Study cohort

We analysed a retrospective cohort of 1374 BC patients who received neoadjuvant chemotherapy (NACT), with or without targeted therapy, and subsequent surgical resection in the years 2013–2019 from 11 institutions (Nottingham University Hospitals NHS Trust; Addenbrookes Hospital, Cambridge; University Hospitals Birmingham NHS Foundation Trust; University Hospitals of Leicester NHS Trust; St. Vincent’s University Hospital, Dublin; University Hospital Galway; St. Helens and Knowsley Teaching Hospital NHS Trust, Liverpool; Guy’s and St. Thomas’ Hospital, London; Ninewells Hospital, Dundee, Leeds Teaching Hospitals NHS Trust, University of Turin, Italy). Inclusion criteria included availability of data on HER2 gene copy number and HER2/CEP17 ratio, NACT and pathological response details with emphasis on Group 2 tumours. As control groups, some cases with IHC scores of HER2 3+ and 0/1+ were included (Table 1). Cases were identified and data collected from all centres based on these defined criteria to avoid sample bias.

Histological grade, details of oestrogen receptor (ER) status and treatment regimen received were also collected. pCR was defined as no residual invasive carcinoma in both the breast and axillary lymph nodes regardless of the presence of residual ductal carcinoma in situ (ypT0/is ypN0).²³ All histopathological information was obtained from the original pathology reports. Anonymised data were analysed centrally. HER2 status was assessed using IHC and ISH, as described in the UK guidelines,¹⁹ with primary IHC, followed by ISH on all borderline (2+) cases. In cases with an unusual ISH pattern, such as the target study cohort, the existing UK guidelines mandate counting 60 instead of 20 cells. According to the IHC and ISH results, the tumours were classified into the ASCO/CAP HER2 Groups³ (Table 1). ER positivity were defined as nuclear staining of ≥1% of invasive tumour cells.²⁴

Patients were considered eligible for HER2 targeted therapies if their tumours showed a HER2 IHC score of 3+, or 2+ with HER2/CEP17 ratio ≥2.0 regardless of the HER2 copy number (i.e. ASCO/CAP Groups 1 and 2) or if the HER2 copy number was ≥6 (Group 3). For subgroup analysis in this study, patients were categorised according to HER2 status into (1) Groups 1, 2 and 3, and (2) Groups 4 and 5. Patients were subdivided into two groups based on the type of neoadjuvant therapy received: (1) chemotherapy in combination with HER2 targeted therapy, trastuzumab alone or with either pertuzumab or lapatinib and (2) patients who received chemotherapy only. Chemotherapy regimens given were in accordance with individual unit protocols and included anthracycline and taxane, anthracycline without taxane or non-anthracycline based regimens.

This study was approved by the Nottingham Research Tissue Bank Access Committee under the IRAS Project ID: 184265. Data were collected as fully anonymised.