Endocrine Abstracts

Introduction: Circulating tumour DNA (ctDNA)–based monitoring in adrenocortical carcinoma (ACC) remains sparsely characterised relative to breast, colorectal, and lung cancers. We evaluated whether copy-number alteration (CNA) profiling of cell-free DNA (cfDNA) using shallow whole-genome sequencing (sWGS) could serve as a minimally invasive biomarker in ACC.

Methods: Serial plasma samples (n = 7) were collected from two patients with adrenocortical carcinoma (ACC) during periods of stable disease, tumour progression, and following surgical intervention. Total cfDNA was isolated from 4 mL plasma using the MagMAX™ Cell-Free DNA Isolation Kit on a KingFisher™ Flex Magnetic Particle Processor. Quantity and fragment-size distributions were assessed on Agilent 4200 TapeStation. Libraries were prepared for sWGS with the ReproSeq PGS kit and sequenced at low coverage. CNA profiling and tumour-fraction estimation were performed with ichorCNA pipeline under default parameters.

Results: Total cfDNA yields across the two patients were a median of 424 pg/µL (range 206-633); within normal limits, and fragment analyses demonstrated typical cfDNA profiles with a predominant mononucleosomal peak and a secondary dinucleosomal peak, supporting suitability for downstream analysis. Unfortunately, sWGS generated uniformly flat copy-number profiles across all 7 samples; hence no CNAs were detected by ichorCNA,and no ctDNA was detected by this method.

Discussion: In these 2 patients with ACC, sWGS did not reveal evidence of ctDNA, consistent with either (i) absence of CNA-bearing ctDNA in circulation or (ii) ctDNA levels below the detection threshold of sWGS/ichorCNA at times of active disease in ACC. To improve analytical sensitivity in ACC, future studies should focus on detection of ctDNA using more sensitive approaches such as next generation sequencing with either cancer mutation assays, or tumour-informed approaches.

Learning points: Reference1. sWGS-based CNA profiling of cfDNA may lack sensitivity for ACC. 2. Negative CNA results should prompt complementary assays (tumour agnostic and/or tumour-informed approaches).