Relevance of tumour-infiltrating lymphocytes, PD-1 and PD-L1 in patients with high-risk, nodal-metastasised breast cancer of the German Adjuvant Intergroup Node–positive study

Highlights

• Tumour infiltrating lymphocyts (TILs) are predictive for dose-dense EPC therapy as compared to dose dense EC-PwX in node-positive breast cancer.

• TILs correlate with PD-1 and PD-L1 expression.

• TILs, PD-1 and PD-L1 show the highest levels in triple-negative breast cancer (TNBC).

• TILs and PD-1–positive immune cells have prognostic impact in TNBC.

• PD-L1 expression is more common in immune than in tumor cells but has no prognostic value.

Abstract

Background

Immune cell infiltration in breast cancer is important for the patient's prognosis and response to systemic therapies including immunotherapy. We sought to investigate the prevalence of tumour-infiltrating lymphocytes (TILs) and their association with immune checkpoints such as programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1) in high-risk, node-positive breast cancer of the adjuvant German Adjuvant Intergroup Node–positive (GAIN-1) trial.

Patients and methods

We evaluated TILs by haematoxylin and eosin staining and PD-1 and PD-L1 (SP263 assay) expression by immunohistochemistry in 1318 formalin-fixed, paraffin-embedded breast carcinomas. The association of TILs with PD-1, PD-L1, molecular intrinsic subtypes, outcome and therapy regimens (dose-dense [dd] epirubicin, paclitaxel and cyclophosphamide [EPC] and dd epirubicin, cyclophosphamide, paclitaxel and capecitabine [EC-PwX]) was statistically tested.

Results

Overall TILs density was significantly associated with the expression of PD-1 and PD-L1 in immune cells (each p < 0.0001) and PD-L1 in tumour cells (p = 0.0051). TILs were more common in triple-negative breast cancer (TNBC) and human epidermal growth factor receptor 2 (HER2)-positive tumours (each p < 0.0001). On multivariate Cox regression analyses, patients with breast cancer without TILs had an unfavourable disease-free survival (DFS) in the EPC arm compared with the EC-PwX arm (hazard ratio [HR] = 0.69 [0.44–1.06], p = 0.0915); but no differences were seen in tumours with TILs (HR = 1.24 [0.92–1.67], p = 0.1566, interaction p = 0.0336). PD-1–positive immune cells in TNBC were associated with a significantly better DFS (HR = 0.50 [0.25–0.99], p = 0.0457). PD-L1 expression had no impact on patient outcome.

Conclusions

TILs predict the benefit of intensified ddEPC compared with ddEC-PwX therapy in node-positive, high-risk breast cancer. TILs, PD-1 and PD-L1 are linked to each other indicating tumour immunogenicity. Moreover, PD-1–positive immune cells have a positive prognostic impact in TNBC.

Clinical trial

NCT00196872.

Introduction

Tumour-infiltrating lymphocytes (TILs) are present in all molecular subtypes of breast cancer (BC) but most common in triple-negative and non-luminal HER2-positive tumours [1], [2], [3]. Several studies have shown that TILs predict prognosis and response to systemic therapies in BC. Patients with triple-negative BC (TNBC) with increased TILs have a better overall survival (OS) in the adjuvant setting compared with patients without TILs [3], [4], [5], [6], [7]. In contrast, there was no prognostic significance for TILs in hormone receptor–positive, HER2-negative BC [3], [4]. In neoadjuvant trials, increased TILs were associated with a higher pathological complete response (pCR) to systemic therapies, especially in TNBC and HER2-positive cancer [1], [8], [9]. Patients who achieve a pCR in these subtypes will have better survival [9], [10], [11], [12].

Furthermore, it was recently demonstrated that TILs and PD-L1–positive immune cells may predict the response to immune checkpoint therapies with pembrolizumab and atezolizumab in BC [13], [14], [15], [16], [17]. So far, additional biomarkers for the identification of patients with BC that will have a benefit of such immuno-oncologic therapies have not been established. In other tumour entities, such as malignant melanoma and non–small cell lung cancer, testing of PD-1 and/or PD-L1 by immunohistochemistry (IHC) is an approach for patient selection [18], [19], [20]. The data on the prevalence of PD-1 and PD-L1 expression in BC are conflicting, and their relation to TILs and molecular subtypes is not well known, especially not in patient cohorts with defined treatment and reasonable follow-up. Previous studies have resulted in variable findings, probably also based on the use of different staining assays, definitions of positivity, and scoring criteria [21], [22], [23]. The objective of this work was to evaluate the clinical validity of TILs and the expression of immune checkpoints such as PD-1 and PD-L1 as prognostic and predictive biomarkers in a prospective cohort of central pathological validated, molecularly diverse BC samples of the adjuvant German Adjuvant Intergroup Node–positive (GAIN-1) trial. Because previous studies have suggested that TILs are related to the response of certain chemotherapies such as carboplatin and anthracycline [3], [24], we also examined the impact of TILs in different chemotherapy regimens.