S2591
Radiobiology - Preclinical biomarkers
ESTRO 2026
Digital Poster 912 Whole genome analysis for identification of non-coding G- quadruplex mutations in Pan-Cancer Amen Shamim 1,2 , Ayesha Shamim 3 , Wanki Yoo 4 , Nazia Perveen 4 , Muhammad Imran Arshad 5,6 , Ananya Choudhury 1,7 , Kyeong Kyu Kim 4 1 Translational Radiobiology Group, Division of Cancer Sciences, School of Medical Sciences, Faculty of Biology, Medicine & Health, University of Manchester, Manchester, United Kingdom. 2 Department of Computer Science, University of Agriculture, Faisalabad, Pakistan. 3 Faculty of Pharmacy, University of Sargodha, Sargodha, Pakistan. 4 Department of Precision Medicine, Graduate School of Basic Medical Sciences (GSBMS), Sungkyunkwan University School of Medicine, Suwon, Korea, Republic of. 5 Department of Veterinary Preventive Medicine, College of Veterinary Medicine, Qassim University, Buraydah, Saudi Arabia. 6 Department of Epidemiology and Public Health, Faculty of Health and Pharmaceutical Sciences, University of Agriculture, Faisalabad, Pakistan. 7 Department of Clinical Oncology, The Christie NHS Foundation Trust, Manchester, United Kingdom Purpose/Objective: Cancer development and progression are primarily driven by the accumulation of cancer-associated mutations, however, the functional impact of mutations in noncoding regions is not well characterised. Given that G-quadruplex (G4) structures within regulatory regions can suppress oncogene expression, as observed in genes such as c-myc1, we hypothesised that G4-disrupting mutations in noncoding regulatory regions may promote cancer progression by enhancing oncogene transcription. Material/Methods: Whole-genome sequencing (WGS) data from 1,667 cancer across 29 cancer types were obtained from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium (1290 samples) and Sanger Institute cohorts (377 samples). Variants were mapped with enhancer- promoter regions and potential G4-forming sequences in the human genome. Hotspot-G4s defined as a G4 region with a mutation probability (Pm) < 0.05, calculated using a Poisson distribution based on the expected mutation rate (E) derived from the background mutation probability (Pb), with multiple testing correction applied using the Bonferroni method. Differential expression of variant-associated genes was calculated using DeSeq22. Experimental validation of the target promoter was performed using CD spectroscopy and EMSA to assess G4 formation and its disruption, followed by luciferase reporter assays to evaluate the impact of G4 mutations on promoter activity. Results: The systematic analysis of cancer mutations revealed a higher mutation frequency in intergenic region compared to coding regions. Hotspot-G4s showed high mutation probability, and their prevalence varied across cancer types. Genes associated with hotspot-G4s showed high expression levels in cancers, suggesting the possible correlation between G4 mutation in the regulatory region and their target gene expression (Figure 1). By predicting the effect of mutation on G4 disruption, we further narrow down 250 cancer genes associated with 445 G4-disrupting mutations and identified 155 genes whose expression are significantly increased by G4 disruption. Experimental validation confirmed that G4 mutations in the promoters of ZM1Z1 (Figure 2), S100A7A, WDR37, LASP1, SH3PXD2B and TRIM37 reduced G4 stability and increased gene expression.
Figure 1.a) Distribution of mutations in whole genome and analysing them in intergenic region by characterising mutation distribution in regulatory region, enhancer, promoter, CTCF, and TFBS. b) Mutation burden probability in each cancer type with background mutation rate. Conclusion: G4-disrupting mutations are enriched in intergenic regions that can significantly enhance the expression of nearby oncogenes. The G4- disrupting mutations may represent an important mechanism driving oncogene activation and could serve as potential biomarkers or therapeutic targets in cancer.
Figure 2 Schematic representation of ZMIZ1 promoter with hotspot- G4 and mutation burden. References: 1. Fekete, A., Kenesi, E., Hunyadi-Gulyas, E., Durgo, H., Berko, B., Dunai, Z.A. and Bauer, P.I. (2012) The guanine-quadruplex structure in the human c-myc gene's promoter is converted into B-DNA form by the human poly(ADP-ribose)polymerase-1. PLoS One, 7, e42690.2. Love, M.I., Huber, W. and Anders, S. (2014) Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol, 15, 550. Keywords: Pan-cancer, Somatic mutations, G-quadruplex
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