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CCR Translation for This Article from Reciprocal and Complementary Role of MET Amplification and EGFR T790M Mutation in Acquired Resistance to Kinase Inhibitors in Lung Cancer

Purpose: In epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) therapy for lung cancer patients, acquired resistance develops almost inevitably and this limits the improvement in patient outcomes. EGFR T790M mutation and MET amplification are the two main mechanisms underlying this resistance, but the relationship between these two mechanisms is unclear. In this study, we explored their relationship using in vitro models and autopsy specimens.Experimental Design: Erlotinib-resistant HCC827 (HCC827ER) cells were developed by chronic exposure to erlotinib at increasing concentrations. HCC827EPR cells were also developed by chronic exposure to erlotinib in the presence of PHA-665,752 (a MET TKI). The erlotinib-resistant mechanisms of these cells were analyzed. In addition, 33 autopsy tumor samples from 6 lung adenocarcinoma patients harboring multiple gefitinib-refractory tumors were analyzed.Results: HCC827ER developed MET amplification, and clinically relevant resistance occurred at ≥4-fold MET gene copy number gain (CNG). By contrast, HCC827EPR developed T790M without MET CNG. Of six patients harboring gefitinib-refractory tumors, three exhibited T790M only, one exhibited MET amplification only, and the other two exhibited T790M and/or MET amplification depending on the lesion sites. In these gefitinib-refractory tumors, T790M developed in 93% (14 of 15) of tumors without MET gene CNGs, in 80% (4 of 5) of tumors with moderate MET gene CNGs (MET amplification (≥4-fold).Conclusions: These results indicate a reciprocal and complementary relationship between T790M and MET amplification and the necessity of concurrent inhibition of both for further improving patient outcomes. Clin Cancer Res; 16(22); 5489–98. ©2010 AACR.

Background Metastases of meningiomas are infrequent and the site of extracranial metastasis such as the bone is extremely rare. Case report A 75-year-old male had a history of five sessions of surgery and gamma-knife treatment for brain meningioma over a period of 29 years. He visited our hospital because he noticed a swelling in his anterior chest 2 years and 6 months after the final treatment. After an open biopsy, histopathological analysis revealed the mass to be a metastatic grade II meningioma. We resected the tumor along with the sternum, ribs, pleura, and pericardium. The patient had recurrences in the thoracic cavity and pericardium postoperatively and received radiation therapy. He also had metastasis in the abdominal cavity, which spread rapidly. Conclusion We report on a rare instance of metastasis to the sternum in a case of atypical meningioma, showing rapid growth and invasion after long-term treatment.

A case of a 64-year-old woman who had undergone resection of a femoral giant cell tumor in another hospital 35 years before was referred to our hospital for diabetes education. Chest radiography and computed tomography (CT) revealed diffuse densely calcified nodular lesions that appear to be in both lung fields. Since pulmonary metastases of the giant cell tumor of the bone was suspected, she underwent a lung biopsy, which showed diffuse pulmonary ossification (DPO) of the dendriform type. It is extremely rare that idiopathic DPO is diagnosed by lung biopsy.

Pigmented paraganglioma of the anterior mediastinum is extremely rare. We report a 72-year-old female patient who was pointed out an anterior mediastinum tumor showed by chest computed tomography(CT) scan during a medical check up. The tumor was 50×35 mm showing gradual enlargement and was slightly homogenously contrasted by CT scan and magnetic resonance imaging(MRI). During the resection of the tumor, sudden elevations of both blood pressure up to 205 mmHg and tachycardia up to 131 beat per minate(bpm)was noted on tumor manipulation. Pathologically, the tumor was diagnosed as pigmented paraganglioma with adjacent lymph node metastasis and microscopic extracapsular invasion. The patient is well 5 years post-surgery without recurrence.

