Accurate expression of PD-L1/L2 in lung adenocarcinoma cells: A retrospective study by double immunohistochemistry

Abstract

The percentage of programmed death ligand 1 (PD-L1) positivity in cancer cells, named as the tumor proportion score, is considered to be a predictive biomarker for anti-PD-1/PD-L1 therapy in lung cancer. PD-L1 is expressed on not only cancer cells but also on immune cells, including macrophages. Although previous studies related to PD-L1/2 expression in cancer tissues have been generally based on single immunohistochemistry (IHC), in the present study, we attempted to evaluate accurate PD-L1/2 expression in cancer cells in lung adenocarcinoma cells using double IHC to also evaluate macrophages. Of the 231 patients, PD-L1 expression was negative in 169 patients (73.2%), 1%-49% positive in 47 patients (20.3%), and ≥50% positive in 15 patients (6.5%). Interestingly, PD-L1 positivity was decreased when using double IHC compared with the estimation by single IHC. High PD-L1 expression was associated with high-grade cancer cells and in higher stage cancer. PD-L2 was negative in 109 patients (47.2%), 1%-49% positive in 50 patients (21.6%), and ≥50% positive in 72 patients (31.2%). The number of PD-L2-positive patients was increased in cases that had an epidermal growth factor receptor (EGFR) mutation and in lower stage cancer. Thirty-five patients (15.2%) were positive for both PD-L1 and PD-L2, whereas 81 patients (35.1%) were negative for both PD-L1 and PD-L2. Log-rank analysis showed that progression-free survival and overall survival were significantly the longest in the PD-L1-negative and PD-L2-positive groups (P < .0001 and P = .0120). We observed lower PD-L1 or PD-L2 expression in lung adenocarcinoma than previously reported. Double IHC for macrophages may help clinicians to evaluate PD-L1 or PD-L2 expression specifically in cancer cells.

Keywords: PD-L1; PD-L2; lung adenocarcinoma; macrophage; tumor proportion score.

Figure 1

Anti‐programmed death ligand 1 (PD‐L1) immunohistochemistry (IHC). A, Single IHC of PD‐L1 (upper panels) and double IHC of PD‐L1 and Iba‐1 (a pan‐macrophage marker) (lower panels). Representative images from PD‐L1‐negative (left side) and PD‐L1‐positive (right side) cases are presented. PD‐L1 and Iba‐1 signals were labeled as brown and green, respectively. B, Tumor proportion score determined by single IHC of PD‐L1 and double IHC of PD‐L1 and Iba‐1. Numbers represent the number of patients for each group

Figure 2

Kaplan‐Meier analysis of PD‐L1 tumor proportion score (TPS) and survival rate. PD‐L1 expression was divided into three or two groups according to TPS. Statistical analyses related to progression‐free survival (A) and overall survival (B) were performed

Figure 3

Anti‐programmed death ligand 2 (PD‐L2) immunohistochemistry (IHC). A, Single IHC of PD‐L2 (upper panels) and double IHC of PD‐L2 and Iba‐1 (lower panels). Representative images from PD‐L2‐negative (left side) and anti‐programmed death ligand 1 (PD‐L1)‐positive (right side) cases are presented. PD‐L2 and Iba‐1 signals were labeled as brown and green, respectively. B, The number of cases for each PD‐L1 and PD‐L2 tumor proportion score (TPS) and percentages of PD‐L2 TPS by double IHC are presented

Figure 4

Kaplan‐Meier analysis of anti‐programmed death ligand 2 (PD‐L2) tumor proportion score (TPS) and survival rate. PD‐L2 expression was divided into three or two groups according to TPS. Statistical analyses related to progression‐free survival (A) and overall survival (B) were performed

Figure 5

Anti‐programmed death ligand 1 (PD‐L1) and anti‐programmed death ligand 2 (PD‐L2) expression in cancer cells. A, PD‐L1 and PD‐L2 expression was evaluated in 231 patients. B, Patients were divided into four groups based on PD‐L1 and PD‐L2 expression and Kaplan‐Meier analysis was performed