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S. Ryan
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P1.04 - Poster Session/ Biology, Pathology, and Molecular Testing (ID 233)
- Event: WCLC 2015
- Type: Poster
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 2
- Moderators:
- Coordinates: 9/07/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P1.04-088 - Lung Cancer Cells Can Alter the Behaviour of Normal Bronchial Epithelial Cells Through Multiple Mechanisms (ID 1312)
09:30 - 09:30 | Author(s): S. Ryan
- Abstract
Background:
Lung cancer is one of the most heterogeneous of all solid cancers. This may in part be due to hi-jacking and additional bystander affects that are exerted on the normal lung cell population by the cancer cells. A number of pathways may be stimulated through soluble factors or effector filled vesicles such as exosomes secreted by cancer cells. The aim of this project was to evaluate the effects of non-small cell lung cancer (NSCLC) cells on an immortalised normal bronchial epithelial cell line.
Methods:
A normal bronchial epithelial cell line (HBEC4) was exposed to adenocarcinoma, large cell and squamous NSCLC cell lines and a number of phenotypic and genotypic characterisations were undertaken. These included cellular proliferation (BrdU ELISA), gene (RT-PCR) and miRNA expression screening (Nanostring). The effect of cancer exosome fractions was also determined.
Results:
Exposure to various subtypes of NSCLC significantly increased the cellular proliferation rate of the immortalised cell line in a number of models. Expression of a number of miRNAs were altered in the normal cells pre- and post exposure to the cancer cells. Various stem cell factor markers (KLF4, Oct, c-myc) were also significantly changed at the mRNA level. In addition, exosome fractions altered the behaviour of the normal cell line, likewise stimulating cell proliferation.
Conclusion:
Lung cancer cells may influence normal cell behaviour in both a direct and indirect manner using multiple mechanisms. Normal bronchial epithelial cells with stem like features may be induced to proliferate and behave in a malignant manner. This, akin to Hodgkin’s lymphoma, may contribute significantly to the composition of the tumour. Furthermore this observation may contribute to the heterogeneity of lung cancer tumours and affect treatment response. Ongoing studies are evaluating these effects in novel 2D and 3D culture systems.
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P1.04-105 - The Development and Assessment of Advanced Cellular Models for the Study of Non-Small Cell Lung Cancer (ID 1335)
09:30 - 09:30 | Author(s): S. Ryan
- Abstract
Background:
The key mechanisms that underlie drug resistance in lung cancer have yet to be fully elucidated. A significant limiting factor is the lack of biologically relevant cellular models for basic laboratory research. To address these issues, many are now turning to three-dimensional (3D) based cellular assay systems that permit the formation of multicellular structures such as tumour spheroids. Depending on their size the internal microenvironment of these structures mimics closely that of those in vivo. In the majority of cases, spheroids with a diameter greater than 100µm exhibit an asymmetry in cellular proliferation and viability - proliferating tumour cells at the periphery; cell-cycle arrested cells at larger distances from the surface. Regions of necrosis associated with reduced oxygen tension and hypoxia have often been reported. This study compared drug resistant models of non-small cell lung cancer (NSCLC) in 3D culture with those in grown in two-dimensional (2D) culture. The behaviour of cells grown in these distinct geometric configurations was monitored and compared by measuring viability, proliferation and oxygen tensions.
Methods:
Happy Cell Advanced Suspension Medium[™] (ASM) was chosen to culture our 3D spheroids. This polymer-based formulation was selected for its ease of use, as well as its compliance with liquid handling, high content imaging and analysis (HCSA) and high throughput screening (HTS) systems. Isogenic NSCLC cell line models of cisplatin resistance were cultured in 2D and 3D cell culture systems. Cisplatin sensitive (Pt) and isogenic cisplatin resistant (CisR) NSCLC sub-types were studied. IC50 values were calculated and a positive control was selected. All cultures were grown in a range of cisplatin concentrations for 72 hours. Subsequently, viability and hypoxia assays were conducted in order to compare the response of Pt and CisR cells in both 2D and 3D culture systems. Morphological analysis was performed via high content analysis (HCA) and confocal microscopy.
Results:
At equivalent cisplatin concentrations 3D spheroids exhibit greater resistance compared with monolayers. Imaging experiments have shown that these 3D structures have a central necrotic core, a feature of the asymmetric growth patterns associated with 3D structures.
Conclusion:
Happy Cell ASM is a novel 3D culture medium for generating multicellular tumour spheroids and has potential for HTS and HCSA. When treated with cisplatin our spheroids exhibited resistance to therapy compared to 2D monolayer cultures. These results suggest that spheroids may provide a more accurate in vitro model to elucidate mechanisms of drug resistance and may aid the identification of novel targets to re-sensitise patient therapy.
