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Sonja Klebe
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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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P1.09-003 - Malignant Mesothelioma Versus Synovial Sarcoma: An Analysis of 19 Cases with Molecular Diagnosis (ID 9390)
09:30 - 09:30 | Presenting Author(s): Sonja Klebe
- Abstract
Background:
Intrathoracic synovial sarcomas (SSas) are well documented in the literature and characterized by a distinctive t(X;18) translocation. The histologic appearances of a monophasic or biphasic SSa can lead to confusion with biphasic or sarcomatoid mesothelioma (MM). The distinction of pleural SSa from pleural MM is important, because SSas may be responsive to ifosfamide-based chemotherapy and have no proven causal relationship to prior asbestos exposure. Demonstration of the t(X;18) by cytogenetics, fluorescence in situ hybridization (FISH) or reverse-transcriptase polymerase chain reaction is the gold standard for diagnosis, but availability of molecular diagnosis can be limited and testing is time consuming. Recently, it has been suggested that immunohistochemistry (IHC) for transducin-like enhancer of split 1 (TLE) is reliable for diagnosis of SSa and may replace molecular diagnosis.
Method:
We reviewed 19 pleura-based malignancies that had either been referred for a potential diagnosis of SSa, or where SSa was a differential diagnosis considered by the authors, based on morphology. Only cases with molecular diagnostics and clinical follow-up and blocks or unstained slides for further IHC studies are included.
Result:
Fourteen (14/19) cases were diagnosed as MM with morphology indistinguishable from SSa, based on lack of the t(X;18) by FISH and/or PCR, whereas 5 cases were diagnosed as SSa based on molecular diagnostics in conjunction with morphology. The mean age at diagnosis was 40.6 and 70.35 years for SSa and MM respectively. In the MM group, 21% of the patients were female, compared to 80% in the SSa group. Median survival after diagnosis was 9 months for MM, whereas all of the SSa patients were alive after follow-ups ranging from 3 months to 21 years. On average, MMs were larger tumours (average size of 97 mm, ranging from 20 to 220 mm), compared to 37 mm (range 20-50 mm) in SSa. Pleural plaques were present in 9 of the MM patients, with no information on plaques for 4 patients, and with 1 patient not having plaques, whereas only one of the SSa patients was confirmed as having pleural plaques. Of note, 7 of the MMs showed positive labelling for TLE1, with 7 MM not showing labelling, whereas all 5 SSas showed positive labelling.
Conclusion:
This indicates that positive labeling for TLE1 in isolation is insufficient for discrimination between MM vs SSa in pleura-based lesions with SSa-like morphology, and that molecular studies remain the gold standard for diagnosis.
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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): Sonja Klebe
- 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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P1.09-012 - A Pre-Clinical Investigation of Intrapleural Curcumin Treatments as an Adjunct Therapy for Malignant Pleural Mesothelioma (ID 9312)
09:30 - 09:30 | Author(s): Sonja Klebe
- Abstract
Background:
Malignant pleural mesothelioma (MPM) is an aggressive malignancy originating in pleural mesothelial cells with median survivals of approximately 12 months following diagnosis. Recently, anti-angiogenic therapies have been trialled with only a modest effect. This may be, in part, due to alternative mechanisms of tumour vascularisation such as vasculogenic mimicry (VM), the ability of tumour cells to form fluid carrying vascular channels. Curcumin, a polyphenol extracted from the spice turmeric, has numerous anti-cancer and anti-inflammatory properties. Our aims were to investigate the effect of curcumin on vasculogenic mimicry and to determine if curcumin acts by disrupting microRNA profiles of mesothelioma cells. In preparation for future clinical trials, we evaluated the safety of curcumin treatments in vivo, when applied to the pleural cavity.
