ABSTRACT Cancer is projected to affect 20M people worldwide in 2023, with new cases increasing almost 30% in the next two decades. Cancer is a complex disease caused by a vast range of genetic and epigenetic defects. Existing technologies often fall short in comprehensively identifying the various genetic aberrations underlying different cancer types. A significant number of cancers remain without detected actionable or causative variants, revealing a critical gap in our understanding, and delaying or preventing the use of targeted treatments. To address this gap, we have developed an innovative Hi-C workflow that we call VariLink™. VariLink overcomes many of the limitations of current genomic technologies and substantially enhances the detection of cancer- driving structural variants (SVs) in the genome. By leveraging the 3D structure of the genome to incorporate long-range linkage information into the sequencing library, VariLink enables highly sensitive detection of large SVs, while maintaining shotgun-like detection of smaller variants such as small nucleotide variants (SNVs) and insertions/deletions (indels). VariLink also overcomes the drawbacks of other commercially available Hi-C technologies with its 1) rapid workflow (<8 hours vs. 2-3 days); and 2) unbiased genome coverage, enabling more accurate variant calling. By combining and exceeding the capabilities of existing methods, VariLink can uncover a broader spectrum of genetic and epigenetic changes in a single assay from a single sample. In our Phase I-equivalent work, we performed initial development of VariLink and demonstrated its performance relative to existing Hi-C assays. We also showed feasibility in clinically relevant tissue types, including blood. Importantly, we showed that VariLink detects SVs at much higher sensitivity than standard whole-genome sequencing technologies. In our Phase II work, we aim to further validate the performance of VariLink across multiple clinical sample types and compare it to standard molecular cytogenetic technologies. We also aim to adapt VariLink for formalin-fixed, paraffin-embedded (FFPE) samples and develop high-throughput automation and data analysis protocols. Successful completion of this project will demonstrate VariLink's performance in comprehensive variant detection and its suitability for commercialization in the translational research and clinical oncology space. By enabling more sensitive detection of genetic alterations in cancer patient samples, VariLink could contribute to improved targeted therapies, drug discovery, and reduced cancer mortality. With a focus on the expansive cancer diagnostics market, we anticipate VariLink's eventual adoption as a diagnostic test, pushing cancer genomic characterization beyond the limitations of current assays.