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M.B. Kirschner
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MINI 24 - Epidemiology, Early Detection, Biology (ID 140)
- Event: WCLC 2015
- Type: Mini Oral
- Track: Thymoma, Mesothelioma and Other Thoracic Malignancies
- Presentations: 1
- Moderators:J. Creaney, M. Carbone
- Coordinates: 9/08/2015, 16:45 - 18:15, 102+104+106
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MINI24.09 - Cell-Free MicroRNA miR-625-3p Is Elevated in the Blood of Patients with Thoracic Malignancies (ID 1282)
17:30 - 17:35 | Author(s): M.B. Kirschner
- Abstract
Background:
In most instances definitive diagnosis of malignant pleural mesothelioma (MPM) requires a tissue biopsy of sufficient size. As a biopsy is not always feasible, the identification of an accurate biomarker easily measured in blood would represent an important step forward. A recent study indicated that microRNA miR-625-3p was present in elevated concentration in plasma or serum of MPM patients compared to healthy controls and asbestosis patients. (Kirschner et al, JTO; 7:1184). In this study, we have further investigated the diagnostic potential of miR-625-3p.
Methods:
MiR-625-3p and other microRNAs were measured by RT-qPCR in two independent series of MPM patients and controls. After exclusion of haemolysed samples and those yielding RNA of insufficient quality, series 1 consisted of serum samples from 73 MPM patients, 69 healthy volunteers and 64 patients with non-small cell lung cancer (NSCLC) collected at the Netherlands Cancer Institute (NKI) between 1994 and 2013. The second series consisted of plasma samples from 29 MPM patients and 35 healthy volunteers collected in Vienna and Hungary (V/H) between 2011 and 2013. Additionally levels of soluble mesothelin-related protein (SMRP) were assessed (ELISA) in the NKI series.
Results:
Analyses of samples from patients and controls in the NKI series revealed that serum miR-625-3p concentrations were on average 5.35-fold higher (p=0.0054) in MPM, and 3.47-fold (p=0.003) in NSCLC than in control samples. Levels in MPM patients were 1.54-fold higher than in NSCLC patients but this did not reach statistical significance (p=0.273). Compared to healthy controls, the areas under the ROC curve (AUC) were 0.82 (95% CI: 0.75-0.89) for MPM and 0.75 (95% CI: 0.67-0.84) for NSCLC. In the samples of the V/H series, plasma miR-625-3p concentrations were on average 1.98-fold (p<0.001) higher in MPM patients than in healthy volunteers, with an AUC of 0.80 (95% CI: 0.69-0.91). Assessment of SMRP in the NKI series revealed AUCs of 0.69 (95% CI: 0.59-0.78) differentiating MPM from healthy individuals and 0.65 (95% CI: 0.54-0.75) separating MPM from NSCLC, comparable to AUC values reported earlier.
Conclusion:
Data from two independent validation series confirms the previously observed increased abundance of miR-625-3p in blood from MPM patients. However, the miR-625-3p levels observed in NSCLC patients show that elevation of the level of this microRNA in plasma/serum is not restricted to MPM. Further studies into combinations of microRNAs and SMRP (diagnostic signature) in MPM are warranted.
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P1.01 - Poster Session/ Treatment of Advanced Diseases – NSCLC (ID 206)
- Event: WCLC 2015
- Type: Poster
- Track: Treatment of Advanced Diseases - NSCLC
- Presentations: 1
- Moderators:
- Coordinates: 9/07/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P1.01-072 - Targeted Delivery of a Synthetic microRNA-Based Mimic to Treat Thoracic Cancers (ID 2911)
09:30 - 09:30 | Author(s): M.B. Kirschner
- Abstract
Background:
MicroRNA expression is commonly suppressed in cancer, contributing to tumor cell biology. Recently we demonstrated that multiple members of the miR-15/16 family are downregulated and have tumor suppressor functions in malignant pleural mesothelioma (MPM), an asbestos-related cancer for which few treatments are available. These results are similar to previous findings in non-small cell lung cancer (NSCLC). As multiple microRNAs from the same family are downregulated in MPM, we investigated whether a single synthetic mimic based on the consensus sequence of the entire family could restore activity of the entire family.
