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  current news   Press   selected story    
     
  20 December 2017  
 
Induced-Decay of Glycine Decarboxylase Transcripts as an Anticancer Therapeutic Strategy for Non-Small-Cell Lung Carcinoma
 
 




Authors
Jing Lin1,2, Jia Hui Jane Lee3, Kathirvel Paramasivam4, Elina Pathak3, Zhenxun Wang3,
Zacharias Aloysius Dwi Pramono5, Bing Lim3, Keng Boon Wee1,2 and Uttam Surana4,6,7

Author Affiliations
1 Bioinformatics Institute, A*STAR, 30 Biopolis Street, Singapore 138671, Singapore
2 Institute of High Performance Computing, A*STAR, 1 Fusionopolis Way, Singapore 138632, Singapore
3 Genome Institute of Singapore, A*STAR, 60 Biopolis Street, Singapore 138672, Singapore
4 Department of Pharmacology, National University of Singapore, 16 Medical Drive, Singapore 117660,   Singapore
5 Department of Research, National Skin Centre, 1 Mandalay Road, Singapore 308205, Singapore
6 Bioprocessing Technology Institute, A*STAR, 20 Biopolis Way, Singapore 138668, Singapore
7 Institute of Molecular and Cellular Biology, A*STAR, 61 Biopolis Drive, Singapore 138673, Singapore

Published in Molecular Therapy Nucleic acids 9:263–273, 15 December 2017

Abstract
Self-renewing tumor-initiating cells (TICs) are thought to be responsible for tumor recurrence and chemo-resistance. Glycine decarboxylase, encoded by the GLDC gene, is reported to be overexpressed in TIC-enriched primary non-small-cell lung carcinoma (NSCLC). GLDC is a component of the mitochondrial glycine cleavage system, and its high expression is required for growth and tumorigenic capacity. Currently, there are no therapeutic agents against GLDC. As a therapeutic strategy, we have designed and tested splicing-modulating steric hindrance antisense oligonucleotides (shAONs) that efficiently induce exon skipping (half maximal inhibitory concentration [IC50] at 3.5–7 nM), disrupt the open reading frame (ORF) of GLDC transcript (predisposing it for nonsense-mediated decay), halt cell proliferation, and prevent colony formation in both A549 cells and TIC-enriched NSCLC tumor sphere cells (TS32). One candidate shAON causes 60% inhibition of tumor growth in mice transplanted with TS32. Thus, our shAONs candidates can effectively inhibit the expression of NSCLCassociated metabolic enzyme GLDC and may have promising therapeutic implications.

Figure

Figure legend
:Transfection of shAONs at 100 nM Leads to Efficient and Specific Skipping of GLDC Target Exon in A549 Cells (A) The number of PTCs generated and the percentage of amino acid residues removed when each of the 14 out-of-frame exons are skipped individually. GLDC has a total of
25 exons. (B) Images of agarose gel electrophoresis of the PCR products demonstrating specific exon skipping induced by shAON transfection in A549 cells. Cells were harvested 24 hr post-transfection. 100 mg/mL cycloheximide was added 5 hr after shAON transfection to inhibit the skipped transcripts from undergoing NMD. (C) The exonskipping efficiency induced by each shAON as determined by densitometry analysis of the PCR products. The y axis shows the skipping efficiency, which is the percentage of the amplicons with exon skipping relative to the total amplicons (skipped + non-skipped). Data are presented as means ± SEM. (D) DNA sequencing of the bands corresponds to the skipped transcript excised from agarose gel (as indicated in B) to confirm skipping of the specific target exon.

For more information on Uttam SURANA 's lab, please click here.