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N. Leonard
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MINI 34 - RNA and miRNA (ID 162)
- Event: WCLC 2015
- Type: Mini Oral
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
- Moderators:C. Mascaux, L.(. Wang
- Coordinates: 9/09/2015, 18:30 - 20:00, 205+207
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MINI34.07 - A Novel microRNA Signature Associated with Cisplatin Resistance in NSCLC (ID 2709)
18:30 - 20:00 | Author(s): N. Leonard
- Abstract
- Presentation
Background:
MicroRNAs (miRNAs) are an abundant class of small non-coding RNAs that range in size from 19 to 25 nucleotides. Alteration in miRNA expression can cause them to act as either tumour suppressor or oncogenes. They have also been shown to regulate a number of processes involved in tumour biology such as metastasis, invasion and angiogenesis. More recently, miRNAs have been linked to chemo- and radio-resistance in many solid tumours, including lung cancer.
Methods:
An isogenic model of cisplatin resistance was established by chronically exposing a panel of NSCLC cell lines (MOR, H460, A549, SKMES-1, H1299) to cisplatin for 12 months, generating cisplatin resistant (CisR) sublines from their corresponding age-matched parental (PT) cells. MicroRNA expression profiling was carried out using 7th generation miRCURY LNA™ microRNA arrays consisting of 1,919 miRNAs (Exiqon). MicroRNAs that were significantly increased in CisR sublines were inhibited using antagomirs (Exiqon), while those that were significantly decreased were over-expressed using pre-miRs (Ambion). Functional studies examining clonogenic survival ability, proliferation (BrdU) and apoptosis (Annexin V/PI) were subsequently carried out in the presence or absence of cisplatin. To examine the translational relevance of these microRNAs, their expression was further examined in a cohort of pre-treatment matched normal and tumour lung tissues from NSCLC patients of different histological subtypes. Validation of this miRNA signature is currently being investigated in serum samples from this same cohort of patients and normal controls.
Results:
MicroRNA profiling analysis identified ten miRNAs which were significantly altered between parental and corresponding cisplatin resistant lung cancer cell lines. Validation of these miRNAs by real-time PCR (qPCR) identified a specific 5-miR signature that was significantly altered in CisR cells relative to their parental counterparts. Modification of these microRNAs altered the response of resistant cells to the cytotoxic effects of cisplatin and decreased the clonogenic survival of CisR cells when treated with increasing doses of cisplatin (0.1µM-10μM). Significant differential expression was found between normal and tumour tissues across each histological subtype, highlighting the potential use of these microRNAs as markers of response to cisplatin therapy in NSCLC patients. Three miRNAs (miR-A, B, C) belonging to the same family were significantly altered in tumour lung tissue of adenocarcinoma and squamous cell histology compared to matched normal lung tissue. MicroRNA-D expression was significantly altered in squamous cell carcinomas while miR-E was differentially expressed in adenocarcinomas only. Data relating to the expression of this novel signature in the circulation of our NSCLC patient cohort and normal controls will be presented at WCLC 2015.
Conclusion:
We have identified and validated a novel miRNA signature associated with cisplatin resistance in a panel of cisplatin resistant cell lines and in patient lung tumours. Genetic manipulation of these specific miRNAs in vitro altered the cisplatin resistant cell response to the cytotoxic effects of cisplatin chemotherapy. The data obtained from this study may provide a basis for the potential development of a companion diagnostic for lung cancer patients who are most likely to benefit, or not, from cisplatin chemotherapy.
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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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P2.04-057 - Targeting PIM Kinase in NSCLC (ID 933)
09:30 - 17:00 | Author(s): N. Leonard
- Abstract
Background:
PIM proteins belong to a family of serine/threonine kinases composed of 3 isoforms, PIM1, PIM2 and PIM3, that play a key role in cell cycle regulation, have potent anti-apoptotic activity and play a role in the homing and migration of metastatic cells. Furthermore, PIM kinases have also been shown to be activated in response to Akt pathway inhibition, indicating a role in adaptive responses to inhibition of this pathway potentially leading to treatment resistance. Thus, there is a strong rationale for combining PIM kinase inhibition with inhibition of the Akt pathway (i.e., inhibitors of EGFR, PI3K, Akt and mTOR). PIM kinase has been recognised as a therapeutic target particularly in haematological malignancies however the role of PIM kinases in solid tumours and NSCLC in particular are less well characterised. This study is the first to elucidate the expression of all 3 PIM isoforms in NSCLC cell lines and patient tumours as well as to examine the effect of Inflection Bioscience Ltd novel dual PI3K/PIM kinase (IBL-202) and triple PI3K/mTOR/PIM kinase (IBL-301) targeted therapies in-vitro and in-vivo.
Methods:
PIM 1/2/3 protein expression was quantified by western blot analysis in a panel of NSCLC cell lines and 40 matched normal/tumour tissues from NSCLC patients (20 adenocarcinoma and 20 squamous cell carcinoma). PIM kinase expression was correlated to patient clinicopathological characteristics and survival data. The effectiveness of IBL-202 and IBL-301 on proliferation and apoptosis in NSCLC cell lines were examined by BrdU and Annexin V/PI FACS analysis, respectively. A head-to-head in-vivo study of IBL-202 vs. IBL-301 in xenograft nude mice formed using H1975 cells is ongoing.
Results:
All 3 isoforms of PIM kinase are highly expressed across a panel of NSCLC cell lines. PIM kinase is expressed in ~ 90% of NSCLC tumour tissues across all stages of the disease. IBL-202 and IBL-301 induced apoptosis and decreased cell proliferation in NSCLC cell lines at micromolar concentrations in-vitro. The in-vivo study is ongoing and results will be presented.
Conclusion:
PIM kinase is a promising new therapeutic target for the treatment of NSCLC patients. Dual PI3K/PIM kinase (IBL-202) and triple PI3K/mTOR/PIM kinase (IBL-301) targeted therapies have demonstrated pro-apoptotic and anti-proliferative activity in-vitro and in-vivo and should be considered in the treatment of NSCLC patients.
