Shinichi Sasaki

Hydration with magnesium and mannitol without furosemide prevents the nephrotoxicity induced by cisplatin and pemetrexed in patients with advanced non-small cell lung cancer

BACKGROUND The aim of this study was to examine the effect of hydration with magnesium and mannitol without furosemide on the nephrotoxocity accompanying combination chemotherapy using cisplatin and pemetrexed in patients with advanced non-small cell lung cancer (NSCLC). METHODS Fifty patients with NSCLC who received cisplatin plus pemetrexed, using either old hydration protocol including normal saline with mannitol and furosemide, or a new one including normal saline with magnesium and mannitol without furosemide were retrospectively analyzed. Nephrotoxicity was compared between patients treated using the old protocol and those treated with the new protocol. Univariate and multivariate analyses were performed to identify the independent factors associated with protection against nephrotoxicity in patients with NSCLC who received cisplatin plus pemetrexed. RESULTS Thirty patients received the old hydration protocol, while 20 patients were treated using the new hydration protocol. The patients treated using the new hydration protocol showed a significantly greater increase in creatinine clearance (P=0.0004) and a decrease in the serum creatinine level (P=0.0148) after one course of chemotherapy compared with those treated using the old hydration protocol. There were no differences in the chemotherapeutic response or overall survival between the groups (P=0.572). The new hydration protocol with supplemented magnesium with mannitol without furosemide was an independent factor for the protection against nephrotoxicity induced by cisplatin and pemetrexed in patients with advanced NSCLC [HR 0.232 (95% CI: 0.055-0.986), P=0.039]. CONCLUSIONS These results demonstrate that the new hydration protocol comprising supplementation with magnesium without furosemide could prevent the nephrotoxicity induced by cisplatin and pemetrexed without affecting the treatment outcome.

Structural organization of pulmonary arteries in the rat lung

The structure of the normal pulmonary arteries in the rat was studied with light and electron microscopy after use of a newly devised technique of perfusion fixation and tissue preparation. We distinguished two main types of artery in the rat lung on the basis of the structure of the media, an elastic artery and a muscular artery. The elastic artery was characterized by an abundance of extracellular matrix in the media and by an oblique arrangement of smooth muscle cells to connect neighboring elastic laminae. It was subdivided into two segments, a classical elastic and a transitional elastic segment. The muscular artery was distinguished by a paucity of extracellular matrix in the media and by a circumferential arrangement of smooth muscle cells (or pericytes) enclosing the lumina, and was subdivided into four segments, a thick muscular, an ordinary muscular, a partially muscular and a nonmuscular segment. The smooth muscle cells in the muscular artery contained well-developed microfilament bundles compared with those in the elastic artery. Structural differences in smooth muscle cells and in extracellular matrix in the media between the elastic and muscular arteries may reflect the functional heterogeneity of pulmonary arteries in response to hypoxic pulmonary vasoconstriction and to vasoactive substances such as endothelium-derived relaxing and hyperpolarizing factors, and endothelin.

Osteopontin is involved in the development of acquired chemo-resistance of cisplatin in small cell lung cancer

Osteopontin (OPN) is a multi-functional cytokine involved in cell survival, migration and adhesion which is associated with tumorigenesis, progression and metastasis. However, the role of OPN in chemo-sensitivity of human lung cancer has not yet been elucidated. The purpose of this study is to investigate the role of OPN in chemo-sensitivity of lung cancer cells. We developed a stable OPN transfectant (SBC-3/OPN) and a control transfectant (SBC-3/NEO) from human small cell lung cancer cell line, SBC-3. SBC-3/OPN cells were more resistant to cisplatin than SBC-3/NEO cells. Multi-drug resistance-associated protein (MRP) does not appear to be involved in the development of acquired chemo-resistance, since MRP inhibitor did not alter chemo-sensitivity. After exposure to cisplatin, the apoptotic SBC-3/OPN cells were reduced in number compared to SBC-3/NEO cells. Treatment with cisplatin revealed that the expression of anti-apoptotic protein, bcl-2, was down-regulated in SBC-3/NEO cells, while that of SBC-3/OPN cells was not altered. In contrast, pro-apoptotic protein, bax, was not altered in both SBC-3/OPN and SBC-3/NEO cells, thus bcl-2/bax ratio was decreased in SBC-3/NEO but not altered in SBC-3/OPN cells. Activation of caspase-3 and caspase-9 was increased in SBC-3/NEO cells, but not in SBC-3/OPN cells. Our results suggest that OPN enhances chemo-resistance of cisplatin in SBC-3 cells by suppressing bcl-2 protein down-regulation, thereby blocking the caspase-9- and caspase-3-dependent cell apoptosis.

