# Improving HER2 Diagnostics with Digital Real‐Time PCR for Ultrafast, Precise Prediction of Anti‐HER2 Therapy Response in Patients with Breast Cancer

## TL;DR

A new digital PCR method improves HER2 testing accuracy and speed, leading to better predictions of anti-HER2 therapy response in breast cancer patients.

## Contribution

A fully automated digital real-time PCR platform is introduced for ultrafast and precise HER2 diagnostics.

## Key findings

- drPCR is faster, simpler, and more accurate than IHC/ISH for HER2 status assessment.
- drPCR identifies patients with specific genomic alterations who respond well to anti-HER2 therapy.
- drPCR achieves high pathological complete response rates in HER2-positive patients.

## Abstract

While human epidermal growth factor receptor (HER2) has emerged as a tumor‐agnostic biomarker, standard HER2 testing for anti‐HER2 therapies using immunohistochemistry (IHC) and in situ hybridization (ISH) assays remains subjective, time‐consuming, and often inaccurate. To address these limitations, an ultrafast and precise HER2 testing method is developed using Lab‐On‐An‐Array (LOAA) digital real‐time PCR (drPCR), a fully automated digital PCR enabling real‐time absolute quantification. A multicenter study involving four independent breast cancer cohorts cross‐validates the high diagnostic accuracy of drPCR‐based HER2 assessment. Comparative analyses with artificial intelligence algorithms, next‐generation sequencing, and droplet digital PCR demonstrate that drPCR is faster, simpler, and more accurate than conventional assays for assessing HER2 status, while IHC/ISH frequently yields false positives. Importantly, in patients initially diagnosed as HER2‐positive and treated with neoadjuvant anti‐HER2 therapy, the HER2 drPCR(+)/IHC‐ISH(+) group achieves high pathological complete response rates, while HER2 drPCR(‐)/IHC‐ISH(+) cases exhibit poor treatment responses, highlighting the superior predictive accuracy of drPCR for anti‐HER2 therapy response. Additionally, drPCR identifies patients with chromosome 17 centromere abnormalities, HER2‐zero/ERBB2 hemizygous deletion, and ERBB2 hyperamplification who respond favorably to anti‐HER2 therapy. Collectively, these findings establish drPCR as a clinically feasible, standardized, and ultrafast HER2 testing method for improved prediction of anti‐HER2 therapy response in patients with cancer.

This study presents a novel, clinically feasible HER2 testing method using a fully automated digital real‐time PCR platform. It enables real‐time absolute quantification of ERBB2 copy number alterations for rapid, simple, and accurate assessment of HEgR2 status, thereby overcoming the limitations of conventional testing and improving prediction of anti‐HER2 therapy response.