Abstract A14: Changes in miRNA profile in a murine model of lung adenocarcinoma are attenuated by increased prostacyclin levels
Authors: New Melissa L. , Tennis Meredith A. , Nield Lori D. , McArthur Debbie , Keith Robert L. 
Published: February 2019

Rationale: Lung cancer has a high mortality even when it is diagnosed at an early stage. Intercepting these early changes using chemoprevention could dramatically improve survival. Increased prostacyclin provides effective chemoprevention in multiple murine models and its oral analogue, iloprost, increased regression of dysplastic lesions in former smokers in clinical trials. MicroRNAs are short, noncoding RNA that are important post-transcriptional regulators during carcinogenesis. miRNA stability makes them attractive as potential biomarkers of premalignancy and response to chemoprevention. We hypothesize that alterations in the miRNA profile in early tumors and at-risk lung tissue are attenuated by increased levels of prostacyclin.

Methods: Wild-type and transgenic prostacyclin synthase overexpressing (PGIS-OE) FVB/N mice were sacrificed at time = 0 as well as 8 and 16 weeks after intraperitoneal injection with urethane, which is known to induce Kras mutations resulting in adenocarcinoma. Transgenic mice and wild-type littermates were sacrificed at multiple timepoints (baseline, 8 weeks and 16 weeks post-urethane). Visible tumors were dissected and separated from remaining lung tissue. Tissue was lysed and homogenized and total RNA was extracted using the QIAGEN AllPrep DNA/RNA/miRNA Universal Kit. Samples from time = 0 and 16 weeks were analyzed using the Nanostring nCounter analysis system, which analyzed 550 murine miRNAs per sample. Quantitative real-time PCR (qRT-PCR) was performed using primers from QIAGEN.

Results: miRNAs from lungs of naive WT and PGIS-OE mice were compared using Nanostring nCounter, without significant differences. From the 16-week group, miRNAs from tumors were compared with uninvolved lung tissue, with significant changes identified in over 80% of miRNAs tested. These data were cross-referenced with published human data, and a subset of biologically relevant miRNAs was chosen for further investigation with qRT-PCR in the earlier, 8-week timepoint: miR-143 (fold change=1.84), miR-146b (1.41), miR-154 (1.29), miR-410 (0.57), miR-429 (2.01), miR-451 (0.47). The miRNA fold-changes observed in the urethane-treated WT mice at 8 weeks were attenuated in the PGIS-OE mice.

Conclusions: There are important changes to the miRNA profile in adenomas (early tumors) at 16 weeks, and these miRNAs are aberrant in the lungs at an even earlier time point when tumors have not yet developed (8 weeks). The changes to the miRNA profile in early carcinogenesis are attenuated by the presence of increased prostacyclin, even though prostacyclin does not cause significant changes to the miRNA profile at baseline. Ongoing investigation includes the evaluation of additional miRNAs in these samples, some from later time points (24 weeks), in cancer cell lines exposed to iloprost and in bronchial epithelial cells from a clinical trial of high-risk subjects treated with inhaled iloprost.

Citation Format: Melissa L. New, Meredith A. Tennis, Lori D. Nield, Debbie McArthur, Robert L. Keith. Changes in miRNA profile in a murine model of lung adenocarcinoma are attenuated by increased prostacyclin levels [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr A14.