The role of immunohistochemistry in patient stratification and personalised medicine

Not all patients respond well to treatment and drugs have failed in the clinic due to “all patient” designs. As a result drugs that could help subpopulations of patients may never make it to market. Identifying patients and stratifying them according to their likelihood as responders can guide clinical trial design, leading to more success in the clinic and ultimately increasing the range and personalisation of treatments available.

Immunohistochemistry (IHC) could be used to group patients according to the expression levels of a specific biomarker. With more emphasis on standardisation, reliability, validation and quantification, IHC assays can play this role in patient stratification[2].

IHC assays are not easily validated - hindered not least by the lack of standardisation in tissue collection, handling and fixation[1,2,3]. Availability of specific and reliable antibodies is the crux of successful assay development and much time can be spent identifying the best reagent. Furthermore, Moulis[1] highlights that IHC assays are generally optimised to the strongest level of expression when they actually need to be optimised around the dynamic range of the biomarker (i.e. the range of possible levels of expression) in order for them to be useful and fit-for-purpose. In personalised medicine, the dynamic range of expression needs to be known to determine cut-off points and subsequently stratify patients[2]. Therefore, more emphasis and evaluation of assay performance on low, medium and high expressing controls in the form of cell lines and tissues would be beneficial if not necessary.

Traditionally IHC is a qualitative rather than quantitative assay but has always been considered valuable due to its tissue-rich context and ability to demonstrate expression in situ. Accurate and reproducible quantification of IHC could fill the gap and leave IHC an unrivalled assay in comparison to the inherently quantitative but liquid-based assays like Western Blotting and ELISAs. Conventionally, semi-quantitative scoring is generally performed visually by a pathologist and while good correlation has been demonstrated[4], quantification achieved through image analysis can produce much more precise measurements (See Figs 1 and 2). With successful validation and quantification, IHC assays can play a vital role in drug development as a method of patient stratification.

titration of a primary antibody on two tumours

Figure 1 shows the titration of a primary antibody on two tumours with different levels of expression. Tumour A has much higher expression than Tumour B. Based on Tumour A alone, the lowest concentration of antibody would be chosen as the most optimal for the assay as it maximises the signal: noise ratio. Based on Tumour B alone, the highest concentration of antibody would be chosen as the most optimal for the assay.

Hypothetically where this expression indicates a responder to treatment, if both tumours are labelled with the highest concentration both patients will be treated but if both tumours are labelled with the lowest concentration, only patient with tumour A would be treated.

percentage of tumour positively labelled

Figure 2: Image analysis data: percentage of tumour positively-labelled

Hypothetically, if the cut off level for positive treatment response was tumour expression level of over 0.5%, then patient A would only be seen as a responder if the assay was performed with a primary antibody concentration of 2µg/ml.


[1] Definiens Webinar: Clinical Trial Patient Stratification with Quantitative IHC Assays Patient. Sharon Moulis 13 May 2015

[2] Definiens: Redefining IHC Assays for Powerful, Quantification, Patient Stratification and Companion Diagnostics

[3] Biomarkers for Patient Stratification. Brussels 10 – 11 Jun 2010

[4] Quantitative comparison of immunohistochemical staining measured by digital image analysis versus pathologist visual scoring. Rizzardi AE, Johnson AT, Vogel RI, Pambuccian SE, Henriksen J, Skubitz AP, Metzger GJ, Schmechel SC. Diagn Pathol. 2012 Jun 20;7:42. doi: 10.1186/1746-1596-7-42.

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Epistem is your ideal partner for biomarker discovery research. We have extensive experience optimising and validating histologic and IHC staining in our GCLP-accredited laboratories and have established a range of SOPs for immediate analysis of some markers. We offer both automated and manual image analysis using the Aperio Scanscope system and our experienced staff can provide both quantitative and qualitative asessments based on the individual requirements of the project.

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