Oncology Reports

Our medical research library holds 42 studies published in Oncology Reports (2002–2023). Each entry links to the study's record on PubMed and, where there is one, to the publisher's page and a free full text. 42 have an abstract, the authors' own summary as PubMed records it: open "Abstract" to read it.

About the journal

• Publisher: D.A. Spandidos
• Published: 1994 to the present, Greece
• Language: English
• ISSN: 1021-335X (print), 1791-2431 (electronic)
• Its record in the NLM Catalog

2020s

• Concurrent induction of apoptosis and necroptosis in apigenin‑treated malignant mesothelioma cells: Reversal of Warburg effect through Akt inhibition and p53 upregulation. Y.J. Lee, K.S. Park, S.H. Heo, et al. 2023;49(6). PubMed · Publisher · Free full text

  Abstract

  A high dependence on aerobic glycolysis, known as the Warburg effect, is one of the metabolic features exhibited by tumor cells. Therefore, targeting glycolysis is becoming a very promising strategy for the development of anticancer drugs. In the present study, it was investigated whether pre‑adaptation of malignant mesothelioma (MM) cells to an acidic environment was associated with a metabolic shift to the Warburg phenotype in energy production, and whether apigenin targets acidosis‑driven metabolic reprogramming. Cell viability, glycolytic activity, Annexin V‑PE binding activity, reactive oxygen species (ROS) levels, mitochondrial membrane potential, ATP content, western blot analysis and spheroid viability were assessed in the present study. MM cells pre‑adapted to lactic acid were resistant to the anticancer drug gemcitabine, increased Akt activation, downregulated p53 expression, and upregulated rate‑limiting enzymes in glucose metabolism compared with their parental cells. Apigenin treatment increased cytotoxicity, Akt inactivation and p53 upregulation. Apigenin also reduced glucose uptake along with downregulation of key regulatory enzymes in glycolysis, increased ROS levels with loss of mitochondrial membrane potential, and downregulated the levels of complexes I, III and IV in the mitochondrial electron transport chain with intracellular ATP depletion, resulting in upregulation of molecules mediating apoptosis and necroptosis. Apigenin‑induced alterations of cellular responses were similar to those of Akt inactivation by Ly294002. Overall, the present results provide mechanistic evidence supporting the anti‑glycolytic and cytotoxic role of apigenin via inhibition of the PI3K/Akt signaling pathway and p53 upregulation.

• Downregulation of lncRNA PVT1 inhibits proliferation and migration of mesothelioma cells by targeting FOXM1. Y. Fujii, V.J. Amatya, K. Kushitani, et al. 2022;47(2). PubMed · Publisher · Free full text

  Abstract

  Malignant mesothelioma is a highly aggressive tumor, and an effective strategy for its treatment is not yet available. Long non‑coding RNAs (lncRNAs) have been reported to be associated with various biological processes, including the regulation of gene expression of cancer‑related pathways. Among various lncRNAs, plasmacytoma variant translocation 1 (PVT1) acts as a tumor promoter in several human cancers, but its mechanism of action has not yet been elucidated. Increased PVT1 expression was identified in ACC‑MESO‑1, ACC‑MESO‑4, CRL‑5915, and CRL‑5946 mesothelioma cell lines. PVT1 expression was investigated in mesothelioma cell lines by reverse transcription‑quantitative polymerase chain reaction and its functional analysis by cell proliferation, cell cycle, cell migration, and cell invasion assays, as well as western blot analysis of downstream target genes. Knockdown of PVT1 expression in these cell lines by small interfering RNA transfection resulted in decreased cell proliferation and migration and increased the proportion of cells in the G2/M phase. The results of reverse transcription‑quantitative polymerase chain reaction analysis revealed that PVT1 knockdown in mesothelioma cell lines caused the downregulation of Forkhead box M1 (FOXM1) expression, while the results of western blot analysis revealed that this knockdown reduced FOXM1 expression at the protein level. In addition, combined knockdown of PVT1 and FOXM1 decreased the proliferation of mesothelioma cell lines. In conclusion, PVT1 and FOXM1 were involved in the proliferation of cancer cells. Therefore, PVT1‑FOXM1 pathways may be considered as candidate targets for the treatment of malignant mesothelioma.

• Connective tissue growth factor produced by cancer‑associated fibroblasts correlates with poor prognosis in epithelioid malignant pleural mesothelioma. Y. Ohara, A. Enomoto, Y. Tsuyuki, et al. 2020;44(3):838-848. PubMed · Publisher · Free full text

  Abstract

  Malignant mesothelioma is an aggressive neoplasm for which effective treatments are lacking. We often encounter mesothelioma cases with a profound desmoplastic reaction, suggesting the involvement of cancer‑associated fibroblasts (CAFs) in mesothelioma progression. While the roles of CAFs have been extensively studied in other tumors and have led to the view that the cancer stroma contains heterogeneous populations of CAFs, their roles in mesothelioma remain unknown. We previously showed that connective tissue growth factor (CTGF), a secreted protein, is produced by both mesothelioma cells and fibroblasts and promotes the invasion of mesothelioma cells in vitro. In this study, we examined the clinical relevance of CAFs in mesothelioma. Using surgical specimens of epithelioid malignant pleural mesothelioma, we evaluated the clinicopathological significance of the expression of α‑smooth muscle actin (αSMA), the most widely used marker of CAFs, the expression of CTGF, and the extent of fibrosis by immunohistochemistry and Elastica‑Masson staining. We also analyzed the expression of mesenchymal stromal cell‑ and fibroblast‑expressing Linx paralogue (Meflin; ISLR), a recently reported CAF marker that labels cancer‑restraining CAFs and differ from αSMA‑positive CAFs, by in situ hybridization. The extent of fibrosis and CTGF expression in mesothelioma cells did not correlate with patient prognosis. However, the expression of αSMA and CTGF, but not Meflin, in CAFs correlated with poor prognosis. The data suggest that CTGF+ CAFs are involved in mesothelioma progression and represent a potential molecular target for mesothelioma therapy.

• Selection of microRNAs in extracellular vesicles for diagnosis of malignant pleural mesothelioma by in vitro analysis. T. Jotatsu, H. Izumi, Y. Morimoto, et al. 2020;44(5):2198-2210. PubMed · Publisher · Free full text

  Abstract

  Malignant pleural mesothelioma (MPM) is a malignant tumor which is a challenge for diagnosis and is associated with a poor patient prognosis. Thus, early diagnostic interventions will improve the quality of life and life expectancy of these patients. Recently, cellular microRNAs (miRNAs) have been found to be involved in maintaining homeostasis, and abnormal miRNA expression has often been observed in various diseases including cancer. Extracellular vesicles (EVs) released by many cells contain proteins and nucleic acids. miRNAs are secreted from all cells via EVs and circulate throughout the body. In this study, culture media were passed sequentially through membrane filters 220‑50 nm in size, and EVs with diameters of 50 to 220 nm (EVcap50/220) were collected. miRNAs (EV50‑miRNAs) in EVcap50/220 were purified, and microarray analysis was performed. EV50‑miRNA expression profiles were compared between MPM cells and a normal pleural mesothelial cell line, and six EV50‑miRNAs were selected for further investigation. Of these, hsa‑miR‑193a‑5p and hsa‑miR‑551b‑5p demonstrated higher expression in MPM‑derived EVcap50/220. These miRNAs reduced the expression of several genes involved in cell‑cell interactions and cell‑matrix interactions in normal pleural mesothelial cells. Our data suggest that hsa‑miR‑193a‑5p and hsa‑miR‑551b‑5p in EVcap50/220 could be diagnostic markers for MPM.

2010s

• Aberrant expression of FoxP3 in a human T cell line possessing regulatory T cell‑like function and exposed continuously to asbestos fibers. M. Maeda, H. Matsuzaki, S. Yamamoto, et al. 2018;40(2):748-758. PubMed · Publisher

  Abstract

  Prompted by the known carcinogenic activity of asbestos, our investigations revealed that asbestos causes a reduction in antitumor immunity. One mechanism involves the enhancement of regulatory T (Treg) cell function and volume assayed using MT‑2 original cells (Org), an HTLV‑1 immortalized human T cell line which possesses Treg‑like function. Continuous and relatively low‑dose exposure of MT‑2 to asbestos fibers yielded sublines resistant to asbestos‑induced apoptosis and enhanced Treg function via cell‑cell contact mechanisms and increased the production of soluble factors such as interleukin (IL)‑10 and transforming growth factor (TGF)‑β. Additionally, cell cycle progression was accelerated in these sublines. Subsequently, the status of the Treg‑specific transcription factor FoxP3 was examined. Unexpectedly, FoxP3 mRNA levels decreased in the sublines, although significant changes in protein expression were absent. Methylation analysis of CpG sites located in the promoter region of FoxP3 in original MT‑2 cells and sublines showed almost complete methylation in Org and slight hypomethylation in the sublines. Although treatment with the demethylating agent 5‑aza‑deoxycytidine tended to upregulate FoxP3 expression, the methylation status did not match the mRNA expression and enhanced function. Additionally, the expression of other transcription factors related to Treg did not differ between Org and subline CB1. Collectively, aberrant expression and methylation patterns of FoxP3 were detected in human T cells continuously exposed to asbestos, although cell function was enhanced by asbestos exposure. Future analyses to identify factors responsible for Treg functional enhancements induced by asbestos, such as the investigation of surface molecules, are needed for the development of strategies to prevent the occurrence of asbestos‑induced cancers.

