International Journal of Oncology

About the journal

Publisher: D.A. Spandidos
Published: 1992 to the present, Greece
Language: English
ISSN: 1019-6439 (print), 1791-2423 (electronic)
Its record in the NLM Catalog

2020s

Telomeres and telomerase in mesothelioma: Pathophysiology, biomarkers and emerging therapeutic strategies (Review). D. Andreikos, D.A. Spandidos, V.E. Georgakopoulou. 2025;66(3). Review PubMed · Publisher · Free full text

Abstract

Malignant mesothelioma (MM) is a rare but aggressive cancer linked to asbestos exposure and characterized by advanced‑stage disease at presentation. Despite advances in treatment, prognosis remains abysmal, highlighting the imperative for the development of novel biomarkers and treatment approaches. Telomere biology plays a pivotal role in the tumorigenic process and has emerged as a key area in oncology research. Short telomeres have been associated with genomic instability, and substantially shorter telomere length (TL) has been identified in MM, showcasing the potential of TL in risk assessment, early detection, and disease progression monitoring. MM predominantly maintains TL through telomerase activity (TA), which in research has been identified in >90% of MM cases, underscoring the potential of TA as a biomarker in MM. Telomerase reverse transcriptase (TERT) polymorphisms may serve as valuable biomarkers, with research identifying associations between single nucleotide polymorphisms (SNPs) and the risk and prognosis of MM. Additionally, TERT promoter mutations have been associated with poor prognosis and advanced‑stage disease, with the non‑canonical functions of TERT hypothesized to contribute to the development of MM. TERT promoter mutations occur in ~12% of MM cases; C228T, C250T and A161C are the most common, while the distribution and frequency differ depending on histological subtype. Research reveals the promise of the various approaches therapeutically targeting telomerase, with favorable results in pre‑clinical models and inconclusive findings in clinical trials. The present review examines the role of telomere biology in MM and its implications in diagnosis, prognosis, and therapy.
Ingredients such as trehalose and hesperidin taken as supplements or foods reverse alterations in human T cells, reducing asbestos exposure-induced antitumor immunity. S. Yamamoto, S. Lee, T. Ariyasu, et al. 2021;58(4). PubMed · Publisher · Free full text

Abstract

Exposure of human immune cells to asbestos causes a reduction in antitumor immunity. The present study aimed to investigate the recovery of reduced antitumor immunity by several ingredients taken as supplements or foods, including trehalose (Treh) and glycosylated hesperidin (gHesp). Peripheral blood CD4+ cells were stimulated with IL‑2, anti‑CD3 and anti‑CD28 antibodies for 3 days, followed by further stimulation with IL‑2 for 7 days. Subsequently, cells were stimulated with IL‑2 for an additional 28 days. During the 28 days, cells were cultured in the absence or presence of 50 <em>µ</em>g/ml chrysotile asbestos fibers. In addition, cells were treated with 10 mM Treh or 10 <em>µ</em>M gHesp. Following culture for 28 days, reverse transcription‑quantitative PCR was performed to assess the expression levels of transcription factors, cytokines and specific genes, including matrix metalloproteinase‑7 (MMP‑7), nicotinamide nucleotide transhydrogenase (NNT) and C‑X‑C motif chemokine receptor 3, in unstimulated cells (fresh) and cells stimulated with PMA and ionomycin (stimuli). The results demonstrated that compared with the control group, chrysotile‑exposure induced alterations in MMP‑7, NNT and IL‑17A expression levels were not observed in the 'Treh' and 'gHesp' groups in stimulated cells. The results suggested that Treh and gHesp may reverse asbestos exposure‑induced reduced antitumor immunity in T helper cells. However, further investigation is required to confirm the efficacy of future trials involving the use of these compounds with high‑risk human populations exposed to asbestos, such as workers involved in asbestos‑handling activities.
Effects of selected deubiquitinating enzyme inhibitors on the proliferation and motility of lung cancer and mesothelioma cell lines. T. Mirzapoiazova, A. Pozhitkov, A. Nam, et al. 2020;57(1):80-86. PubMed · Publisher · Free full text

Abstract

The post‑translational modification of proteins by ubiquitinating enzymes plays a central role in a number of cellular functions, such as cell proteolysis, DNA repair, and cell signaling and communication. Deubiquitinating enzymes (DUBs) disassemble ubiquitin chains and remove ubiquitin moieties from proteins. Targeting DUBs in cancer models has revealed an important role for these enzymes in tumorigenesis, and they therefore have emerged as attractive therapeutic targets. In the present study, the effects of three DUB inhibitors, PR‑619, RA‑9 and LDN‑91946, on a non‑small cell lung cancer cell line (A549) and a mesothelioma cell line (H2373) were investigated. PR‑619 significantly inhibited cell adhesion and the proliferation of both cell lines. RA‑9 exerted an inhibitory effect on the adhesion and proliferation of H2373 cells, whereas it had no effect on A549 cells. Notably, however, while PR‑619 attenuated the proliferation of both cell lines, it exerted an opposite effect on cell motility; in the case of A549 cells, there was a significant increase in cell motility, while for the H2373 cells, there was a significant decrease. Furthermore, protein phosphorylation kinetic analyses revealed that the effects were cell line‑specific. In H2373 cells, the phosphorylation of only one peptide corresponding to the P85A protein was significantly affected, and while LDN‑91946 treatment increased phosphorylation, treatment with RA‑9 or PR‑619 decreased its phosphorylation compared to the DMSO control. By contrast, in the case of A549 cells, the phosphorylation of 21 peptides was significantly affected by the same compounds. In light of the potential for the negative side‑effects of DUB inhibition, such as increased cancer cell motility, the data presented herein underscore the dire need for the development of specific DUB inhibitors and to elucidate the individual role of DUB family members in cancer biology before they can be specifically pharmacologically targeted.

2010s

Droplet digital PCR as a novel system for the detection of microRNA‑34b/c methylation in circulating DNA in malignant pleural mesothelioma. H. Sato, J. Soh, K. Aoe, et al. 2019;54(6):2139-2148. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is a rare malignancy arising from the pleura that is difficult to diagnose, contributing to its dismal prognosis. Previously, we reported that the degree of microRNA (miR)‑34b/c methylation in circulating DNA is associated with the development of MPM. Herein, we present a newly developed droplet digital PCR (ddPCR)‑based assay for the detection of miR‑34b/c methylation in circulating DNA in patients with MPM. We originally prepared two probes within a short amplicon of 60 bp, designing one from the positive strand and the other from the complementary strand. The two probes functioned cooperatively, and our established assay detected DNA methylation accurately in the preliminary validation. We subsequently verified this assay using clinical samples. Serum samples from 35 cases of MPM, 29 cases of pleural plaque and 10 healthy volunteers were collected from 3 different institutions and used in this study. We divided the samples into 2 groups (group A, n=33; group B, n=41). A receiver‑operating characteristic curve analysis using the samples in group A determined the optimal cut‑off value for the diagnosis of MPM, with a sensitivity of 76.9% and a specificity of 90%. On the other hand, the use of the same criterion yielded a sensitivity of 59.1% and a specificity of 100% in group B, and corresponding values of 65.7 and 94.9% for the entire cohort, indicating a moderate sensitivity and a high specificity. In addition, when the analysis was focused on stage II or more advanced MPM, the sensitivity improved to 81.8%, suggesting the possibility that the methylated allele frequency in MPM may be associated with the stage of disease progression. On the whole, the findings of this study indicate that miR‑34b/c methylation in circulating DNA is a promising biomarker for the prediction of disease progression in patients with MPM.
Role of evaluating tumor‑infiltrating lymphocytes, programmed death‑1 ligand 1 and mismatch repair proteins expression in malignant mesothelioma. L. Losi, F. Bertolini, G. Guaitoli, et al. 2019;55(5):1157-1164. PubMed · Publisher

Abstract

The tumor immune microenvironment (TME) and immune checkpoints have been reported to serve a role in the pathogenesis of malignant mesothelioma (MM) and treatment outcome. Additionally, mismatch Repair (MMR) deficiency appears to enhance the response to checkpoints blockade in several tumors. The aim of the present study was to analyze programmed death‑1 ligand 1 (PD‑L1) expression in MM and to characterize the TME. This could help to understand the immune response, and evaluate its prognostic and predictive values. We also investigated MMR protein expression. We retrospectively analyzed 55 mesotheliomas to determine PD‑L1, CD4+, CD8+, mutL homolog 1 (MLH1), mutS homolog 2 (MSH2), mutS homolog 6 (MSH6) and PMS1 homolog 2, mismatch repair system component (PMS2) expression. We used an immunoscore (1+, 2+ and 3+) to evaluate tumor‑infiltrating lymphocytes (TILs). TILs were observed in all but two samples (53/55); the majority had an immunoscore 1+ (30/53), while 2+/3+ was reported for 23/53 samples. A predominance of CD8+ was highlighted in 8 cases (15%). PD‑L1 expression of ≥1% on tumor cells was displayed in 40 cases; in 9 of these, ≥50% expression was reported. Of note, alterations in MMR staining was not observed. In addition, survival analysis revealed that epithelioid subtype was associated with better prognosis. We observed a trend towards poorer prognosis for ≥50% PD‑L1 expression on tumor cells, lower immunoscore (1+) and CD8+ TIL predominance. The present study highlighted the importance of exploring the TME and the standardization of PD‑L1 assessment guidelines to apply in the field of immunotherapy.
Curcumin induces apoptosis and inhibits angiogenesis in murine malignant mesothelioma. C. Zhang, Y. Hao, L. Wu, et al. 2018;53(6):2531-2541. PubMed · Publisher · Free full text

