Autophagy Correlates with the Therapeutic Responsiveness of Malignant Pleural Mesothelioma in 3D Models.

Malignant pleural mesothelioma is a highly chemoresistant solid tumor. We have studied this apoptotic resistance using in vitro and ex vivo three-dimensional models, which acquire a high level of chemoresistance that can be reduced by PI3K/mTOR inhibitors. Here, we investigate the activity of GDC-0980, a novel dual PI3K/mTOR inhibitor, which has been proposed to be effective in mesothelioma. In this work, we aimed to identify mechanisms and markers of efficacy for GDC-0980 by utilizing 3D models of mesothelioma, both in vitro multicellular spheroids and ex vivo tumor fragment spheroids grown from patient tumor samples. We found that a subset of mesothelioma spheroids is sensitive to GDC-0980 alone and to its combination with chemotherapy. Unexpectedly, this sensitivity did not correlate with the activation of the Akt/mTOR pathway. Instead, sensitivity to GDC-0980 correlated with the presence of constitutive ATG13 puncta, a feature of autophagy, a cellular program that supports cells under stress. In tumor fragment spheroids grown from 21 tumors, we also found a subset (n = 11) that was sensitive to GDC-0980, a sensitivity that also correlated with the presence of ATG13 puncta. Interference with autophagy by siRNA of ATG7, an essential autophagic protein, increased the response to chemotherapy, but only in the sensitive multicellular spheroids. In the spheroids resistant to GDC-0980, autophagy appeared to play no role. In summary, we show that GDC-0980 is effective in mesothelioma 3D models that display ATG13 puncta, and that blockade of autophagy increases their response to chemotherapy. For the first time, we show a role for autophagy in the response to chemotherapy of 3D models of mesothelioma and propose ATG13 as a potential biomarker of the therapeutic responsiveness of mesothelioma.

The association of 18F-FDG PET/CT parameters with survival in malignant pleural mesothelioma.

PURPOSE: Malignant pleural mesothelioma (MPM) is a disease with poor prognosis despite multimodal therapy but there is variation in survival between patients. Prognostic information is therefore potentially valuable in managing patients, particularly in the context of clinical trials where patients could be stratified according to risk. Therefore we have evaluated the prognostic ability of parameters derived from baseline 2-[(18)F]fluoro-2-deoxy-D-glucose positron emission tomography/computed tomography ((18)F-FDG PET/CT). METHODS: In order to determine the relationships between metabolic activity and prognosis we reviewed all (18)F-FDG PET/CT scans used for pretreatment staging of MPM patients in our institution between January 2005 and December 2011 (n = 60) and measured standardised uptake values (SUV) including mean, maximum and peak values, metabolic tumour volume (MTV) and total lesion glycolysis (TLG). Overall survival (OS) or time to last censor was recorded, as well as histological subtypes. RESULTS: Median follow-up was 12.7 months (1.9-60.9) and median OS was 14.1 months (1.9-54.9). By univariable analysis histological subtype (p = 0.013), TLG (p = 0.024) and MTV (p = 0.038) were significantly associated with OS and SUVmax was borderline (p = 0.051). On multivariable analysis histological subtype and TLG were associated with OS but at borderline statistical significance (p = 0.060 and 0.058, respectively). No statistically significant differences in any PET parameters were found between the epithelioid and non-epithelioid histological subtypes. CONCLUSION: (18)F-FDG PET/CT parameters that take into account functional volume (MTV, TLG) show significant associations with survival in patients with MPM before adjusting for histological subtype and are worthy of further evaluation to determine their ability to stratify patients in clinical trials.

Imaging in pleural mesothelioma: a review of the 11th International Conference of the International Mesothelioma Interest Group.

Imaging of malignant pleural mesothelioma (MPM) is essential to the diagnosis, assessment, and monitoring of this disease. The complex morphology and growth pattern of MPM, however, create unique challenges for image acquisition and interpretation. These challenges have captured the attention of investigators around the world, some of whom presented their work at the 2012 International Conference of the International Mesothelioma Interest Group (iMig 2012) in Boston, Massachusetts, USA, September 2012. The measurement of tumor thickness on computed tomography (CT) scans is the current standard of care in the assessment of MPM tumor response to therapy; in this context, variability among observers in the measurement task and in the tumor response classification categories derived from such measurements was reported. Alternate CT-based tumor response criteria, specifically direct measurement of tumor volume change and change in lung volume as a surrogate for tumor response, were presented. Dynamic contrast-enhanced CT has a role in other settings, but investigation into its potential use for imaging mesothelioma tumor perfusion only recently has been initiated. Magnetic resonance imaging (MRI) and positron-emission tomography (PET) are important imaging modalities in MPM and complement the information provided by CT. The pointillism sign in diffusion-weighted MRI was reported as a potential parameter for the classification of pleural lesions as benign or malignant, and PET parameters that measure tumor activity and functional tumor volume were presented as indicators of patient prognosis. Also reported was the use of PET/CT in the management of patients who undergo high-dose radiation therapy. Imaging for MPM impacts everything from initial patient diagnosis to the outcomes of clinical trials; iMig 2012 captured this broad range of imaging applications as investigators exploit technology and implement multidisciplinary approaches toward the benefit of MPM patients.