Two 50s female patients with the taste disorder of sweet taste loss and stage IV a type B2 invasive thymoma underwent surgery at our hospital. One patient with myasthenia gravis (MG) developed postoperative myasthenic crisis and recovered by the treatment with plasma apheresis and steroid pulse therapy. Her taste disorder fully recovered together with her MG symptom. The taste disorder of the other patient without MG had persisted for 3 years after the surgery. The taste disorder of sweet taste loss was reported as one of non-motor symptoms caused by MG-related autoimmune mechanisms associated with thymoma, improving with the therapy for MG. Anti-Kv 1.4 antibody was reported to be positive in nearly half patients with the taste disorder and MG and is speculated to affect selectively the sweet taste receptor.

A paravertebral block (PVB) given via the surgical field can be safer and technically simpler than an epidural block (EP) for postoperative pain control. We conducted this clinical trial to confirm the effectiveness of PVB after thoracotomy.In this non-inferiority trial, patients were randomly assigned to receive PVB (n = 35) or EP (n = 35). The primary endpoint was the pain assessed using the visual analog scale (VAS) at rest, 2, 24, and 48 h after thoracotomy, with the non-inferiority margin set at 15 mm. The secondary end points were the pain assessed using the VAS on exercising and on coughing, 2, 24, and 48 h after surgery, respectively, and the complications and need for additional analgesic agents.This trial revealed that PVB was not inferior to EP with respect to the primary end point: The mean VAS scores at rest, 2, 24, and 48 h after thoracotomy were 26.3, 10.8, and 8.3 mm in the PVB group and 23.6, 12.4, and 12.6 mm in the EP group, respectively (P0.01 for non-inferiority at all points). There were no significant differences between the groups in the incidence of complications or the need for additional analgesic agents.PVB may replace EP for postoperative pain control because of its technical simplicity and safety.

Objective We previously reported the noninferiority of paravertebral block (PVB) to epidural block. In this study, we assessed whether PVB via an intrathoracic approach was also safe for the patients ineligible for epidural block because of, for example, anticoagulation or antiplatelet therapy.

Background Patients with lung adenocarcinoma who carry epidermal growth factor receptor ( EGFR ) gene mutations respond remarkably well to EGFR tyrosine kinase inhibitor (EGFR-TKI), gefitinib, or erlotinib. However, the effect of EGFR-TKI treatment on the prolongation of overall survival (OS) of these patients remains uncertain, although several recent studies have shown prolongation of progression free survival compared with cytotoxic chemotherapy. Methods A total of 304 patients with lung adenocarcinoma who had postoperative recurrent disease were studied. To eliminate potential biases as possible, the matching of four potential predictive factors of responsiveness to EGFR-TKI led to the identification of 81 pairs of patients (those who were treated with gefitinib and those who were not). A deletion mutation in exon 19 and a point mutation (L858R) in exon 21 of the EGFR gene were also analyzed. We compared the OS between the two groups. Results OS in the gefitinib group was significantly longer than in the control group (median, 63 vs. 41 months; p = 0.015). EGFR mutations were detected in 65 out of 129 patients (50%) in the whole sample. EGFR mutational status was not an independent prognostic factor of gefitinib benefit; rather, it was a predictive factor. Conclusions This study strongly suggested that gefitinib treatment improved OS of lung adenocarcinoma patients who had postoperative recurrence, especially those carrying EGFR mutations.

Purpose: In epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) therapy for lung cancer patients, acquired resistance develops almost inevitably and this limits the improvement in patient outcomes. EGFR T790M mutation and MET amplification are the two main mechanisms underlying this resistance, but the relationship between these two mechanisms is unclear. In this study, we explored their relationship using in vitro models and autopsy specimens. Experimental Design: Erlotinib-resistant HCC827 (HCC827ER) cells were developed by chronic exposure to erlotinib at increasing concentrations. HCC827EPR cells were also developed by chronic exposure to erlotinib in the presence of PHA-665,752 (a MET TKI). The erlotinib-resistant mechanisms of these cells were analyzed. In addition, 33 autopsy tumor samples from 6 lung adenocarcinoma patients harboring multiple gefitinib-refractory tumors were analyzed. Results: HCC827ER developed MET amplification, and clinically relevant resistance occurred at ≥4-fold MET gene copy number gain (CNG). By contrast, HCC827EPR developed T790M without MET CNG. Of six patients harboring gefitinib-refractory tumors, three exhibited T790M only, one exhibited MET amplification only, and the other two exhibited T790M and/or MET amplification depending on the lesion sites. In these gefitinib-refractory tumors, T790M developed in 93% (14 of 15) of tumors without MET gene CNGs, in 80% (4 of 5) of tumors with moderate MET gene CNGs ( Conclusions: These results indicate a reciprocal and complementary relationship between T790M and MET amplification and the necessity of concurrent inhibition of both for further improving patient outcomes. Clin Cancer Res; 16(22); 5489–98. ©2010 AACR.

