Abstract
Tumour budding (TB) is an established prognostic feature in multiple cancers but is not routinely assessed in pathology practice. Efforts to standardise and automate assessment have shifted from haematoxylin and eosin (H&E)-stained images towards cytokeratin immunohistochemistry. The aim of this study was to compare manual H&E and cytokeratin assessment methods with a semi-automated approach built within QuPath open-source software. TB was assessed in cores from the advancing tumour edge in a cohort of stage II/III colon cancers (n = 186). The total numbers of buds detected with each method were as follows: manual H&E, n = 503; manual cytokeratin, n = 2290; and semi-automated, n = 5138. More than four times the number of buds were identified manually with cytokeratin assessment than with H&E assessment. One thousand seven hundred and thirty-four individual buds were identified with both manual and semi-automated assessments applied to cytokeratin images, representing 75.7% of the buds identified manually (n = 2290) and 33.7% of the buds detected with the semi-automated method (n = 5138). Higher semi-automated TB scores were due to any discrete area of cytokeratin immunopositivity within an accepted area range being identified as a bud, regardless of shape or crispness of definition, and to the inclusion of tumour cell clusters within glandular lumina ('luminal pseudobuds'). Although absolute numbers differed, semi-automated and manual bud counts were strongly correlated across cores (ρ = 0.81, P < 0.0001). All methods of TB assessment demonstrated poorer survival associated with higher TB scores. We present a new QuPath-based approach to TB assessment, which compares favourably with established methods and offers a freely available, rapid and transparent tool that is also applicable to whole slide images.
Highlights
Tumour budding (TB) is the histological manifestation of local tumour cell dissemination, usually most evident at the invasive front region of a tumour mass
Encouraging data was emerging at that time regarding TB assessment by cytokeratin (CK) immunohistochemistry (IHC), most of the established evidence was based on haematoxylin and eosin (H&E) assessment
The consensus preference from International Tumour Budding Consensus Conference (ITBCC) was for H&E staining in conjunction with a three-tier scoring system within a “hot spot” field area normalised to 0.785 mm2
Summary
Tumour budding (TB) is the histological manifestation of local tumour cell dissemination, usually most evident at the invasive front region of a tumour mass. TB is an established prognostic factor in a number of solid tumours [1], it has been most extensively studied in colorectal cancer (CRC). TB has been shown to have prognostic value in all other stages of CRC, with most evidence reported for stage II disease [1,3,4]. In an attempt to address this issue in 2016, the International Tumour Budding Consensus Conference (ITBCC) established a consensus definition of a tumour bud, namely a single tumour cell or tumour cell cluster of up to four cells, and an agreed histopathological method of assessment [8]. Encouraging data was emerging at that time regarding TB assessment by cytokeratin (CK) immunohistochemistry (IHC), most of the established evidence was based on haematoxylin and eosin (H&E) assessment. The consensus preference from ITBCC was for H&E staining in conjunction with a three-tier scoring system within a “hot spot” field area normalised to 0.785 mm
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