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Michaela B Kirschner
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MA 19 - Mesothelioma: Bench to Bedside (ID 680)
- Event: WCLC 2017
- Type: Mini Oral
- Track: Mesothelioma
- Presentations: 1
- Moderators:Dean A Fennell, Hedy Lee Kindler
- Coordinates: 10/18/2017, 11:00 - 12:30, Room 315
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MA 19.06 - Multiple Mechanisms Contribute to Downregulation of Tumour Suppressor microRNAs in Malignant Pleural Mesothelioma (ID 9745)
11:35 - 11:40 | Author(s): Michaela B Kirschner
- Abstract
- Presentation
Background:
Malignant pleural mesothelioma (MPM) is a disease with an almost invariably fatal diagnosis with limited therapeutic options. Characteristic patterns of deregulated microRNA expression have been demonstrated in MPM, and many downregulated microRNAs have been shown to have tumour suppressor activity. However, apart from silencing of miR-34b/c by promoter hypermethylation and co-deletion of miR-31 with the CDKN2A locus, the mechanisms responsible for downregulation of other tumour suppressor miRNAs such as miR-16 are yet to be elucidated.
Method:
Tumour samples (n=60) were from MPM patients undergoing extrapleural pneumonectomy, and samples of pleura (n=23) collected from patients undergoing cardiac surgery were used as normal controls. MPM cells lines were obtained from the ATCC. Expression levels of mature microRNAs in MPM tumour samples and cell lines, and pri-miRs and miRNA host genes in cell lines, were determined by RT-qPCR. Copy number variation (CNV) was analysed by droplet digital PCR (ddPCR), and methylation was inferred by miRNA expression following decitabine treatment. MYC was analysed by Western blot, and expression modulated by siRNAs.
Result:
Analysis of microRNA expression in tumour samples revealed a consistent and significant downregulation of miR-15a (4-fold, P<0.01), 15b (10-fold, P<0.01), 16 (22-fold, P<0.05), 34a (1.6-fold, P<0.05), 34b (1.8-fold, P<0.01), 34c (2.3-fold, P<0.0001) and 193a (3.1-fold, P<0.001) compared with normal pleura. Copy number variation analysis showed evidence of heterozygous loss for miR-193a (4 of 5 cell lines) and miR-15a/16-1 (2 of 5), but no change in miR-15b/16-2. Treating cell lines with the demethylating agent decitabine resulted in dramatic upregulation only in the case of miR-34c. RNAi-mediated knockdown of c-MYC led to upregulation of miR-15b and 16, and to a lesser extent miR-15a, as well as a consistent increase in the miR-15b/16-2 host gene SMC4 and the miR-15a/16-1 host gene DLEU2. Analysing the expression of these microRNAs in the tumour samples revealed a strong correlation between miR-15b and 16 (R[2]=0.793) and miR-34b and 34c (R[2]=0.753), but not between others.
Conclusion:
Our data suggest that a combination of deletion, hypermethylation and transcriptional regulation contribute to the downregulation of miR-15a/b, 16, 34a/b/c and 193a. In MPM, unlike other cancers, the downregulation of miR-15a/16-1, miR-15b/16-2 appears to be due to transcriptional changes rather than deletion or promoter hypermethylation. MYC appears to contribute to miR-16 downregulation primarily via control of SMC4 and the miR-15b/16-2 locus, suggesting that the transcriptional control of miR-16 expression by c-Myc contributes to the malignant phenotype of MPM.
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P1.09 - Mesothelioma (ID 695)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Mesothelioma
- Presentations: 3
- Moderators:
- Coordinates: 10/16/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P1.09-006 - JMJ and BRD Domain Family Members in Malignant Pleural Mesothelioma: Potential Therapeutic Targets or Not? (ID 9919)
09:30 - 09:30 | Author(s): Michaela B Kirschner
- Abstract
Background:
Malignant pleural mesothelioma (MPM) is an aggressive rare cancer affecting the pleura and is predominantly associated with prior exposure to asbestos. Treatment options are limited, and most patients die within 24 months of diagnosis. There is an urgent unmet need to identify new therapeutic options for the treatment of MPM. Asbestos fibres contain transition metals such as iron, and 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. Bromodomain containing proteins a family of chromatin reader proteins, have potential therapeutic efficacy against various malignancies. Long non-coding RNAs (lncRNAs) have also been shown to play a role as oncogenic molecules in different cancers. Several such lncRNAs have now been shown to locate to the same chromosomal region as various KDMs. We therefore examined the expression of various JmjC and Brd members (along with any associated lncRNAs) in MPM and assessed some for their clinical potential using existing small molecule inhibitors.
