotecan and SN-38 concentrations in the tumors were measured at 24 hours and 72 hours. After transarterial chemoembolization, levels of liver enzymes aspartate aminotransferase and alkaline phosphatase were significantly higher in group 1 compared with groups 2 and 3. Histologic findings showed microspheres had deeply penetrated into tumors. Significantly higher tumor necrosis ratios were observed in groups 1 (86.6%-90.0%) and 3 (90.0%-100%) compared with group 2 (63.3%-70%) (P = .031 and P = .016). CONCLUSIONS Slow drug release with high drug concentration in tumors can be provided with 40 μm TANDEM-IRI. When complete arterial embolization is performed, the dose of irinotecan loaded on 40 μm TANDEM microspheres can be reduced while maintaining efficacy.

Epidermal growth factor receptor mutations in malignant pleural and peritoneal mesothelioma

Background Epidermal growth factor receptor (EGFR) gene mutation at the kinase domain and EGFR gene amplification are reported to be predictors of the response to EGFR tyrosine kinase inhibitors in lung cancer cases. In malignant mesothelioma (MM), the role of EGFR is less clear. Methods Thirty-eight MM specimens were submitted to EGFR mutation evaluation, and compared with the results of immunohistochemical staining and fluorescence in situ hybridization (FISH) analysis. DNA was extracted from paraffin blocks and PCR was performed to amplify exon regions 18–21 of the EGFR gene. Direct sequencing of the purified PCR products was performed. Results Five EGFR missense mutations were detected in six of the 38 patients (16%); two of these mutations were novel, two were originally detected in non-small cell lung carcinoma, and one resembled a location previously noted for malignant peritoneal mesothelioma. Conclusion As far as the authors are aware there has been no report of the EGFR mutation of MM in Japanese cases, but in this study EGFR missense mutations were detected in some cases. EGFR mutation results were not related to immunohistochemical and FISH analysis.

A comparison of epidermal growth factor receptor expression in malignant peritoneal and pleural mesothelioma.

An evaluation of epidermal growth factor receptor (EGFR) phenotypic expression in malignant pleural and peritoneal mesothelioma was undertaken, using immunohistochemical (IHC) and fluorescence in situ hybridization (FISH) analysis. Thirty‐eight malignant mesothelioma (MM) specimens were subjected to IHC staining and FISH to evaluate the expression of EGFR protein and gene status. Overall positive IHC reaction was detected in 20/38 (53%) cases, in 11/22 (50%) pleural MM, and in 9/16 (56%) peritoneal MM. Our study confirmed that EGFR membranous expression is a common feature in MM, but not in benign mesothelial lesion. Thirty‐seven cases did not show a gene copy number gain. Only one case showed a copy number gain. The protein overexpression of EGFR was not related to a gene copy number gain.

Autopsy case of primary myelofibrosis in which myeloid sarcoma was the initial manifestation of tumor progression

Myeloid sarcoma (MyS) is defined as an extramedullary tumor‐forming neoplasm consisting of immature myeloid cells with/without maturation. We experienced a case involving a 68‐year‐old Japanese male patient who had been followed‐up for four years with a diagnosis of chronic idiopathic myelofibrosis/primary myelofibrosis (PMF) and noticed a painful mass in his left axilla. A wedge biopsy characterized the lesion as MyS that displayed megakaryoblastic/megakaryocytic differentiation. As his complete blood count included a few myeloid blasts (1% of WBC) and a bone marrow biopsy detected fibrosis without evidence of acute myelogenous leukemia (AML), a diagnosis of extramedullary blastic transformation of PMF was made, which was confirmed later by V617F mutation in Janus kinase‐2 in both initial bone marrow biopsy and axillary tumor biopsy specimens. The patient died of pneumonia eight months after developing the axillary tumor. At autopsy, multiple MyS masses were detected in his soft tissue, but his bone marrow only contained fibrosis. Although MyS rarely develops before the leukemic transformation of PMF, no evidence of AML could be found in the patients bone marrow at any point during the course of his disease. Thus, it is possible that the blasts in his peripheral blood were derived from the remaining MyS. Furthermore, the present case indicates that extramedullary blastic transformation, which is occasionally seen in CML, can also occur in PMF. Therefore, it is important to recognize that there is a wide variation in the pathogeneses of MyS and PMF.

