Analytical validation of a standardized scoring protocol for Ki67: phase 3 of an international multicenter collaboration

Abstract

Pathological analysis of the nuclear proliferation biomarker Ki67 has multiple potential roles in breast and other cancers. However, clinical utility of the immunohistochemical (IHC) assay for Ki67 immunohistochemistry has been hampered by unacceptable between-laboratory analytical variability. The International Ki67 Working Group has conducted a series of studies aiming to decrease this variability and improve the evaluation of Ki67. This study tries to assess whether acceptable performance can be achieved on prestained core-cut biopsies using a standardized scoring method. Sections from 30 primary ER+ breast cancer core biopsies were centrally stained for Ki67 and circulated among 22 laboratories in 11 countries. Each laboratory scored Ki67 using three methods: (1) global (4 fields of 100 cells each); (2) weighted global (same as global but weighted by estimated percentages of total area); and (3) hot-spot (single field of 500 cells). The intraclass correlation coefficient (ICC), a measure of interlaboratory agreement, for the unweighted global method (0.87; 95% credible interval (CI): 0.81–0.93) met the prespecified success criterion for scoring reproducibility, whereas that for the weighted global (0.87; 95% CI: 0.7999–0.93) and hot-spot methods (0.84; 95% CI: 0.77–0.92) marginally failed to do so. The unweighted global assessment of Ki67 IHC analysis on core biopsies met the prespecified criterion of success for scoring reproducibility. A few cases still showed large scoring discrepancies. Establishment of external quality assessment schemes is likely to improve the agreement between laboratories further. Additional evaluations are needed to assess staining variability and clinical validity in appropriate cohorts of samples.

Analytical validation of a standardised scoring protocol for Ki67 immunohistochemistry on breast cancer excision whole sections: an international multicentre collaboration

“…Therefore, in Phase IIIA, we assessed whether acceptable performance could be achieved on core‐cut biopsies using a standardised method with two distinct methods of scoring field selection: global (four representative fields, counting 100 nuclei each) and hot‐spot (one field with highest Ki67, counting 500 nuclei). The global method achieved acceptable interobserver reproducibility (ICC = 0.87; 95% CI = 0.81–0.93) according to our prespecified criteria, whereas the hot‐spot method did not (ICC = 0.84; CI = 0.77–0.92) …”

“…The scoring methods used were the same as those employed in the Phase IIIA study: (i) a global assessment that is weighted according to the estimated percentage of the total cancer area covered by each of high, medium, low or negligible Ki67 staining levels; (ii) an unweighted global assessment; and (iii) assessment of Ki67 only in a ‘hot‐spot’ area.…”

“…The International Ki67 Working Group (IKWG) has undertaken a systematic multiphase programme to determine whether Ki67 scoring can be standardised and analytically validated throughout laboratories . In Phase I, as assessed by the intraclass correlation coefficient (ICC) estimate of interobserver reproducibility, differences in pathologists’ visual interpretation were the main source of variability (ICC = 0.71, 95% credible interval (CI) = 0.47–0.78) .…”

Assessing Tumor-infiltrating Lymphocytes in Solid Tumors: A Practical Review for Pathologists and Proposal for a Standardized Method From the International Immunooncology Biomarkers Working Group: Part 1: Assessing the Host Immune Response, TILs in Invasive Breast Carcinoma and Ductal Carcinoma In Situ, Metastatic Tumor Deposits and Areas for Further Research

“…Many potentially useful biomarkers such as the Ki67 proliferative index in breast cancer suffer from a plethora of non-comparable methods, which affects the level of evidence that can be obtained and prevents uniform clinical implementation. International efforts at standardization are also being explored for this important biomarker [45]. …”

Neoadjuvant Trials in ER+ Breast Cancer: A Tool for Acceleration of Drug Development and Discovery

“…The interobserver concordance is higher for low and high Ki67 values, which manifest the difficulty of establishing a cut-off on the intermediates Ki67 values for making clinical decisions. In order to decrease this variability, the International Ki67 Working Group has conducted several studies to analytically validate and standardize Ki67 evaluation across laboratories (63,64), and recommended that an improved inter-observer reproducibility can be achieved on centrally stained tissue sections after training observers on a standardized visual scoring method. Another source of variability, which cannot be diminished through adoption of standard operating procedures, is intratumor heterogeneity of Ki67 levels (65,66).…”