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Greg L Stewart



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    P1.03 - Biology (ID 161)

    • Event: WCLC 2019
    • Type: Poster Viewing in the Exhibit Hall
    • Track: Biology
    • Presentations: 1
    • Moderators:
    • Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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      P1.03-29 - Cellular Biology Determines Ability of XIST to Act as a miRNA Sponge in Non-Small Cell Lung Cancer (ID 2631)

      09:45 - 18:00  |  Presenting Author(s): Greg L Stewart

      • Abstract
      • Slides

      Background

      XIST, the female-specific lncRNA canonically involved in silencing the X chromosome, has been suggested by many studies to function as a miRNA sponge. This sponge mechanism has been suggested to affect many cancer types, including lung adenocarcinoma (LUAD). However, determination of miRNA:lncRNA interaction is largely based on sequence homology, impeding the identification of functional gene pairs. The sex-specificity of XIST provides the opportunity to study the biological relevance of these interactions. Here we take a comprehensive approach by considering factors that affect preferential regulation through XIST-miRNA sponging in LUAD.

      Method

      Sequencing data from 568 LUAD and non-malignant samples (304 female and 264 male) were downloaded from CancerBrowser and processed to provide gene expression for mRNAs, miRNAs, and XIST. 3’UTR sequences of all candidate genes were run through the miRanda binding prediction algorithm. Spearman’s tests were performed to identify positively correlated candidate sponged genes.

      Result

      To identify the best sponge-regulated candidates, we considered genes that (1) were positively correlated with XIST expression and (2) are targeted by shared miRNA with XIST, and (3) expressed in lung adenocarcinoma. This revealed a robust set 128 of genes potentially positively regulated by XIST through the sequestration of 804 shared miRNAs. As XIST is sex-specific, we compared the changes in miRNA-target gene relationships in XIST-high and XIST-low systems to identify a high-confidence set of 13 miRNA-gene pairs. In order to interact with the exclusively nuclear expressed XIST,miRNAs must also be expressed in the nucleus. We validate the nuclear presence of several of these high confidence miRNAs using RT-qPCR, confirming the co-localization required for XIST to interact with these species.

      Conclusion

      We use a biology-driven approach to identify genes defended from miRNA-based inhibition by the lncRNA XIST in LUAD. Importantly, we identify that the miRNAs that mediate the XIST-target gene axis are enriched in the nucleus, co-localizing with XIST in lung cancer cell models. Our results reinforce the necessary consideration of biological features in future studies of lncRNA:miRNA interactions.

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    P2.03 - Biology (ID 162)

    • Event: WCLC 2019
    • Type: Poster Viewing in the Exhibit Hall
    • Track: Biology
    • Presentations: 1
    • Moderators:
    • Coordinates: 9/09/2019, 10:15 - 18:15, Exhibit Hall
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      P2.03-47 - Deregulation of a Novel Cis-Acting lncRNA in Non-Small Cell Lung Cancer May Control HMGA1 Expression (ID 2727)

      10:15 - 18:15  |  Presenting Author(s): Greg L Stewart

      • Abstract
      • Slides

      Background

      Since the discovery of long non-coding RNAs (lncRNAs), they have been increasingly implicated in cancer-associated phenotypes. Recently, some lncRNAs have been shown to regulate the expression of neighbouring protein-coding genes, including oncogenes and tumour suppressor genes. High mobility group A1 (HMGA1) is aberrantly expressed in several aggressive cancer types, including non-small cell lung cancer (NSCLC), where high HMGA1 expression has been associated with poor overall survival and chemotherapy resistance. While HMGA1 is known to be deregulated in lung cancer, the mechanisms that mediate its expression remain unknown. These lncRNAs, known as cis-acting, may represent undiscovered therapeutic action points in cancer driving pathways.

      Here we investigate the deregulation of a putative cis-acting lncRNA in NSCLC, and it’s relationship with the oncogene HMGA1.

      Method

      LncRNA expression was generated from RNA-sequencing data from 36 microdissected tumour and matched non-malignant tissues. Normalized sequence read counts were used to identify transcripts with significantly deregulated expression (Wilcoxon Signed-Rank Test, BH-p<0.05). Validation was performed in sequencing data obtained from The Cancer Genome Atlas (TCGA). SiRNA-mediated knockdown of lncRNA candidates were performed in a non-malignant epithelial lung cell line (BEAS-2B). Quantitative real-time PCR was used to observe the effects of lncRNA knockdown on the expression of neighbouring protein-coding genes.

      Result

      Our analyses identified a lncRNA neighbour to HMGA1, RP11.513I15.6, to be significantly downregulated in 2 cohorts of LUAD samples. Conversely, we found HMGA1 expression to be significantly overexpressed in LUAD tumours, and was found to be anticorrelated with RP11.513I15.6. Additionally while RP11.513I15.6 decreased with tumour stage, HMGA1 expression increased with stage. In vitro experiments demonstrated siRNA-mediated inhibition of RP11.513I15.6 in immortalized lung epithelial cells resulted in a significant increase in HMGA1 expression.

      Conclusion

      Our results suggest that RP11.513I15.6 is a novel cis-acting lncRNA that negatively regulates HMGA1, and may contribute mechanistically to the maintenance of lung cancer phenotypes. Further characterization of this oncogene regulatory mechanism may uncover a novel therapeutic intervention point for tumours driven by HMGA1.

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