• Berberine induces mitochondrial‑mediated apoptosis and protective autophagy in human malignant pleural mesothelioma NCI‑H2452 cells. Z. Yao, Y. Wan, B. Li, et al. 2018;40(6):3603-3610. PubMed · Publisher

  Abstract

  Increasing evidence shows that berberine has antitumor effects against a number of tumor cells. In the present study, we evaluated the effect of berberine on the proliferation of the human malignant pleural mesothelioma (MPM) cell line NCI‑H2452, and explored the therapeutic potential and underlying mechanisms of this agent. Our results showed that berberine inhibited the proliferation of NCI‑H2452 cells in a dose‑ and time‑dependent manner and could induce apoptosis, possibly through a caspase‑9‑dependent intrinsic mitochondrial pathway. In addition, autophagy was induced by berberine, which was characterized by the accumulation of LC3‑II and decreased p62 expression. We used inhibitors of apoptosis and autophagy, and an inducer of autophagy, to evaluate the significance of autophagy in berberine‑induced cell death. The results demonstrated that apoptosis is the primary route through which berberine induces NCI‑H2452 cell death. Berberine‑induced autophagy may be an adaptive response to antitumor agents and have a protective role in MPM cells. Inhibition of autophagy enhanced berberine‑induced apoptosis. Therefore, inhibition of autophagy may be an effective treatment strategy in the management of MPM. In conclusion, berberine is a potent antitumor agent for treating MPM, and it induces mitochondrial‑mediated apoptosis and protective autophagy in human NCI‑H2452 MPM cells.

• Inhibition of ornithine decarboxylase 1 facilitates pegylated arginase treatment in lung adenocarcinoma xenograft models. S.K. Lam, K.P. U, Y.Y. Li, et al. 2018;40(4):1994-2004. PubMed · Publisher · Free full text

  Abstract

  Arginine depletion has shown anticancer effects among arginine auxotrophic cancers. An anti‑proliferative effect of pegylated arginase (BCT‑100) has been shown in acute myeloid leukaemia, hepatocellular carcinoma and mesothelioma. The aim of the present study was to evaluate the effect of BCT‑100 in lung adenocarcinoma. A panel of lung adenocarcinoma cell lines and xenograft models were used to investigate the effect of BCT‑100. Protein expression, arginine level, putrescine level, spermidine level and apoptosis were analyzed by western blotting, ELISA, high performance liquid chromatography, dot blot and TUNEL assay, respectively. BCT‑100 converts arginine to ornithine. BCT‑100 reduced in vitro cell viability across different lung adenocarcinoma cell lines and suppressed tumour growth in an HCC4006 xenograft, while paradoxical growth stimulation was observed in H358, HCC827, H1650 and H1975 xenografts. Upon BCT‑100 treatment, ornithine decarboxylase 1 (ODC1) was induced in two solid tumour xenografts (H1650 and H1975). It was postulated that the accumulated ornithine could be channeled via ODC1 to produce polyamines that promoted tumour growth. The action of an ODC1 inhibitor (α‑difluoromethylornithine, DFMO) was studied in the restoration of the anticancer effects of BCT‑100 in lung adenocarcinoma. In both H1650 and H1975 xenografts, a combination of DFMO and BCT‑100 significantly suppressed tumour growth, resulting in doubled median survival compared with the control. Putrescine was decreased in almost all treatment arms in the H1650, H1975 and HCC4006 xenografts. Nonetheless spermidine was reduced only following DFMO/BCT‑100 treatment in the H1650 and H1975 xenografts. Apoptosis was enhanced in the combined treatment arm in both H1650 and H1975 xenografts. In the HCC4006 xenograft, addition of DFMO did not alter the tumour suppressive effect of BCT‑100. In conclusion, inhibition of ODC1 by DFMO was crucial in facilitating BCT‑100 treatment in lung adenocarcinoma that was partially mediated by depleting arginine and polyamines with consequent apoptosis.

• A panel of markers for identification of malignant and non-malignant cells in culture from effusions. F.P. Carneiro, M.I. Muniz-Junqueira, F. Pittella-Silva, et al. 2017;38(6):3538-3544. PubMed · Publisher

  Abstract

  The aim of the present study was to identify cell types in primary culture from malignant and non-malignant effusions. Effusion samples were subjected to cytology and culture. Immunocytochemistry was performed in cytological slides to evaluate malignancy (positivity for malignancy markers) and in culture slides for identification of cell types in growth. A total of 143 effusion samples (pleural n=76; peritoneal n=37; pericardial n=4; and peritoneal lavage n=26) were analyzed. Cell growth was observed in 34.9% of all samples and immunocytochemistry for identification of cell types in culture slides was conclusive in 90% of them. In non-malignant samples (n=28), growth of mesothelial cells, macrophages and of both cell types was identified in 82.14, 10.71 and 7.14%, respectively. In malignant samples (n=17, all carcinomas), growth of malignant epithelial cells and of both malignant epithelial and mesothelial cells was identified in 41.17 and 23.52%, respectively. In the remaining 35.29% of malignant samples, the only cells in growth were mesothelial and/or macrophages instead of malignant epithelial cells. In conclusion, in culture of malignant effusions, mesothelial cells may be simultaneously identified with malignant epithelial cells. Besides, mesothelial cells and macrophages may be the only cells identified in malignant effusion culture. Therefore, a broad panel of cell markers should be used for unmistakable identification of cells in studies of effusion primary culture. The ideal malignant effusion sample to obtain culture of neoplastic cells should be that without the presence of mesothelial cells and macrophages.

• EpCAM-independent capture of circulating tumor cells with a 'universal CTC-chip'. Y. Chikaishi, K. Yoneda, T. Ohnaga, et al. 2017;37(1):77-82. PubMed · Publisher

  Abstract

  Capture of circulating tumor cells (CTCs), which are shed from the primary tumor site and circulate in the blood, remains a technical challenge. CellSearch® is the only clinically approved CTC detection system, but has provided only modest sensitivity in detecting CTCs mainly because epithelial cell adhesion molecule (EpCAM)-negative tumor cells may not be captured. To achieve more sensitive CTC‑capture, we have developed a novel microfluidic platform, a 'CTC-chip' comprised of light-curable resins that has a unique advantage in that any capture antibody is easily conjugated. In the present study, we showed that EpCAM-negative tumor cells as well as EpCAM-positive cells were captured with the novel 'universal CTC-chip' as follows: i) human lung cancer cells (PC-9), with strong EpCAM expression, were efficiently captured with the CTC-chip coated with an anti-EpCAM antibody (with an average capture efficiency of 101% when tumor cells were spiked in phosphate‑buffered saline (PBS) and 88% when spiked in blood); ii) human mesothelioma cells (ACC-MESO-4), with no EpCAM expression but with podoplanin expression, were captured with the CTC-chip coated with an anti-podoplanin antibody (average capture efficiency of 78% when tumor cells were spiked in PBS and 38% when spiked in blood), whereas ACC-MESO-4 cells were not captured with the CTC-chip coated with the anti-EpCAM antibody. These results indicate that the novel 'CTC-chip' can be useful in sensitive EpCAM-independent detection of CTCs, which may provide new insights into personalized medicine.

• Auranofin induces mesothelioma cell death through oxidative stress and GSH depletion. B.R. You, W.H. Park. 2016;35(1):546-51. PubMed · Publisher

  Abstract

  Mesothelioma is an aggressive tumor associated with asbestos exposure. Auranofin as an inhibitor of thioredoxin reductase (TrxR) affects many biological processes such as inflammation and proliferation. In the present study, we investigated the cellular effects of auranofin on patient-derived mesothelioma cells in relation to reactive oxygen species (ROS) and glutathione (GSH) levels. Basal TrxR1 levels have no difference between mesothelial cells and certain mesothelioma cells. In particular, ADA, CON and Hmeso mesothelioma cells showed lower levels of TrxR1 expression. Auranofin inhibited the proliferation of mesothelioma cells in a dose-dependent manner. Among mesothelioma cells were ADA and CON cells sensitive to auranofin. This agent also induced caspase-independent apoptosis and necrosis in ADA cells. In addition, auranofin increased ROS levels including O2(•-) and induced GSH depletion in mesothelioma cells. While N-acetyl cysteine (NAC) prevented cell death and decreased ROS levels in auranofin-treated mesothelioma cells, L-buthionine sulfoximine (BSO) intensified apoptosis and GSH depletion in these cells. In conclusion, auranofin induced mesothelioma cell death through oxidative stress and the death was regulated by the status of GSH content.