Methods:
Novel microRNA mimics based on the consensus sequence of the miR-15/107 group were designed (con15/107.1 to 4). The effects on growth, migration, target regulation, drug sensitivity and angiogenesis of the con15/107 mimics were compared with native miR-15 and miR-16 mimics using standard assays in MPM and lung cancer cell lines in vitro. The regulation of specific target genes was assessed by RT-qPCR, Western blot and luciferase reporter assays. Global gene regulation was assessed by proteomics. Activity of con15/107 mimics was investigated in vivo in xenograft models in nude mice.
Results:
The consensus mimics inhibited growth and migration of MPM and lung cancer cell lines in vitro, and effects were greater than with native miR-15 or miR-16 mimics. Growth inhibition was associated with an induction of apoptosis, and downregulation of predicted targets of the mimics. Target gene interactions were confirmed with 3’UTR reporter constructs, and proteomics identified a number of candidate genes involved in consensus mimic-induced growth inhibition. Consensus mimics also sensitized multiple MPM and lung cancer cell lines to chemotherapy agents, and inhibited angiogenic activity in endothelial cells. In a xenograft model, the consensus mimic con15/107.2, packaged in bacterially-derived, EGFR antibody-targeted, EDV[TM]nanocells, inhibited MPM tumor growth in vivo.
Conclusion:
The novel con15/107 mimics based on the consensus sequence of the miR-15/107 group have greater activity than native miR-15 or miR-16 mimics in vitro and are active in vivo. Increased activity correlates to greater target gene downregulation. These preclinical studies support a Phase I clinical trial has been initiated for patients with MPM or NSCLC failing standard therapy. This represents the first trial of microRNA replacement as a therapy for thoracic cancer.
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P2.08 - Poster Session/ Thymoma, Mesothelioma and Other Thoracic Malignancies (ID 225)
- Event: WCLC 2015
- Type: Poster
- Track: Thymoma, Mesothelioma and Other Thoracic Malignancies
- Presentations: 1
- Moderators:
- Coordinates: 9/08/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P2.08-006 - The JmjC Family of Lysine Demethylases Are Overexpressed and Potential Therapeutic Targets in Malignant Pleural Mesothelioma (ID 1753)
09:30 - 09:30 | Author(s): M.B. Kirschner
- Abstract
Background:
Malignant pleural mesothelioma (MPM) is an aggressive rare cancer affecting the pleura and is predominatly associated with prior exposure to asbestos. Treatment options are limited, and most patients die within 24 months of diagnosis. The current standard of care for MPM patients is a combination of cisplatin and pemetrexed (or alternatively cisplatin and raltitrexed), yet most patients die within 24 months of diagnosis. There is therefore an urgent unmet need to identify new therapeutic options for the treatment of MPM. Asbestos fibres contain of transition metals and their ability to both adsorb and accumulate these metals was one of the first mechanisms suggested for explaining the toxic and particularly carcinogenic effects of asbestos. One of the transition metals in asbestos fibres is iron, and therefore asbestos fibres may cause an alteration of iron homeostasis in the tissue. In addition, asbestos fibres have also been shown to have high affinity for histones, and therefore may result in high accumulation of iron around chromatin. Lysine Demethylases (KDMs) containing a JmjC domain require both Fe2+ and 2-oxoglutarate as co-factors to regulate gene expression by “erasing or removing” methylation on histones in chromatin. Members of this family are frequently found to have aberrant expression in cancer and currently are actively pursued as candidate pharmaceutical therapeutic targets. Given that asbestos increases iron levels, this may result in aberrant KDM activity, and these KDMs could therefore be novel candidate targets in mesothelioma. We therefore examined the expression of several JmjC containing KDMs in MPM and assessed their potential for therapeutic intervention in mesothelioma using existing small molecule inhibitors.
Methods:
A panel of MPM cell lines were screened for expression of KDM4A-D, KDM5A/B and KDM6A/B by RT-PCR. mRNA levels were subsequently examined by RT-PCR in a cohort of snap-frozen patient samples isolated at surgery comprising benign, epithelial, biphasic, and sarcomatoid histologies. IHC was performed for KDM4A on a cohort of FFPE specimens. The effects of treatments with small molecule inhibitors targeting these proteins on both cellular health and gene expression were assessed.
