Exploring per-base quality scores as a surrogate marker of cell-free DNA fragmentome

Abstract Per-base quality scores are widely treated as technical metadata in next-generation sequencing. Here, we show that in rigorously controlled whole-genome sequencing of cell-free DNA, quality profiles may carry fragmentomic-associated signal that enables classification of cancer samples against matched controls. Analyzing four independent batches (23 cancer samples: pancreatic and breast; 22 matched controls) sequenced in a within-lane regime and further normalized per flow-cell tile to reduce technical confounders, we demonstrate through unsupervised analysis that boundary-enriched dynamics captured in these quality scores consistently separate cancer from control samples. A leave-one-batch-out classifier trained on quality-derived scores achieved a pooled area under the curve of 0.81. Furthermore, we show that the quality-derived metric correlates with short-fragment enrichment and tumor-associated 5′-end motifs, performing comparably to established, motif-based orthogonal methods. These results provide initial evidence that quality scores could serve as a low-cost, alignment-free surrogate signal for cfDNA-based cancer detection.