• Constitutive expression and anticancer potency of a novel immunotoxin onconase-DV3. M. Sun, H. Tang, Y. Gao, et al. 2016;35(4):1987-94. PubMed · Publisher

  Abstract

  Onconase is an RNase of the ribonuclease A superfamily that is purified from the Northern leopard frog (Rana pipiens). It targets several types of malignant tumors, digests cytoplasmic transfer RNA (tRNA), and causes tumor cell apoptosis. Onconase has been employed in clinical trials as an antitumor drug, and has revealed its valuable clinical activity in several types of tumors, particularly pleural mesothelioma. However, its inefficiency in targeting tumor cells and its non‑specific toxicity in normal tissues have diminished its clinical benefits. Furthermore, cyclization of the N-terminal glutamine residue (Gln), possesses more RNase activity than the structure of Met ahead of Glu in the N-terminal (99:1), which is more difficult for producing onconase by Pichia pastoris. Under the guidance of α-mating factor-pre (α-MF-pre) secretion signal, the secretion of the recombinant protein can reach a high level. In the present study, we constructed a constitutive expression vector for onconase-(DV3)2 (Onc-DV3) production in yeast Pichia pastoris with the GAP promoter, in which the Onc-DV3 gene is inserted downstream of the truncated Saccharomyces cerevisiae α-mating factor-pre (α-MF-pre) secretion signal. The immuno-RNase Onc-DV3 expressed a high level of production and bioactivity and possessed enhanced capability to deliver the Onc molecule to tumor cell monomeric counterparts. Notably, Onc-DV3 showed strong cytotoxicity to highly metastatic tumor cells, weak cytotoxicity to lowly metastatic tumor cells and no toxicity to normal cells. These results demonstrate that the specific toxicity to highly metastatic tumor cells has made Onc-DV3 a promising antitumor drug by using two copies of DV3 for the targeted delivery of onconase.

• Manumycin A induces apoptosis in malignant pleural mesothelioma through regulation of Sp1 and activation of the mitochondria-related apoptotic pathway. K.H. Kim, J.I. Chae, H. Oh, et al. 2016;36(1):117-24. PubMed · Publisher

  Abstract

  Manumycin A (Manu A) is a natural product isolated from Streptomyces parvulus and has been reported to have anti-carcinogenic and anti-biotic properties. However, neither its molecular mechanism nor its molecular targets are well understood. Thus, the aim of the present study was to explore the possibility that Manu A has cancer preventive and chemotherapeutic effects on malignant pleural mesothelioma (MPM) through regulation of Sp1 and induction of mitochondrial cell death pathway. Manu A inhibited the cell viability of MSTO-211H and H28 cells in a concentration‑dependent manner as determined by MTS assay. IC50 values were calculated as 8.3 and 4.3 µM in the MSTO-311H and H28 cells following 48 h incubation, respectively. Manu A induced a significant increase in apoptotic indices as shown by DAPI staining, Annexin V assay, multi-caspase activity and mitochondrial membrane potential assay. The downregulation of Sp1 mRNA and protein expression by Manu A led to apoptosis by suppressing Sp1-regulated proteins (cyclin D1, Mcl-1 and survivin). Manu A decreased the protein levels of BID, Bcl-xL and PARP while it increased Bax levels. Manu A caused depolarization of the mitochondrial membrane with induction of CHOP, DR4 and DR5. Our results demonstrated that Manu A exerted anticancer effects by inducing apoptosis via inhibition of the Sp1-related signaling pathway in human MPM.

• Combinatorial anti-angiogenic gene therapy in a human malignant mesothelioma model. S. Kubo, M. Takagi-Kimura, N. Kasahara. 2015;34(2):633-8. PubMed · Publisher

  Abstract

  Anti-angiogenic gene therapy represents a promising strategy for cancer; however, it has rarely been tested in malignant mesothelioma, a highly aggressive tumor associated with asbestos with poor prognosis. In the present study, we investigated whether anti-angiogenic factors such as angiostatin, endostatin and the soluble form of vascular endothelial growth factor receptor 2 (sFlk1) were able to inhibit endothelial cell proliferation via lentivirus-mediated gene transfer into malignant mesothelioma cells in culture. We also assessed whether a dual-agent strategy had greater therapeutic benefit. Human malignant pleural mesothelioma MSTO-211H cells were transduced using lentiviral vectors that individually expressed angiostatin, endostatin and sFlk1 and linked to enhanced green fluorescent protein (EGFP) marker gene expression via an internal ribosome entry site. The lentivirus expressing EGFP alone was used as a control. The resultant cells designated as MSTO-A, MSTO-E, MSTO-F and MSTO-C were confirmed by western blot analysis and fluorescence microscopy to stably express the corresponding proteins. No differences were observed in the in vitro growth rates between any of these cells. However, co-culture of MSTO-A, MSTO-E and MSTO-F showed significant suppression of human umbilical endothelial cell growth in vitro compared with that of MSTO-C. Furthermore, a combination of any two among MSTO-A, MSTO-E and MSTO-F significantly enhanced efficacy. These results suggest that combinatorial anti-angiogenic gene therapy targeting different pathways of endothelial growth factor signaling has the potential for greater therapeutic efficacy than that of a single-agent regimen.

• Inhibition of NADPH oxidase 4 induces apoptosis in malignant mesothelioma: Role of reactive oxygen species. M. Tanaka, Y. Miura, H. Numanami, et al. 2015;34(4):1726-32. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) is an aggressive tumor that is characterized by dysregulated growth and resistance to apoptosis. Reactive oxygen species (ROS)-generating NADPH oxidase (Nox) family enzymes have been suggested to be involved in neoplastic proliferation. Both the antioxidant N-acetylcysteine (NAC) and the inhibitor of flavoprotein-dependent oxidase, diphenylene iodonium (DPI), inhibited the cell viability of MPM cells in a dose-dependent manner. To examine whether Nox-mediated ROS generation confers antiapoptotic activity and thus a growth advantage to MPM cells, we analyzed the mRNA expression of Nox family members using quantitative RT-PCR in 7 MPM cell lines and a normal mesothelial cell line. Nox4 mRNA was expressed in all of the examined MPM cell lines, whereas little or no Nox2, Nox3 and Nox5 mRNA expression was detected. In 2 MPM cell lines, Nox4 mRNA expression was significantly higher than that in a normal mesothelial cell line. siRNAs targeting Nox4 suppressed ROS generation and cell viability in the MPM cell lines. In addition, DPI treatment and knockdown of Nox4 attenuated phosphorylation of AKT and ERK. Taken together, our results indicate that Nox4-mediated ROS, at least in part, transmit cell survival signals and their depletion leads to apoptosis, thus highlighting the Nox4-ROS-AKT signaling pathway as a potential therapeutic target for MPM treatment.

• Inhibition of Src family kinases overcomes anoikis resistance induced by spheroid formation and facilitates cisplatin-induced apoptosis in human mesothelioma cells. R. Eguchi, Y. Fujita, C. Tabata, et al. 2015;34(5):2305-10. PubMed · Publisher

  Abstract

  Malignant mesothelioma is an aggressive tumor arising from mesothelial cells of serous membranes, and forms spheroid-like cell aggregates in pleural and peritoneal effusions. We examined the levels of anoikis, apoptosis induced by the detachment of cells from the extracellur matrix, in suspension culture in the human mesothelioma cell line NCI-H2052. NCI-H2052 cells were adherent in conventional monolayer cultures, but were found to form spheroids in suspension cultures using dishes with ultra-low cell binding capacity. NCI-H2052 cells proliferated in both cultures, but the proliferation rate was markedly lower in suspension cultures than in monolayer cultures. In addition, NCI-H2052 cells in suspension cultures showed little apoptosis, suggesting that the suspension culture induces anoikis resistance. Western blot analysis revealed that suspension cultures induced activation of Src family kinases (SFK) after spheroid formation. Dasatinib, an inhibitor of multi-tyrosine kinases including SFK, abolished anoikis resistance in suspension cultures, indicating that SFK activated by spheroid formation are responsible for anoikis resistance. Cisplatin induced apoptosis in NCI-H2052 cells, but the apoptotic rate was significantly lower in suspension cultures than in monolayer cultures, suggesting that spheroid formation is involved in cisplatin resistance. Furthermore, a combination of dasatinib and cisplatin induced apoptosis more significantly than either alone in suspension cultures. These results suggest that spheroid formation induces resistance to anoikis and to cisplatin through SFK activation and that dasatinib facilitates cisplatin-induced apoptosis in human mesothelioma cells.