Introduction In clinical practice, peripheral small-sized lung cancers with positive mediastinal lymph nodes are sometimes detected. To understand the characteristics of these aggressive tumors, we reviewed the clinicopathological features of small-sized non-small cell lung cancer patients with mediastinal lymph node metastasis resected in our institution. Methods We studied 360 patients with small-sized lung lesions with a maximum diameter of 2 cm or less. The clinicopathological characteristics of each patient were reviewed and compared among the subgroups, which were stratified according to pathological nodal status. Results 21 patients (5.8%) had a positive mediastinal lymph node. Among them, 17 patients had lung lesions larger than 1.5 cm. No mediastinal nodal involvement was found in patients with squamous cell carcinomas. In contrast, mediastinal nodal involvement was significantly common in patients with poorly differentiated carcinoma (P = 0.004) and high serum carcinoembryonic antigen levels detected during preoperative evaluation (P = 0.006). None of the 14 patients with upper lobe tumor had a positive subcarinal lymph node. Lower lobe tumors frequently developed extensive multiple-level involvement, which included the upper mediastinum. Radiographic evaluation of pN2 patients using computed tomography revealed a total absence of ground-glass opacity, or the presence of a small area of ground-glass opacity. Conclusions Most small-sized non-small cell lung cancer cases with mediastinal lymph node metastasis were invasive adenocarcinoma with poor differentiation, which usually showed a solid shadow without ground-glass opacity on computed tomography.

Summary Evapotranspiration above a Japanese cypress ( Chamaecyparis obtusa Sieb. et Zucc.) forest with complex topography in central Japan were observed for 3 years using the eddy covariance method. The observations describe the magnitude and seasonal and inter-annual variations in sensible and latent heat fluxes, evapotranspiration and the parameters describing bulk canopy characteristics. Average annual evapotranspiration over the period was approximately 735 mm. Despite large fluctuations in precipitation (1179–1971 mm) during the 3 years, inter-annual fluctuations in evapotranspiration were small. Midday average dry canopy surface conductance was 6.7 mm s −1 during midseason and decreased in winter. The midday average dry canopy decoupling factor ( Ω ) was 0.21 during midseason and decreased in winter. This forest is characterized by relatively small values of surface conductance and decoupling factor compared to other forests. However, several other coniferous forests show smaller values. Surface conductance, and thus stomatal regulation, is important in controlling dry-canopy transpiration in this forest. Inter-annual fluctuations of bulk parameters were small; surface conductance characteristics affected by soil moisture did not vary between dry and wet years.

目的．非小細胞肺癌患者は術後補助化学療法が有効であると示されつつある．IB期患者で再発に影響を与える因子を解析し，どのような患者について化学療法を省くことができるかを検討した．対象と方法．縦隔リンパ節郭清を行い完全切除した病理病期IB期非小細胞肺癌患者225名を対象とした．年齢，性別，喫煙歴，術前CEA値，左右，術式，腫瘍サイズ，組織型，分化度，胸膜浸潤の有無の臨床病理学的因子から再発に影響を与える因子を解析した．結果．5年，10年全生存割合は，各々73％，57％であった．再発は87名（39％）にみられた．5年，10年無再発生存率は，各々63％，51％であった．単変量解析では腫瘍サイズが有意な因子で，4 cm未満，4 cm以上の5年無再発生存率は各々70％，57％であった（P＝0.018）．組織型では腺変扁平上皮癌が扁平上皮癌，大細胞癌と比べて有意に予後不良であった．多変量解析では腫瘍サイズが有意な予後因子であった．結論．肺癌術後IB期患者で再発がより少ない要因は，腫瘍サイズが4 cm未満でかつ，大細胞癌または扁平上皮癌であった．この患者群に対しては化学療法を省ける可能性がある．