Method:
A panel of MPM cell lines and a cohort of snap-frozen patient samples isolated at surgery comprising benign, epithelial, biphasic, and sarcomatoid histologies were screened for expression of various BRD and JmjC members and associated lncRNAs by RT-PCR. IHC for KDM4A was performed 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.
Result:
The expression of the various KDMs was detectable across our panel of cell lines. In primary tumours the expression of many of these genes 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 various KDMs, BRD genes and associated lncRNAs are significantly altered in MPM. Small molecule inhibitors directed against these 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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P1.09-008 - A 4-microRNA Signature in Serum Can Discriminate Between Non-Small-Cell Lung Cancer and Malignant Pleural Mesothelioma (ID 9956)
09:30 - 09:30 | Presenting Author(s): Michaela B Kirschner
- Abstract
Background:
Differential diagnosis of malignant pleural mesothelioma (MPM) and non-small cell lung cancer (NSCLC) can be difficult. For both malignancies various studies have in recent years investigated circulating cell-free microRNAs (miRs) as potential diagnostic markers, with most of these focusing on NSCLC. One of the most recent studies has suggested a 4-miR signature consisting of miR-141, miR-200b, miR-193b and miR-301 in serum to be specific for NSCLC patients. Here, we investigated the value of this 4-miR signature in discriminating serum samples from NSCLC and MPM patients.
Method:
RNA was extracted from a series of serum samples from 98 NSCLC, 98 MPM and 96 healthy controls collected at the Netherlands Cancer Institute between 1995 and 2011. MicroRNA-specific TaqMan assays were used to quantify serum microRNA levels in the t groups which after initial quality control consisted of 65, 68 and 58 samples in the NSCLC, MPM and control groups, respectively. Expression levels of individual microRNAs between the different groups were analysed using one-way ANOVA with Tukey-Kramer Posthoc Test. Binary logistic regression modelling was used to generate the 4-miR-signature, for which accuracy in discriminating NSCLC from MPM or healthy was analysed by ROC curve analysis.
Result:
Analysis of the signature microRNAs showed for all 4 miRs trends towards higher abundance in serum from NSCLC patients. Statistical significance was however only reached for miR-141, which was found to be increased by 4.4-fold in NSCLC compared to healthy controls (p=0.014) and by 5.6-fold in NSCLC vs MPM (p=0.004). Although we did not observe significant differences in abundance for all microRNAs, ROC curve analysis of the 4-miR signature confirmed the discriminatory potential with an AUC 0.73 (95% CI: 0.62-0.85) for NSCLC vs healthy controls. When applying the best achievable Youden Index as cut-off point, the signature showed a sensitivity of 91.4% and a specificity of 44.4%. In addition to being able to discriminate NSCLC from healthy controls, the 4-miR-signature also proved to be valuable for discriminating NSCLC from MPM, where an AUC of 0.77 (95% CI: 0.66-0.89), a sensitivity of 74.3% and a specificity of 80.4% could be observed.
Conclusion:
Initial analyses have confirmed the diagnostic potential of the previously described NSCLC-specific serum-based microRNA signature for distinguishing NSCLC from healthy controls. In addition, we have shown that the same signature can also discriminate between NSCLC and MPM.
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P1.09-009 - Evaluation of a Combined MicroRNA-Clinical Score as Prognostic Factor for Malignant Pleural Mesothelioma (ID 9245)
09:30 - 09:30 | Presenting Author(s): Michaela B Kirschner
- Abstract
Background:
In 2015, a 6-microRNA signature (miR-Score, Kirschner et al 2015) was demonstrated to show high prognostic accuracy in a series of surgical specimens (with and without induction chemotherapy) from patients with malignant pleural mesothelioma. In-depth analysis of matching pre- and post-chemotherapy tissue specimens has recently shown that a refined 2-miR-Score appears more suitable for use in diagnostic chemo-naïve specimens (Kirschner et al, WCLC 2016). Here, in addition to continued validation, we also aimed to further improve the prognostic accuracy by combining the 2-miR-Score with known clinical prognostic factors.