Results:
The expression of the various KDMs was detectable across our panel of cell lines. In primary tumours the expression of these KDMs were significantly elevated in malignant MPM compared to benign pleura (p<0.05), and significant differences were also observed when samples were analysed across different histological subtypes. Treatment of mesothelioma cell lines with various small molecule inhibitors caused significant effects on cellular health and on the expression of a panel of genes.
Conclusion:
The expression of KDMs are significantly altered in MPM. Small molecule inhibitors directed against these KDMs show potential therapeutic efficacy with significant anti-proliferative effects. We continue to assess the effects of these compounds on gene expression and cellular health to confirm their potential utility as novel therapies for the treatment of MPM.
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P3.08 - Poster Session/ Thymoma, Mesothelioma and Other Thoracic Malignancies (ID 226)
- Event: WCLC 2015
- Type: Poster
- Track: Thymoma, Mesothelioma and Other Thoracic Malignancies
- Presentations: 1
- Moderators:
- Coordinates: 9/09/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P3.08-007 - Fibulin-3: A Potential Prognostic Biomarker in Malignant Pleural Mesothelioma? (ID 1668)
09:30 - 09:30 | Author(s): M.B. Kirschner
- Abstract
Background:
Malignant pleural mesothelioma (MPM) is a highly aggressive asbestos-induced cancer arising from the mesothelium lining the thoracic cavities. The definitive diagnosis of MPM in most instances depends on the availability of a biopsy. A number of biomarkers have been proposed to assist in making the MPM diagnosis but none of them has yet reached the accuracy required for routine clinical use. Among the candidates is the secreted extracellular glycoprotein Fibulin-3 (FBLN3) (Pass et al, NEJM 2012; 367(15)). In this study, we have further investigated the potential of FBLN3 to serve as a biomarker for MPM.
Methods:
Cellular and secreted FBLN3 was measured (ELISA) in MPM and normal mesothelial cell lines, plasma of xenograft tumour-bearing mice, plasma from two independent series of MPM and non-MPM patients, and in malignant and non-malignant pleural effusions. The diagnostic and prognostic potential of FBLN3 was assessed by receiver operating characteristics curve analysis and the Kaplan-Meier method, respectively.
Results:
FBLN3 levels were significantly higher in MPM cells than in mesothelial cells, with a strong correlation between secreted and cellular levels. Human FBLN3 was also detectable in the plasma of tumour-bearing mice, suggesting that MPM cells were the origin of circulating FBLN3. Plasma FBLN3 levels found in MPM patients were lower than previously reported (Pass et al, NEJM 2012; 367(15)), but were comparable to those appearing in subsequent validation studies (Creaney et al, Thorax 2014; 69(10), Corradi et al, Anticancer Res 2013; 33(12)). Plasma FBLN3 was significantly elevated in MPM patients from a Sydney cohort, but far less in a Vienna cohort and the diagnostic accuracy of FBLN3 was insufficient in both cohorts [63%, (95%CI: 50.1-76.4) and 56% (95%CI: 41.5-71.0), respectively]. FBLN3 levels found in pleural effusions were comparable to those reported in previous studies, but the difference between cases and controls did not reach significance. In our series low levels of pleural effusion FBLN3 were again associated (p=0.002) with prolonged survival. In multivariate analysis taking histological subtype, age and gender into account FBLN3 remained significant with a hazard ratio of 9.92 (95%CI: 2.14–45.93).
Conclusion:
FBLN3 is overexpressed in MPM cell lines and may point to a potential oncogenic role for this protein. In contrast to the initial report linking FBLN3 to diagnosis in MPM, the levels of FBLN3 measured in plasma and pleural fluid of our series of MPM patients lacked diagnostic accuracy. However, the potential prognostic value of FBLN3 levels measured in pleural fluid was confirmed and in line with previous validation studies. These data underline the importance of validation studies for newly proposed biomarkers.
