Jamin Wu¹, Eva Perak¹, Neha Dhasmana¹, Chanel Schroff¹, Eloise Freitag¹, Yiying Yang¹, Matija Snuderl¹

¹Department of Pathology, NYU Langone Health, New York, USA

Background: Hi-C is a sequencing-based 3D genomics assay which captures pairwise chromatin proximity genome-wide, enabling global structural variant reconstruction in formalin-fixed paraffin-embedded (FFPE) tissue^1,2. We present a case using Hi-C to investigate a 43-year-old male with glioblastoma with NTRK3-fusion (detected by NYU Langone's RNA-based FUSION-SEQer) who showed poor response to larotrectinib.
Methods: FFPE-compatible Hi-C was performed retrospectively on tissue from unstained slides and sequenced to 64 million paired-end reads. Structural variants were reconstructed from chromosomal segments with aberrant interaction frequencies on Hi-C contact maps.
Results: Hi-C identified the known PHF21B::NTRK3 fusion as part of a complex rearrangement involving 1p, 15q, and 22q, and revealed loss of CDKN2A and CDKN2B due to recombined fragments from 9p, 11q, 14q, and 22q.  Hi-C also identified >1Mb amplifications on 1q and 7p containing MDM4 and EGFR with 3D genomic interactions suggestive of heterogeneous amplicons of circularised extra-chromosomal DNA (ecDNA), revealing a possible mechanism of rapid adaptive evolution and therapeutic resistance³.
Conclusion: This case demonstrates Hi-C's ability to reconstruct complex structural variants (unresolvable using FISH) in FFPE tissue (unsuitable for conventional karyotyping or long-read sequencing), highlighting Hi-C as a tool for precision diagnostics to identify concurrent oncogenic drivers with potential implications for prognosis or therapy.  

1. Dixon JR, Xu J, Dileep V, Zhan Y, Song F, Le VT, et al. Integrative detection and analysis of structural variation in cancer genomes. Nat Genet. 2018 Oct;50(10):1388–98.

2. Schmitt AD, Sikkink K, Ahmed AA, Melnyk S, Reid D, Van Meter L, et al. Evaluation of Hi-C Sequencing for Detection of Gene Fusions in Hematologic and Solid Tumor Pediatric Cancer Samples. Cancers. 2024 Jan;16(17):2936.

3. Lange JT, Rose JC, Chen CY, Pichugin Y, Xie L, Tang J, et al. The evolutionary dynamics of extrachromosomal DNA in human cancers. Nat Genet. 2022 Oct;54(10):1527–33. 

The author (J.W.) performed the bioinformatic analysis, structural variant reconstruction, interpretation, and write-up of the Hi-C findings for this case. E.P. performed manual review of Hi-C breakpoint calls and literature search of involved genes. N.D. and C.S. performed the wet-lab Hi-C library preparation. E.F. and Y.Y. ran the automated Hi-C sequencing pipeline. M.S. is the laboratory principal investigator and supervisor of this work. 
The abstract content is original content written by the author and approved by all co-authors.