Abstract

Malignant pleural mesothelioma (MPM) is a rare form of cancer that is associated with asbestos exposure. Unfortunately, current therapies have limited efficacy. Previous studies have indicated that curcumin exerts antiproliferative and antitumor effects, and has low toxicity. The present study aimed to evaluate the anticancer effects of curcumin on the RN5 MPM cell line. The inhibitory effects of curcumin on cell viability were determined using the sulforhodamine B assay. In addition, cell cycle progression was analyzed by propidium iodide (PI) staining and flow cytometry, and curcumin‑induced apoptosis was measured by Annexin V/PI double staining. The translocation of apoptosis-inducing factor (AIF) was assessed by western blotting and immunofluorescence, and the expression levels of the phosphoinositide 3-kinase (PI3K)-AKT serine/threonine kinase (Akt)‑mammalian target of rapamycin (mTOR) signaling pathway proteins and mitochondria-associated proteins were evaluated by western blotting. In vivo antitumor effects were evaluated in a subcutaneous murine model. Briefly, tumors were harvested from the mice, and immunohistochemistry was conducted to evaluate cell proliferation, apoptosis and angiogenesis. The results indicated that curcumin inhibited RN5 cell viability and induced apoptotic cell death. In addition the findings suggested that curcumin-induced cell apoptosis occurred via the mitochondrial pathway, and caspase‑independent and AIF-dependent pathways. Further analysis revealed that curcumin may act as a PI3K-Akt-mTOR signaling pathway inhibitor by downregulating PI3K, p-Akt, p-mTOR and p-p70 ribosomal protein S6 kinase. Furthermore, curcumin inhibited tumor angiogenesis in vivo. In conclusion, curcumin may be potent enough to be developed as a novel therapeutic agent for the treatment of MPM.
Preclinical investigation of folate receptor-targeted nanoparticles for photodynamic therapy of malignant pleural mesothelioma. T. Kato, C.S. Jin, D. Lee, et al. 2018;53(5):2034-2046. PubMed · Publisher · Free full text

Abstract

Photodynamic therapy (PDT) following lung-sparing extended pleurectomy for malignant pleural mesothelioma (MPM) has been investigated as a potential means to kill residual microscopic cells. High expression levels of folate receptor 1 (FOLR1) have been reported in MPM; therefore, targeting FOLR1 has been considered a novel potential strategy. The present study developed FOLR1‑targeting porphyrin-lipid nanoparticles (folate-porphysomes, FP) for the treatment of PDT. Furthermore, inhibition of activated epidermal growth factor (EGFR)-associated survival pathways enhance PDT efficacy. In the present study, these approaches were combined; FP-based PDT was used together with an EGFR-tyrosine kinase inhibitor (EGFR-TKI). The frequency of FOLR1 and EGFR expression in MPM was analyzed using tissue microarrays. Confocal microscopy and a cell viability assay were performed to confirm the specificity of FOLR1‑targeting cellular uptake and photocytotoxicity in vitro. In vivo fluorescence activation and therapeutic efficacy were subsequently examined. The effects of EGFR-TKI were also assessed in vitro. The in vivo combined antitumor effect of EGFR-TKI and FP-PDT was then evaluated. The results revealed that FOLR1 and EGFR were expressed in 79 and 89% of MPM samples, respectively. In addition, intracellular uptake of FP corresponded well with FOLR1 expression. When MPM cells were incubated with FP and then irradiated at 671 nm, there was significant in vitro cell death, which was inhibited in the presence of free folic acid, thus suggesting the specificity of FPs. FOLR1 targeting resulted in disassembly of the porphysomes and subsequent fluorescence activation in intrathoracic disseminated MPM tumors, as demonstrated by ex vivo tissue imaging. FP-PDT resulted in significant cellular damage and apoptosis in vivo. Furthermore, the combination of pretreatment with EGFR-TKI and FP-PDT induced a marked improvement of treatment responses. In conclusion, FP-based PDT induced selective destruction of MPM cells based on FOLR1 targeting, and pretreatment with EGFR-TKI further enhanced the therapeutic response.
Accelerated cell cycle progression of human regulatory T cell-like cell line caused by continuous exposure to asbestos fibers. S. Lee, H. Matsuzaki, M. Maeda, et al. 2017;50(1):66-74. PubMed · Publisher · Free full text

Abstract

Asbestos exposure causes malignant tumors such as lung cancer and malignant mesothelioma. Based on our hypothesis in which continuous exposure to asbestos of immune cells cause reduction of antitumor immunity, the decrease of natural killer cell killing activity with reduction of NKp46 activating receptor expression, inhibition of cytotoxic T cell clonal expansion, reduced CXCR3 chemokine receptor expression and production of interferon-γ production in CD4+ T cells were reported using cell line models, freshly isolated peripheral blood immune cells from health donors as well as asbestos exposed patients such as pleural plaque and mesothelioma. In addition to these findings, regulatory T cells (Treg) showed enhanced function through cell-cell contact and increased secretion of typical soluble factors, interleukin (IL)-10 and transforming growth factor (TGF)-β, in a cell line model using the MT-2 human polyclonal T cells and its sublines exposed continuously to asbestos fibers. Since these sublines showed a remarkable reduction of FoxO1 transcription factor, which regulates various cell cycle regulators in asbestos-exposed sublines, the cell cycle progression in these sublines was examined and compared with that of the original MT-2 cells. Results showed that cyclin D1 expression was markedly enhanced, and various cyclin-dependent kinase-inhibitors were reduced with increased S phases in the sublines. Furthermore, the increase of cyclin D1 expression was regulated by FoxO1. The overall findings indicate that antitumor immunity in asbestos-exposed individuals may be reduced in Treg through changes in the function and volume of Treg.
DCLK1 is correlated with MET and ERK5 expression, and associated with prognosis in malignant pleural mesothelioma. H. Wang, Y.Y. Dai, W.Q. Zhang, et al. 2017;51(1):91-103. PubMed · Publisher · Free full text

Abstract

Malignant pleural mesothelioma (MPM) is an aggressive cancer for which more effective treatments are needed. In this study, strong to moderate staining of MET and ERK5 was detected in 67.1 and 48% of the analyzed 73 human mesothelioma tumors, and significant correlation of MET and ERK5 expression was identified (P<0.05). We evaluated the doublecortin-like kinase 1 (DCLK1) expression in human mesothelioma tumors. Our results showed that 50.7% of the immunohistochemistry analyzed human mesothelioma tumors have strong to moderate staining of DCLK1, and its expression is significantly correlated with MET or ERK5 expression (P<0.05). Also, the upregulation of DCLK1 is correlated with poor prognosis in MPM patients (P=0.0235). To investigate whether DCLK1 is downstream of MET/ERK5 signaling in human mesothelioma, the effect of DCLK1 expression was analyzed after treatments with either the MET inhibitor XL184 or the ERK5 inhibitor XMD8-92 in human mesothelioma cell lines. Our results showed that the MET inhibitor XL184 reduced the expression of phospho‑ERK5 and DCLK1 expression in human mesothelioma cell lines. In addition, the ERK5 inhibitor XMD8-92 reduced the expression of phospho-ERK5 and DCLK1 expression in human mesothelioma cell lines. Furthermore, XML184 and XMD8-92 treatment impaired invasion and tumor sphere formation ability of H290 mesothelioma cells. These results suggest that DCLK1 is regulated by MET/ERK5 signaling in human mesothelioma, and the MET/ERK5/DCLK1 signaling cascade could be further developed into a promising therapeutic target against mesothelioma.
Induction of IL-17 production from human peripheral blood CD4+ cells by asbestos exposure. M. Maeda, Y. Chen, S. Lee, et al. 2017;50(6):2024-2032. PubMed · Publisher

Abstract

We have previously reported that chronic, recurrent and low-dose exposure to asbestos fibers causes a reduction in antitumor immunity. Investigation of natural killer (NK) cells using an in vitro cell line model and comprising in vitro activation using freshly isolated NK cells co-cultured with chrysotile fibers, as well as NK cells derived from asbestos-exposed patients with pleural plaque (PP) or malignant mesothelioma (MM), revealed decreased expression of NK cell activating receptors such as NKG2D, 2B4 and NKp46. An in vitro differentiation and clonal expansion model for CD8+ cytotoxic T lymphocytes (CTLs) showed reduced cytotoxicity with decreased levels of cytotoxic molecules such as granzyme B and perforin, as well as suppressed proliferation of CTLs. Additionally, analysis of T helper cells showed that surface CXCR3, chemokine receptor, and the productive potential of interferon (IFN)γ were reduced following asbestos exposure in an in vitro cell line model and in peripheral CD4+ cells of asbestos-exposed patients. Moreover, experiments revealed that asbestos exposure enhanced regulatory T cell (Treg) function. This study also focused on CXCR3 expression and the Th-17 cell fraction. Following activation with T-cell receptor and co-culture with various concentrations of chrysotile fibers using freshly isolated CD4+ surface CXCR3 positive and negative fractions, the intracellular expression of CXCR3, IFNγ and IL-17 remained unchanged when co-cultured with chrysotile. However, subsequent re-stimulation with phorbol 12-myristate 13-acetate (PMA) and ionomycin resulted in enhanced IL-17 production and expression, particularly in CD4+ surface CXCR3 positive cells. These results indicated that the balance and polarization between Treg and Th-17 fractions play an important role with respect to the immunological effects of asbestos and the associated reduction in antitumor immunity.
Kdm6a and Kdm6b: Altered expression in malignant pleural mesothelioma. S. Cregan, M. Breslin, G. Roche, et al. 2017;50(3):1044-1052. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is a rare aggressive cancer of the pleura primarily associated with prior exposure to asbestos. The current standard of care for patients suffering from MPM is a combination of cisplatin and pemetrexed (or alternatively cisplatin and raltitrexed). Most patients, however, die within 24 months of diagnosis. New therapies are therefore urgently required for this disease. Inflammation is thought to be a key element in the pathogenesis of MPM, and recently Kdm6 family members (Kdm6a and Kdm6b) have been identified as playing important roles in inflammatory processes. As such these genes could potentially represent novel candidate targets for intervention in MPM. Using RT-PCR we examined the expression of Kdm6aA and Kdm6b in a panel of MPM cell lines and in a cohort of snap-frozen patient samples isolated at surgery comprising benign, epithelial, biphasic and sarcomatoid histologies. Both Kdm6a and Kdm6b were found to be significantly overexpressed in MPM at the mRNA level. However, tests examining if targeting therapeutically Kdm6a/b using a specific small molecule inhibitor (GSK-J4) was potentially useful for treating MPM, revealed that anti-proliferative activity was higher at lower drug concentrations in cell lines derived from normal mesothelial cells compared to those derived from malignant cells. Treatments with GSK-J4 were found to be associated with the induction of apoptosis and increased expression of pro-inflammatory cytokines. As such our results demonstrate that whilst members of the Kdm6 family are overexpressed in MPM they may not be suitable candidates for therapy and may elicit a cytokine storm.
PIM1 knockdown inhibits cell proliferation and invasion of mesothelioma cells. A.S. Mawas, V.J. Amatya, R. Suzuki, et al. 2017;50(3):1029-1034. PubMed · Publisher

