Three Japanese patients (mother and two children) with familial Mediterranean fever associated with compound heterozygosity for L110P/E148Q/M694I and an autosomal true dominant inheritance pattern.

Familial Mediterranean fever (FMF) is characterized by repeated episodes of fever, peritonitis, pleuritis, and synovitis. We describe here 3 Japanese patients (a mother and 2 children) in whom FMF was diagnosed on analysis of MEFV. A 40-year-old woman presented with fever and abdominal pain. The patient had had these symptoms on and off since childhood and consulted many hospitals. A 38-year-old man had abdominal pain and fever since the age of 30 years. A 59-year-old woman had had episodes of fever, abdominal pain, and chest pain for more than 20 years. MEFV gene analysis showed compound heterozygosity for L110P, E148Q, and M694I in all three patients. In Japanese patients with FMF, this mode of autosomal true dominant inheritance has not yet been reported. FMF is difficult to diagnose unless it is included in the differential diagnosis by physicians. We hope that our valuable experience will promote increased awareness and understanding of FMF.

A population-based study of gefitinib in patients with postoperative recurrent non-small cell lung cancer.

There is no standard treatment and there are no clearly defined guidelines for the treatment of postoperative recurrent non-small-cell lung cancer (NSCLC). We performed a retrospective population-based study to assess the benefits of treatment with gefitinib in patients with a postoperative recurrence of NSCLC in general clinical practice. This retrospective population-based study was conducted on patients with postoperative recurrent NSCLC who had been treated with gefitinib at 14 institutions in Ibaraki Prefecture between July 2002 and September 2007. The objective response rate to gefitinib therapy was 37.6% for local and distant recurrence. The median survival time following the start of gefitinib therapy was 12 months, and the one-year and two-year survival rates were 48.9 and 28.9%, respectively. The median survival time of the females was 19 months, and the median survival time of the males was 9 months (p=0.002). Univariate analysis showed that female gender, adenocarcinoma, a performance status (PS) of 0-1 and absence of smoking history were favorable prognostic factors. Only female gender and a PS of 0-1 were independent statistically significant prognostic factors in the multivariate analysis. The rate of greater than grade 1 interstitial lung damage as an adverse event was 3.5%. Gefitinib is a feasible treatment for postoperative recurrent NSCLC in general clinical practice, and a good response and prolonged survival were obtained, similar to the findings reported in published clinical studies that were conducted on highly selected patients.

Expression of phosphorylated Akt in patients with small cell carcinoma of the lung indicates good prognosis.

Patients with pulmonary small cell carcinoma are well known to have a poor prognosis. However, predictors of prognosis and treatment outcome have not been reported for this type of cancer. We examined whether excision repair cross-complementation group 1 (ERCC1), phosphorylated Akt (p-Akt), phosphorylated mammalian target of rapamycin (p-mTOR), and the uridine diphosphate glucuronosyl transferase 1A1 (UGT1A1) genetic polymorphism were useful indicators of prognosis in cases of small cell carcinoma of the lung. We investigated 45 patients with advanced small cell lung cancer who received chemotherapy with irinotecan combined with cisplatin or carboplatin. Staining results showed that 12 of 45 cases (27%) were positive for p-Akt, while 18 (40%) were positive for p-mTOR and 16 (36%) were positive for ERCC1. As for UGT1A1, more than one polymorphism was found in 30 cases (67%). There was a significant relationship observed between the expressions of p-Akt and p-mTOR (P= 0.0006). Univariate analysis indicated a significantly better survival rate for patients positive for p-Akt or p-mTOR, while multivariate analysis using a Cox test showed p-Akt to be the strongest prognostic factor. Our results indicate that p-Akt is a reliable prognostic factor in patients with small cell carcinoma of the lung.

[Detection of ALK positive pulmonary adenocarcinoma using immunostaining].

Although ALK-positive lung cancer cases have been recently reported, it is impossible to detect using only morphology technique. Furthermore, though RT-PCR and FISH techniques can be used for detection, they are not practical for screening. We investigated whether ALK-positive lung cancer could be detected using a conventional immunostaining method. Resected lung adenocarcinoma samples from 88 nonsmoker cases were selected and screening was performed using ALK immunostaining in 24 cases that did not have the EGFR or k-ras mutation. We found that the optimal staining condition was treatment using a water bath and detection with a Novo Link Polymer Detection System (Leica microsystems). Of the 24 cases examined, ALK expression was found in 4, of which ALK separated signals were found in 3 using a FISH method. No separated signals were seen in cases with negative immunostaining findings. Detection by immunostaining was found useful for ALK mutated lung cancer cases, though the pretreatment and detection methods utilized are important.

Expression of ERCC1 and class IIIbeta tubulin for predicting effect of carboplatin/paclitaxel in patients with advanced inoperable non-small cell lung cancer.