A retrospective study of patients with follicular lymphoma (FL): identification of in situ FL or FL-like B cells of uncertain significance in lymph nodes resected at the time of previous surgery for carcinomas

Background and aims In situ follicular lymphoma (iFL)/intrafollicular neoplasia and follicular lymphoma (FL)-like B cells of uncertain significance represent proliferation of Bcl-2/t(14;18)-positive B cells solely in the germinal centres. The condition is interpreted as an early event in the multi-step lymphomagenesis of FL. The aim of this study is to examine the issue more specifically. Methods We reviewed medical history of FL patients in whom thoracoabdominal surgery with lymphadenectomy had been performed for management of carcinomas. These previously resected lymph nodes as well as the current FL nodes were analysed by immunohistochemistry, fluorescent in situ hybridisation, and PCR amplification with direct sequencing. Results We studied four such FL patients from a total of 150 patients with FL; all had iFL in the previously resected lymph nodes. Clonal relation was verified and suggested in one case each. The time from lymphadenectomy to the diagnosis of FL was 23–120 months. There appeared to be a reverse correlation between the rate of Bcl-2-positive follicle proliferation and the time from surgery to diagnosis of FL. Conclusions Although the rate for development of FL in individuals having iFL has been reported to be low from prospective studies, the present data indicate that follow-up studies for a longer period is necessary; the rate of Bcl-2-positive follicle proliferation could be a factor to predict development of FL in prospective studies. Such a retrospective study may contribute to elucidate mechanism(s) involved in lymphomagenesis of FL.

An esophageal gastrointestinal stromal tumor with regional lymph node metastasis

We report a case of a 72-year-old woman with an esophageal gastrointestinal stromal tumor (GIST) with regional lymph node metastasis. Endoscopy and barium esophagography revealed a large submucosal tumor in the lower esophagus. Computed tomography showed a solid 8-cm tumor, suggesting an esophageal mesenchymal tumor. Endoscopic ultrasonography-guided fine-needle aspiration biopsy was positive for c-KIT and CD34, and negative for desmin and S-100. The patient underwent middle and lower esophagectomy via left thoracotomy, followed by gastric tube reconstruction. The tumor was completely resected, but a metastasis in the right paracardial lymph node was observed. Pathological examination confirmed the tumor to be high risk. We are carefully following up the patient.

Multivariate analyses of Ki-67, cytokeratin 13 and cytokeratin 17 in diagnosis and prognosis of oral precancerous lesions

BACKGROUND Ki-67, cytokeratin 13, and/or cytokeratin 17 detection by immunohistochemistry has been reported to be useful for the diagnosis of oral precancerous lesions. However, the use of these markers remains controversial because of the lack of appropriately designed statistical studies. We assessed the hypothesis that Ki-67, cytokeratin 13, or cytokeratin 17 immunohistochemistry could facilitate the diagnosis of oral precancerous lesions and/or predict prognosis. METHODS Epithelial dysplasia was classified as low grade (none or mild dysplasia) or high grade (moderate dysplasia, severe dysplasia, or carcinoma in situ). This study included 58 low-grade and 36 high-grade dysplasia cases. We used logistic regression to assess the diagnostic values of Ki-67, cytokeratin 13, and cytokeratin 17 for high-grade dysplasia. Correlations between these markers and the prognosis of oral atypical epithelium were assessed using the Cox proportional hazards model. RESULTS Ki-67 overexpression and cytokeratin 13 loss were independent diagnostic markers for high-grade dysplasia (odds ratios, 1.92 and 2.53; 95% confidence intervals, 1.03-3.58, and 1.19-5.38, respectively). The area under the curve of Ki-67 was 0.73 and that of cytokeratin 13 was 0.72. However, the combination of Ki-67 and cytokeratin 13 yielded the area under the curve of 0.78. Ki-67 overexpression was significantly associated with recurrence and/or malignant transformation of oral atypical epithelium (hazard ratio, 7.25; 95% confidence interval, 1.07-48.92). CONCLUSIONS Ki-67 overexpression and cytokeratin 13 loss may be useful for distinguishing oral precancerous lesions from reactive atypical epithelium. Moreover, Ki-67 overexpression may be a risk factor for recurrence and/or malignant transformation of oral atypical epithelium.