Abstract

Malignant mesothelioma is a major asbestos-related cancer with prolonged time lapse from the first exposure of asbestos to the development of mesothelioma. Most of mesothelioma patients show very poor prognosis, thus, an urgent improvement of its treatment is required by development of novel therapeutic strategies. RNA interference (RNAi) is a powerful tool in post-genomic research and cancer therapy through inhibition of gene expression. In the present study, we analyzed the function of PIM1 on mesothelioma cell lines with its knockdown by siRNA transfection. Here, we report that the downregulation of PIM1 led to suppression of cell proliferation by cell cycle arrest at G1 phase and suppression of cell invasion and migration. Considering the mesothelioma as rapidly growing invasive cancer, downregulation of PIM1 may have a potential role for therapeutic management of malignant mesothelioma.
The Wnt regulator SFRP4 inhibits mesothelioma cell proliferation, migration, and antagonizes Wnt3a via its netrin-like domain. V. Perumal, A.M. Dharmarajan, S.A. Fox. 2017;51(1):362-368. PubMed · Publisher

Abstract

Secreted frizzled related proteins (SFRPs) are a family of Wnt regulators which are frequently downregulated in cancers. In malignant mesothelioma (MM), downregulation of SFRP4 has been reported as a mechanism which contributes to aberrant activation of oncogenic Wnt signaling. Here we investigated the biological consequences of SFRP4 in two mesothelioma cell models where this protein is downregulated. We used recombinant SFRP4 and transient overexpression to study changes in proliferation, migration and downstream signaling. We found that recombinant SFRP4 inhibited both proliferation and migration of MM cells as well as abrogating the stimulatory effect of recombinant Wnt3a. Morphologically SFRP4 induced a cytotoxic effect distinct from apoptosis and consistent with mitotic catastrophe. Overexpression of SFRP4 in these cell lines displayed similar effects as endogenous protein on cell viability, migration and nuclear morphology. We also used expression constructs to examine the role of the SFRP4 cysteine rich domain (CRD) and a netrin-like domain (NLD) in these effects. Interestingly, we found it was the NLD which mediated the biological effects of SFRP4 in these cells. Our results indicate that SFRP4 inhibits mesothelioma proliferation, migration and activates alternative cell death pathways. The finding that the NLD is responsible for these has broader implications for this protein family. Overall this study suggests that the Wnt pathway may prove a promising target for therapy in mesothelioma.
Differential microRNA expression profiling of mesothelioma and expression analysis of miR-1 and miR-214 in mesothelioma. V.J. Amatya, A.S. Mawas, K. Kushitani, et al. 2016;48(4):1599-607. PubMed · Publisher

Abstract

Malignant mesothelioma is a highly aggressive cancer with poor prognosis and refractory to currently available therapies. Most of the patients with advanced invasive nature are not amenable to surgical resection and/or available anticancer therapy, thus development of novel effective therapeutic regimes is needed. Aberrant expression of microRNAs (miRNAs) has been proposed to contribute to carcinogenesis and aggressiveness of mesothelioma. We analyzed miRNA expression in mesothelioma cell lines using TaqMan miRNA expression array and found significant number of miRNAs, which showed increased or lost expression. We validated the increased expression of miR-182, and miR-183 in mesothelioma cell lines by individual miRNA assays and SmartFlare miRNA probes. We further investigated the miR-1, and miR-214, which were not expressed in mesothelioma cells by real-time RT-PCR. Transfection of mesothelioma cells, ACC-Meso-1 and CRL5915, with miRNA mimic (hsa-miR-1 mimic and hsa-miR-214 mimic) led to inhibition of cell growth, invasion and migration. We paid attention to PIM1, the target gene of both miR-1 and miR-214 miRNAs and which was found overexpressed in mesothelioma cells, and miR-1 and miR-214 mimic transfection of mesothelioma cell lines showed downregulation of PIM1 by western blot analysis. The miRNAs, miR-1 and miR-214, may play a role in carcinogenesis of mesothelioma thus might be considered as candidate therapeutic targets in mesothelioma.
Downregulation of thymidylate synthase by RNAi molecules enhances the antitumor effect of pemetrexed in an orthotopic malignant mesothelioma xenograft mouse model. A.S. Abu Lila, C. Kato, M. Fukushima, et al. 2016;48(4):1399-407. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is an incurable cancer with an increasing incidence. Currently, pemetrexed (PMX)-based chemotherapy is the mainstay of chemotherapy for MPM, however, the outcome of PMX-based chemotherapy in patients with MPM is dismal. RNA interference (RNAi) technology has been considered as an effective tool to substantially enhance the therapeutic efficacy of chemotherapeutic agents in many preclinical and clinical settings. In this study, therefore, we investigated whether non-viral anti-thymidylate synthase RNAi embedded liposome (TS shRNA lipoplex) would effectively guide the downregulation of TS in human malignant mesothelioma MSTO-211H cells. Consequently, it enhanced the antitumor effect of PMX both in vitro and in vivo. TS shRNA effectively enhanced the in vitro cell growth inhibition upon treatment with PMX via downregulating TS expression in the MSTO-211H cell line. In in vivo orthotopic tumor model, the combined treatment of PMX and TS shRNA lipoplex efficiently combated the progression of orthotopic thoracic tumors and as a result prolonged mouse survival, compared to each single treatment. Our findings emphasize the pivotal relevance of RNAi as an effective tool for increasing the therapeutic efficacy of PMX, a cornerstone in the treatment regimens of MPM, and thereby, raising the possibility for the development of a novel therapeutic strategy, combination therapy of TS-shRNA and PMX, that can surpass many of the currently applied, but less effective, therapeutic regimens against lethal MPM.
Genomic profiling of the genes on chromosome 3p in sporadic clear cell renal cell carcinoma. Y. Togo, Y. Yoshikawa, T. Suzuki, et al. 2016;48(4):1571-80. PubMed · Publisher

Abstract

Somatic mutations of the BRCA1 associated protein-1 (BAP1) gene, which maps to 3p21, have been found in several tumors including malignant mesothelioma, uveal melanoma, and renal cell carcinoma (RCC). The role of BAP1 inactivation in tumor development remains unclear. It has been reported that Vhl knock-out mice did not develop RCC, but Vhl knock-out mice with single allele loss of Bap1 in nephron progenitor cells developed RCC, indicating that Bap1 inactivation may be essential in murine renal tumorigenesis. To clarify the role of BAP1 in human RCC development, we performed mutation analyses, including copy number detection of BAP1 and assessment of allelic imbalance using microsatellite polymorphisms on 3p, in 45 RCC samples derived from 45 patients without VHL or BAP1 germline mutation. Additionally, we analyzed the sequences of the VHL, PBRM1, and SETD2 genes, and examined promoter methylation of VHL. Using immunostaining, we also checked for expression of BAP1 protein, which is normally located in the nuclei. None of the RCCs had biallelic deletion of BAP1, but five (11.1%) showed a biallelic mutation (four with a sequence-level mutation with monoallelic loss and one with a biallelic sequence-level mutation); these cells were negative for nuclear BAP1 staining. These patients had worse recurrence-free survival than the patients without a biallelic mutation (p=0.046). However, there were no significant differences in worse outcome by multivariate analysis combined with age, T stage, histological subtype, infiltration and vascular invasion. In 35 RCCs (77.8%), monoallelic loss of BAP1 was accompanied by VHL biallelic mutation or VHL promoter hypermethylation. In five RCCs (11.1%), we detected 3p loss-of-heterozygosity, but the copy number of BAP1 was normal. Surprisingly, nuclear staining of BAP1 was negative in 10 out of 31 tumors (32.3%) with hemizygous normal BAP1, suggesting that haploinsufficiency may relate to RCC development.
KAT5 (Tip60) is a potential therapeutic target in malignant pleural mesothelioma. S. Cregan, L. McDonagh, Y. Gao, et al. 2016;48(3):1290-6. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is a rare aggressive cancer of the pleura. Asbestos exposure (through inhalation) is the most well established risk factor for mesothelioma. The current standard of care for patients suffering from MPM is a combination of cisplatin and pemetrexed (or alternatively cisplatin and raltitrexed). Most patients, however, die within 24 months of diagnosis. New therapies are therefore urgently required for this disease. Lysine acetyltransferases (KATs) including KAT5 have been linked with the development of cisplatin resistance. This gene may therefore be altered in MPM and could represent a novel candidate target for intervention. Using RT-PCR screening the expression of all known KAT5 variants was found to be markedly increased in malignant tumors compared to benign pleura. When separated according to histological subtype, KAT5 was significantly overexpressed in both the sarcomatoid and biphasic subgroups for all transcript variants. A panel of MPM cell lines including the normal pleural cells LP9 and Met5A was screened for expression of KAT5 variants. Treatment of cells with a small molecule inhibitor of KAT5 (MG-149) caused significant inhibition of cellular proliferation (p<0.0001), induction of apoptosis and was accompanied by significant induction of pro-inflammatory cytokines/chemokines.
Kinesin family members KIF11 and KIF23 as potential therapeutic targets in malignant pleural mesothelioma. T. Kato, D. Lee, L. Wu, et al. 2016;49(2):448-56. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is a rare and aggressive form of cancer commonly associated with asbestos exposure that stems from the thoracic mesothelium with high mortality rate. Currently, treatment options for MPM are limited, and new molecular targets for treatments are urgently needed. Using quantitative reverse transcription-polymerase chain reaction (RT-PCR) and an RNA interference-based screening, we screened two kinesin family members as potential therapeutic targets for MPM. Following in vitro investigation of the target silencing effects on MPM cells, a total of 53 MPMs were analyzed immunohistochemically with tissue microarray. KIF11 and KIF23 transcripts were found to be overexpressed in the majority of clinical MPM samples as well as human MPM cell lines as determined by quantitative RT-PCR. Gene knockdown in MPM cell lines identified growth inhibition following knockdown of KIF11 and KIF23. High expression of KIF11 (KIF11-H) and KIF23 (KIF23-H) were found in 43.4 and 50.9% of all the MPM cases, respectively. Patients who received curative resection with tumors displaying KIF23-H showed shorter overall survival (P=0.0194). These results provide that inhibition of KIF11 and KIF23 may hold promise for treatment of MPMs, raising the possibility that kinesin-based drug targets may be developed in the future.
SORORIN and PLK1 as potential therapeutic targets in malignant pleural mesothelioma. T. Kato, D. Lee, L. Wu, et al. 2016;49(6):2411-2420. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is an aggressive type of cancer of the thoracic cavity commonly associated with asbestos exposure and a high mortality rate. There is a need for new molecular targets for the development of more effective therapies for MPM. Using quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) and an RNA interference-based screening, we examined the SORORIN gene as potential therapeutic targets for MPM in addition to the PLK1 gene, which is known for kinase of SORORIN. Following in vitro investigation of the effects of target silencing on MPM cells, cell cycle analyses were performed. SORORIN expression was analyzed immunohistochemically using a total of 53 MPM samples on tissue microarray. SORORIN was found to be overexpressed in the majority of clinical MPM samples and human MPM cell lines as determined by qRT-PCR. Gene suppression of each SORORIN and PLK1 led to growth inhibition in MPM cell lines. Knockdown of SORORIN showed an increased number of G2M-phase population and a larger nuclear size, suggesting mitotic arrest. High expression of SORORIN (SORORIN-H) was found in 50.9% of all the MPM cases, and there is a tendency towards poorer prognosis for the SORORIN-H group but the difference is not significant. Suppression of SORORIN with PLK1 inhibitor BI 6727 showed a combinational growth suppressive effect on MPM cell growth. Given high-dose PLK1 inhibitor induced drug-related adverse effects in several clinical trials, our results suggest inhibition SORORIN-PLK1 axis may hold promise for the treatment of MPMs.
The levels of HDAC1 and thioredoxin1 are related to the death of mesothelioma cells by suberoylanilide hydroxamic acid. B.R. You, W.H. Park. 2016;48(5):2197-204. PubMed · Publisher