目的．現行肺癌病期分類ではIB期とIIA期の予後が分離せず，かつIIA期の症例数が少ない．腫瘍径と胸膜浸潤からT因子を再評価し新病期分類を考案する．方法．非小細胞肺癌切除1245例を対象とした．病理病期T2を腫瘍径と胸膜浸潤（P因子）で規定し予後別に分け，IB期のうち予後の悪いT2群をIIA期に組み入れた．結果．現行病理病期別5年生存率と症例数はIA 82.2％（n＝354），IB 64.6％（n＝311），IIA 69.2％（n＝39），IIB 45.5％（n＝145）であった．T2を腫瘍径で3群（≦3 cm，＞3～≦5，＞5），胸膜浸潤で2群（P0-1，P2）に分けると，5年生存率は＞3～≦5 cmかつP0-1群で59.5％，＞5 cmまたはP2群で37.5～47.3％であり，前者をT2a，後者をT2bとした．T2aN0M0を新IB，T2bN0M0をT1N1M0とともに新IIAに分類すると，生存率と症例数はIB 70.6％（n＝163），IIA 60.4％（n＝187）となった．結論．IB期のうち＞5 cmまたはP2群は予後が悪く，IIA期に組み入れると病期別に予後が分離し症例数が分散する．

To examine the characteristics of carbon exchange in coniferous forests, we analysed the seasonal and diurnal patterns of CO2 exchange, as measured using the eddy covariance method, in a Japanese cypress forest in the Kiryu Experimental Watershed (KEW) in central Japan. The net CO2 exchange data during periods of low-friction velocity conditions and during periods of missing data were interpolated. The daily CO2 uptake was observed throughout the year, with maximum values occurring in early summer. Periods of low carbon uptake were seen in late summer owing to high respiratory CO2 efflux. The diurnal and seasonal patterns of daytime CO2 exchange at KEW were compared with those in a cool-temperate deciduous forest of the Tomakomai Experimental Forest (TOEF) in Japan. The environmental differences between evergreen and deciduous forests affected the seasonal patterns of carbon uptake. Although there were great differences in the mean monthly air temperatures between the sites, the mean monthly daytime carbon uptake was almost equal at both sites during the peak growing period. The carbon-uptake values at the same PAR level were greater before noon than after noon, especially at TOEF, suggesting the stomatal regulation of carbon uptake.

左肺癌症例に対して左肺全摘術を施行したが, 術後肺血栓塞栓症を生じた. 血栓吸引や薬物治療を行い救命できた. 肺血栓塞栓症発症後に施行した全身CTで左内頸静脈に血栓を認め, これが原因血栓と考えられた. 術後肺血栓塞栓症の原因血栓は通常, 手術中から形成されることが多いが, 本症例で認められた左内頸静脈血栓は術中に行った無名静脈のテーピングによる血流鬱滞が原因になったと推察され, この様な術中操作には細心の注意をはらわなければならない. また本症例では肺血栓塞栓症発症直後の血液ガス分析において肺血栓塞栓症で特徴的とされる低CO2血症は認められず, 逆に高CO2血症を示した. これは左肺全摘術後の片肺状態に生じた肺血栓塞栓症であるため肺血管床の減少が高度となりCO2排出能が激減し, 低CO2血症ではなく, 逆に高CO2血症を示したものと考えられた. このように肺切除後に肺血栓塞栓症が発症した場合は低CO2血症を示さず高CO2血症を示すことがあり診断上注意を要する.