BAK and NOXA are critical determinants of mitochondrial apoptosis induced by bortezomib in mesothelioma.

Based on promising preclinical efficacy associated with the 20S proteasome inhibitor bortezomib in malignant pleural mesothelioma (MPM), two phase II clinical trials have been initiated (EORTC 08052 and ICORG 05-10). However, the potential mechanisms underlying resistance to this targeted drug in MPM are still unknown. Functional genetic analyses were conducted to determine the key mitochondrial apoptotic regulators required for bortezomib sensitivity and to establish how their dysregulation may confer resistance. The multidomain proapoptotic protein BAK, but not its orthologue BAX, was found to be essential for bortezomib-induced apoptosis in MPM cell lines. Immunohistochemistry was performed on tissues from the ICORG-05 phase II trial and a TMA of archived mesotheliomas. Loss of BAK was found in 39% of specimens and loss of both BAX/BAK in 37% of samples. However, MPM tissues from patients who failed to respond to bortezomib and MPM cell lines selected for resistance to bortezomib conserved BAK expression. In contrast, c-Myc dependent transactivation of NOXA was abrogated in the resistant cell lines. In summary, the block of mitochondrial apoptosis is a limiting factor for achieving efficacy of bortezomib in MPM, and the observed loss of BAK expression or NOXA transactivation may be relevant mechanisms of resistance in the clinic.

In vivo loss of expression of argininosuccinate synthetase in malignant pleural mesothelioma is a biomarker for susceptibility to arginine depletion.

PURPOSE: Malignant pleural mesothelioma (MPM) is an increasing health burden on many societies worldwide and, being generally resistant to conventional treatment, has a poor prognosis with a median survival of <1 year. Novel therapies based on the biology of this tumor seek to activate a proapoptotic cellular pathway. In this study, we investigated the expression and biological significance of argininosuccinate synthetase (AS), a rate-limiting enzyme in arginine production. EXPERIMENTAL DESIGN: Initially, we documented down-regulation of AS mRNA in three of seven MPM cell lines and absence of AS protein in four of seven MPM cell lines. We confirmed that the 9q34 locus, the site of the AS gene, was intact using a 1-Mb comparative genomic hybridization array; however, there was aberrant promoter CpG methylation in cell lines lacking AS expression, consistent with epigenetic regulation of transcription. To investigate the use of AS negativity as a therapeutic target, arginine was removed from the culture medium of the MPM cell lines. RESULTS: In keeping with the cell line data, 63% (52 of 82) of patients had tumors displaying reduced or absent AS protein, as assessed using a tissue microarray. Cell viability declined markedly in the AS-negative cell lines 2591 and MSTO but not in the AS-positive cell line, 28. This response was apparent by day 4 and maintained by day 9 in vitro. Arginine depletion induced BAX conformation change and mitochondrial inner membrane depolarization selectively in AS-negative MPM cells. CONCLUSIONS: In summary, we have identified AS negativity as a frequent event in MPM in vivo, leading to susceptibility to cytotoxicity following restriction of arginine. A phase II clinical trial is planned to evaluate the role of arginine depletion in patients with AS-negative MPM.

Prognostic factors in mesothelioma.

Prognostic factors can help clinicians and patients when deciding a treatment plan. Patients in the best prognostic groups can be considered for more intensive or experimental therapy. Alternatively, patients in the best prognostic groups might prefer a period of observation prior to commencement of therapy. For patients with mesothelioma prognostic factors are potentially especially important because of the lack of a widely applicable anatomical staging system. Both the International Mesothelioma Interest Group (IMIG) and Brigham staging systems are of limited relevance to patients not undergoing radical debulking surgery. Radiological prediction of IMIG or Brigham stage is of little value. Review of the best-known prognostic scoring systems from the EORTC and CALGB has shown that the most important predictors of poor prognosis are: poor performance status; non-epithelioid histology; male gender; low hemoglobin; high platelet count; high white blood cell count; and high lactate dehydrogenase (LDH). The EORTC model was validated at St Bartholomew's Hospital in a group of 145 patients treated in sequential phase II chemotherapy trials. For 70 patients treated with vinorelbine, those having the best EORTC prognosis had a median survival of 19.2 months [95% C.I.=14.7-23.7] compared to 9.9 months [95% C.I.=8.5-11.3] for those in the worst group. The suggestion is that all clinical and biological factors relevant to prognosis should be recorded prospectively in mesothelioma patients selected for clinical trials.