Abstract

Mesothelioma is an aggressive tumor which is mainly derived from the pleura of lung. In the present study, we evaluated the anticancer effect of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor on human mesothelioma cells in relation to the levels of HDAC1, reactive oxygen species (ROS) and thioredoxin (Trx). While 1 µM SAHA inhibited cell growth in Phi and ROB cells at 24 h, it did not affect the growth in ADA and Mill cells. Notably, the level of HDAC1 was relatively overexpressed among Phi, REN and ROB cells. SAHA induced necrosis and apoptosis, which was accompanied by the cleavages of PARP and caspase-3 in Phi cells. This agent also increased the loss of mitochondrial membrane potential (MMP, ΔΨm) in Phi cells. All the tested caspase inhibitors attenuated apoptosis in SAHA-treated Phi cells whereas HDAC1 siRNA enhanced the apoptotic cell death. SAHA increased intracellular ROS levels including O2•- in Phi cells. N-acetyl cysteine (NAC) and vitamin C (Vit.C) significantly reduced the growth inhibition and death of Phi cells caused by SAHA. This drug decreased the mRNA and protein levels of Trx1 in Phi and ROB cells. Furthermore, Trx1 siRNA increased cell death and O2•- level in SAHA-treated Phi cells. In conclusion, SAHA selectively inhibited the growth of Phi and ROB mesothelioma cells, which showed the higher basal level of HDAC1. SAHA-induced Phi cell death was related to oxidative stress and Trx1 levels.
Combination effect of photodynamic therapy using NPe6 with pemetrexed for human malignant pleural mesothelioma cells. S. Maehara, J. Usuda, T. Ishizumi, et al. 2015;46(2):741-9. PubMed · Publisher

Abstract

To identify a possible new treatment modality for malignant pleural mesothelioma (MPM), we examined whether combination treatment consisting of pemetrexed chemotherapy and photodynamic therapy (PDT) using the photosensitizer NPe6, enhanced the antitumor effect in both in vitro and in vivo models. We also investigated preclinical treatment schedules. Four human malignant mesothelioma cell lines (MSTO‑211H, H2052, H2452 and H28) were assayed using the WST assay after treatment with pemetrexed and NPe6‑PDT. The treatment schedule for the combination treatment was examined using nude mice. Pemetrexed pre‑treatment enhanced the lethal effect of NPe6‑PDT in the four malignant mesothelioma cell lines, but NPe6‑PDT followed by pemetrexed treatment did not enhance cell lethality in the in vitro assay. Pemetrexed pre‑treatment did not enhance the intracellular accumulation of NPe6, which is one of the determinants of the antitumor effect of PDT. In nude mice injected with MSTO‑211H cells and then treated using a combination of pemetrexed and NPe6‑PDT (10 mg/kg NPe6, 10 J/cm(2) laser irradiation), the tumor volume decreased by 50% but subsequently increased, reaching the pre‑treatment value after 14 days. Pemetrexed treatment followed by NPe6‑PDT resulted in an 80% reduction in the tumor size and inhibited re‑growth. NPe6‑PDT followed by pemetrexed treatment resulted in a 60% reduction in tumor size but did not inhibit re‑growth. NPe6‑PDT induced the expression of thymidylate synthase (TS), which confers resistance to pemetrexed, and NPe6‑PDT followed by pemetrexed treatment did not enhance the treatment outcome in vivo. In conclusion, combination treatment, consisting of pemetrexed followed by NPe6‑PDT, should be further investigated as a new treatment modality for MPM. In the future, this combination treatment may contribute to a reduction in local recurrence and a prolonged survival period in patients with MPM.
Downregulation of thymidylate synthase and E2F1 by arsenic trioxide in mesothelioma. S.K. Lam, Y.Y. Li, C.Y. Zheng, et al. 2015;46(1):113-22. PubMed · Publisher

Abstract

Malignant pleural mesothelioma is a global health issue. Arsenic trioxide (ATO) has been shown to suppress thymidylate synthase (TYMS) in lung adenocarcinoma and colorectal cancer, and induce apoptosis in acute promyelocytic leukemia. With TYMS as a putative therapeutic target, the effect of ATO in mesothelioma was therefore studied. A panel of 5 mesothelioma cell lines was used to study the effect of ATO on cell viability, protein expression, mRNA expression and TYMS activity by MTT assay, western blot, qPCR and tritium-release assay, respectively. The knockdown of TYMS and E2F1 was performed with a specific siRNA. Phosphatidylserine externalization and mitochondrial membrane depolarization were measured by Annexin V and JC-1 staining respectively. The in vivo effect of ATO was studied using a nude mouse xenograft model. Application of ATO demonstrated anticancer effects in the cell line model with clinically achievable concentrations. Downregulation of TYMS protein (except H226 cells and 1.25 µM ATO in H2052 cells) and mRNA expression (H28 cells), pRB1 (H28 cells) and E2F1 and TYMS activity (except H226 cells) were also evident. E2F1 knockdown decreased cell viability more significantly than TYMS knockdown. In general, thymidine kinase 1, ribonucleotide reductase M1, c-myc and skp2 were downregulated by ATO. p-c-Jun was downregulated in H28 cells while upregulated in 211H cells. Phosphatidylserine externalization, mitochondrial membrane depolarization, downregulation of Bcl-2 and Bcl-xL, and upregulation of Bak and cleaved caspase-3 were observed. In the H226 xenograft model, the relative tumor growth was aborted, and E2F1 was downregulated while cleaved caspase-3 was elevated and localized to the nucleus in the ATO treatment group. ATO has potent antiproliferative and cytotoxic effects in mesothelioma in vitro and in vivo, partially mediated through E2F1 targeting (less effect through TYMS targeting). There is sound scientific evidence to support the clinical application of ATO in treatment of mesothelioma.
Licochalcone A induces apoptosis in malignant pleural mesothelioma through downregulation of Sp1 and subsequent activation of mitochondria-related apoptotic pathway. K.H. Kim, G. Yoon, J.J. Cho, et al. 2015;46(3):1385-92. PubMed · Publisher

Abstract

Licochalcone A (LCA) is a natural product derived from the roots of Glycyrrhiza inflata exhibiting a wide range of bioactivities such as antitumor, anti-oxidant and anti-bacterial effects. Malignant pleural mesothelioma (MPM) is an extremely aggressive type of cancer with a poor prognosis because of its rapid progression. However, LCA has not been investigated concerning its effects on MPM. Preliminarily, we observed that LCA negatively modulated not only cell growth, but also specificity protein 1 (Sp1) expression in MSTO-211H and H28 cell lines. It was found that IC50 values of LCA for growth inhibition of MSTO-211H and H28 cells were approximately 26 and 30 µM, respectively. Consistent with downregulation of Sp1, expression of Sp1 regulatory proteins such as Cyclin D1, Mcl-1 and Survivin was substantially diminished. Mechanistically, LCA triggered the mitochondrial apoptotic pathway by affecting the ratio of mitochondrial proapoptotic Bax to anti-apoptotic Bcl-xL. Bid induced loss of mitochondrial membrane potential, eventually leading to multi-caspase activation and increased sub-G1 population. Moreover, nuclear staining with DAPI highlighted nuclear condensation and fragmentation of apoptotic features. Flow cytometry analyses after staining cells with Annexin V and propiodium iodide corroborated LCA-mediated apoptotic cell death of MPM cells. In conclusion, these results present that LCA may be a potential bioactive material to control human MPM cells by apoptosis via the downregulation of Sp1.
AHNAK is highly expressed and plays a key role in cell migration and invasion in mesothelioma. H. Sudo, A.B. Tsuji, A. Sugyo, et al. 2014;44(2):530-8. PubMed · Publisher