滋賀県南部桐生水文試験地のヒノキ(Chamaecyparis obtusa)人工林において土壌呼吸量の時間的, 空間的変動と，それらに対する地温，土壌水分の影響に関する調査を行った。調査は，流域内に内部を1 m間隔に区切った4 m×4 mのプロットを4箇所設置し，その格子点上の土壌呼吸量，地温，土壌水分の測定を行った。土壌呼吸量は，時間的には地温に大きく依存すること，各プロット内では，そのプロットの平均値周辺を中心とした頻度分布を示すこと，プロット間の比較から，土壌水分の高いプロットや，夏場の乾燥の影響が大きいプロットで小さくなることが示された。各プロットにおける年間土壌呼吸量は，地温との関係から，454-566 gC/m2/yrと推定された。

We appreciate the comments and concerns expressed by Bronshteyn and Modest regarding our article titled ‘‘Safety of the paravertebral block in patients ineligible for epidural block undergoing pulmonary resection’’ [1]. A catheter is certainly inserted in the operative field by surgeons; catheter placement usually induces bleeding, which is directly visualized and monitored several times for approximately 10 min before closing the chest cavity. If bleeding occurs within this time, it can be directly stopped by applying pressure or coagulation with a cautery knife. As rightly pointed out by them [2], the paravertebral and epidural spaces are contiguous. Hence, we believe that our procedure via an intrathoracic approach is one of the safest methods to place a paravertebral catheter with respect to bleeding risk. In our procedure, the pleura are separated to the paravertebral space starting from the main incision at the side of the chest cavity. Therefore, the worst possible complication that can arise due to postoperative hemorrhage from the paravertebral space will be hemothorax and not spinal hematoma (and also epidural hematoma) as with the existing procedures. Thus, we consider that this procedure is certainly useful rather than being an absolute contraindication for postoperative pain control among patients on anticoagulation therapy. As per their claim [2], although our study size remained small despite the subsequent accumulation of over 100 cases, spinal hematoma (and also epidural hematoma) was still a relatively rare complication. While spinal hematoma can potentially occur, its pathophysiology remains mostly unknown [3]. Furthermore, the absolute safety of patients on anticoagulation therapy with respect to bleeding risk cannot be stated. Therefore, we provide detailed information to such patients and their relatives/guardians regarding bleeding risk and attempt to obtain informed consent. Currently, this paravertebral catheter placement trial is still carefully performed as a clinical study under approval from our hospital’s research ethics committee.

The dual puncture is one of the diseaseful complications at the induction of the epidural anesthesia, which causes severe symptoms of intracranial hypotension such as headache and nausea. The clinical courses of 3 patients with the dual puncture symptoms after pulmonary resections were retrospectively reviewed, and the effect of the continuous epidural saline infusion treatment (CESI) for the dual puncture was evaluated. Pneumococcal empyema developed in 1 patient who had been treated with conservative management. In contrast, the symptoms of the others who were treated with the CESI were quickly recovered or were effectively prevented. This report strongly suggested that the CESI was convenient and effective treatment for dual punctune symptoms by suppressing the cerebrospinal fluid leakage by elevation of the fluid pressure in the extradural space.

Introduction The efficacy of high-dose (1250 mg/d) gefitinib for the treatment of leptomeningeal metastasis in a patient with lung cancer harboring a mutation in the epidermal growth factor receptor (EGFR) gene was previously reported. We speculate that erlotinib, instead of high dose of gefitinib, may be also effective for the treatment of central nervous system (CNS) lesions, as trough serum concentration of erlotinib is nine times higher than that of gefitinib. Patients and Methods Patients with lung cancer in whom CNS lesions developed after an initial good response to gefitinib for extra CNS lesions were enrolled in the study. Tumor response, performance status, neurologic symptoms, and survival were retrospectively evaluated. Results All seven patients had EGFR mutations in their primary tumors except one patient. The median interval between gefitinib withdrawal and erlotinib administration was 5 days. Three patients showed partial response, three had stable disease, and one had progressive disease. Performance status and symptoms improved in five patients. The overall survival from the initiation of erlotinib treatment ranged from 15 to 530 days (median, 88 days). Conclusions Erlotinib was a reasonable option for the treatment of CNS diseases that appeared after a good initial response of extra CNS disease to gefitinib.

