Integrating extracellular vesicle and circulating cell‐free DNA analysis using a single plasma aliquot improves the detection of HER2 positivity in breast cancer patients

Abstract

Multi‐analyte liquid biopsies represent an emerging opportunity for non‐invasive cancer assessment. We developed ONCE (One Aliquot for Circulating Elements), an approach for the isolation of extracellular vesicles (EV) and cell‐free DNA (cfDNA) from a single aliquot of blood. We assessed ONCE performance to classify HER2‐positive early‐stage breast cancer (BrCa) patients by combining EV‐associated RNA (EV‐RNA) and cfDNA signals on n = 64 healthy donors (HD) and non–metastatic BrCa patients. Specifically, we isolated EV‐enriched samples by a charge‐based (CB) method and investigated EV‐RNA and cfDNA by next‐generation sequencing (NGS) and by digital droplet PCR (ddPCR). Sequencing of cfDNA and EV‐RNA from HER2‐ and HER2+ patients demonstrated concordance with in situ molecular analyses of matched tissues. Combined analysis of the two circulating analytes by ddPCR showed increased sensitivity in ERBB2/HER2 detection compared to single nucleic acid components. Multi‐analyte liquid biopsy prediction performance was comparable to tissue‐based sequencing results from TCGA. Also, imaging flow cytometry analysis revealed HER2 protein on the surface of EV isolated from the HER2+ BrCa plasma, thus corroborating the potential relevance of studying EV as companion analyte to cfDNA. This data confirms the relevance of combining cfDNA and EV‐RNA for HER2 cancer assessment and supports ONCE as a valuable tool for multi‐analytes liquid biopsies’ clinical implementation.

An electro-optical platform for the ultrasensitive detection of small extracellular vesicle sub-types and their protein epitope counts

“…Validating the quantity of CD81 proteins in spiked plasma was not possible since the plasma of healthy donors naturally has CD81 + sEVs while HER2 + sEVs are restricted to patients with elevated HER2 levels. 24 …”

Neuroendocrine gene subsets are uniquely dysregulated in prostate adenocarcinoma

“…Along the same lines, a study showed that BRCA patient plasma-derived EVs or cell-free DNA are both informative biomarker sources in BRCA, specifically for HER2 positive patients. 44 Moreover, the data demonstrated that combining EV and cell-free DNA assessments increased detection performance. 44 Therefore, we can speculate that aligning findings from plasma-derived EV content and cell-free DNA from PRAD and NEPC patients may increase the performance of liquid biopsies.…”

“… 44 Moreover, the data demonstrated that combining EV and cell-free DNA assessments increased detection performance. 44 Therefore, we can speculate that aligning findings from plasma-derived EV content and cell-free DNA from PRAD and NEPC patients may increase the performance of liquid biopsies.…”