Abstract

The worldwide incidence of the highly aggressive tumor mesothelioma is expected to increase. Mesothelioma is classified into three main histological subtypes: epithelioid, sarcomatoid and biphasic. Although the pathological diagnostic markers for epithelioid are established, to date no adequate marker for sarcomatoid mesothelioma has been found. Thus, a reliable diagnostic marker of sarcomatoid mesothelioma is necessary. In this study, to identify an unknown protein with 120 kDa expressed only in the mesothelioma cell line 211H, we conducted proteomic analysis and found five candidate proteins. One such protein, AHNAK, was highly expressed in all seven mesothelioma cell lines (211H, H28, H226, H2052, H2452, MESO1 and MESO4), but not in the mesothelial cell line MeT-5A by RT-PCR and immunofluorescence staining. Furthermore, we confirmed high AHNAK expression not only in xenografts but also in human mesothelioma specimens including sarcomatoid, epithelioid and biphasic mesothelioma using immunohistochemical staining. These findings suggest that AHNAK has the potential to be a new marker for detecting mesothelioma. Since AHNAK is involved in cell migration and invasion in other metastatic tumor cells, we conducted migration and invasion assays in mesothelioma cell lines. The number of migrating cells in six of seven mesothelioma cell lines and the number of invading cells in all seven cell lines were significantly increased compared with those in MeT-5A. Knockdown of AHNAK significantly reduced the cell migration and invasion ability in all seven mesothelioma cell lines. These results support further clinical evaluation of the association of AHNAK and metastasis in mesothelioma.
Chronic exposure to asbestos enhances TGF-β1 production in the human adult T cell leukemia virus-immortalized T cell line MT-2. M. Maeda, Y. Chen, H. Hayashi, et al. 2014;45(6):2522-32. PubMed · Publisher

Abstract

Asbestos exposure causes various tumors such as lung cancer and malignant mesothelioma. To elucidate the immunological alteration in asbestos-related tumors, an asbestos-induced apoptosis-resistant subline (MT-2Rst) was established from a human adult T cell leukemia virus-immortalized T cell line (MT-2Org) by long-term exposure to asbestos chrysotile-B (CB). In this study, transforming growth factor-β1 (TGF-β1) knockdown using lentiviral vector-mediated RNA interference showed that MT-2Rst cells secreted increased levels of TGF-β1, and acquired resistance to TGF-β1-mediated growth inhibition. We showed that exposure of MT-2Org cells to CB activated the mitogen-activated protein kinases (MAPKs), ERK1/2, p38 and JNK1. Furthermore, TGF-β1-knockdown cells and treatment with MAPK inhibitors revealed that MT-2Rst cells secreted a high level of TGF-β1 mainly through phosphorylation of p38. However, an Annexin V assay indicated that TGF-β1 resistance in MT-2Rst cells was not directly involved in the acquisition of resistance to apoptosis that is triggered by CB exposure. The overall results demonstrate that long-term exposure of MT-2Org cells to CB induces a regulatory T cell-like phenotype, suggesting that chronic exposure to asbestos leads to a state of immune suppression.
Enhanced apoptosis by pemetrexed and simvastatin in malignant mesothelioma and lung cancer cells by reactive oxygen species-dependent mitochondrial dysfunction and Bim induction. K.E. Hwang, Y.S. Kim, Y.R. Hwang, et al. 2014;45(4):1769-77. PubMed · Publisher

Abstract

Pemetrexed is a multitarget antifolate currently used for the treatment of malignant mesothelioma and non-small cell lung cancer (NSCLC). Statins, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors used primarily for hyperlidpidemia, have been studied for their antiproliferative and pro-apoptotic effects. However, the effects of simvastatin on pemetrexed-induced apoptosis have not been investigated. In this study, we investigated whether combination treatment with pemetrexed and simvastatin potentiates the apoptotic activity above that is seen with either drug alone in malignant mesothelioma and NSCLC cells. We found that the combination of pemetrexed and simvastatin induced more extensive caspase-dependent apoptosis than either drug alone in malignant mesothelioma cells (MSTO-211) or NSCLC cells (A549). In addition, reactive oxygen species (ROS) generation in cells treated with both pemetrexed and simvastatin was markedly increased compared to cells treated with either pemetrexed or simvastatin alone. Combination treatment also increased the loss of mitochondrial membrane potential, increased cytosolic release of cytochrome c, and altered expression of inhibitor of apoptosis proteins (IAP) and B-cell lymphoma-2 (Bcl-2) families of apoptosis related proteins. On the other hand, pretreatment with N-acetylcysteine (NAC) prevented apoptosis and mitochondrial dysfunction by pemetrexed and simvastatin. In addition, Bim siRNA conferred protection against apoptosis induced by pemetrexed and simvastatin. These results suggest that combination of pemetrexed and simvastatin potentiates their apoptotic activity beyond that of either drug alone in malignant mesothelioma and lung cancer cells. This activity is mediated through ROS-dependent mitochondrial dysfunction and Bim induction.
Sialic acid-binding lectin (leczyme) induces apoptosis to malignant mesothelioma and exerts synergistic antitumor effects with TRAIL. T. Tatsuta, M. Hosono, K. Takahashi, et al. 2014;44(2):377-84. PubMed · Publisher · Free full text

Abstract

Malignant mesothelioma is a highly aggressive tumor with poor prognosis. An effective drug for treatment of malignant mesothelioma is greatly needed. Sialic acid-binding lectin (SBL) isolated from oocytes of Rana catesbeiana is a multifunctional protein which has lectin activity, ribonuclease activity and antitumor activity, so it could be developed as a new type of anticancer drug. The validity of SBL for treatment of malignant mesothelioma was assessed using three malignant mesotheliomas and a non-malignant mesothlial cell line. Effectiveness of combinatorial treatment of SBL and tumor necrosis factor-related apoptosis inducing ligand (TRAIL) was also elucidated and characterized. SBL induced tumor-selective cytotoxicity that was attributed to induction of apoptosis. Combinatorial treatment of SBL and TRAIL showed synergistic apoptosis-inducing effect. Additional experiments revealed that Bid was the mediating molecule for the synergistic effect in SBL and TRAIL. These results suggested that SBL could be a promising candidate for the therapeutics for malignant mesothelioma. Furthermore, the combinatorial treatment of SBL and TRAIL could be an effective regimen against malignant mesothelioma.
Significance of osteopontin in the sensitivity of malignant pleural mesothelioma to pemetrexed. S. Takeuchi, M. Seike, R. Noro, et al. 2014;44(6):1886-94. PubMed · Publisher

Abstract

Pemetrexed (PEM) is currently recommended as one of the standard anticancer drugs for malignant pleural mesothelioma (MPM). However, the mechanism of the sensitivity of MPM to PEM remains unclear. We analyzed the antitumor effects of PEM in six MPM cell lines by MTS assay. To identify genes associated with drug sensitivity, we conducted gene expression profiling on the same set of cell lines using GeneChips and pathway analysis. Three cell lines were sensitive to PEM. A total fo 18 transcripts and 14 genes identified by GeneChips were significantly correlated with sensitivity to PEM. Pathway analysis revealed that osteopontin (SPP1/OPN) was an important target in PEM sensitivity. Overexpression of SPP1/OPN was observed in the sensitive cells by quantitative PCR and western blot analysis. Introduction of SPP1/OPN by lentiviral vector significantly enhanced the invasion activities of MPM cells. PEM treatment with SPP1/OPN knockdown inhibited the PEM-induced cell growth-inhibitory effect in PEM-sensitive cells. Expression of SPP1/OPN and AKT phosphorylation significantly decreased after PEM treatment of the PEM-sensitive cells. High immunohistochemical expression of SPP1/OPN was observed in two of three MPM patients who had a partial response to PEM-based chemotherapy. PEM has antitumor effects in MPM cells dependent on SPP1/OPN overexpression resulting in AKT activation. Our results suggest that SPP1 may be used as a single predictive biomarker of the effectiveness of PEM treatment in MPM.
The small GTPase RhoA regulates the expression and function of the sodium channel Nav1.5 in breast cancer cells. C. Dulong, Y.J. Fang, C. Gest, et al. 2014;44(2):539-47. PubMed · Publisher

Abstract

Voltage-gated Na+ channels (VGSCs) are highly expressed in several types of carcinomas including breast, prostate and lung cancers as well as in mesothelioma and cervical cancers. Although the VGSCs activity is considered crucial for the potentiation of cancer cell migration and invasion, the mechanisms responsible for their functional expression and regulation in cancer cells remain unclear. In the present study, the role of the small GTPase RhoA in the regulation of expression and function of the Nav1.5 channel in the breast cancer cell lines MDA-MB 231 and MCF-7 was investigated. RhoA silencing significantly reduced both Nav1.5 channel expression and sodium current indicating that RhoA exerts a stimulatory effect on the synthesis of an active form of Nav1.5 channel in cancer cells. The inhibition of Nav1.5 expression dramatically reduced both cell invasion and proliferation. In addition, a decrease of RhoA protein levels induced by Nav1.5 silencing was observed. Altogether, these findings revealed: i) the key role of the small GTPase RhoA in upregulation of Nav1.5 channel expression and tumor aggressiveness, and ii) the existence of a positive feedback of Nav1.5 channels on RhoA protein levels.
Mechanisms of cisplatin-induced cell death in malignant mesothelioma cells: role of inhibitor of apoptosis proteins (IAPs) and caspases. I.L. Cregan, A.M. Dharmarajan, S.A. Fox. 2013;42(2):444-52. PubMed · Publisher · Free full text