It was recently reported that expression of excision repair cross-complementation group 1 (ERCC1), a DNA repair protein, predicts sensitivity to platinum-based chemotherapy drugs. Microtubule inhibitors such as paclitaxel demonstrate anticancer effects by inhibiting spindle fibers during mitosis; and class IIIbeta tubulin (IIIbeta tubulin), a microtubule component, is thought to be resistant to microtubule inhibitors. The purpose of the present study was to examine the correlation between prognosis and expression of these proteins using biopsy tissues obtained from 40 patients with advanced inoperable non-small cell lung cancer who had been treated with carboplatin plus Taxol. On immunostaining 27 patients (68%) were positive for ERCC1 and 22 (55%) were positive for IIIbeta tubulin. The prognosis of the ERCC1-negative group was significantly better than that for the ERCC1-positive group (P= 0.014). As for IIIbeta tubulin, the prognosis for the negative group was also significantly better than that for the positive group (P= 0.025). Multivariate analysis showed that the expression of ERCC1 was an independent predictor of prognosis (hazard ratio: 3.485; 95% confidence interval: 1.123-10.818, P= 0.031). It was concluded that determination of the expression of these proteins is useful to predict the effects of platinum-based anticancer drugs.

[Epidermal growth factor receptor aberrations in malignant mesotheliomas].

Recently, the incidence of malignant mesothelioma has been increasing from such causes as asbestos exposure. In this study, we compared Epidermal Growth Factor Receptor (EGFR) aberrations in malignant mesotheliomas to those in lung cancers, to determine whether gefitinib may be useful for treatment of a malignant mesothelioma. We investigated 15 cases with malignant mesothelioma, 5 with lung cancer, and 10 with cell block from pleural effusion. In the malignant mesothelioma and lung cancer cases, we also examined the expression of EGFR by immunostaining using two kinds of anti-EGFR antibodies. In addition, we searched for ser473 p-Akt (p-Akt), which is activated by EGFR, and Epithelial Membrane Antigen (EMA), which is generally used as differential diagnosis marker of mesothelial cells. Furthermore, we examined the expressions of EGFR and EMA in benign mesothelial cells in cell blocks. Numerical abnormalities of the EGFR gene were examined by chromosome in situ hybridization, while the exonl9 and exon21 gene mutations of EGFR were examined using PCR-heteroduplex and PCR-RFLP methods, respectively. Our results showed that the EGFR protein was expressed in most of the malignant mesotheliomas in the same manner as in the lung cancer specimens. On the other hand, p-Akt was expressed in all cases (100%) of lung cancer, whereas that expression was seen in only 1 of 15 (7%) of malignant mesothelioma cases. As for EGFR gene abnormalities, a mutation was found in 1 (20%) and gene amplification in 2 (40%) lung cancer cases, while in malignant mesothelioma cases, amplification was found in 2 cases (13%) and no mutation was detected in any. Moreover, in staining results with one of the antibodies, the expression of EGFR was not different between malignant mesothelioma and non-neoplastic mesothelial cells. We observed EGFR expression at a high frequency in both the malignant mesothelioma and lung cancer specimens. However, distinct differences were detected between them in regard to EGFR gene abnormalities and the expression of p-Akt. These results suggest that it would be rare for a patient with malignant mesothelioma to benefit from gefitinib treatment.

Rapid detection of matrix metalloproteinase-7 and carcinoembryonic antigen mRNA expression in intraoperative pleural lavage samples using the reverse transcriptase loop-mediated isothermal amplification method.

OBJECTIVE: To detect the RNA of the tumor origin in intraoperative pleural lavage accurately by examining using rapid nucleic acid amplification method because patients with lung cancer often have recurrence detected by a cytologic examination of cancer cells in intraoperative pleural lavage. STUDY DESIGN: We used reverse transcriptase loop-mediated isothermal amplification (RT-LAMP), which enables rapid gene amplification, and examined the expression of matrix metalloproteinase-7 (MMP-7) and carcinoembryonic antigen (CEA) mRNA in intraoperative pleural lavage samples obtained from 79 consecutive surgical patients with lung adenocarcinoma. RESULTS: MMP-7 mRNA was detected in cancer tissues from 76 (96%) of those cases, while it was also detected in the intraoperative pleural lavage samples of 23 (30%) of those 76 cases. In addition, CEA mRNA was detected in cancer tissues from 55 (70%) cases and in 8 (15%) of the lavage samples. MMP-7 mRNA-positive cases showed a significantly positive correlation with the p and n factors, and moderate to poor differentiation. Three cases shown to be positive in cytology examinations were also positive for both MMP-7 and CEA mRNA by RT-LAMP. CONCLUSION: This detection method can be utilized during an operation and may provide useful information.

[S-1 activity in non-small cell lung cancer in clinical practice].