• Lapatinib enhances trastuzumab-mediated antibody-dependent cellular cytotoxicity via upregulation of HER2 in malignant mesothelioma cells. R. Okita, K. Shimizu, Y. Nojima, et al. 2015;34(6):2864-70. PubMed · Publisher · Free full text

  Abstract

  EGFR/HER2 are frequently expressed in MPM tissues, however, no studies have shown the clinical benefit of using EGFR/HER2-targeting drugs in patients with malignant pleural mesothelioma (MPM). It was reported that the tyrosine kinase inhibitor (TKI) lapatinib enhanced trastuzumab-mediated antibody-dependent cellular cytotoxicity (ADCC) in HER2-positive breast cancer, suggesting that this combination is a promising strategy for MPM treatment. The aim of the present study was to explore the possibility of a TKI combined with trastuzumab to enhance ADCC in MPM cells. Five MPM cell lines were used to test the effects of TKIs targeting EGFR (gefitinib, afatinib and lapatinib) on cell proliferation and the expression of the HER family receptor. The combined effects of TKI with trastuzumab on ADCC were evaluated using the LDH release assay. Additionally, MPM cells were isolated from patients and evaluated for lapatinib-induced upregulation of HER family receptors and trastuzumab- or cetuximab‑mediated ADCC. In MPM cell lines, HER2 expression was upregulated by lapatinib, downregulated by afatinib and unaffected by gefitinib. As expected, more trastuzumab bound to MPM cells pretreated with lapatinib than untreated cells, resulting in the enhancement of trastuzumab-mediated ADCC in MPM cells. In patient-derived MPM cells, both HER2 and EGFR were upregulated by lapatinib, resulting in the enhancement of both trastuzumab- and cetuximab-mediated ADCC. Of the three TKIs, only lapatinib enhanced trastuzumab-mediated ADCC via the upregulation of HER2 expression in MPM cells, suggesting that sequential combination of lapatinib and trastuzumab may be a promising strategy for MPM treatment.

• Pemetrexed induces apoptosis in malignant mesothelioma and lung cancer cells through activation of reactive oxygen species and inhibition of sirtuin 1. K.E. Hwang, Y.S. Kim, Y.R. Hwang, et al. 2015;33(5):2411-9. PubMed · Publisher

  Abstract

  Pemetrexed is a multitargeted antifolate used for the treatment of malignant mesothelioma and non-small cell lung cancer (NSCLC). However, the mechanism by which pemetrexed induces apoptosis remains unclear. In the present study, we investigated the involvement of reactive oxygen species (ROS) and sirtuin 1 (SIRT1) in pemetrexed-induced apoptosis in MSTO-211 malignant mesothelioma cells and A549 NSCLC cells. Pemetrexed enhanced caspase-dependent apoptosis, induced intracellular ROS generation, and downregulated SIRT1 in the MSTO-211 and A549 cells. Pemetrexed-induced apoptosis, which was prevented by pretreatment with N-acetyl-cysteine (NAC), was mediated by effects on the mitochondria, including mitochondrial membrane potential transition (MPT) and cytosolic release of cytochrome c, and also involved regulation of SIRT1 expression. Interference with SIRT1 expression using siRNA enhanced pemetrexed-induced apoptosis through mitochondrial dysfunction and ROS generation, whereas resveratrol, an activator of SIRT1, protected against pemetrexed-induced apoptosis. These results show that pemetrexed induces apoptosis in MSTO-211 mesothelioma cells and A549 NSCLC cells through mitochondrial dysfunction mediated by ROS accumulation and SIRT1 downregulation.

• Wnt7A is a putative prognostic and chemosensitivity marker in human malignant pleural mesothelioma. T. Hirata, Q. Zheng, Z. Chen, et al. 2015;33(4):2052-60. PubMed · Publisher · Free full text

  Abstract

  Malignant pleural mesothelioma (MPM) is a highly aggressive tumor that has a poor prognosis, limited treatment options, and a worldwide incidence that is expected to increase in the next decade. We evaluated Wnt7A expression in 50 surgically resected tumor specimens using quantitative PCR. The expression values, were assessed by clinicopathological factors and K-M and Cox's regression with OS. The mean level of Wnt7A expression had a significant correlation with International Mesothelioma Interest Group (IMIG) stage (P<0.034), gender, smoking history and ethnicity, respectively (P=0.020, P=0.014, P=0.039). In the univariate analysis, low Wnt7A expression was a significant negative factor for overall survival (P=0.043, HR=2.30). However, multivariate Cox's regression revealed no significant factors for overall survival (low Wnt7A: P=0.051, HR=2.283; non-epithelioid subtype: P=0.050, HR=2.898). In patients with epithelioid tumors, those with low Wnt7A expression had significantly worse prognosis (P=0.019, HR=2.98). In patients with epithelioid tumors, females had significantly better prognosis than males (P=0.035). In patients who did not have neoadjuvant chemotherapy, prognosis was significantly more favorable for patients with high Wnt7A expression than for those with low Wnt7A expression (P=0.031). Among the patients with low Wnt7A-expressing tumors, those who received neoadjuvant chemotherapy had better prognosis than those who did not (P=0.024). The results of our study suggest that Wnt7A expression is a putative prognostic factor and a predictor of chemosensitivity.

• Antitumor effect and antiangiogenic potential of the mTOR inhibitor temsirolimus against malignant pleural mesothelioma. M. Moriya, T. Yamada, M. Tamura, et al. 2014;31(3):1109-15. PubMed · Publisher

  Abstract

  The mTOR inhibitor temsirolimus has antitumor and antiangiogenic activity against several carcinomas, yet few reports document the efficacy of temsirolimus against malignant pleural mesothelioma (MPM). Therefore, we evaluated the efficacy of temsirolimus and the antiangiogenic effect of temsirolimus in the treatment of MPM. We examined the efficacy of temsirolimus alone and the efficacy of the combination of temsirolimus and cisplatin or pemetrexed against four MPM cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The effect of temsirolimus on the production of proangiogenic cytokines by MPM cell lines was examined by enzyme-linked immunosorbent assay (ELISA). Expression of mTOR and proangiogenic cytokines in clinical specimens from MPM patients was determined by immunohistochemistry. Temsirolimus inhibited cell viability and suppressed cell proliferation of all MPM cell lines. Combined treatment with temsirolimus and cisplatin inhibited the viability of all MPM cell lines more effectively than temsirolimus alone. Temsirolimus strongly inhibited the phosphorylation of p70s6k, a downstream molecule of mTOR, in all MPM cell lines and led to an increase in the levels of cleaved caspase-3 in the H226 and Y-meso14 cells. Temsirolimus also inhibited the production of vascular endothelial growth factor (VEGF) and platelet-derived growth factor-AA (PDGF-AA). Phosphorylated mTOR and high expression of VEGF and PDGF were detected in 2 and 3, respectively, out of the 5 MPM specimens. These results suggest that temsirolimus has activity against MPM cells by inhibition of cell proliferation and angiogenesis, and may be beneficial for a subset of MPM patients with high mTOR expression.

• Combined effects of asbestos and cigarette smoke on the development of lung adenocarcinoma: different carcinogens may cause different genomic changes. K. Inamura, H. Ninomiya, K. Nomura, et al. 2014;32(2):475-82. PubMed · Publisher · Free full text

  Abstract

  The carcinogens in cigarette smoke are distinct from asbestos. However, an understanding of their differential effects on lung adenocarcinoma development remains elusive. We investigated loss of heterozygosity (LOH) and the p53 mutation in 132 lung adenocarcinomas, for which asbestos body burden (AB; in numbers per gram of dry lung) was measured using adjacent normal lung. All cases were classified into 9 groups based on a matrix of cumulative smoking (CS in pack‑years; CS=0, 0<CS<25, ≥25 CS) and AB (AB=0, 0<AB<1,000, ≥1,000 AB). AB=0 indicates a lower level than the detection limit of ~100. LOH frequency increased only slightly with the elevation of CS in the AB=0 groups. In the AB>0 groups, LOH frequency increased as AB and/or CS was elevated and was significantly higher in the ≥1,000 AB, ≥25 CS group (p=0.032). p53 mutation frequency was the lowest in the AB=0, CS=0 group, increased as AB and/or CS rose, and was significantly higher in the ≥1,000 AB, ≥25 CS group (p=0.039). p53 mutations characteristic of smoking were frequently observed in the CS>0 groups contrary to non-specific mutations in the CS=0, AB>0 groups. Combined effects of asbestos and smoking were suggested by LOH and p53 analyses. Sole exposure to asbestos did not increase LOH frequency but increased non‑specific p53 mutations. These findings indicate that the major carcinogenic mechanism of asbestos may be tumor promotion, acting in an additive or synergistic manner, contributing to the genotoxic effect of smoking. Since this study was based on a general cancer center's experience, the limited sample size did not permit the consideration that the result was conclusive. Further investigation with a large sample size is needed to establish the mechanism of asbestos-induced lung carcinogenesis.