Abstract

Malignant mesothelioma (MM) is an aggressive and highly chemoresistant tumour. Although cisplatin is used in frontline therapy of this disease treatment remains palliative at best. The biochemical pathways activated by cisplatin and the mechanisms of resistance in mesothelioma cells are poorly understood. Overexpression of inhibitor of apoptosis proteins (IAPs) has been described in clinical mesothelioma tumours and proposed as therapeutic targets. In this study, we examined cisplatin-induced cell death pathways and IAPs in three mesothelioma-derived cell lines. Cisplatin induced cell death in mesothelioma cell lines was characterised by biochemical mechanisms classically associated with apoptosis including: mitochondrial depolarisation, phosphatidylserine translocation and caspase activation. Surprisingly mRNA expression of IAPs in mesothelioma was not upregulated relative to primary mesothelial cells except for survivin which was higher in the most resistant cell line. In contrast, protein expression of both XIAP and survivin was upregulated in all mesothelioma cells, consistent with post-translational regulation. Knockdown of either XIAP or survivin by RNAi did not affect the sensitivity to cisplatin in any of the cell lines. Survivin RNAi did, however, inhibit proliferation in the highest expressing cell line, ONE58. The pan-caspase inhibitor z-VAD and the more selective caspase 3/7 inhibitor z-DEVD had no effect upon the sensitivity of any of the cell lines to cisplatin indicating that caspase-independent pathways predominate. The findings of the present study provide insights into cisplatin-induced mechanisms in mesothelioma cells and show that alternative pathways are operating which may provide new options for targeting this extremely resistant tumour.
Modulation of u-PA, MMPs and their inhibitors by a novel nutrient mixture in human lung cancer and mesothelioma cell lines. M.W. Roomi, T. Kalinovsky, A. Niedzwiecki, et al. 2013;42(6):1883-9. PubMed · Publisher · Free full text

Abstract

Lung cancer, the most prevalent cancer worldwide and malignant mesothelioma are highly aggressive tumors that are characterized by high levels of matrix metalloproteinase (MMP)-2 and -9 secretion. Proteases play a key role in tumor cell invasion and metastasis by digesting the basement membrane and ECM components. Strong clinical and experimental evidence demonstrates association of elevated levels of u-PA and MMPs with cancer progression, metastasis and shortened patient survival. MMP activities are regulated by specific tissue inhibitors of metalloproteinases (TIMPS). Our main objective was to study the effect of a nutrient mixture (NM) on the activity of u-PA, MMPs and TIMPs on human lung and malignant mesothelioma (MM) cell lines. Human lung cancer (A-549 and Calu-3) and malignant mesothelioma (MSTO-211H) cell lines were cultured in their respective media and treated at confluence with NM at 0, 50, 100, 250, 500 and 1000 µg/ml. Analysis of u-PA activity was carried out by fibrin zymography, MMPs by gelatinase zymography and TIMPs by reverse zymography. Both lung cancer cell lines expressed u-PA, which was inhibited by NM in a dose-dependent manner. However, no bands corresponding to u-PA were detected for the MSTO-211H MM cell line. On gelatinase zymography, A-549 cells showed one band corresponding to MMP-2 and induction of MMP-9 with PMA (100 ng/ml) treatment. MSTO-211H showed two bands, an intense band corresponding to MMP-2 and a faint band corresponding to MMP-9; MMP-9 was enhanced significantly with PMA treatment. NM inhibited their expression in both cell lines in a dose-dependent manner. Calu-3 showed no MMP-2 or MMP-9 expression. Activity of TIMPs was upregulated by NM in all cancer cell lines in a dose-dependent manner. Analysis revealed a positive correlation between u-PA and MMPs and a negative correlation between u-PA/MMPs and TIMPs. These findings suggest the therapeutic potential of NM in the treatment of lung and mesothelioma cancers.
Proteomic (antibody microarray) exploration of the molecular mechanism of action of the specific COX-2 inhibitor DuP 697. V. Agarwal, V.C. Hodgkinson, G.L. Eagle, et al. 2013;42(3):1088-92. PubMed · Publisher

Abstract

We have previously shown that specific COX-2 inhibitors, including DuP 697, have anti-proliferative effects on mesothelioma cells and potentiate the cytotoxicity of pemetrexed. Here, we used a novel proteomic approach to explore the mechanism of action of this agent. COX-2-positive cell lines MSTO-211H (mesothelioma) and A549 (lung cancer) were exposed to DuP 697 for 72 h. Drug carrier only was added to control cells. Extracted proteins from treated and control cells were analysed using a comparative proteomic platform. Differentially expressed proteins, identified by the Panorama Xpress Profiler725 antibody microarray were submitted to Ingenuity Pathway Analysis. A total of 32 unique differentially expressed proteins were identified with a significant (>1.8-fold) difference in expression between treated and untreated cells in at least one cell line. Five molecules, BCL2L1 (Bcl-xL), BID, CHUK (IKK), FASLG and RAF1, were mapped to the Apoptosis Signaling pathway following Ingenuity Pathway Analysis. BCL2L1 (Bcl-xL) and BID were analysed using immuno-blotting and differential expression was confirmed. Proteomic (antibody microarray) analysis suggests that the mechanism of action of DuP 697 may be exerted via the induction of apoptosis. The antibody microarray platform can be utilised to explore the molecular mechanism of action of novel anticancer agents.
Antitumor activity of MEK and PI3K inhibitors against malignant pleural mesothelioma cells in vitro and in vivo. S. Miyoshi, H. Hamada, N. Hamaguchi, et al. 2012;41(2):449-56. PubMed · Publisher · Free full text

Abstract

Malignant pleural mesothelioma (MPM) is an aggressive malignancy for which there is no approved targeted therapy. We examined the therapeutic efficacy of the mitogen-activated protein kinase kinase (MEK) and phosphatidylinositol 3-kinase (PI3K) inhibitors against human MPM cell lines both in vitro and orthotopically inoculated into severe combined immunodeficient (SCID) mice. In addition, the molecular mechanisms of these agents were confirmed in vitro and in vivo. The MEK or the PI3K inhibitor suppressed MPM cell growth in vitro in a dose-dependent manner via induction of G1 cell cycle arrest and apoptosis. In addition, combined use of the MEK and PI3K inhibitors showed an additive or synergistic inhibitory effect on MPM cell growth compared to treatment with either individual drug. Treatment with MEK or PI3K inhibitor suppressed the production of thoracic tumors and pleural effusion and prolonged the survival time of EHMES-10 cell-bearing SCID mice. The combination therapy more effectively prolonged the survival time compared to treatment with either individual drug. Immunohistochemical and western blot analysis of thoracic tumors suggested that these agents induced cell cycle arrest, apoptosis and inhibition of tumor angiogenesis. Our results suggest that a combination of MEK and PI3K inhibitors is a promising therapeutic strategy for MPM.
Nitric oxide (NO) enhances pemetrexed cytotoxicity via NO‑cGMP signaling in lung adenocarcinoma cells in vitro and in vivo. H. Nagai, H. Yasuda, Y. Hatachi, et al. 2012;41(1):24-30. PubMed · Publisher

Abstract

Pemetrexed (PEM) is a novel, multitargeted, antifolate, antineoplastic agent for the treatment of non-small cell lung cancer and malignant pleural mesothelioma. Additional effects of nitric oxide (NO) donors on the chemosensitivity of cancers have been reported. However, the effects of an NO donor on PEM-induced cytotoxicity remain unknown. In this study, we investigated the effects of the NO donors, NOC-18 on the cytotoxicity in A549 cells in vitro and of nitroglycerin (GTN), on the tumor growth of Lewis lung carcinoma cells in a murine syngraft model treated with PEM. The effects of NO donors on the expression of proteins associated with PEM metabolism, including thymidylate synthase (TS), reduced folate carrier 1 (RFC1), folylpolyglutamate synthase (FPGS), γ-glutamyl hydrolase (GGH) and multidrug resistance-related protein (MRP)5, and the effects of cyclic guanosine mono-phosphate (cGMP) signaling on these proteins were examined in A549 cells. Treatment with 100 nM NOC-18 for 3 days significantly enhanced PEM-induced cytotoxicity and increased the expression of RFC1 and FPGS in A549 cells. Treatment with 10 nM 8-bromo-cGMP (8-Br-cGMP) for 3 days also increased the expression of RFC1 and FPGS in A549 cells. 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (10 µm) significantly reversed the increase in RFC1 and FPGS expression induced by 100 nM NOC-18 in A549 cells. Combination therapy with GTN and PEM significantly reduced tumor growth compared with PEM alone in the syngraft model. The enhanced antitumor effect of GTN plus PEM was significantly reversed by the concomitant addition of ODQ. These findings suggest that NO donors, such as NOC-18 and GTN, enhance the anticancer effects of PEM by increasing the RFC1 and FPGS expression and stimulating cGMP signaling pathways in cancer cells.
Novel role of c-jun N-terminal kinase in regulating the initiation of cap-dependent translation. M.R. Patel, A.A. Sadiq, J. Jay-Dixon, et al. 2012;40(2):577-82. PubMed · Publisher · Free full text

Abstract

Initiation of protein translation by the 5' mRNA cap is a tightly regulated step in cell growth and proliferation. Aberrant activation of cap-dependent translation is a hallmark of many cancers including non-small cell lung cancer. The canonical signaling mechanisms leading to translation initiation include activation of the Akt/mTOR pathway in response to the presence of nutrients and growth factors. We have previously observed that inhibition of c-jun N-terminal kinase (JNK) leads to inactivation of cap-dependent translation in mesothelioma cells. Since JNK is involved in the genesis of non-small cell lung cancer (NSCLC), we hypothesized that JNK could also be involved in activating cap-dependent translation in NSCLC cells and could represent an alternative pathway regulating translation. In a series of NSCLC cell lines, inhibition of JNK using SP600125 resulted in inhibition of 4E-BP1 phosphorylation and a decrease in formation of the cap-dependent translation complex, eIF4F. Furthermore, we show that JNK-mediated inhibition of translation is independent of mTOR. Our data provide evidence that JNK is involved in the regulation of translation and has potential as a therapeutic target in NSCLC.
Therapeutic antitumor efficacy of anti-epidermal growth factor receptor antibody, cetuximab, against malignant pleural mesothelioma. J. Kurai, H. Chikumi, K. Hashimoto, et al. 2012;41(5):1610-8. PubMed · Publisher · Free full text

