PD-L1 protein expression in breast cancer.

AIMS: The immune checkpoint marker, Programmed cell death-ligand 1 (PD-L1), is expressed by both cancer epithelial cells and tumour-infiltrating immune cells (TICs) thus constituting a potential target for immunotherapy. This is of particular interest in triple negative breast cancer. In this study, we assessed the prognostic value of PD-L1 expression in tumour epithelial cells and TICs in a series of patients with breast cancer with long-term follow-up, and associations between PD-L1 expression and histopathological type and grade, proliferation and molecular subtype. METHODS: Using immunohistochemistry for PD-L1 in tissue microarrays, we assessed PD-L1 expression in 821 tumours. Expression of PD-L1 was assessed separately in the epithelial and stromal compartments and classified as <1%, ≥1% to <10% or ≥10% positive staining cells. We correlated PD-L1 expression in tumour epithelial cells and TICs with tumour characteristics using Pearson's χ2 test, and prognosis by cumulative incidence of death from breast cancer and Cox regression analyses. RESULTS: We found membranous staining in ≥1% of tumour epithelial cells in 53/821 cases (6.5%). Of these, 21 (2.6%) were ≥10%. Among TICs, staining (≥1%) was seen in 144/821 cases (17.6%). Of these, 62 were ≥10% (7.6%). PD-L1 was associated with high histopathological grade and proliferation, and the medullary and metaplastic patterns. In TICs, PD-L1 ≥1% found in 22/34 (34.4%) human epidermal growth factor receptor 2 type and 29/58 (50%) basal phenotype. An independent association between PD-L1 expression and prognosis was not observed. CONCLUSIONS: PD-L1 is expressed more frequently in TICs than tumour epithelial cells. Expression in TICs is associated with aggressive tumour characteristics and non-luminal tumours but not with prognosis.

Oestrogen receptor low positive breast cancer: associations with prognosis.

PURPOSE: In this study of oestrogen receptor (ER) Low Positive breast cancers (BC) in three large cohorts of BC patients, we assess associations between levels of ER expression and tumour characteristics and prognosis. METHODS: Cases were stratified into patients unlikely to have received adjuvant therapy according to treatment guidelines at time of diagnosis (before 1995), and those who could have received adjuvant therapy (diagnosed in 1995 or later). ER status was divided into < 1%; ≥ 1 < 10%; ≥ 10%. Results were correlated with time of diagnosis, histopathological grade, proliferation status, and molecular subtypes, using Pearson's Chi-square test. For prognosis, hazard ratios and cumulative incidence of death from BC were used. RESULTS: Of the 1955 tumours, 65 (3.3%) were ER Low Positive (ER ≥ 1 < 10%). Overall, the highest proportion of ER Low Positive tumours was observed among Luminal B (HER2 +) subtype (9.4%) and grade 3 tumours (4.3%). The risk of death from BC was lower in ER Low Positive and ER ≥ 10% compared to ER-negative cases. Compared to patients diagnosed before 1995, women diagnosed in 1995 or later showed a higher proportion of ER Low Positive BCs, and their tumours were of smaller size, lower grade, and lower proliferative status. There was no significant difference in prognosis compared to those with ER ≥ 10% tumours. CONCLUSION: Women with ER Low Positive tumours diagnosed in a time period when adjuvant therapy was available had tumours of smaller size, lower grade, and lower proliferative status, and similar prognosis to those with ER ≥ 10% compared to women diagnosed earlier.

PAK1 copy number in breast cancer-Associations with proliferation and molecular subtypes.