• ERC/mesothelin is expressed in human gastric cancer tissues and cell lines. T. Ito, K. Kajino, M. Abe, et al. 2014;31(1):27-33. PubMed · Publisher · Free full text

  Abstract

  ERC/mesothelin is expressed in mesothelioma and other malignancies. The ERC/mesothelin gene (MSLN) encodes a 71-kDa precursor protein, which is cleaved to yield 31-kDa N-terminal (N-ERC/mesothelin) and 40-kDa C-terminal (C-ERC/mesothelin) proteins. N-ERC/mesothelin is a soluble protein and has been reported to be a diagnostic serum marker of mesothelioma and ovarian cancer. Gastric cancer tissue also expresses C-ERC/mesothelin, but the significance of serum N-ERC levels for diagnosing gastric cancer has not yet been studied. We examined the latter issue in the present study as well as C-ERC/mesothelin expression in human gastric cancer tissues and cell lines. We immunohistochemically examined C-ERC/mesothelin expression in tissue samples from 50 cases of gastric cancer, and we also assessed the C-ERC/mesothelin expression in 6 gastric cancer cell lines (MKN-1, MKN-7, MKN-74, NUGC-3, NUGC-4 and TMK-1) using reverse transcription-polymerase chain reaction, flow cytometry, immunohistochemistry and immunoblotting. We also examined the N-ERC/mesothelin concentrations in the supernatants of cultured cells and in the sera of gastric cancer patients using an enzyme-linked immunosorbent assay (ELISA). N-ERC/mesothelin was detected in the supernatants of 3 gastric cancer cell lines (MKN-1, NUGC-4 and TMK-1) by ELISA, but its concentration in the sera of gastric cancer patients was almost same as that observed in the sera of the normal controls. In the gastric cancer tissues, C-ERC/mesothelin expression was associated with lymphatic invasion. N-ERC/mesothelin was secreted into the supernatants of gastric cancer cell lines, but does not appear to be a useful serum marker of gastric cancer.

• Lymph node involvement and metastatic lymph node ratio influence the survival of malignant pleural mesothelioma: a French multicenter retrospective study. I. Hysi, F. Le Pimpec-Barthes, M. Alifano, et al. 2014;31(1):415-21. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) is a rare tumor with disastrous evolution. The prognostic value of nodal involvement is still debated. We analyzed the impact of nodal involvement on overall survival (OS) in patients treated by multimodal therapy including extra pleural pneumonectomy (EPP). We evaluated the role, as a prognostic factor, of the metastatic lymph node ratio (LNR), corresponding to the number of involved nodes out of the total number of removed nodes. In this retrospective multicentric study, we reviewed the data of 99 MPM patients. Information regarding lymph node involvement was assessed from the final pathology reports. N1-N3 patients were pooled as N+ group. The OS, calculated by the Kaplan-Meier method, was compared using the log-rank test. A multivariate Cox proportional hazards model was used to identify independent prognostic factors. For the whole cohort, median OS was 18.3 months and 5-year survival was 17.5%. N+ status reduced significantly the median survival (22.4 months for N0 patients vs 12.7 months for N+ patients, P=0.002). A lower metastatic LNR (≤13%) was associated with a significantly improved median survival (19.9 vs. 11.7 months, P=0.01). OS was not related to the number of involved or total removed lymph nodes. In multivariate analysis, only adjuvant radiotherapy (P=0.001) was identified as an independent positive prognostic factor. Metastatic LNR is a more reliable prognostic factor than the number of involved lymph nodes or the total number of removed nodes. However, it could not be identified as an independent prognostic factor.

• MiR-379/411 cluster regulates IL-18 and contributes to drug resistance in malignant pleural mesothelioma. K. Yamamoto, M. Seike, S. Takeuchi, et al. 2014;32(6):2365-72. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) is a rapidly fatal malignancy that is increasing in incidence in Japan. In this study, we performed gene and microRNA (miRNA) expression profiling to identify novel therapeutic targets in MPM cells. Based on relative sensitivities to pemetrexed (PEM) and the histone deacetylase (HDAC) inhibitor, vorinostat (SAHA), 211H cells were determined to be the only sensitive MPM cell line out of the 6 tested. On the same series of cell lines, we performed whole genome transcriptomic profiling via DNA microarrays and pathway analysis of the derived data. Of particular note, IL-18 gene expression levels were significantly higher in the cell lines that were either drug resistant or displayed intermediate sensitivity, compared to the sensitive 211H cell line. Pathway analysis revealed IL-18 as an important gene associated with drug sensitivity of MPM cells. A relationship between IL-18 overexpression and drug resistance was also observed following targeted assessment of 10 cytokine genes using quantitative RT-PCR. miRNA expression profiles were evaluated in the MPM cell line panel in order to discern the mechanism of IL-18 induction in the drug-resistant lines. We found that miR-379 and miR-411 belonged to the same cluster of miRNAs located on chromosome 14q32 that commonly target the IL-18 gene. Luciferase reporter assays revealed that miR-379 and miR-411 directly target the IL-18 gene. Introduction of miR-379 plus miR-411, as well as IL-18 silencing, significantly suppressed the invasive capacity of MESO1 cells in vitro. Furthermore, the use of either PEM or SAHA together with miR-379 plus miR-411 mimics mediated increased sensitivity to these drugs in MESO1 cells. These results suggest that the miR-379/411 cluster may provide new therapeutic opportunities for advanced MPM patients, depending on the nature of IL-18 gene expression.

• ¹⁸F-FDG uptake on PET is a predictive marker of thymidylate synthase expression in patients with thoracic neoplasms. K. Kaira, N. Yamamoto, M. Endo, et al. 2014;31(1):209-15. PubMed · Publisher

  Abstract

  The aim of this study was to investigate the relationship between the expression levels of thymidylate synthase (TS) and 2-[¹⁸F]-fluoro-2-deoxy-D-glucose (¹⁸F-FDG) uptake on positron emission tomography (PET) in various thoracic neoplasms. In total, 392 patients [non-small cell lung cancer (NSCLC) (n=140), malignant pleural mesothelioma (MPM) (n=21), pulmonary metastatic tumors (PMT) (n=148), thymic epithelial tumors (n=49) and pulmonary neuroendocrine (NE) tumor (n=34)] who underwent ¹⁸F-FDG PET before treatment were included in this study. Tumor sections were stained using immunohistochemistry for determination of TS, orotate phosphoribosyltransferase (OPRT), dihydropyrimidine dehydrogenase (DPD), vascular endothelial growth factor (VEGF), microvessel density (MVD), CD34 and p53. The expression of TS in thoracic neoplasms had a positivity of 58% (230/392), and the positive rates of TS expression in NSCLC, PMT, thymic epithelial tumor, NE tumor and MPM samples were 56, 57, 57, 85 and 47%, respectively. The positivity of TS expression was significantly higher in NE tumors compared to that in other thoracic tumors. A statistically significant correlation between TS expression and ¹⁸F-FDG uptake was observed in thoracic neoplasms, in particular primary lung adenocarcinomas, high-grade NE tumors, thymomas and MPMs. Moreover, TS expression was closely associated with angiogenesis, DPD, OPRT and p53. Our results indicated that SUV(max) by ¹⁸F-FDG uptake may be an alternative biomarker for predicting TS expression in patients with primary lung adenocarcinoma, high-grade NE tumor, thymoma and MPM.

• CD9 expression as a favorable prognostic marker for patients with malignant mesothelioma. V.J. Amatya, Y. Takeshima, K. Aoe, et al. 2013;29(1):21-8. PubMed · Publisher · Free full text

  Abstract

  CD9 is involved in cell growth, adhesion and motility and its expression is reported to be of prognostic significance in various types of human malignancies. We found increased cell migration in the mesothelioma cell lines MSTO-211H and TUM1 following in vitro shRNA-mediated knockdown of CD9 expression. We investigated CD9 expression in 112 malignant pleural mesotheliomas. CD9 expression was observed in 62 of 71 epithelioid, 13 of 20 biphasic and only 1 of 21 sarcomatoid mesotheliomas. Among the epithelioid mesotheliomas (EMs), CD9 expression was observed in all of the 33 cases with a differentiated type (EM-D) and in 29 of the 38 cases with a less-differentiated type (EM-LD). Patients with CD9 expression showed higher 1- and 2-year survival rates (63 and 25%) compared to the patients without CD9 expression (39 and 11%). Univariate analysis revealed that patients with CD9 expression demonstrated a more favorable survival (P=0.0025) along with other clinicopathological factors, including age younger than 60 years, IMIG stage I-II, epithelioid histology, EM-D and patients who underwent extrapleural pneumonectomy or received chemotherapy. Multivariate analysis identified CD9 expression as an independent prognostic factor with a hazard ratio (HR) of 1.99 in the analysis of all mesotheliomas (P=0.0261) and an HR of 2.60 in the analysis of EMs (P=0.0376). CD9 expression is an independent favorable prognostic marker of malignant mesothelioma.

• Downregulation of microRNA-34 induces cell proliferation and invasion of human mesothelial cells. N. Tanaka, S. Toyooka, J. Soh, et al. 2013;29(6):2169-74. PubMed · Publisher

  Abstract

  Malignant mesothelioma (MM) is an aggressive tumor with a dismal prognosis, and the molecular alterations involved in this disease remain unknown. We previously reported that microRNA-34s (miR-34s) are methylated and downregulated in MM and may play an important role in the carcinogenesis of MM. In this study, we downregulated miR-34s in human mesothelial cells to investigate the cellular effect of miR-34 knockdown. For the cell study, we used LP-9, a human mesothelial cell line, and three human primary-cultured mesothelial cell lines. RNA-based miR-34a, -34b and -34c inhibitors were transfected into these cells, and their effects on proliferation and invasion were evaluated. A scramble RNA oligonucleotide was used as a control. The protein expression status was estimated using western blotting. After miR-34 inhibitor transfection, miR-34a, -34b and -34c were downregulated in all the examined mesothelial cell lines. miR-34 inhibitor transfection significantly increased cell proliferation in all of the mesothelial cell lines, compared with the scramble control. The invasive ability also increased in the miR-34 inhibitor transfectants, compared with the scramble control, in the LP-9 cell line. Western blotting confirmed the upregulation of c-MET, phospho-c-MET, and bcl-2 proteins in LP-9 cells after miR-34 inhibitor transfection. In conclusion, our study showed that the downregulation of miR-34s induced an oncogenic phenotype in non-malignant mesothelial cells. The present study, together with the results of our previous report, strongly suggest that miR-34s play an important role in the early carcinogenic process involved in the transformation of human mesothelial cells to MM.

