Effect of Digoxin on clusters of circulating tumor cells in patients with metastatic breast cancer: a phase 1 trial

The presence of circulating tumor cell (CTC) clusters is associated with disease progression, new metastasis formation and reduced survival in a variety of cancer types. In breast cancer, pre-clinical studies showed that inhibitors of the Na+/K+-ATPase can suppress CTC clusters shedding and block metastasis. Here, we conducted a prospective, open-label, phase I study in patients with metastatic breast cancer, where the primary endpoint was to determine whether a short (one week) treatment with the Na+/K+-ATPase inhibitor digoxin could reduce mean CTC cluster size. Mechanistically, transcriptome profiling of CTCs highlighted downregulation of cell-cell adhesion and cell cycle-related genes upon treatment with digoxin, in line with its cluster-dissolution activity. ClinicalTrials.gov identifier: NCT03928210. Overall design: We isolated CTCs from a breast cancer patient (patient 5) at different time points. Single CTCs, CTC clusters and CTC-white blood cell (WBC)-clusters were pooled into tubes for molecular characterisation using next-generation sequencing. Amplified cDNA was prepared on-bead according to the Smart-seq2 protocol. Libraries were prepared using Nextera XT (Illumina) and sequenced on an Illumina NextSeq2000 instrument in 101-base-pair single-read mode. This yielded a median raw sequencing depth of 7.8 million reads per sample.