HISTOPATHOLOGY
Improving HER2 classification with molecular testing
Recent changes in HER2 scoring in breast cancer have further complicated an already challenging diagnosis. Vinicio Tassani examines current workflows and makes the case for adding molecular testing to the mix.
Human epidermal growth factor receptor 2 (HER2) scoring in breast cancer traditionally labelled tumours as either positive or negative but, more recently, HER2-low breast cancer has been recognised as a clinically relevant subset. This shift introduces challenges for diagnosis, as immunohistochemistry (IHC) – the gold standard diagnostic method – is prone to subjectivity and poor reproducibility when distinguishing between HER2 0, 1+ and 2+. Borderline cases can be ambiguous, and studies from across the UK and Europe have shown poor agreement in HER2-low scoring between different pathologists, raising concerns over consistency in routine practice. This article explores the limitations of current diagnostic workflows and discusses the role of RT-qPCR-based molecular subtyping as a complementary approach to improve reproducibility and support IHC interpretation. HER2 is a transmembrane receptor
tyrosine kinase that is vital for cell growth and survival. It was first identified as a breast cancer biomarker in 2005, and soon became the primary indicator for stratifying patients at initial diagnosis.3 HER2 status went on to also be used to determine the best treatment option, primarily to determine whether HER2- targeted agents such as pertuzumab, trastuzumab and trastuzumab emtansine (T-DM1) would be effective.4-6
traditionally classed into two categories: HER2-negative or HER2-positive, which guided the use of these therapies.
Current testing protocols HER2 protein expression on tumour cells
Breast infiltrating ductal carcinoma showing HER2-positive cell membrane staining (immunohistochemistry with DAB chromogen).
is currently detected using IHC, which is considered the gold standard for assessing HER2 status. IHC results are scored to reflect the intensity and completeness of membrane staining,7
where 0 or 1+
is HER2-negative, 2+ is equivocal and 3+ is HER2-positive. Reflex testing is performed if a tumour scores IHC 2+, using in situ hybridisation (ISH) techniques – fluorescence (FISH) or chromogenic (CISH) – to determine whether the HER2 gene is amplified in the patient,7
and they Patients were
are therefore eligible for HER2-targeted therapies (Table 1).
Paradigm shift in HER2 classification This traditional classification of HER2 status as either positive or negative
is now being contested. For example, the DESTINY-Breast04 trial revealed the relevance of a third group, HER2- low breast cancer, which is defined by IHC scores of 1+ or 2+ without HER2 gene amplification.8
This is particularly
significant because the antibody drug conjugate (ADC) trastuzumab deruxtecan (T-DXd) has shown clinical benefits in HER2-low tumours, a group that was previously considered HER2-negative, and was therefore deemed to be ineligible for therapy.8
Researchers have also
recognised a potential fourth category – HER2-ultralow – encompassing tumours marginally above the IHC 0 threshold.10 These tumours are not yet formally outlined in clinical guidelines, but it is thought that they may also respond
February 2026
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