• Downregulation of Sp1 is involved in honokiol-induced cell cycle arrest and apoptosis in human malignant pleural mesothelioma cells. J.I. Chae, Y.J. Jeon, J.H. Shim. 2013;29(6):2318-24. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) is an extremely aggressive type of cancer and is associated with a poor patient prognosis due to its rapid progression. Novel therapeutic agents such as honokiol (HNK) improve the clinical outcomes of cancer therapy, yet the mechanisms involved have not been fully elucidated. The present study examined the regulatory effects of HNK on the growth and apoptosis of MSTO-211H mesothelioma cells and investigated its anticancer mechanism. The results revealed that HNK significantly reduced the cell viability and increased the sub-G1 population in MSTO-211H cells and suppressed the expression of the specificity protein 1 protein (Sp1). HNK reduced the transcriptional activity of Sp1 regulatory proteins, including cyclin D1, Mcl-1 and survivin, and, thus, induced apoptosis signaling pathways by increasing Bax, reducing Bid and Bcl-xl and activating caspase-3 and PARP in mesothelioma cells. The results suggest that Sp1, a novel molecular target of HNK, may be related to cell cycle arrest and apoptosis induction through the modulation of signal transduction pathways in MPM.

• Overexpression of Numb suppresses tumor cell growth and enhances sensitivity to cisplatin in epithelioid malignant pleural mesothelioma. Y. Kang, M. Ding, G. Tian, et al. 2013;30(1):313-9. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) is a highly aggressive and conventional treatment-resistant tumor with a dismal prognosis. Among the three histological subtypes of MPM, the epithelioid is the most common type. Numb is considered as a tumor suppressor playing a critical role in controlling asymmetric cell division, maintenance of stem cell compartments, ubiquitination of specific substrates and regulating Notch-, Hedgehog- and TP53-activated pathways. The present study was designed to analyze the role of Numb in epithelioid MPM. We investigated the expression of Numb in 39 epithelioid MPM and 22 normal pleural tissues by immunohistochemistry. Furthermore, we overexpressed Numb in NCI-H2452, an epithelioid human MPM cell line, and investigated the effect of Numb overexpression on the proliferation, apoptosis and sensitivity to cisplatin in cells. The expression of Numb was significantly lower in MPM compared to the control group and Numb had an inverse correlation with the ki-67 labeling index. Loss of Numb expression was associated with poor prognosis in epithelioid MPM. Overexpression of Numb in NCI-H2452 cells significantly inhibited proliferation, promoted apoptosis and enhanced sensitivity to cisplatin. Moreover, Numb overexpression activated caspase-9 and caspase-3 through release of cytochrome c as well as downregulation of XIAP and survivin. We speculate that cytochrome c/caspase signaling is a possible mechanism through which Numb enhances the apoptosis of NCI-H2452 cells. These results suggest that Numb may be involved in epithelioid MPM development, and its upregulation may confer sensitivity to cisplatin, suggesting potential therapeutic options for MPM.

• Podoplanin overexpression in human mesothelioma cell lines enhances the tumorigenic phenotype. E. Yamaki, T. Yajima, T. Kosaka, et al. 2013;29(3):932-40. PubMed · Publisher

  Abstract

  Podoplanin, a small type I integral membrane mucin-type sialoglycoprotein, serves as a useful marker for diagnosing malignant pleural mesothelioma (MPM); however, the physiological function of podoplanin in mesothelioma cells is not known. To elucidate the role of podoplanin in the pathogenesis of MPM, we generated two mesothelioma cell lines (PODO1 and PODO2) that stably express high levels of podoplanin. Although PODO1 cells proliferated to the same extent in culture or in nude mice, the survival rate of the mice was significantly reduced compared with that of the controls. We demonstrated that PODO1 and PODO2 cells had increased invasive ability in in vitro assays and induced upregulation of matrix metalloproteinase-1. PODO1 and PODO2 cultures could not be induced to undergo apoptosis when starved or treated with cis-diamminedichloroplatinum(II) (CDDP) compared with the controls. Moreover, silencing of podoplanin expression using RNA interference restored the ability of CDDP to induce apoptosis. Consistent with their growth properties, we detected constitutive activation of extracellular signal-regulated kinase in PODO1 and PODO2 cultures. These findings suggest that constitutive expression of podoplanin contributes to the invasive growth properties of mesothelioma cells and their resistance to apoptosis. Moreover, our data suggest that podoplanin or components of its signaling pathway, or both, may serve as important targets for developing novel treatments for MPM.

• A somatic TSHR mutation in a patient with lung adenocarcinoma with bronchioloalveolar carcinoma, coronary artery disease and severe chronic obstructive pulmonary disease. J.W. Kim, S. Lee, N. Lui, et al. 2012;28(4):1225-30. Case report PubMed · Publisher

  Abstract

  In a screen for thoracic malignancy-associated markers, thyroid stimulating hormone receptor (TSHR) was identified as a candidate as it binds to the previously-characterized lung cancer marker NKX2-1. We screened for mutations in all coding regions of the TSHR gene in 96 lung adenocarcinoma samples and their matched adjacent normal lung samples. We found one patient with a somatic mutation at codon 458 (exon 10), which is located at the transmembrane domain where most TSHR mutations have been found in thyroid-related diseases. This patient had lung adenocarcinoma with BAC (bronchioloalveolar carcinoma) features in the setting of a prior medical history significant for carotid stenosis and severe chronic obstructive pulmonary disease (COPD). In order to characterize the genetic features of TSHR in lung cancer, we checked for TSHR expression and copy number in the 96 lung cancer tissues. TSHR protein expression was generally overexpressed in multiple thoracic malignancies (adenocarcinoma, squamous cell carcinoma and malignant pleural mesothelioma) by immunohistochemistry. Our data suggest that aberrant TSHR function may contribute to lung cancer development or a subgroup of lung cancer with specific clinical phenotypes.

• Clinical implications of 18F-fluorodeoxyglucose positron emission tomography/computed tomography at delayed phase for diagnosis and prognosis of malignant pleural mesothelioma. Y. Abe, K. Tamura, I. Sakata, et al. 2012;27(2):333-8. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) has a poor prognosis, and conventional imaging modalities do not reflect the prognosis of MPM. In this study, the clinical significance of 18F-fluorodeoxyglucose positron emission tomography/computed tomography fusion imaging (18F-FDG PET/CT) was evaluated for the differential diagnosis, staging and prognosis in MPM patients. Ninety patients who underwent 18F-FDG PET/CT scanning due to a clinical diagnosis or suspicion of MPM prior to therapy were reviewed. Of 90 patients, 31 were pathologically diagnosed as MPM. Maximum standardized uptake values (SUVmax) were semi-quantitatively obtained from PET/CT 60 min (early phase) and 120 min (delayed phase) after injection of 18F-FDG, and the clinicopathological correlations with the level of SUVmax obtained from PET/CT were examined. The survival curves of MPM patients were plotted according to the methods of Kaplan-Meier. The prognostic implications of the level of SUVmax were estimated by t-test. PET/CT scan showed intense abnormal FDG uptake (SUVmax>2.0) in the pleural lesions of all 31 MPM patients at delayed phase, while it showed abnormal FDG uptake in 30 (97%) patients at early phase. In all 31 MPM patients, the values of SUVmax at delayed phase were higher than those at the early phase. PET/CT also indicated metastasis in the lymph node in 7 patients (23%) and in the systemic lesions in 8 patients (26%) with MPM. Twenty-three MPM patients with high SUVmax, whose prognosis was apparent, showed significantly poorer prognosis in both early and delayed phase (respectively, p=0.03 and p=0.01, t-test). The results showed that 18F-FDG PET/CT at delayed phase is very useful for the diagnosis of pleural diseases, and SUVmax on PET/CT in the delayed phase is a more reliable prognostic factor than that in the early phase. High uptake of 18F-FDG PET/CT may be a predictive factor of prognosis in MPM patients.

