What causes therapy-related malignancies in childhood cancer survivors? Dissecting the etiology of second cancers

Projektinformationen

SecondCANCERinKIDS

ID Finanzhilfevereinbarung: 864499

DOI

10.3030/864499

Projekt abgeschlossen

EK-Unterschriftsdatum 19 März 2020

Startdatum 1 April 2020

Enddatum 31 März 2025

Finanziert unter

• EXCELLENT SCIENCE - European Research Council (ERC)

Gesamtkosten

€ 2 000 000,00

EU-Beitrag

€ 2 000 000,00

Koordiniert durch

PRINSES MAXIMA CENTRUM VOOR KINDERONCOLOGIE BV
Netherlands

Periodic Reporting for period 4 - SecondCANCERinKIDS (What causes therapy-related malignancies in childhood cancer survivors? Dissecting the etiology of second cancers)

Berichtszeitraum: 2024-10-01 bis 2025-03-31

Zusammenfassung vom Kontext und den Gesamtzielen des Projekts

The survival of children with cancer has increased tremendously during the last decades. For example, the 15 years survival rate of children with acute lymphoblastic leukemia (ALL), which is the most common type of cancer in children, has increased from less than 10% in the 1960s to more than 90% now. This success in childhood cancer survival can be attributed to increasingly higher doses of chemo- and/or radiotherapy. Although high-intensity therapy more efficiently eradicates tumor cells, it can also be genotoxic to normal, that is noncancerous, cells. Indeed, because of their life-saving therapy, childhood cancer survivors are confronted with a variety of chronic health conditions later in life, including an increased risk for developing a novel (second) malignancy unrelated to the initial cancer. Compared to the general population, childhood cancer survivors not only show a higher incidence in novel malignancies, but also develop the disease earlier in life. In fact, second malignancies are one of the major causes for long-term mortality among survivors of childhood cancer. Importantly, the incidence of second malignancies is expected to rise in parallel with the increase of cancer survivors worldwide. Ideally, therapeutic strategies should be developed, which are aimed at preventing carcinogenesis in patients at risk for second malignancies.

Our overall aim is to determine the mechanisms and rate-limiting steps underlying the genesis of second malignancies in childhood cancer survivors. For this, we will study mutation accumulation at the single cell level in longitudinally collected blood samples of children who developed a therapy-related hematopoietic malignancy. Instead of focusing on identifying cancer driver mutations, we will utilize the entire set of (mostly passenger) mutations in these cells to i) track down the cellular origin of second malignancies by retrospective lineage tracing, ii) identify the mechanisms causing second malignancies by in-depth mutational analyses, and iii) study phenotypic effects of cancer treatment on population dynamics of blood.

Arbeit, die ab Beginn des Projekts bis zum Ende des durch den Bericht erfassten Berichtszeitraums geleistet wurde, und die wichtigsten bis dahin erzielten Ergebnisse

In this project, we investigated how chemotherapy contributes to the development of therapy-related myeloid neoplasms (t-MNs) in children. We assembled and analyzed a unique cohort of over 60 pediatric patients who developed t-MN after successful treatment of their first cancer. This includes 20 patients from our in-house biobank and 42 from international collaborators through the International BFM study group. We performed whole-genome sequencing (WGS) on all leukemic samples and, for a subset, also analyzed non-malignant hematopoietic stem and progenitor cells (HSPCs).

We found that most pediatric t-MNs are driven by structural variants, particularly involving the KMT2A locus. Normal HSPCs exposed to chemotherapy showed an increased mutation burden equivalent to several decades of natural aging, mostly due to acceleration of clock-like mutational processes. Phylogenetic reconstruction revealed that in most cases, the leukemic clone originated after the start of therapy and expanded during or shortly after treatment. This work was published in Cancer Discovery.

We also discovered a novel mutational signature caused by the antiviral drug ganciclovir, used to treat cytomegalovirus reactivation after hematopoietic stem cell transplantation (HSCT). Using a newly developed in vitro genotoxicity assay, we validated this mutational footprint and identified the same signature in pan-cancer datasets. This work, which has implications for long-term drug safety, was published in Cell Stem Cell. A patent application was filed on the methodology (Means and methods for assessing genotoxicity), and a related valorization trajectory was initiated via an ERC Proof of Concept grant (MUTAPREDICT).

Subsequently, we completed analysis of the full cohort and discovered that the expansion of t-MN clones is suppressed during platinum-based treatment and begins only after cessation of therapy. In children with TP53 germline mutations (Li-Fraumeni syndrome), this suppression does not occur, highlighting a TP53-dependent resistance mechanism. These findings were published in Nature Communications.

We also investigated prenatal chemotherapy exposure by sequencing fetal HSPCs from cord blood of neonates born to mothers treated for cancer during pregnancy. We showed that chemotherapy induces a measurable mutational burden in fetal cells, including platinum-related signatures, indicating transplacental genotoxicity (Cancer Discovery, 2025).

Finally, using a single-cell multi-omics approach combining genome and transcriptome sequencing of the same cells, we uncovered cell-intrinsic mechanisms that may protect a subset of HSPCs from mutagenesis. These "resilient" or potentially “super” stem cells may repopulate the hematopoietic system post-treatment and explain why mutation burdens normalize over time.

All data and protocols have been shared through open-access publications and our GitHub repository (https://github.com/ToolsVanBox).

Fortschritte, die über den aktuellen Stand der Technik hinausgehen und voraussichtliche potenzielle Auswirkungen (einschließlich der bis dato erzielten sozioökonomischen Auswirkungen und weiter gefassten gesellschaftlichen Auswirkungen des Projekts)

At the start of the project, the mechanisms by which chemotherapy contributes to secondary cancers in children were poorly understood. We significantly advanced the field by showing that only a subset of chemotherapeutic agents induces direct mutagenesis in hematopoietic stem and progenitor cells (HSPCs), while others mainly accelerate endogenous aging-related mutational processes. This concept of "mutational aging" in response to therapy represents a major shift in our understanding of long-term treatment effects.

We also discovered that the widely used antiviral drug ganciclovir is highly mutagenic, capable of inducing known cancer-driving mutations such as RAS gene alterations. This finding was unexpected and has important implications for the safety of antiviral treatment in immunocompromised patients, especially children. Using our newly developed genotoxicity assay, we identified alternative compounds with similar antiviral efficacy but without mutagenic activity. This work can potentially directly informs future clinical decision-making and drug development.

We completed the analysis of a unique cohort of over 60 pediatric t-MN patients and mapped their clonal evolution using phylogenetic inference. We found that platinum-based chemotherapy temporarily suppresses leukemic clone expansion, with outgrowth occurring only after cessation of treatment. This dynamic is absent in children with TP53 germline mutations, highlighting a critical interaction between genetic predisposition and therapeutic pressure. These insights refine our understanding of timing and selection in therapy-related cancer evolution.

Looking forward, our data offer the potential to identify biomarkers that predict which children are most at risk of developing t-MN, enabling earlier intervention or treatment modification. In particular, our inclusion of patients with genetic cancer predisposition provides a valuable framework for exploring personalized approaches to reduce long-term risk. The single-cell multi-omics framework developed in this project also offers a powerful platform for further investigating resilience and damage repair mechanisms in stem cells, which could be leveraged to design less genotoxic therapies in the future.

Graphical abstract of paper or mutagenicity of antiviral agents