Diagnostic significance of cerebrospinal fluid EGFR mutation analysis for leptomeningeal metastasis in non-small-cell lung cancer patients harboring an active EGFR mutation following gefitinib therapy failure

BACKGROUND Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been successfully used to treat patients with non-small cell lung cancer (NSCLC) harboring EGFR mutations. However, despite an initial excellent response, recurrence within one or two years is common. Diagnosis and treatment of leptomeningeal metastasis (LM), a form of NSCLC recurrence, remains particularly difficult. Here, we analyzed the EGFR mutation status of cerebrospinal fluid (CSF) directly using real-time polymerase chain reaction (PCR) and evaluated the efficacy of therapy with erlotinib, an EGFR TKI. PATIENTS AND METHODS Seven NSCLC patients harboring activating EGFR mutations who had developed LM during or after therapy with gefitinib, an EGFR TKI, were retrospectively analyzed. CSF was obtained and subjected to cytological examination and EGFR mutation analysis, including detection of the resistance-associated T790M mutation, using real-time PCR. RESULTS In all seven cases, the EGFR mutation detected in the CSF was the same as that detected in the primary tumor (sensitivity, 100%). Conversely, cytology results were positive in only two patients (sensitivity, 28.6%). No additional T790M mutations were detected. Erlotinib was efficacious in all cases, and improved performance status was achieved for five of the seven patients. The effect of erlotinib treatment was temporary, however, with time to treatment failure (TTF) ranging from 29 to 278 days (median, 65 days) and the interval between commencement of erlotinib treatment and death ranging from 45 to 347 days (median, 168 days). CONCLUSIONS Analysis of EGFR mutations in CSF using a highly sensitive real-time PCR assay is a potentially powerful diagnostic method for LM.

Signal transduction mediated by endostatin directly modulates cellular function of lung cancer cells in vitro

Endostatin (ED) is a carboxyl‐terminal fragment of collagen XVIII with strong antiangiogenic activity. ED has been considered as a highly specific inhibitor of endothelial cell proliferation and migration through interaction with its receptor on the surface of endothelial cells. Recently, direct antitumor effects of ED in colon cancer cells and head and neck squamous cell carcinoma cells has been reported. However, its effect on lung cancer cells has not been clarified. The purpose of the present study was to determine the effect of ED on in vitro lung cancer cell function and to identify its receptor on lung cancer cells. We revealed that α5 integrin is capable of being a functional ED receptor among several integrins that are expressed on murine lung cancer (Lewis lung cancer [LLC]) cells. We further demonstrated that the ED–integrin interaction modulates various in vitro biological functions of LLC cells as we revealed that immobilized ED helps in LLC cell adhesion and migration in an integrin‐dependent manner. Furthermore, ED inhibited LLC cell proliferation and induced apoptosis. Interestingly, ED did not demonstrate any antiproliferative activity against the other murine lung cancer cell line, KLN205, that lacks α5 integrin but binds to immobilized ED through the β1 integrin. In addition, the binding of ED to α5 integrin on LLC cells induced phosphorylation of focal adhesion kinase. Taken together, these results suggest that the interaction between ED and α5 integrin may play an important role in lung cancer cell function. (Cancer Sci 2007; 98: 830–837)

High prevalence of gene abnormalities in young patients with lung cancer

BACKGROUND Recently, driver oncogenes in adenocarcinoma of the lung were identified, and several molecular target agents were introduced in the clinical setting. However, there are few reports on the frequency of gene abnormalities in young patients with lung cancer. MATERIALS AND METHODS Twelve patients with lung adenocarcinoma aged 40 or younger at Juntendo University Urayasu Hospital or Juntendo University Hospital from July 2004 to March 2010 were analyzed for driver oncogene status including EGFR activating mutation, EML4-ALK fusion gene, and K-ras mutation. RESULTS Four patients showed EGFR gene mutation. Five out of 7 EGFR mutation-negative patients showed positive results for EML4-ALK gene fusion. One case whose EGFR mutation was indeterminate. CONCLUSIONS Driver oncogene including EGFR mutation and EML4-ALK fusion gene was identified in 9 of 12 cases (75%). Examination of gene abnormalities is essential in young patients with non-small cell lung cancer to provide the best treatment.

Oct4 plays a crucial role in the maintenance of gefitinib-resistant lung cancer stem cells.

