13 November 2018 from 14:30 to 15:30

PhD Defence: Decoding the mutational history of human stem cells: From patterns to mechanisms

The extreme variation in cancer risk across tissues was recently proposed to depend on the lifetime number of adult stem cell (ASC) divisions, owing to unavoidable random mutations that arise during DNA replication. However, the rates and patterns of mutations in normal human ASCs remain unknown. In this thesis, a new method is presented for cataloging mutations in individual human ASCs. Single ASCs are expanded in vitro into clonal organoid cultures to generate sufficient DNA for whole-genome sequencing (WGS) analysis, which allows detection of the mutations that accumulated in vivo in the original ACS. Using this method, we determined the genome-wide mutation patterns in ASCs of the small intestine, colon and liver of human donors with ages ranging from 3 to 87 years.

Our results show that mutations accumulate steadily over time in all of the assessed tissue types, at a rate of approximately 40 novel mutations per year, despite the large variation in cancer incidence among these tissues. However, mutational signature analysis suggests that different mutational processes contribute to mutation accumulation in liver ASCs compared to colon and small intestinal ASCs. In the intestinal ASCs the majority of the mutations are a result of spontaneous deamination of methylated cytosine residues. In liver, a mutational signature with an as-yet-unknown underlying mechanism is predominant.

In addition to adult stem cells, we directly measured the base substitutions that have accumulated in single stem cells of second-trimester human fetuses in the liver and intestine. We detected on average ~48 somatic base substitutions per fetal stem cell, indicating that the mutation accumulation rate is at least 5-fold higher during fetal development than during postnatal life. Furthermore, stem cells of the fetal liver and intestine show distinct mutational profiles, suggesting that different DNA damage and/or repair mechanisms are at play in these tissues during development. Intriguingly, the mutational landscape of the fetal intestinal stem cells closely resembled that observed in intestinal ASCs from postnatal tissue.

We used the CRISPR-Cas9 gene-editing technology to delete key DNA repair genes in human colon organoids and asses the mutational consequences in vitro. We found that organoids deficient in base excision repair harbor a mutational footprint  similar to signature 30, which is previously observed in a breast cancer cohort. Furthermore, nucleotide excision repair (NER) deficient ASCs show an increased contribution of Signature 8 mutations, which is a pattern with unknown etiology that is recurrently observed in human cancers. The presence of these signatures in cancer genomes could hold diagnostic and prognostic value, and may improve personalized cancer treatment strategies.. For example, tumors deficient in NER are more sensitive to cisplatin treatment.

Start date and time
13 November 2018 14:30
End date and time
13 November 2018 15:30
PhD candidate
Francis Blokzijl
Dissertation
Decoding the mutational history of human stem cells: From patterns to mechanisms
PhD supervisor(s)
Prof.dr. Edwin CuppenDr. Ruben van Boxtel
Entrance fee
Free