We have been using a silastic drain [Blake drain (BD)] after pulmonary resection by different placement methods and reviewed the daily amount of drainage in each patient. A 19-Fr BD was placed for each of 110 patients. First, a drain was inserted from the anterior chest wall and the tip reached the dorsal part of the diaphragm [anterior-to-posterior (AP)]. For the others [posterior-to-anterior (PA); n=37], we inserted a drain from the lower intercostal space, turned it around the apex and placed its tip in the lower front. Patients in the AP group included those placed under a water seal (AP-WS; n=43) or suction (AP-SC; n=30). The reference group consisted of 68 patients with a 32-Fr plastic drain during the same period [conventional drains (CD)]. The amount of drainage on the day of surgery in the PA group was significantly higher than that in the AP-WS group (P

We report a relatively rare surgical treatment for two cases of inflammatory pseudotumors of the lung. In case 1, a 52-year-old male with a history of left chest pain was admitted to our hospital for an abnormal nodule with an irregular margin that was detected in the left upper lung field. The nodule, measuring 15 mm in diameter, was larger than the one observed six months earlier, which had been removed by a thoracoscopic resection. In case 2, a 64-year-old female with a history of chronic cough and hemoptysis was admitted to our hospital, and an abnormal nodule with pleural indentation was detected in the lower left lung field. The nodule, measuring 8 mm in diameter, was also removed by a thoracoscopic resection. In both cases, the histologic examination enabled us to diagnose the lesion as an inflammatory pseudotumor. In general, it is very difficult to differentiate inflammatory pseudotumors from malignant tumors of the lung. The best treatment for inflammatory pseudotumors is usually early and complete surgical resection, since it can lead to improved survival. Therefore, we consider thoracoscopy-aided surgery to be less invasive and more useful than other surgical methods in the diagnosis and treatment of inflammatory pseudotumor of the lung.

Carcinoembryonic antigen (CEA) is a tumor marker widely used for nonsmall cell lung cancer (NSCLC). The aim of this study was to evaluate changes in serum CEA levels as a surrogate marker for tumor response to chemotherapy in NSCLC.From 1995 through 2005, we retrospectively analyzed 24 NSCLC patients who had high serum CEA levels (5 ng/ml) and who received chemotherapy followed by surgery. We compared serum CEA levels with tumor response, as defined by Response Evaluation Criteria in Solid Tumors (RECIST) or World Health Organization (WHO) criteria, as well as with histological response.Serum CEA levels after chemotherapy significantly decreased in patients who achieved partial response, defined by RECIST or WHO criteria (p = 0.004 and p = 0.008, respectively), when compared with the CEA levels before chemotherapy. In contrast, there was no significant difference in CEA levels in patients with either stable disease or no response to chemotherapy. They decreased significantly, however, in patients in whom less than one-third of tumor cells was viable by pathological examination, but not in patients in whom more than a third was viable (p = 0.008). Using the receiver-operating characteristic (ROC) curve analysis, we found that a 60% reduction of CEA levels was an appropriate cutoff value in predicting a good response to chemotherapy. When the value was set at that level, the sensitivity of CEA for RECIST was 82%, and the specificity was 69%.Serum CEA concentration was a useful surrogate marker for the evaluation of tumor response to chemotherapy and seemed to be comparable with RECIST in NSCLC patients who had elevated CEA levels prior to treatment.

Although PDZK1 is a well-known adaptor protein, the mechanisms for its role in transcriptional regulation are largely unknown. The peroxisome proliferator-activated receptor alpha (PPARalpha) is a ligand-activated transcription factor that plays an important role in the regulation of lipid homeostasis. Previously, we established a tetracycline-regulated human cell line that can be induced to express PPARalpha and identified candidate target genes, one of which was PDZK1. In this study, we cloned and characterized the promoter region of the human pdzk1 gene and determined the PPAR response element. Finally, we demonstrate that endogenous PPARalpha regulates PDZK1 expression.

