Mutagenic DNA Lesions in Normal Hematopoietic Stem Cells Persist for Years Causing Distinct Patterns of Mutation

BACKGROUND

Hematopoietic stem cells (HSCs) constantly acquire somatic mutations at a rate of ~17 per cell per year. Mutations in specific genes offer a growth advantage that may result in clonal hematopoiesis, and eventually, hematological malignancy. Understanding the processes causing these somatic mutations is fundamental to understanding the early origins of malignancy and informing interventions to reduce cancer risk. While some mutations in HSCs are from well-characterised processes, most are of unknown cause. Mutations are usually preceded by DNA damage. In fact, DNA suffers continual damage, with thousands of individual lesions at any moment. The efficiency of DNA repair means that most known classes of lesions have a half-life of minutes to hours, but might DNA damage persist longer? Understanding the dynamics of DNA lesions may help elucidate their origins and mitigate their consequences.

METHODS

We hypothesised that examining high-resolution phylogenetic trees of somatic cells would let us infer the persistence of DNA lesions across multiple cell cycles. A given lesion that persisted across several cell divisions would have potential to generate a mutation each time that strand was replicated, and these separate mutations could be detectable in the phylogeny. If different bases were misincorporated opposite the lesion during sequential rounds of DNA replication, closely related clones would carry two alternative mutations at the same position in the genome (‘multi-allelic’ variants; MAVs). If a persistent lesion has the same (incorrect) base misincorporated opposite during different rounds of replication, those mutations may be recognised by their contravention of the consensus phylogeny (‘phylogeny-violating’ variants; PVVs).

We used seven published sets of somatic phylogenies from whole-genome sequencing of single-cell-derived colonies, organoids or laser-capture microdissections (LCM). The dataset comprised 103 phylogenies from 89 individuals, in total 11,429 genomes. Each phylogeny was generated from a single tissue type: hematopoietic stem and progenitor cells (HSPCs, n=39), bronchial epithelial cells (n=16) or liver parenchyma (n=48). The HSPC phylogenies were either from foetal and cord blood (n=4), healthy adults (n=13), stem cell transplant donor/recipient pairs (n=10), patients with myeloproliferative neoplasms (n=10) or chemotherapy-exposed patients (n=2). We examined all phylogenies to identify MAVs and PVVs caused by persistent DNA lesions.

RESULTS

• We identified mutations arising from 816 DNA lesions that persisted across multiple cell cycles in normal human stem cells from the blood, liver and bronchial epithelium.
• In HSCs, persistent DNA lesions, likely from endogenous sources, generated a characteristic mutational signature, so-called SBS19.
• The causative lesions predominantly affected guanines in an ApG context.
• The persistent lesions occurred steadily throughout life, enduring 2.2 years on average, with 25% lasting 3+ years.
• We estimate that a single HSC has ~8 such lesions at any moment in time, half of which will generate a mutation with each cell cycle.
• Overall, 16% of mutations in blood cells are attributable to SBS19, and similar proportions of driver mutations in blood cancers exhibit this signature.

CONCLUSION

A vast array of DNA lesions emerges from the quotidian chemistry of life coupled with the rather more elective chemistry of our lifestyles. While many lesions are generated frequently and efficiently repaired, our data imply the existence of a family of DNA lesions present in low numbers, but persisting for months to years, generating sizable fractions of cells’ mutation burdens.

HSCs are unusual amongst normal tissues in that a substantial proportion of mutations have a pattern corresponding to SBS19. Our data explain this, as HSCs feature long-lived lesions giving rise specifically to mutations with this profile. Intriguingly, some patients with sickle cell disease have been shown to have increased rates of SBS19, suggesting there is variability in the degree of damage caused.

Recognizing the unusual dynamics of this DNA damage in HSCs, future work will be well-placed to identify the causative lesion and the reason for its persistence, informing strategies to reduce the mutations it causes. In so doing, we may be able to prevent the earliest origins of blood cancer.