Abstract

Epidermal growth factor receptor (EGFR) is commonly overexpressed in malignant pleural mesothelioma (MPM). Cetuximab is a chimeric mouse-human antibody targeted against EGFR and induces potent antibody-dependent cellular cytotoxicity (ADCC). The action of cetuximab against MPM cells has not been well studied. Therefore, in this study, we investigated the antitumor activity of cetuximab against MPM cell lines, particularly with respect to ADCC activity in vitro and in vivo. EGFR expression of MPM cells was measured by a quantitative flow cytometric analysis and immunohistochemistry. The effect of cetuximab on growth inhibition was assessed using a modified MTT assay. The ADCC activity was measured by a 4-h 51Cr release assay using fresh or IL-2-activated peripheral blood mononuclear cells. In vivo antitumor activity of cetuximab was evaluated using an orthotopic implantation mouse model. Cetuximab-mediated ADCC activity against MPM cells was observed at low concentration (0.25 mg/ml) and was enhanced by IL-2, whereas no direct effect on growth inhibition was detected. A logarithmic correlation was observed between the number of EGFRs on MPM cells and ADCC activity. Low EGFR expression on the MPM cells, which was weakly detectable by immunohistochemistry, was sufficient for maximum ADCC activity. In the mouse model, cetuximab treatment with or without IL-2 significantly inhibited intrathoracic tumor growth and prolonged their survival. Our study shows that cetuximab has potent anti-MPM activity both in vitro and in vivo, mainly through the immunologic mechanism of ADCC. Cetuximab has the potential to be used as a novel therapy for MPM patients.
Frequent deletion of 3p21.1 region carrying semaphorin 3G and aberrant expression of the genes participating in semaphorin signaling in the epithelioid type of malignant mesothelioma cells. Y. Yoshikawa, A. Sato, T. Tsujimura, et al. 2011;39(6):1365-74. PubMed · Publisher

Abstract

Array-based comparative genomic hybridization analysis was performed on 21 malignant mesothelioma (MM) samples (16 primary cell cultures and 5 cell lines) and two reactive mesothelial hyperplasia (RM) primary cell cultures. The RM samples did not have any genomic losses or gains. In MM samples, deletions in 1p, 3p21, 4q, 9p21, 16p13 and 22q were detected frequently. We focused on 3p21 because this deletion was specific to the epithelioid type. Especially, a deletion in 3p21.1 region carrying seven genes including SEMA3G was found in 52% of MM samples (11 of 14 epithelioid samples). The allele loss of 3p21.1 might be a good marker for the epithelioid MM. A homozygous deletion in this region was detected in two MM primary cell cultures. A heterozygous deletion detected in nine samples contained the 3p21.1 region and 3p21.31 one carrying the candidate tumor suppressor genes such as semaphorin 3F (SEMA3F), SEMA3B and Ras association (RalGDS/AF-6) domain family member 1 (RASSF1A). SEMA3B, 3F and 3G are class 3 semaphorins and inhibit growth by competing with vascular endothelial growth factor (VEGF) through binding to neuropilin. All MM samples downregulated the expression of more than one gene for SEMA3B, 3F and 3G when compared with Met5a, a normal pleura-derived cell line. Moreover, in 12 of 14 epithelioid MM samples the expression level of SEMA3A was lower than that in Met5a and the two RM samples. An augmented expression of VEGFA was detected in half of the MM samples. The expression ratio of VEGFA/SEMA3A was significantly higher in the epithelioid MMs than in Met5a, RMs and the non-epithelioid MMs. Our data suggest that the downregulated expression of SEMA3A and several SEMA3s results in a loss of inhibitory activities in tumor angiogenesis and tumor growth of VEGFA; therefore, it may play an important role on the pathogenesis of the epithelioid type of MM.
Potent anti-tumor killing activity of the multifunctional Treg cell line HOZOT against human tumors with diverse origins. T. Inoue, Y. Tashiro, M. Takeuchi, et al. 2011;38(5):1299-306. PubMed · Publisher

Abstract

The T cell line HOZOT has a unique FOXP3+CD4+ CD8+CD25+ phenotype, exhibits suppressive activity in allogeneic mixed lymphocyte reactions (MLR), and produces IL-10, defining HOZOT as regulatory T cells (Tregs). Interestingly, in addition to possessing a suppressive Treg ability, HOZOT was also found to show cytotoxicity against certain representative human cancer cell types. In order to disclose the range of anti-tumor activity by HOZOT, we screened it by using a panel of twenty human tumor cell lines with different origins. Consequently, HOZOT showed potent cytocidal effects against a wide spectrum of neoplastic cells including carcinomas, sarcomas, mesotheliomas and glioblastomas except for hematopoietic malignancies. Its anti-tumor activity was strong enough with an E:T ratio of 4:1, which is considered to be more effective than that by conventional CTLs. Furthermore, an in vivo representative mouse tumor model by implanting human colon adenocarcinoma cells revealed that adoptive transfer of HOZOT almost completely eradicated disseminated lesions on peritoneum, markedly reduced metastases in lung and liver, and dramatically decreased bloody ascites caused by peritoneal carcinomatosis. Treatment of the tumor model mice by HOZOT with an E:T ratio of 2:1 even indicated the prolongation of their survival, although not reaching obvious statistical significance. In vitro blocking experiments using antibodies and inhibitors suggested that the cytotoxic mechanism of HOZOT against tumors is different from conventional cytotoxic cells such as CTL, NK or NKT cells. Altogether, our studies demonstrated the potent killing activity of HOZOT against a broad range of human malignancies, which indicates that HOZOT is a powerful tool in immunotherapy for advanced stage tumors.
Transforming growth factor-β-induced protein (TGFBI) suppresses mesothelioma progression through the Akt/mTOR pathway. G. Wen, M. Hong, B. Li, et al. 2011;39(4):1001-9. PubMed · Publisher · Free full text

Abstract

As an uncommon cancer, mesothelioma is very hard to treat with a low average survival rate owing to its usual late detection and being highly invasive. The link between asbestos exposure and the development of mesothelioma in humans is unequivocal. TGFBI, a secreted protein that is induced by transforming growth factor-β in various human cell types, has been shown to be associated with tumorigenesis in various types of tumors. It has been demonstrated that TGFBI expression is markedly suppressed in asbestos-induced tumorigenic cells, while an ectopic expression of TGFBI significantly suppresses tumorigenicity and progression in human bronchial epithelial cells. In order to delineate a potential role of TGFBI in mediating the molecular events that occur in mesothelioma tumorigenesis, we generated stable TGFBI knockdown mutants from the mesothelium cell line Met-5A by using an shRNA approach, and secondly created ectopic TGFBI overexpression mutants from the mesothelioma cell line H28 in which TGFBI is absent. We observed that in the absence of TGFBI, the knockdown mesothelial and mesothelioma cell lines exhibited an elevated proliferation rate, enhanced plating efficiency, increased anchorage-independent growth, as well as an increased cellular protein synthesis rate as compared with their respective controls. Furthermore, cell cycle regulatory proteins c-myc/cyclin D1/phosphor-Rb were upregulated; a more active PI3K/Akt/mTOR signaling pathway was also detected in TGFBI-depleted cell lines. These findings suggest that TGFBI may repress mesothelioma tumorigenesis and progression via the PI3K/Akt signaling pathway.
Putative cancer stem cells in malignant pleural mesothelioma show resistance to cisplatin and pemetrexed. L. Cortes-Dericks, G.L. Carboni, R.A. Schmid, et al. 2010;37(2):437-44. PubMed · Publisher

Abstract

Malignant pleural mesothelioma (MPM) is a lethal cancer of the mesothelium with high chemotherapeutic resistance via unknown mechanisms. A prevailing hypothesis states that cancer stem cells (CSCs) persist in tumors causing relapse after chemotherapy, thus, rendering these cells as critical targets responsible for tumor resistance and recurrence. We selected candidate CSC markers based on expansion under hypoxic conditions, a hallmark for the selection of chemoresistant cells; and investigated the expression of CSC markers: CD133, Bmi-1, uPAR and ABCG2 in three MPM cell lines and normal mesothelial cells by quantitative RT-PCR. Furthermore, we evaluated the chemotherapeutic resistance associated with each CSC marker by determining the change in CSC marker-mRNA levels as an index of drug-resistance following treatment with either cisplatin or pemetrexed. We demonstrate the expression of CSC markers: CD133, Bmi-1, uPAR and ABCG2 in both normal and MPM cell lines. Bmi-1+, uPAR+ and ABCG2+ cells show a distinct role in conferring chemoresistance to cisplatin and pemetrexed in the malignant setting. By contrast, these markers have no apparent participation in chemoresistance to drug treatments in normal mesothelial cells. Intriguingly, CD133 revealed chemoresistant properties in both normal mesothelial and malignant pleural mesothelioma cells. This study provides evidence of putative CSCs conferring drug-resistance to cisplatin and pemetrexed in MPM cell lines. Specific targeting of these drug-resistant cells, while considering the functional heterogeneity of the MPM subtypes, may contribute to more focused and effective chemotherapeutic regimens for malignant pleural mesothelioma.

2000s

Mineral fiber-mediated activation of phosphoinositide-specific phospholipase c in human bronchoalveolar carcinoma-derived alveolar epithelial A549 cells. C. Loreto, M.L. Carnazza, V. Cardile, et al. 2009;34(2):371-6. PubMed

Abstract

Given the role of phosphoinositide-specific phospholipase C (PLC) isozymes in the control of cell growth and differentiation we were prompted to analyze the expression of some of these PLC in human bronchoalveolar carcinoma-derived alveolar epithelial A549 cells. The effects of several fluoro-edenite fibers were compared with those of tremolite, a member of the calcic amphibole group of asbestos that originates from Calabria (Italy), and crocidolite, that, due to its high toxicity, is one of the most studied asbestos amphiboles. Our data show an increased expression of both PLC beta1 and PLC gamma1 in A549 cells treated with asbestos-like fibers, hinting at a role of PLC signalling in those cancerous cells.
Defining high-risk individuals in a population-based molecular-epidemiological study of lung cancer. A. Cassidy, J.P. Myles, T. Liloglou, et al. 2006;28(5):1295-301. PubMed