OBJECTIVE: We evaluated retrospectively single-agent S-1 chemotherapy in non-small cell lung cancer patients in clinical practice. METHODS: Sixteen consecutive patients treated with single-agent S-1 for NSCLC between July 2005 and June 2007 at the Department of Thoracic Surgery, Tsuchiura Kyodo General Hospital. The treatment schedule comprised oral administration of S-1 at 80-120 mg/day. One cycle of S-1 consisted of consecutive administration to 14 (10 cases)or 28(6 cases)days followed by a 14-day rest. RESULTS: Patients profiles were: M/F: 11/5, median age 68 years old(range 51-83), PS 0/1/2/3: 2/6/5/3, adeno/squamous/large: 13/2/1, clinical stage 3A/3B/4: 3/4/9, prior chemotherapy regimens 0/1/2/3/4: 2/3/4/5/2, prior surgery/radiation: 12/5 were performed. Median number of delivered cycles was 5 cycles(range 1-13). Grade 3 hematological toxicities were anemia(6%)and thrombocytopenia(6%). Grade 3 non-hematological toxicities were nausea(6%)and vomiting(6%). Response of 13 patients could be evaluated after 2-4 cycles of S-1. Four partial responses were observed, for a response rate of 31%. The survival time was 67-852 days(average 14.0 months), 1-year survival rate was 74.0%, median time to progression was 4.6 m, and 1- year progression free survival was 25.0%. CONCLUSION: Single-agent S-1 chemotherapy has modest activity and is the one of the important regimens and tolerable for elderly, poor-PS, recurrent patients with NSCLC in clinical practice.

Detection of gene point mutation in paraffin sections using in situ loop-mediated isothermal amplification.

For pathological diagnoses, visualization of genetic status using routine tissue sections is important to determine the relationships between histopathological findings and genetic alterations. Loop-mediated isothermal amplification (LAMP) has been reported to have high levels of specificity and amplification efficiency. An in situ LAMP method was used, along with an amplification refractory mutation system (ARMS) to directly detect a specific point mutation, L858R, which is a mutation of epidermal growth factor receptor (EGFR), useful for the prediction of the effects of the anti-lung cancer drug gefitinib. The investigation was done using two types of cultured cells as well as paraffin-sectioned specimens collected from 26 cases of surgically resected lung cancer. Twelve of the specimens had an L858R mutation and in situ LAMP showed reactions in the nuclei of all cancer cells present in those. Such reactions were also shown on in situ LAMP in three of the remaining 14 cases that were without the L858R mutation. In addition, a few cases showed responses in the nuclei of bronchial epithelium cells located in non-cancerous areas in the vicinity of a positive tumor, which suggested that the mutation had already occurred in the tumorigenic early stage. It is concluded that the present method is useful for pathological and genetic diagnoses.

Correlation between EGFR gene mutation pattern and Akt phosphorylation in pulmonary adenocarcinomas.

The detection of gene mutation of epithelial growth factor receptor (EGFR) is important to predict the therapeutic effect of gefitinib. Recently, it was reported that examination of the activation of the downstream protein of EGFR is useful in the same way as the EGFR mutation. Therefore the purpose of the present paper was to determine whether activation of Akt and Erk, which are downstream proteins, and the EGFR gene mutation pattern was correlated. A total of 130 pulmonary adenocarcinomas were studied for the gene mutations of EGFR in exon 19 and 21, and the phosphorylation of Akt and Erk was investigated by immunostaining. The EGFR mutation was detected in 32%, the positivity of p-Akt was 51%, and the rate of p-Erk was 27%. The EGFR mutation-positive cases were the minority in p-Akt-negative cases, and the p-Akt expression was significantly associated with the mutation of EGFR (P=0.0014). In addition, there was a significant correlation between the L858R mutation and the expression of p-Akt (P=0.040). It is suggested that the activation of Akt is dependent on EGFR mutation pattern.

The characterization of gefitinib sensitivity and adverse events in patients with non-small cell lung cancer.

BACKGROUND: Factors predicting gefitinib sensitivity and adverse events in non-small cell lung cancer (NSCLC) remain controversial. PATIENTS AND METHODS: Correlations among clinicopathological characteristics, gefitinib sensitivity and adverse events were studied in 154 patients with NSCLC, whereas epidermal growth factor receptor (EGFR) mutations were analyzed in 44 patients. RESULTS: Female, non-smoker, adenocarcinoma of stage I-II, and gefitinib effectiveness correlated with longer time to progression (TTP) and overall survival (OS), while the rate of interstitial lung disease in patients undergoing thoracic radiotherapy and stomatitis in females or those who never smoked were significantly higher. EGFR mutations were identified in 18 cases, and among 34 gefitinib-treated patients, 16 patients harboring mutations tended to do better, both in terms of TTP and OS. The results of the mutation analysis from surgical and non-surgical specimens were identical. CONCLUSION: Certain clinicopathological characteristics and EGFR mutations can be either predictive of gefitinib sensitivity or adverse events.