• Extracellular protease imaging for cell mass tracking of xenografted human malignant pleural mesothelioma. L. Zhou, W. El-Deiry, W. Wang, et al. 2012;28(3):883-8. PubMed · Publisher

  Abstract

  Malignant pleural mesothelioma (MPM) is locally aggressive and challenging to quantitate non-invasively in vivo, particularly in orthotopic models of disease. We describe imaging of extracellular protease activity, typically elevated in locally aggressive tumors, as a novel method for tracking MPM in vivo. Mice bearing human MPM subcutaneous flank xenografted tumors were imaged with ProSense 680, an optical imaging agent of extracellular cysteine protease activity. The relative contribution of extracellular cysteine proteases to the ProSense tumor signal was estimated using RT-PCR quantitation of cysteine protease RNA expression of the MPM cell lines and compared to ArrayExpress microarray RNA expression data from human MPM tumors. Feasibility of orthotopic intraperitoneal MPM cell mass tracking with fluorescence signal was evaluated using CellVue Maroon-coated MSTO-211H and compared to bioluminescent signal using luciferase-transfected MSTO-211H cells. ProSense 680 yielded a robust tumor signal in MPM subcutaneous grafts, primarily resulting from MPM secretion of cathepsin L demonstrated not only by RT-PCR data on MPM cell lines but also by microarray expression data from resected human patient tumors. CellVue Maroon intraperitoneal tumor signal was robust and durable indicating feasibility of intraperitoneal cell mass tracking of orthotopically-xenografted MPM. Optical imaging of extracellular cysteine protease activity is useful for tracking MPM tumor cell mass in vivo. Intraperitoneal MPM cell mass tracking of fluorescently labeled tumor is feasible.

• Interaction and cross-resistance of cisplatin and pemetrexed in malignant pleural mesothelioma cell lines. M. Kitazono-Saitoh, Y. Takiguchi, S. Kitazono, et al. 2012;28(1):33-40. PubMed · Publisher

  Abstract

  Although cisplatin and pemetrexed are key drugs in the treatment of malignant pleural mesothelioma, their drug-drug interactions, cross-resistance and resistance mechanisms in malignant pleural mesothelioma are not well understood. In the present study, the interaction of these 2 agents was determined by clonogenic assays followed by isobologram analysis of 4 human malignant pleural mesothelioma cell lines. The cell lines were exposed to the agents using a stepwise dose-escalation method to establish drug-resistant sublines. Thymidylate synthase mRNA expression was evaluated in the drug-resistant sublines. As a consequence, cisplatin and pemetrexed had synergistic effects in 3 cell lines and an additive effect in the fourth cell line. The former 3 cell lines showed similar pemetrexed sensitivity in the parental cells and their cisplatin-resistant sublines, whereas the fourth cell line exhibited cross-resistance. In contrast, cisplatin had diverse effects on pemetrexed-resistant sublines. High thymidylate synthase expression did not correlate with natural pemetrexed resistance. Elevated thymidylate synthase expression correlated with acquired pemetrexed resistance in 2 sublines. In conclusion, cisplatin and pemetrexed showed synergistic activity and no cross-resistance in 3 of the 4 malignant pleural mesothelioma cell lines, suggesting the clinical relevance of their combination in chemotherapy. Thymidylate synthase expression did not necessarily correlate with pemetrexed resistance. The information together with the experimental model presented here would be useful for further investigating therapeutic targets of malignant mesothelioma.

• Patients with peritoneal mesothelioma lack epidermal growth factor receptor tyrosine kinase mutations that would make them sensitive to tyrosine kinase inhibitors. N. Kalra, A. Ashai, L. Xi, et al. 2012;27(6):1794-800. PubMed · Publisher · Free full text

  Abstract

  The epidermal growth factor receptor (EGFR) is a promising target for cancer therapy. The presence of certain somatic mutations in the tyrosine kinase (TK) domain of the EGFR gene is associated with clinical response to TK inhibitors (TKI) in patients with lung adenocarcinoma. In this study we evaluated the status of somatic mutations in the entire TK domain of the EGFR gene by direct sequencing using early passage peritoneal mesothelioma cells, established cell lines as well as 33 peritoneal mesothelioma tumor samples. No novel mutations were found in the cell lines. Sequence analysis of the EGFR TK domain revealed the presence of a silent polymorphism (c.2607G → A, Q787Q) at exon 20 of both peritoneal mesothelioma cell lines as well as tumor specimens. The frequency of genotypes AA and GA was 42.8 and 57.2% in the cell lines and 33.3 and 57.6% in tumor specimens, respectively. The TKI erlotinib showed an IC50 in the range of 10-50 µM in five out of the seven cell lines with a GA genotype while all five cell lines with the AA genotype had an IC50 >50 µM. Of the 33 peritoneal mesothelioma tumor samples analyzed none had an EGFR TKI sensitizing mutation and only one specimen showed an earlier reported somatic mutation at codon 850 in exon 21 of the EGFR gene. Our data show that patients with peritoneal mesothelioma do not harbor somatic mutations in the EGFR TK domain that would make them sensitive to EGFR TKI.

• The bone morphogenetic protein antagonist Gremlin is overexpressed in human malignant mesothelioma. D.J. Wang, X.Y. Zhi, S.C. Zhang, et al. 2012;27(1):58-64. PubMed · Publisher

  Abstract

  Gremlin is a member of the bone morphogenetic protein (BMP) antagonist family and its antagonistic effect is likely through direct binding to BMP proteins. As an antagonist of BMP, Gremlin plays a role in regulating organogenesis, body patterning and tissue differentiation. Recent studies have shown a deregulation of Gremlin in several types of human cancers. However, the role of Gremlin in human malignant mesothelioma (MM) is still unknown. In this study, we investigated the expression of Gremlin in human MM. We found that Gremlin mRNA and protein were both overexpressed in the majority of primary MM tissue samples that we examined. We also observed high level expression of the Gremlin gene in 4 of the 6 MM cell lines. Consistently, we found that the Gremlin promoter activity was significantly elevated in those MM cell lines expressing the Gremlin gene. On the other hand, no activity of the Gremlin promoter was detected in the two MM cell lines lacking Gremlin expression. Moreover, to examine the functional significance of the Gremlin overexpression in MM, we used shRNA to knock down Gremlin expression in MM cell lines expressing Gremlin and found that inhibition of Gremlin expression significantly suppressed proliferation of those MM cells. Taken together, our results suggest that the BMP antagonist Gremlin is overexpressed in MM and that aberrant activation of Gremlin may play a critical role in the tumorigenesis of human MM.

• Yes is a central mediator of cell growth in malignant mesothelioma cells. A. Sato, M. Sekine, N. Virgona, et al. 2012;28(5):1889-93. PubMed · Publisher

  Abstract

  The constitutive activation of the Src family kinases (SFKs) has been established as a poor prognostic factor in malignant mesothelioma (MM), however, the family member(s) which contribute to the malignancy have not been defined. This study aimed to identify the SFK member(s) contributing to cell growth using RNA interference in various MM cell lines. Silencing of Yes but not of c-Src or Fyn in MM cells leads to cell growth suppression. This suppressive effect caused by Yes silencing mainly depends on G1 cell cycle arrest and partly the induction of apoptosis. Also, the knockout of Yes induces the inactivation of β-catenin signaling and subsequently decreases the levels of cyclin D necessary for G1-S transition in the cell cycle. In addition, Yes knockout has less effect on cell growth suppression in β-catenin-deficient H28 MM cells compared to other MM cells which express the catenin. Overall, we conclude that Yes is a central mediator for MM cell growth that is not shared with other SFKs such as c-Src.

• Overexpression of CD26/DPPIV in mesothelioma tissue and mesothelioma cell lines. V.J. Amatya, Y. Takeshima, K. Kushitani, et al. 2011;26(6):1369-75. PubMed · Publisher

  Abstract

  Mesothelioma, a highly aggressive cancer with poor prognosis and refractory to currently available therapies show increasing trends of its incidence in Japan and other developing countries. Although surgery is a gold standard for patients with early mesothelioma, most patients with advanced disease are not suitable for surgical resection and have option of palliative chemotherapy alone. One of the new treatment strategies for mesothelioma, the humanized anti-CD26 monoclonal antibody therapy is under development. CD26, a 110-kDa transmembrane glycoprotein with known dipeptidyl peptidase IV activity, plays a role in tumor development and its expression was reported in various human malignancies. This study determined the preliminary selection criteria for humanized monoclonal anti-CD26 antibody therapy. Eighty-one epithelioid (49 differentiated and 32 less differentiated), 34 sarcomatoid, 19 biphasic mesothelioma and 8 mesothelioma cell lines were immunohistochemically examined using 8 different commercially available anti-CD26 antibodies for membranous and cytoplasmic expression. The cytoplasmic expression of CD26 was observed in all histological types of mesothelioma, while the membranous expression of CD26 was found in 88% of differentiated and 69% of less differentiated epithelioid mesothelioma, and none of sarcomatoid mesothelioma with anti-CD26 antibodies with rabbit polyclonal anti-DPP4 antibody and similar results were also obtained with goat polyclonal anti-DPP4/CD26 antibody. These antibodies absorbed with soluble human CD26 proteins do not show CD26 expression in mesothelioma tissue, suggesting these two antibodies localize true CD26 protein. Seven mesothelioma cell lines, including sarcomatoid types, also showed membranous expression of CD26 in cellblock preparation. CD26 vector transfection to CD26-negative MSTO-211H cells showed membranous expression of CD26 by flow cytometry, but not in tumor developed in NOD/SCID mice with inoculation of CD26 vector transfected MSTO-211H cells. We found that both rabbit and goat polyclonal antibodies are suitable for immunohistochemical evaluation of membranous expression of CD26 in mesothelioma.