Method:
Mesothelioma cell lines NCl-H226 and NCI-H28, as well as patient-derived primary mesothelioma cells isolated from pleural effusions, were used for in vitro experiments. Matrigel tube formation assays were performed to assess if curcumin could inhibit VM in vitro. Small RNAseq was performed to determine if 6 h curcumin (20 mM) treatments had an effect on microRNA expression. Curcumin (80 mg/kg) was injected into the pleural cavity of Fischer 344 rats (n=6) and blood was taken at 1.5 h, 24 h, 48 h, 7 days, 14 days and 21 days. Rats were euthanized at 48 h, 1 week and 3 weeks (n=2). Parietal pleura, lung, kidney, liver brain and heart tissues were obtained and examined for signs of gross tissue damage and histopathological changes such as inflammation, and necrosis. Curcumin plasma and concentrations were measured using UPLC-MS to determine systemic distribution of curcumin following intrapleural treatments.
Result:
Non-cytotoxic curcumin treatments (20-10 mM) significantly inhibited the ability of mesothelioma cells to perform vasculogenic mimicry in vitro in a dose dependent manner. The microRNA expression profiles differed greatly between each mesothelioma sub-type. Minimal curcumin-induced change was observed, however differential expression analysis revealed some potential microRNA targets. No adverse effects were observed following intrapleural curcumin administration. Encapsulated curcumin deposits were observed in the pleural cavity of rats at 1 and 3 weeks following curcumin administration. Histological analysis revealed focal reactive mesothelial hyperplasia and a histiocytic response towards curcumin. Lung, liver, heart, brain and kidney tissues all display normal histological appearances. Curcumin was detected in the plasma samples of rats receiving intrapleural curcumin with peak concentration observed at 1.5 h post curcumin treatment (387-100 µg/ml).
Conclusion:
Intrapleural curcumin treatments may be suitable as adjunct treatment for MPM.
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P3.02 - Biology/Pathology (ID 620)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Biology/Pathology
- Presentations: 1
- Moderators:
- Coordinates: 10/18/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P3.02-078 - Establishing Malignant Pleural Mesothelioma Primary Cell Lines Using the 3D Spheroid Method Produces a Model with Better Tumour Architecture (ID 10456)
09:30 - 09:30 | Author(s): Sonja Klebe
- Abstract
Background:
Malignant pleural mesothelioma (MPM) is an aggressive malignancy with no effective treatment options. Poor prognosis and drug resistance are the main challenges of this deadly disease. There is also no simple distinctive diagnosis tool for identification of MPM. Better diagnostic markers may also provide better biological information for newer treatment option development. In this study we have established primary MPM cell lines and characterised them with current biomarkers. Our ultimately goal is to use these cell lines for better identification of diagnostic biomarkers.
Method:
MPM cell lines were either established from tissue specimens or pleural effusion from patients with pathologically confirmed MPM. Cells were cultured in standard (2D) and spheroid (3D) versions for characterisation. Cells prepared in 2D and 3D were stained with H&E and analysed with a diagnostic biomarker panel (CK-8/18, Calretinin, CK5/6, CD141, WT-1, D2-40, EMA, CEA, Tag-72, BG8, CD15, TTF-1 and BAP1). Scoring and comments were provided by pathologists experienced in MPM diagnosis (KL, SK). Established cell lines were also analysed for ploidy (flow cytometry) and interphase (fluorescent microscope) for chromosome number. PBMC from healthy donor was used as a control diploid.
Result:
We successfully established nine cell lines from MPM patient specimens. The original tumour histological sub-types were: three epithelioid, four biphasic, one desmoplastic and one not otherwise specified. Cells grown in 3D with H&E staining revealed better tumour architecture, cell-cell contacts and morphology when compared to cells grown in standard 2D culture. Mesothelioma positive markers were more distinctive and intense in biphasic cell lines grown 3D culture. Other sub-types showed similar staining when grown in both formats. Results from ploidy showed no distinctive difference between sub-types, however, 5 out of 9 cell lines established had tetraploid chromosome content.
Conclusion:
Cells grown in 3D provide more tumour architecture when compared with 2D cells. 3D cells also provide more intensity and greater percentage of positive MPM markers
