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D. Richard
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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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P1.04-076 - CDCA3 Is a Novel Cell Cycle Regulator in Lung Cancer (ID 1630)
09:30 - 09:30 | Author(s): D. Richard
- Abstract
Background:
Progression through the mammalian cell cycle relies upon coordination of a complex network of proteins. Following genomic insult, checkpoints during each stage of the cell cycle are engaged to halt cell cycle progression to allow faithful DNA repair. Failure to arrest cell cycle may lead to genomic instability and cancer development. However, the molecular basis for the loss of genome integrity during cancer development remains to be determined. Cell division cycle associated 3 (CDCA3) is a key regulator of the normal cell cycle. CDCA3 modulates this process by enabling cell entry into mitosis through degradation of the mitosis-inhibitory factor WEE1. CDCA3 itself is also degraded in G1 yet re-expressed in G2/M phase, to allow successful progression through the cell cycle. Here we describe for the first time a novel function for CDCA3 in maintaining effective cell cycle progression in lung cancer.
Methods:
To examine the role of CDCA3 in modulating the cell cycle of lung cancer cells, CDCA3 was depleted using an siRNA approach in A549, SKMES and H460 cell lines. CDCA3 depletion was assessed using Western blot analysis. Cell proliferation assays were performed on control and CDCA3 knockdown cells over a period of 96 h using the Promega CellTitre-Glo cell viability assay. Cell cycle progression was assessed on propidium iodide stained cells using a Beckman Coulter Gallios flow cytometer. To determine if CDCA3 expression is associated with lung cancer progression, a tissue microarray (TMA) with cores from 600 patients was stained with an anti-CDCA3 antibody. Correlation of CDCA3 staining with clinical data and patient prognosis is ongoing.
Results:
As a cell cycle related protein, we tested if CDCA3 is required for effective proliferation of a range of lung cancer cell lines. CDCA3 depletion reduced the proliferation of U2OS (osteosarcoma), A549, MOR (lung adenomcarcenoma) and SKMES cancer cells (squamous lung cancer). Interestingly, depletion of CDCA3 did not affect proliferation of H460 cells (large neuroendocrine lung cancer). We next tested the cell cycle progression and noted that knockdown of CDCA3 induced an increase of all cell lines in G1 arrest with the exception of H460 cells. To observe if CDCA3 expression is linked with disease progression, TMA staining of lung cancer biopsies was performed. Accordingly, elevated expression of CDCA3 was identified specifically in tumour cells. These data highlight the potential prognostic value of CDCA3 expression.
Conclusion:
Our data point to a potential role for CDCA3 in the progression of lung cancer. While the precise mechanism for CDCA3-dependent cell cycle regulation remains unknown, it is possible that elevated CDCA3 levels modulate tumour cell proliferation. Identifying the molecular basis may yield novel therapeutic avenues worth targeting during aberrant cell cycle in cancer.
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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): D. Richard
- 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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P2.04 - Poster Session/ Biology, Pathology, and Molecular Testing (ID 234)
- Event: WCLC 2015
- Type: Poster
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
- Moderators:
- Coordinates: 9/08/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P2.04-102 - Targeting Inflammatory Mediators to Overcome Intrinsic and Acquired Cisplatin Resistance in Non-Small Cell Lung Cancer (ID 1314)
09:30 - 09:30 | Author(s): D. Richard
- Abstract
Background:
Cisplatin based doublet-chemotherapy is commonly used in non-small cell lung cancer (NSCLC) treatment with an initial objective response rate of 40-50%. However, intrinsic and acquired chemo-resistance constitutes a major clinical obstacle. The mechanisms of resistance have yet to be fully understood. We have previously demonstrated that NF-κB levels are elevated in cisplatin resistant cells (CisR) and that the use of an NF-κB inhibitor, DHMEQ, resulted in greater CisR cell death. The goal of this project is to elucidate the mechanistic links between NF-κB regulated pathways and the development of cisplatin resistant NSCLC.
Methods:
The expression of NF-κB mediators and immune regulators were assessed in an isogenic NSCLC cell line model of cisplatin resistance using qPCR arrays (252 genes). A number of targets were identified and validated using PCR. The effect of drug combinations (Cisplatin and DHMEQ) was also determined. Comet assays (DNA damage) were also performed to determine the effect of DHMEQ alone or in combination with irradiation (6 Gy).
Results:
Various chemokines and their receptors were elevated in cisplatin resistant (CisR) cells compared with cisplatin sensitive (PT). In addition, a number of key TLRs and regulators of the innate immune pathway were altered. DHMEQ enhanced cellular sensitivity to cisplatin in both PT and CisR cell lines (p<0.05). This drug also overcame the chemo-protective effect of a number of chemokines and enhanced irradiation induced DNA damage. An animal study will commence shortly using DHMEQ alone and in combination with cisplatin.
Conclusion:
Immune-modulators such as DHMEQ may be a novel viable option in addressing inflammatory mediated acquired and intrinsic NSCLC chemo-resistance. In addition, immune regulators identified in this project may provide innovative targets for immuno-oncology therapy.