The patient was an 84-year-old male. About 7 months after surgery for a non-small cell lung cancer StageIII B, granular shadows were noted on his bilateral lungs on CT; and the level of CA 19-9 , a tumor marker, had increased. These findings led to a diagnosis of intrapulmonary tumor metastasis. Treatment with a platinum-based antineoplastic agent was recommended,but the patient refused it because of his age. The patient was thus placed under observation. Because the tumor appeared to increase in size, treatment was initiated using S-1 capsules (hereafter abbreviated to S-1) alone. Each course consisted of S-1 at a dosage of 100 mg/body/day, twice daily for 21 days, followed by 14 days of drug withdrawal. After the completion of one course, the level of the tumor marker was reduced to normal. After 6 courses were completed, CT indicated a marked reduction or elimination of the intrapulmonary metastatic foci. Malignant phenomenon exceeding grade 3 was no longer recognized,and it was possible to deliver the scheduled dosage of S-1 throughout the entire series of courses. At the end of the 14th course, no signs of exacerbation or recurrence were recognized. It has been about 18 months after the S-1 single therapy was started. This therapy is still being continued.

Background Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors and commonly play an important role in the regulation of lipid homeostasis. To identify human PPARs-responsive genes, we established tetracycline-regulated human hepatoblastoma cell lines that can be induced to express each human PPAR and investigated the gene expression profiles of these cells. Results The expression of each introduced PPAR gene was investigated using the various concentrations of doxycycline in the culture media. We found that the expression of each PPAR subtype was tightly controlled by the concentration of doxycycline in these established cell lines. DNA microarray analyses using these cell lines were performed with or without adding each subtype ligand and provided much important information on the PPAR target genes involved in lipid metabolism, transport, storage and other activities. Interestingly, it was noted that while ligand-activated PPARδ induced target gene expression, unliganded PPARδ repressed these genes. The real-time RT-PCR was used to verify the altered expression of selected genes by PPARs and we found that these genes were induced to express in the same pattern as detected in the microarray analyses. Furthermore, we analysed the 5'-flanking region of the human adipose differentiation-related protein (adrp) gene that responded to all subtypes of PPARs. From the detailed analyses by reporter assays, the EMSAs, and ChIP assays, we determined the functional PPRE of the human adrp gene. Conclusion The results suggest that these cell lines are important tools used to identify the human PPARs-responsive genes.

2'-O,4'-C-methylene bridged nucleic acid (2',4'-BNA) is a nucleic acid analogue that has high affinity binding to its complementary RNA. Peroxisome proliferator-activated receptor (PPAR) y belongs to the nuclear hormone receptor superfamily and is a drug target in the treatment of type 2 diabetes. Here, we show the antisense effects of 2',4'-BNA oligonucleotides against PPARy in the human monocytic leukaemia cell line THP-1 and the human colorectal tumor cell line HCT116.

The peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that belong to the nuclear hormone receptor superfamily. PPARalpha is mainly expressed in the liver, kidney, heart and muscle. PPARalpha activates fatty acid catabolism, stimulates gluconeogenesis and ketone body synthesis and is involved in the control of lipoprotein assembly. Although PPARalpha is well characterized in the liver, its physiological function is unknown in the kidney. To investigate the intimate function of PPARalpha in the kidney, we analyzed the target gene expression in human metastatic renal cell carcinoma cell line, Caki-1, using small interfering RNA (siRNA) against PPARalpha and real-time RT-PCR methods. We found that some selected genes (long-chain fatty-acid-CoA ligase (FACL1), carnitine palmitoyltransferase 1A (CPT1A), adipose differentiation-related protein (ADRP) and aquaporin 3 (AQP3)) were down-regulated by PPARalpha siRNA. These results suggest that PPARalpha regulates fatty acid metabolism and body water homeostasis in this cell line.

The influence of the addition of a foreign element on the superlattice formation of the AuCu3 type, especially the change of transformation temperature, was studied on the basis of the Bragg-Williams theory of superlattice. Calculations were carried out to several simple cases. The results were discussed in connection with the real ternary alloys.