title: An examination of the impact of cellular age on leukaemic transformation to understand biological differences in paediatric and adult acute myeloid leukaemia
Creators: name: Chaudhury, Shahzya Shahrin
Publisher: name: University of Glasgow
Year: 2018

description: Acute myeloid leukaemia (AML) is a heterogeneous disorder and patient age is an independent prognostic factor for clinical outcome. Treatment of paediatric AML is largely extrapolated from adult trial data, despite age related disease heterogeneity. Normal haemopoietic stem and progenitor cells (HSPCs) display differences in cellular behaviour and molecular landscape with age, which may influence leukaemic transformation. Using a murine model to represent infant (foetal liver (FL), childhood (3 week (w)), young adult (10w) and middle aged adult (&gt;52w) age groups, the impact of HSPC age on oncogene mediated leukaemogenesis was investigated. 

Assessment of transformation in vitro suggests that FL HSPCs are more resistant to leukaemia transformation. Using the NUP98-HOXA9 (NH9) fusion oncogene leukaemia model it was demonstrated that cells negative for cell surface lineage makers but expressing stem cell antigen 1 and cKit (LSKs) from all 4 ages transformed in vitro. However, a trend towards delayed transformation in FLLSKs was observed. While post-foetal (3w, 10w and &gt;52w) common myeloid progenitors (CMP) and granulocyte macrophage progenitors (GMP) also transformed, FL-CMPs and GMPs did not. Other oncogenes including the fusion oncogene AML1-ETO (A1E) and internal tandem duplications of the FLT3 gene (FLT3-ITD) also transformed FL-LSKs but consistently failed to transform FL-CMP and GMPs. This suggests that, independent of the oncogenic insult, foetal transformation relies on specific features of the LSK that are absent from FL committed myeloid progenitors. 

These results were corroborated by in vivo data, showing that recipients transplanted with NH9 expressing LSKs derived from young (FL and 3w) donors were less susceptible to leukaemia with a longer latency to disease and incomplete penetrance compared to recipients transplanted with adult (10w and &gt;52w) LSKs. This suggests that young stem cells possess foetal specific protective mechanisms, which prevent or delay the development of AML in vivo, regardless of self-renewal in vitro. Furthermore, despite pre-transplant expansion in myeloid conditions, a small proportion of young, but not adult pre-leukaemic LSKs, gave rise to acute lymphoblastic leukaemia (ALL) or mixed phenotype acute leukaemia (MPAL) suggesting foetal/young LSKs retain a lymphoid bias that can overcome myeloid programmes. Both results from in vivo studies are in line with clinical data, showing a lower incidence of AML in childhood and predominance of ALL.

The observations from in vitro and in vivo data were complemented by gene expression analysis. Targeted gene expression analysis demonstrated the expression of multiple lineage pathways in young (FL and 3w) pre-leukaemic and leukaemic cells, while adult (10w and &gt;52w) pre-leukaemic and leukaemic cells exhibited myeloid restricted transcription. In contrast, adult pre-leukaemic and leukaemic cells exhibited stem cell programmes, which have been implicated in aggressive disease. Upregulation of the BMP pathway in adult pre-leukaemic cells implicates a mechanistic role for the bone marrow microenvironment driving aggressive and myeloid restricted disease in adult LSKs. Global gene expression analysis using RNA-sequencing on in vivo generated AML cells mirrored these findings. This analysis revealed, in addition, apoptotic and tumour suppressive programmes were enriched in young-AML while proleukaemic programmes, including pathways related to the oncogene Myc, were enriched in adult-AML. Therefore, despite using the same oncogenic model to drive AML, cellular age discriminated disease at the transcriptional level and adult-AML expressed transcriptional programmes associated with a more aggressive disease. 

The results of this thesis demonstrate that cellular age does indeed influence oncogene mediated transformability and leukaemia phenotype. Age defined transcriptional pathways that may impact pathogenesis, response to therapy and therapeutic targets have also been identified. In the era of personalised medicine in which a patient’s mutational, transcriptional and epigenetic profile is considered when making therapy decisions, the consequence of cellular ontogeny on leukaemia biology should also inform treatment decisions. In particular, novel therapeutic targets must be specifically sought in the paediatric context as potential targetable dysregulated pathways in paediatric AML are not necessarily the same as adult AML.