Method:
Binary logistic regression modelling was used to build a combined score consisting of the 2-miR-Score and the clinical prognostic factors age (<60 years vs >60 years at diagnosis), gender and histological subtype (epithelioid vs non-epithelioid). In addition, microRNA analysis (RT-qPCR) was performed in an additional 33 pairs of chemo-naïve (diagnostic biopsy) and chemo-treated (EPP) specimens. Accuracy of the investigated scores in predicting a good prognosis (>20 months survival post-surgery) was evaluated by ROC curve analysis.
Result:
Combining the refined 2-miR-Score with the clinical prognostic factors histological subtype and age at diagnosis, increased the overall accuracy of the 2-miR-Score in both chemo-naïve diagnostic (AUC=0.80; 95% CI: 0.65-0.95) and post-chemotherapy (AUC=0.86; 95% CI: 0.73-0.98) specimens. Addition of gender as clinical prognostic factor, did not result in further increases, hence this factor was not included in the combined score. Investigation of an additional set of 33 matched pairs of chemo-naïve and post-chemotherapy tissue samples, confirmed the improved prognostic accuracy of the combined score, with AUCs of 0.76 (95% CI: 0.59-0.92) and 0.79 (95% CI: 0.64-0.95) for chemo-naïve and post-Chemotherapy specimens, respectively. Furthermore, addition of the clinical factors resulted in an increase in specificity of the prognostic score from previously 55-65% to now 65-75%, while keeping sensitivities at the previous levels of 75-85%. Importantly, the combined microRNA-clinical Score did not only outperform the 2-miR-Score, but also the clinical factors alone.
Conclusion:
This validation has confirmed the prognostic potential of the novel 2-miR-Score. Furthermore, addition of known clinical prognostic factors was shown to result in a combined Score with increased prognostic accuracy. In addition to continued validation, in currently ongoing analyses we are also investigating combining the 2-miR-Score with our previously proposed multimodality prognostic score (MMPS; Opitz et al 2015).
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P3.09 - Mesothelioma (ID 725)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Mesothelioma
- Presentations: 1
- Moderators:
- Coordinates: 10/18/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P3.09-010 - 18 Years Single Center Experience of Surgical Resection of Malignant Pleural Mesothelioma After Induction Chemotherapy (ID 10010)
09:30 - 09:30 | Presenting Author(s): Michaela B Kirschner
- Abstract
Background:
Surgical resection of malignant pleural mesothelioma is discussed controversially. Using our data from nearly 2 decades of single center experience, and focusing on the shift from extrapleural pneumonectomy (EPP) to (extended) pleurectomy/decortication ((e)P/D) we compared the peri- and longterm outcomes of EPP and (e)P/D after induction chemotherapy.
Method:
In a retrospective analysis (September 1999 - June 2016) of our prospective database of mesothelioma patients 196 patients received mentioned multimodality therapy: 149 treated with EPP, 34 with eP/D and 13 with P/D. Major morbidity was defined as bleeding necessitating reoperation, patch failure, chylothorax, empyema, bronchopleural fistula (BPF), pulmonary embolism and acute respiratory distress syndrome (ARDS).
Result:
Both groups did not differ significantly in pT stage, hisotype, but in age and lymph node stadium. Overall 30-day and 90-day mortality were 4% and 8%, respectively. However, patients treated with (e)P/D the 30- and 90-day mortality was 0. Major morbidity was not significantly different between both groups with 37% (EPP) and 23% ((e)P/D), respectively. Patient’s characteristics, freedom from recurrence (FFR) and overall survival (OS) are demonstrated in figure 1.Figure 1
Conclusion:
Multimodality treatment with radical surgery is perfromed safely, (e)P/D known as the less invasive procedure than EPP, shows a longer OS whilst having a shorter FFR.
