Abstract 3894: Three-dimensional whole tumor genomic analysis redefine intratumor heterogeneity of gliomas, with implications for early tumorigenesis

Abstract Intratumor heterogeneity and evolution underlie treatment failures but are poorly understood due to a limited number of tumor biopsies. In this study, we examine IDH mutant gliomas, utilizing a 3D whole tumor sampling approach and exome sequencing (32 patients, 345 samples), along with RNAseq (12 of the 32 patients, 88 samples). We investigate and redefine intratumoral heterogeneity and evolution of insertion/deletion (indel) and single nucleotide variant (SNV) in driver genes on a whole tumor scale. In the tumor suppressor capicua transcriptional repressor (CIC), a canonical driver in IDH mutant 1p/19q co-deleted (IDHmut-Codel) tumors, our exome and RNAseq data analysis highlights a wide spectrum of heterogeneity and inactivation by SNVs and indels in independent subclonal expansions. For instance, in nine samples from one patient’s recurrent tumor, five showed distinct CIC SNVs and eight exhibited indels. Notably, all samples showed one or more CIC SNVs or indels. Among five samples harboring SNVs, three had 2 to 3 co-occurring SNVs, and among the 8 samples with indels, three had 2 and one had 5 indels. None had the same combinations of indels and SNVs, indicating extreme heterogeneity and subclonality. Additionally, TA-cloning of a long-range PCR from one sample suggested that each individual SNV and indel were in different tumor cells, raising the possibility of early diversification of subclonal expansions driven by independent CIC alterations. Similar patterns were observed in IDH-mutant and 1p/19q non-codeleted tumors involving tumor suppressor ATRX. These findings demonstrate that lower grade IDH-mutant gliomas (LGGs) exhibit significant intratumoral heterogeneity, a trait typically associated with glioblastoma. To determine if CIC alterations were inactivating or simply passengers, we analyzed gene expression in those spatially mapped samples. Within the ETS-domain transcription factor family, ETS1, ETV1, ETV4, and ETV5 were upregulated in CIC mutant samples versus wild type samples. We also identified novel candidate genes potentially regulated by CIC, suggesting possible mechanisms contributing to tumorigenesis and new treatment targets for IDHmut-Codel tumors. This study elucidates the substantial intratumor heterogeneity and evolutionary dynamics in the drivers of LGGs on a whole tumor scale, highlighting their important role in tumorigenesis. By utilizing spatially mapped samples within the same tumors, our research reveals convergence on CIC loss of function through many mutually exclusive SNVs and indels in IDHmut-Codel tumors. Our findings demonstrate the critical nature of CIC inactivation across entire tumors, emphasizing the importance of indels and SNVs in tumor progression. This research also identifies candidate genes upregulated by CIC inactivation as possible treatment targets for IDHmut-Codel tumors. Citation Format: Lee H. Chen, Nicholas O. Stevers, Chibo Hong, Ivan V. Smirnov, Nadia R. Grishanina, Joshua S. Wu, Anny Shai, Susan M. Chang, Mitchel S. Berger, Shawn L. Hervey-Jumper, Joanna J. Phillips, Joseph F. Costello. Three-dimensional whole tumor genomic analysis redefine intratumor heterogeneity of gliomas, with implications for early tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3894.