Several recent studies have suggested that cancer stem cells (CSCs) are involved in resistance to gefitinib in non-small cell lung cancer (NSCLC). Oct4, a member of the POU-domain transcription factor family, has been shown to be involved in CSC properties of various cancers. We previously reported that Oct4 and the putative lung CSC marker CD133 were highly expressed in gefitinib-resistant persisters (GRPs) in NSCLC cells, and GRPs exhibited characteristic features of the CSCs phenotype. The aim of this study was to elucidate the role of Oct4 in the resistance to gefitinib in NSCLC cells with an activating epidermal growth factor receptor (EGFR) mutation. NSCLC cell lines, PC9, which express the EGFR exon 19 deletion mutation, were transplanted into NOG mice, and were treated with gefitinib in vivo. After 14-17 days of gefitinib treatment, the tumors still remained; these tumors were referred to as gefitinib-resistant tumors (GRTs). PC9-GRTs showed higher expression of Oct4 and CD133. To investigate the role of Oct4 in the maintenance of gefitinib-resistant lung CSCs, we introduced the Oct4 gene into PC9 and HCC827 cells carrying an activating EGFR mutation by lentiviral infection. Transfection of Oct4 significantly increased CD133-positive GRPs and the number of sphere formation, reflecting the self-renewal activity, of PC9 and HCC827 cells under the high concentration of gefitinib in vitro. Furthermore, Oct4-overexpressing PC9 cells (PC9-Oct4) significantly formed tumors at 1 × 10 cells/injection in NOG mice as compared to control cells. In addition, PC9-Oct4 tumors were more resistant to gefitinib treatment as compared to control cells in vivo. Finally, immunohistochemical analysis revealed that Oct4 was highly expressed in tumor specimens of EGFR-mutant NSCLC patients with acquired resistance to gefitinib. Collectively, these findings suggest that Oct4 plays a pivotal role in the maintenance of lung CSCs resistant to gefitinib in EGFR mutation-positive NSCLC.

Successful resection of dermatomyositis associated with thymic carcinoma: Report of a case

We report a case of thymic carcinoma associated with dermatomyositis (DM) in a 53-year-old man. The patient presented with the characteristic features of a skin rash with Gottron’s papules, proximal muscle weakness, and increased serum levels of the muscle-associated enzymes. Comprehensive clinical examinations revealed an anterior mediastinal tumor. We resected the tumor and histological examination confirmed squamous cell carcinoma of the thymus. Thereafter, his clinical symptoms improved dramatically and his serum levels of muscleassociated enzymes dropped, indicating that the DM was a paraneoplastic phenomenon. Our search of the literature found only one other case report of DM accompanied by thymic carcinoma, and to our knowledge, this is the fi rst documented case of dramatic improvement of DM after resection of thymic carcinoma. We propose that thymic carcinoma should be added to the list of malignancies that can complicate DM as a paraneoplastic disease.

The N-ERC index is a novel monitoring and prognostic marker for advanced malignant pleural mesothelioma

BACKGROUND Although N-ERC/mesothelin (N-ERC) is an attractive diagnostic and treatment monitoring biomarker for malignant pleural mesothelioma (MPM), its clinical utility for predicting the prognosis has not yet been clarified. The aim of this study is to investigate whether the serum N-ERC level can accurately predict the outcome in patients with MPM. METHODS Twenty-six patients with MPM were enrolled. Serum N-ERC level was measured before and after chemotherapy. The N-ERC index was determined by the logarithm of the division of the N-ERC level after two courses of chemotherapy by the prior level. RESULTS The median N-ERC index in the partial response (PR) group was significantly lower than that in patients with the stable disease (SD) plus the progressive disease (PD) group. The overall survival in the group whose median N-ERC index was lower than its median value was significantly longer than the group whose median N-ERC index was higher than its median value. CONCLUSIONS The N-ERC index is therefore considered to be a useful biomarker for predicting not only the chemotherapeutic response, but also the prognosis in patients with advanced MPM.

Clinical features of squamous cell lung cancer with anaplastic lymphoma kinase (ALK)-rearrangement: a retrospective analysis and review

Anti-anaplastic lymphoma kinase (ALK)-targeted therapy dramatically improves therapeutic responses in patients with ALK-rearranged lung adenocarcinoma (Ad-LC). A few cases of squamous cell lung carcinoma (Sq-LC) with ALK rearrangement have been reported; however, the clinicopathological features and clinical outcomes following treatment with ALK inhibitors are unknown. We addressed this in the present study by retrospectively comparing the clinical characteristics of five patients with ALK-rearranged Sq-LC with those of patients with ALK-rearranged Ad-LC and by evaluating representative cases of ALK inhibitor responders and non-responders. The prevalence of ALK rearrangement in Sq-LCs was 1.36%. Progression-free survival (PFS) after initial treatment with crizotinib was significantly shorter in Sq-LC than in Ad-LC with ALK rearrangement (p = 0.033). Two ALK rearrangements assayed by fluorescence in situ hybridization (FISH)-positive/immunohistochemistry-negative cases did not respond to crizotinb, and PFS decreased following alectinib treatment of ALK-rearranged Sq-LC (p = 0.045). A rebiopsy revealed that responders to ceritinib harbored the L1196M mutation, which causes resistance to other ALK inhibitors. However, non-responders were resistant to all ALK inhibitors, despite the presence of ALK rearrangement in FISH-positive circulating tumor cells and circulating free DNA and absence of the ALK inhibitor resistance mutation. These results indicate that ALK inhibitors remain a reasonable therapeutic option for ALK-rearranged Sq-LC patients who have worse outcomes than ALK-rearranged Ad-LC patients and that resistance mechanisms are heterogeneous. Additionally, oncologists should be aware of the possibility of ALK-rearranged Sq-LC based on clinicopathological features, and plan second-line therapeutic strategies based on rebiopsy results in order to improve patient outcome.