Application of sensitive sequencing methods for comprehensive characterisation of cell-free tumour DNA in plasma and urine of patients with renal tumours - 11th International Symposium on Circulating Nucleic Acids in Plasma and Serum (CNAPS)

Christopher Smith ¹ Tina Moser ² Anja Riediger ¹ Matthew Eldridge ¹ Florent Mouliere ³ Thomas Mitchell ^4,6 Anne Warren ⁵ Ellen Heitzer ² Nitzan Rosenfeld ¹ Grant Stewart ⁶

¹CRUK Cambridge Institute, Cancer Research UK Cambridge Centre, UK
²Institute of Human Genetics, Medical University of Graz, Austria
³Department of Pathology, Amsterdam UMC, Vrije Universiteit Amsterdam, UK
⁴Wellcome Sanger Institute, Wellcome Sanger Institute, UK
⁵Dept. of Pathology, Cambridge University Hospitals NHS Foundation Trust, UK
⁶Dept. of Surgery, Cambridge University Hospitals NHS Foundation Trust, UK

Renal cell cancer (RCC) is the most lethal urological malignancy. RCC includes a range of pathological subtypes with different biological and genetic characteristics. Tumours often show genetic heterogeneity. Cell-free tumour derived DNA (ctDNA) allows non-invasive detection and monitoring of various cancers. However, its utility in RCC has not been well established.

We characterised the levels and composition of ctDNA in plasma and urine from patients with renal tumours including 43 with metastatic RCCs from the MonReC study, and 47 with a range of renal tumours from the DIAMOND study. For MonReC we utilised a de novo mutation calling approach, targeting 10 genes frequently mutated in RCC. For DIAMOND, a novel personalised approach termed INtegration of VAriant Reads from TArgeted PAnel Sequencing, INVAR-TAPAS was applied. For selected patients we assessed longitudinal samples taken throughout disease course, and explored representation of tumour heterogeneity.

For MonReC, despite an expected analytical sensitivity of 5x10^-3 mAF, ctDNA was detected in only 8/43 (18.6%) patients. For DIAMOND, INVAR-TAPAS improved detection in plasma (12/22, 54.5%) and urine (7/14, 50.0%), detecting down to mAF of 5.2x10^-5 and 2.5x10^-5 respectively. Whilst detection in plasma was more likely in patients with larger tumours (p=0.02), or those with venous tumour thrombus (p<005), ctDNA was also detected in early stage and benign tumours.

Longitudinal sampling of >200 plasma samples revealed that ctDNA can track disease course. Furthermore, we demonstrate that plasma and, for the first time, urine ctDNA is capable of overcoming genetic heterogeneity.

These findings confirm matched data generated by untargeted sequencing (see partner abstract, Moser et al) showing that levels of ctDNA in RCC are low. Nonetheless ctDNA can be detected in blood and urine of RCC patients, even in benign lesions, suggesting potential for clinical utility. However, improved isolation and detection methods are needed before this can be realised.