INTRODUCTION: P21-activated kinase 1 (PAK1) is known to be overexpressed in several human tumour types, including breast cancer (BC). It is located on chromosome 11 (11q13.5-q14.1) and plays a significant role in proliferation in BC. In this study we aimed to assess PAK1 gene copy number (CN) in primary breast tumours and their corresponding lymph node metastases, and associations between PAK1 CN and proliferation status, molecular subtype, and prognosis. In addition, we aimed to study associations between CNs of PAK1 and CCND1. Both genes are located on the long arm of chromosome 11 (11q13). METHODS: Fluorescence in situ hybridization for PAK1 and Chromosome enumeration probe (CEP)11 were used on tissue microarray sections from a series of 512 BC cases. Copy numbers were estimated by counting the number of fluorescent signals for PAK1 and CEP11 in 20 tumour cell nuclei. Pearson's x2 test was performed to assess associations between PAK1 CN and tumour features, and between PAK1 and CCND1 CNs. Cumulative risk of death from BC and hazard ratios were estimated in analysis of prognosis. RESULTS: We found mean PAK1 CN ≥4<6 in 26 (5.1%) tumours, and CN ≥ 6 in 22 (4.3%) tumours. The proportion of cases with copy number increase (mean CN ≥4) was highest among HER2 type and Luminal B (HER2-) tumours. We found an association between PAK1 CN increase, and high proliferation, and high histological grade, but not prognosis. Of cases with PAK1 CN ≥ 6, 30% also had CCND1 CN ≥ 6. CONCLUSIONS: PAK1 copy number increase is associated with high proliferation and high histological grade, but not with prognosis. PAK1 CN increase was most frequent in the HER2 type and Luminal B (HER2-) subtype. PAK1 CN increase is associated with CN increase of CCND1.

Visual and digital assessment of Ki-67 in breast cancer tissue - a comparison of methods.

BACKGROUND: In breast cancer (BC) Ki-67 cut-off levels, counting methods and inter- and intraobserver variation are still unresolved. To reduce inter-laboratory differences, it has been proposed that cut-off levels for Ki-67 should be determined based on the in-house median of 500 counted tumour cell nuclei. Digital image analysis (DIA) has been proposed as a means to standardize assessment of Ki-67 staining in tumour tissue. In this study we compared digital and visual assessment (VA) of Ki-67 protein expression levels in full-face sections from a consecutive series of BCs. The aim was to identify the number of tumour cells necessary to count in order to reflect the growth potential of a given tumour in both methods, as measured by tumour grade, mitotic count and patient outcome. METHODS: A series of whole sections from 248 invasive carcinomas of no special type were immunohistochemically stained for Ki-67 and then assessed by VA and DIA. Five 100-cell increments were counted in hot spot areas using both VA and DIA. The median numbers of Ki-67 positive tumour cells were used to calculate cut-off levels for Low, Intermediate and High Ki-67 protein expression in both methods. RESULTS: We found that the percentage of Ki-67 positive tumour cells was higher in DIA compared to VA (medians after 500 tumour cells counted were 22.3% for VA and 30% for DIA). While the median Ki-67% values remained largely unchanged across the 100-cell increments for VA, median values were highest in the first 1-200 cells counted using DIA. We also found that the DIA100 High group identified the largest proportion of histopathological grade 3 tumours 70/101 (69.3%). CONCLUSIONS: We show that assessment of Ki-67 in breast tumours using DIA identifies a greater proportion of cases with high Ki-67 levels compared to VA of the same tumours. Furthermore, we show that diagnostic cut-off levels should be calibrated appropriately on the introduction of new methodology.

The association of women's birth size with risk of molecular breast cancer subtypes: a cohort study.

BACKGROUND: Because birth size appears to be positively associated with breast cancer risk, we have studied whether this risk may differ according to molecular breast cancer subtypes. METHODS: A cohort of 22,931 women born 1920-1966 were followed up for breast cancer occurrence from 1961 to 2012, and 870 were diagnosed during follow-up. Archival diagnostic material from 537 patients was available to determine molecular breast cancer subtype, specified as Luminal A, Luminal B (human epidermal growth factor receptor 2 (HER2)-), Luminal B (HER2+), HER2 type, and Triple negative (TN) breast cancer. Information on the women's birth weight, birth length and head circumference at birth was used to estimate hazard ratios (HR) with 95% confidence intervals (CI) for each molecular subtype, applying Cox regression, and stratified by maternal height. RESULTS: Birth length (per 2 cm increments) was positively associated with Luminal A (HR = 1.2, 95% CI, 1.0-1.3), Luminal B (HER2+) (HR = 1.3, 95% CI, 1.0-1.7), and TN breast cancer (HR = 1.4, 95% CI, 1.0-1.9). No clear association was found for birth weight and head circumference. The positive associations of birth length were restricted to women whose mothers were relatively tall (above population median). CONCLUSION: We found a positive association of birth length with risk of Luminal A, Luminal B (HER2+) and TN breast cancer that appears to be restricted to women whose mothers were relatively tall. This may support the hypothesis that breast cancer risk is influenced by determinants of longitudinal growth and that this finding deserves further scrutiny.