Abstract

Within the framework of the Liverpool Lung Project (LLP), population-based case-control and prospective cohort studies are in progress to identify molecular and epidemiological risk factors and define populations and individuals most at risk of developing lung cancer. This report describes a strategy for selection of a high-risk population and further provides support for the inclusion of occupational and genetic risk factors in future models. Data from the case-control study (256 incident cases and 314 population controls) were analysed to define a high-risk population. Detailed lifestyle and occupational information were collected during structured interviews. Models were constructed using conditional logistic regression and included terms for age, tobacco consumption and previous respiratory disease. Smoking duration was chosen as the most important predictor of lung cancer risk [>50 years (OR 15.65, 95% CI 6.10-40.15)]. However, such a model would preclude younger individuals. Several combinations of previous respiratory disease were also considered, of which a history of bronchitis, emphysema or pneumonia (BEP) was the most significant (OR 1.86, 95% CI 1.28-2.69). A high-risk subset (based on combinations of smoking duration and BEP) was identified, which have a 4.5-fold greater risk of developing lung cancer (OR 4.5, 95% CI 2.33-8.68). Future refinement of the risk model to include individuals occupationally exposed to asbestos and with the p21 genotypes is discussed. There is real potential for environmental and genetic factors to improve on risk prediction and targeting of susceptible individuals beyond the traditional models based only on smoking and age. The development of a molecular-epidemiological model will inform the development of effective surveillance, early detection and chemoprevention strategies.
Interleukin-1beta and tumour necrosis factor-alpha promote the transformation of human immortalised mesothelial cells by erionite. Y. Wang, S.P. Faux, G. Hallden, et al. 2004;25(1):173-8. PubMed

Abstract

Asbestos fails to induce the transformation of human mesothelial cells in vitro although it has been known as a potential carcinogen to human mesothelial cells. Interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNF-alpha) are major cytokines released by macrophages after inhalation of asbestos. These cytokines can regulate mesothelial cell proliferation both in vitro and in vivo. In the present study, we used the growth in soft agar as an index of transformation and investigated the role of IL-1beta and TNF-alpha during the process of human mesothelial cell carcinogenesis. Both IL-1beta and TNF-alpha were demonstrated to enhance erionite-induced transformation of the immortalised, non-tumorigenic human mesothelial cell line (MeT-5A) in vitro. The MeT-5A cells could only be transformed when the cells were exposed to a combination of cytokines and erionite, or at least two cytokines together without erionite, for at least 4 months in vitro. The findings presented here suggest that IL-1beta and TNF-alpha play a significant role in the pathogenesis of mesothelioma, and that it might be desirable to block or inhibit cytokine secretion in high risk populations to prevent mesothelioma.
Reduced Fhit protein expression and loss of heterozygosity at FHIT gene in tumours from smoking and asbestos-exposed lung cancer patients. L. Pylkkanen, H. Wolff, T. Stjernvall, et al. 2002;20(2):285-90. PubMed

Abstract

The FHIT gene, at 3p14.2, has been suggested to form a molecular target to damage induced by human lung carcinogens. We examined aberrant expression of the Fhit protein and allele loss at the FHIT gene in a series of lung cancer cases, mainly of non-small cell carcinoma (NSCLC) histology. We had detailed data on tobacco smoke exposure and occupational asbestos exposure available for the cases. The principal aim of the present study was to investigate whether absent or reduced Fhit expression or FHIT allele loss was associated with exposure to these pulmonary carcinogens. We detected reduced Fhit expression in 62% (33/53) of the cases analysed. Prevalence of allele loss at the FHIT locus was 22% (20/89). Reduced protein expression was common both in the asbestos-exposed (67%) and non-exposed cases (59%); [odds ratio (OR) 1.4, 95% confidence interval (CI) 0.4-4.9]. LOH frequencies differed somewhat between the two groups and were 25% vs. 16%, respectively (OR 1.8; 95% CI 0.5-5.9). Absent or reduced expression was common in smokers, with no significant difference found between current smokers and non-smokers (mainly former smokers) (OR 1.4, 95% CI 0.5-4.5). NSCLCs with squamous cell histology exhibited both aberrant expression (OR 3.1, 95% CI 0.9-10.3) and allele loss (OR 3.3, 95% CI 0.9-12.7) more frequently than adenocarcinoma. Finally, we found that FHIT allele loss was increased in stage II or more advanced disease (OR 2.5, 95% CI 0.9-7.4), and in poorly differentiated tumours (grade 3, OR 2.6, 95% CI 0.8-8.1). In conclusion, our present data support significance of FHIT inactivation in development of lung cancer.
K-ras and rho A mutations in malignant pleural effusion. M. Nakamoto, H. Teramoto, S. Matsumoto, et al. 2001;19(5):971-6. PubMed · Publisher

Abstract

Mutations of the Kristen ras (K-ras) gene have been implicated in the pathogenesis of human lung cancer, especially adenocarcinoma, and have been proposed to be a prognostic factor. The K-ras mutation in codon 12 is detectable even in cell-free fluids by using the enriched polymerase chain reaction (PCR) technique. On the other hand, based on experimental results, the rho A mutation in codon 14 is also proposed to be oncogenic as observed in the K-ras mutation. Malignant pleural effusion is a common complication of lung cancer. We studied the point mutation of K-ras codon 12 and rho A codon 14 using enriched PCR in specimens of pleural effusion. Forty patients with pleural effusion were enrolled in this study. The causes of pleural effusion were non-small cell lung cancer (18 cases), small cell lung cancer (6 cases), malignant mesothelioma (2 cases), metastatic lung tumor (5 cases), thymoma (1 case), malignant lymphoma (1 case), and pleuritis tuberculosa (7 cases). The K-ras mutation was detected in 4 of 14 cases with adenocarcinoma, 1 of 3 cases with squamous cell carcinoma, 1 of 1 case with large cell carcinoma, and 1 of 5 cases with metastatic lung tumor, respectively. The rho A mutation was not detected in any pleural effusion examined in this study. Our study demonstrates the usefullness of pleural effusion as a clinical specimen for a search of point mutation of oncogenes. The K-ras codon 12 mutation is readily detected in pleural effusion, and the demonstration of this mutation has potentially important implications for the diagnosis of malignant pleural effusion.

1990s

Activation-induced cell death in human peripheral blood lymphocytes after stimulation with silicate in vitro. T. Aikoh, A. Tomokuni, T. Matsukii, et al. 1998;12(6):1355-9. PubMed · Publisher

Abstract

Silica and related substances such as silicate have been proven to possess "adjuvant effects". We have previously reported a finding of polyclonal human T cell activation induced by silicate as a superantigen in vitro. In this study, we observed activation-induced cell death in human lymphocytes after stimulation with chrysotile, a kind of silicate. Apoptotic cells were detected flow cytometrically using the TUNEL assay, and the maximum appearance of TUNEL positive cells occurred on day 4 of incubation. Simultaneously the manifestation of small-sized cells in the specimens increased implying apoptosis. Fas expression on lymphocytes increased to day 3 of incubation with chrysotile, and then spontaneously decreased on day 4 when remarkable apoptosis could be detected. Based on these results it is conceivable that activation-induced cell death occurred through Fas-Fas ligand interaction in lymphocytes after stimulation with silicate in a concentration with which no acute cytotoxicity has been detected. Whether and how the repeated apoptosis in definite clones of lymphocytes causes the induction of sFas synthesis need clarification.
Comparative analysis of p16/CDKN2, p53 and ras gene alterations in human non-small cell lung cancers, with and without associated pulmonary asbestosis. I. Hayashi, N. Konishi, H. Matsuda, et al. 1996;8(1):85-90. PubMed · Publisher

Abstract

To investigate genetic abnormalities in human non-small cell lung carcinomas (NSCLC) associated with pulmonary asbestosis as compared with nou-asbestos linked lung cancers, twenty-nine primary non-small cell lung carcinomas (NSCLC) were examined for genetic abnormalities of p16/CDKN2, p53 and ras genes by single-strand conformation polymorphism analysis of polymerase chain reaction products (PCR-SSCP) and direct sequencing. Ten specimens were obtained from autopsies in which concurrent pulmonary asbestosis was present, while 19 samples were surgical specimens from asbestosis-free patients. K-ras mutations were detected in 10% each of the cancers from both asbestosis and non-asbestosis cases. p16/CDKN2 deletions or mutations and p53 aberrations were demonstrated in 20% and 10% of tumors from asbestosis cases, whereas, 32% and 21% of the cancers, respectively, from asbestosis-free patients were positive. In conclusion, it is suggested that the enhancement of neoplasia in the lung by asbestos is not dependent on suppression of p16/CDKN2 and p53 or ras activation and therefore, that asbestosis may activate alternate tumorigenic pathways in the development of NSCLC.
Treatment of malignant mesothelioma (review). S. Mikulski. 1995;7(6):1415-20. PubMed · Publisher

Abstract

Malignant mesothelioma (MM) has a very variable natural history. Its known relationship to asbestos exposure is often difficult to establish in a particular patient in view of a long preceding latency period of up to 40+ years. Survival depends on several prognostic factors including histological type, stage, performance status, resectability, and response to radiation and chemotherapy (CT). The assessment of the therapeutic impact on survival is usually difficult due to the different treatment philosophies and selections of patients in various clinical trials. Combined modality treatment approaches seem to offer the best outcomes, with occasional long-term disease-free survivors. Unresectable MM presents a difficult therapeutic challenge. In patients treated with systemic CT the median survival time (MST) in the range of 6 to 12 months and the 2 year survival of <20% are typical. The MST in excess of 20 months and the 2 year survival in the range of 40% have been reported in a few multimodality therapy trials in patients with at least partially resectable MM. Despite some encouraging therapeutic results in selected patients, the overall response to treatment(s) remains unsatisfactory, and new treatments must be sought.
Human cd4+ cd45ra+ lymphocytes-T can be stimulated by crocidolite, anthophyllite and amosite asbestos invitro. Y. Watanabe, M. Yamaguchi, Y. Kawakami, et al. 1993;2(2):209-12. PubMed · Publisher

Abstract

Crocidolite, anthophyllite and amosite asbestos stimulate T lymphocyte subsets. Cell surface CD4 or CD45RA expression in peripheral blood mononuclear cells (PBMC) was downregulated temporarily after incubation with asbestos in vitro. The percentage of CD4+ CD45RA+ cells significantly decreased 12 h after incubation with asbestos, suggesting activation of the cells. An early increase in the intracellular Ca++ level was also indicative of activation. An elevated Ca++ level was observed after incubation in both PBMC and purified T cell fractions.

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