Daniel Renouf

Pancreatic ductal adenocarcinoma (PDAC) can be stratified into distinct transcriptomebased molecular subtypes, with the ‘basal-like’ (or ‘squamous’) subtype being associatedwith worse prognosis, compared to the ‘classical’ subtype. These subtypesare assigned based on Moffitt genes signature scores where scores above a thresholdvalue are indictive of the basal-like subtype. Furthermore, PDAC tumours have uniquemetabolic transcriptomic profiles based on stratification of glycolytic and cholesterogenicgenes which correlate with basal-like and classical gene expression patterns,respectively. The mitochondrial pyruvate complex (MPC) mediates the transport ofpyruvate into the mitochondria. The mitochondrial pyruvate carrier 1 (MPC1) gene,which encodes one of two subunits of MPC, is deleted in over 60% of metastatic PDACand PDAC glycolytic tumours have lowest levels of MPC1 expression. Using PDACtissue microarray data, our group found that reduced MPC1 protein expression correlateswith reduced survival in patients. Therefore, we hypothesized that targetingMPC1 will alter metabolic reprogramming which may modulate tumour aggressivenessin tumour models. Genomically and clinically annotated patient-derived tumourorganoids (PDOs) were generated from metastatic biopsies from patients enrolled inthe PanGen study. Baseline metabolism and metabolic flux were measured usingSeahorse XFe96 based glycolytic and mito stress tests, these testswere adapted for compatibilitywith PDOs. Baseline glycolysis and oxidative phosphorylation (OXPHOS)rates demonstrated high variability in glycolytic reserves highlighting the extent ofmetabolic reprogramming in PDOs. This variability in glycolytic reserve positivelyassociated with Moffitt gene signature scores where PDOs with larger reserves tendedto have higher Moffitt scores. To alter metabolic activity, eight PDOs were treated for48 hours with UK-5099, a MPC1 inhibitor, or SRT-1720. SRT-1720 is an activator ofsirtuin 1 (SIRT1) which deacetylates peroxisome proliferator-activated receptor gammacoactivator 1-alpha (PGC1?), enhancing its activity. PGC1? has been shown to increasetranscription of MPC1. Treatment with UK-5099 raised glycolysis and glycolyticcapacity in four PDOs tested and reduced maximal respiration rates in seven PDOs.Treatment with SRT-1720 reduced glycolytic capacity in two PDOs but did not alter OXPHOSrates. Taken together, these results elicit the variability in metabolic dependencyin PDOs to meet energy demands and the plasticity of metabolic reprogramming.

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Genitourinary malignancies are a diverse group of cancers with an ever expanding array of therapeutic options. This thesis addresses how predictive biomarkers can be derived to help in treatment allocation. First, we survey the challenges around developing predictive biomarkers for checkpoint inhibitors in metastatic urothelial carcinoma through a systematic review. Despite multiple efforts, no single biomarker could be developed. Often, promising biomarkers were developed in single cohorts, and the predictive value of these biomarkers cannot be evaluated without a comparable cohort of patients receiving non-checkpoint inhibitor based treatments.Second, we present a cohort of 103 patients with metastatic urothelial cancer who underwent a liquid biopsy for circulating tumour DNA. This shows that ctDNA can be used to generate clinically relevant genomic data; we correlate specific outcomes to mutations detected in the bloodThird, we study the potential role of whole genome and transcriptome analysis (WGTA). In a cohort of patients enrolled in early phase clinical trials, this approach could suggest trial allocation in 50% of cases, which is a 3-fold increase when compared to the data that would be expected from a DNA panel alone. In addition, a cohort of 7 patients with metastatic adrenocortical carcinoma (ACC) has undergone WGTA. We discuss specific genomic findings, and how these may be used to direct treatment.Overall, this thesis provides preliminary data on the application of genomic profiling on diverse cohorts of genitourinary cancers, and suggests future steps for biomarker development in this setting.

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