• Aberrant promoter methylation of WIF-1 and SFRP1, 2, 4 genes in mesothelioma. H. Kohno, V.J. Amatya, Y. Takeshima, et al. 2010;24(2):423-31. PubMed · Publisher

  Abstract

  WIF-1 is a negative regulator of the Wnt-signaling pathway that may have important implications for tumorigenesis. Microarray analysis of whole genome expression in mesothelioma tissue revealed down-regulation of 491 genes and up-regulation of 167 genes involved mainly in Jak-STAT signaling (8 genes), MAPK signaling (16 genes) and Wnt signaling (13 genes) pathways. Of these, WIF-1 gene was down-regulated in mesothelioma 72-fold compared to normal tissue. We also analyzed WIF-1 and SFRPs promoter methylations in 46 mesothelioma tissues, 8 mesothelioma cell lines by methylation-specific polymerase chain reaction (MSP). WIF-1 promoter methylation was observed in 34 of 46 mesothelioma tissues (73.9%) and in all 8 mesothelioma cell lines. SFRP1, 2 and 4 promoter methylation was observed in 21 of 37 (56.8%), 26 of 42 (61.9%) and 17 of 36 (47.2%) mesothelioma tissues, respectively. Promoter methylation of any WIF-1 and/or SFRP genes was observed in 44 of 46 (95.6%) mesothelioma tissues. The treatment of mesothelioma cell lines with 5-aza-2'-deoxycytidine (5-aza-2dC) showed WIF-1 promoter methylation recovery followed by restoration of WIF-1 expression in 6 of 8 mesothelioma cell lines. The cytoplasmic expression of beta-catenin was observed in 38 of 43 cases of mesothelioma without any nuclear reactivity. The eight mesothelioma cell lines and 27 cases of mesothelioma examined showed no mutation in exon 3 of beta-catenin suggesting no alteration of canonical Wnt signaling pathway. Our data suggest that WIF-1 promoter methylation is a common event in mesothelioma.

  In plain English

  This Japanese laboratory study measured promoter methylation, a chemical DNA change that can silence genes, in four genes that restrain a cell signaling pathway called Wnt: WIF-1 and SFRP1, 2 and 4. Only for WIF-1 did it test whether methylation went with lower gene activity. A gene-activity scan found WIF-1 activity 72-fold lower in tumor than normal tissue. In tumor samples from 46 patients at Hiroshima University, WIF-1 methylation was found in 34 of 46 (73.9%). In comparison, it was found in 4 of 24 (16.7%) non-cancerous lung tissue samples including the lung lining (pleura), a statistically significant difference. Methylation of any of the four genes was found in 44 of 46 tumors (95.6%) and 12 of 24 controls (50%). SFRP methylation was more frequent in tumors, but the differences were not statistically significant. All 8 mesothelioma cell lines showed WIF-1 methylation. A drug that removes methylation restored WIF-1 in 6 of them. No mutations were found in the examined part of the beta-catenin gene. Asbestos exposure was not studied. The authors concluded that WIF-1 promoter methylation is a common event in mesothelioma.

  We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

• Inhibition of activated phosphatidylinositol 3-kinase/AKT pathway in malignant pleural mesothelioma leads to G1 cell cycle arrest. I. Mikami, F. Zhang, T. Hirata, et al. 2010;24(6):1677-81. PubMed · Publisher

  Abstract

  Phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway plays pivotal roles in fundamental cellular functions including cell proliferation and cell survival. Its deregulation has been implicated in many types of human malignancies. We investigated the role of PI3K/AKT signaling pathway in human malignant pleural mesothelioma (MM). Here, we report that aberrant activation of the PI3K/AKT signaling pathway is associated with cell cycle progression in MM cells. Inhibition of the PI3K activity by its small molecule inhibitor LY294002 led to significant G1 cell cycle arrest and suppression of cell proliferation in all MM cell lines that we examined. In addition, we found that the protein level of p27Kip1 was up-regulated and the protein level of cyclin D1 was down-regulated following LY294002 treatment in those MM cell lines. However, no noticeable apoptosis induction was observed following 24 h of LY294002 treatment in those MM cell lines. These results confirm that the PI3K/AKT signaling pathway is aberrantly active and plays a critical role for the cell cycle progression in human MM cells.

2000s

• Plasma proteomic profiling: search for lung cancer diagnostic and early detection markers. I.U. Ali, Z. Xiao, W. Malone, et al. 2006;15(5):1367-72. PubMed

  Abstract

  Environmental and occupational exposure to asbestos is among the established risk factors for lung cancer, the leading cause of cancer-related deaths in the United States. This link between exposure to asbestos and the excessive death rate from lung cancer was evident in a study of former workers of an asbestos pipe insulation manufacturing plant in Tyler, TX. We performed comparative proteomic profiling of plasma samples that were collected from nine patients within 12 months before death and their age-, race- and exposure-matched disease-free controls on strong anion exchange chips using surface-enhanced laser desorption ionization time-of-flight mass spectrometry. A distance-dependent K-nearest neighbor (KNN) classification algorithm identified spectral features of m/z values 7558.9 and 15103.0 that were able to distinguish lung cancer patients from disease-free individuals with high sensitivity and specificity. The high correlation between the intensities of these two peaks (r=0.987) strongly suggests that they are the doubly and singly charged ions of the same protein product. Examination of these proteomic markers in the plasma samples of subjects from >5 years before death from lung cancer suggested that they are related to the early development of lung cancer. Validation of these biomarkers would have significant implications for the early detection of lung cancer and better management of high-risk patients.

• Nitric oxide production in fluoro-edenite treated mouse monocyte-macrophage cultures. V. Cardile, L. Proietti, A. Panico, et al. 2004;12(6):1209-15. PubMed

  Abstract

  In the present study, we investigated the involvement of NO in the cytotoxic and genotoxic effects caused by fluoro-edenite in mouse monocyte-macrophage cell line J774. Fluoro-edenite is a new asbestos-like amphibole present in the benmoreitic lavas recently extracted from stone quarries in Biancavilla, a village located in the Etnean Volcanic Complex (Catania, Italy) of eastern Sicily, in which an epidemiological survey evidenced a cluster of cases of the mortality due to pleural mesothelioma. Fluoro-edenite appears as a probable carcinogenic agent. Nitrite and nitrate concentration (NO) in the supernatant was quantified by colorimetric assay based on the Griess reaction and iNOS (inducible nitric oxide synthetase) expression was determined by immunostaining in mouse monocyte-macrophage cell line J774 treated with different concentrations of fluoro-edenite (5, 50 and 100 microg/ml) for 24, 48, 72 and 96 h. Parallel experiments were performed treating the cultures also with lipopolysaccharides (LPS), used as inflammation-inducing molecule. In our experimental conditions, fluoro-edenite did not modify the level of NO and the expression of iNOS at the experimental used concentrations for 24, 48 and 72 h. These parameters were significantly modified at the higher doses (50 and 100 microg/ml) of fluoro-edenite for 96 h and were further more increased, in concentration- and time-dependent manner, when cell cultures were treated with fluoro-edenite and LPS. These findings provide convincing evidence that NO is involved in the fluoro-edenite-induced cytotoxicity and geno-toxicity in mouse monocyte-macrophage cell line J774 when the fiber remain for longer times and particularly in cultures treated with LPS, demonstrating that inflammatory disorders appear to increase the risk for lung cancer induced by fluoro-edenite probably by the involvement of NO.

• Concurrent LOH at multiple loci in human malignant mesothelioma with preferential loss of NF2 gene region. L. Pylkkänen, M. Sainio, T. Ollikainen, et al. 2002;9(5):955-9. PubMed

  Abstract

  Human malignant mesothelioma (MM) is a highly aggressive neoplasm related to occupational asbestos exposure and characterised by a long latency period between the exposure and onset of disease. Previous studies indicate that losses at different genomic regions are present in MM. We examined allele loss at three known tumour suppressor gene regions (22q/NF2 gene, 9p/p16 gene, and 3p/FHIT gene) and at two other frequently deleted areas (14q and 6q) in MM. Loss of heterozygosity (LOH) was investigated in cell cultures and primary tumours with several highly polymorphic markers for each site. To study if LOH of the NF2 gene is a consistent feature in MM, we performed a more detailed analysis of chromosome 22q that included a NF2 marker (NF2CA3). We observed a high frequency of LOH occurring simultaneously at multiple loci. In particular, 100% of the cultured MM cells exhibited LOH at the NF2 gene region. From the other chromosomal sites analysed, recurrent allele loss was detected at 9p (5/7; 71%), 3p (4/7; 57%), 14q (3/7; 43%), and 6q (3/7; 43%). Of the 32 tumours, even those trimmed to exclude normal tissue, few showed LOH, suggesting consielment by normal cells within MM tumours, whereas tumour cells in primary cultures showed LOH already in passages 1-2. In conclusion, our present LOH data indicate that MM cells exhibit allele losses at multiple tumour suppressor gene sites concurrently, involving NF2 gene preferentially. This supports the view that the accumulation of multiple genetic hits is characteristic to malignant transformation of MM cells.

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