Past exposure to genotoxic cancer therapies is associated with elevated local mutagenesis at CTCF binding sites in metastatic tumors

Many cancer therapies are known to have mutagenic effects, but their extent and potential functional implications in surviving cancer cells are incompletely understood. In particular, it is unclear how regulatory elements that are vulnerable to mutations are affected by these therapies. CTCF is a master regulator of chromatin organization; it binds DNA at TAD boundaries and other key regulatory elements. Previous studies have observed that CTCF binding sites (CBS) are enriched in mutations in cancer, but the underlying mechanisms are incompletely understood. We asked whether prior treatments were associated with local mutagenesis at CBS in metastatic tumors. Utilizing a cohort of 4,870 whole-genome-sequenced metastatic tumors from the Hartwig Medical Foundation, we systematically analyzed associations between mutational processes at CBS across 17 cancer types and 45 types of cancer therapies with our computational method RM2. We found 21 associations involving 6 cancer types and 14 therapy types. Most notably, past exposure to radiotherapy or trifluridine was associated with increased CBS mutation rates in metastatic colorectal cancer. Both radiotherapy and trifluridine are associated with increased SNV burden without a strong trinucleotide preference, consistent with a generally weaker DNA repair in these sites. Therapy-associated mutation enrichment occurred primarily in subsets that display the canonical CTCF binding DNA motif, relatively late replication timing or association with lower gene expression, suggesting that specific chromatin environments enable treatment-associated mutagenesis in CBS. We further asked whether the presence of known driver mutations influences therapy-associated local mutagenesis at CBS. This revealed that mutations in DNA damage response and chromatin regulation pathway genes, including BRCA2 and FBXW7, further elevate CBS mutation rate in radiotherapy-exposed tumors. This suggests higher vulnerability to radiotherapy-associated mutations when DNA damage response pathways, especially homologous recombination, are compromised. In conclusion, this study provides, to our knowledge, a first analysis of association between cancer therapies and mutational burden at CTCF binding sites. Our findings highlight a previously unknown side effect of genotoxic cancer therapies on key regulatory elements. This underscores the need for a deeper understanding of therapy-induced mutations in regulatory regions and their potential implications for cancer biology and treatment outcomes.