Expression of PD1/PDL1 in gastric cancer at different microsatellite status and its correlation with infiltrating immune cells in the tumor microenvironment

Abstract

Objective: The microsatellite status and tumor immune microenvironment have a remarkable influence on tumor immunotherapy. This study was performed to investigate programmed cell death protein 1/programmed death ligand 1 (PD1/PDL1) expression and their correlations with CD8+ T cell/CD68+ macrophage (CD68+ M) densities in gastric cancer (GC) at different microsatellite statuses. Methods: The expression of MLH1, PMS2, MSH2, and MSH6 was detected via immunohistochemistry (IHC) to determine the microsatellite status in 215 GC samples obtained from surgical resections. Furthermore, the expression of PD1, PDL1, CD8, and CD68 was detected in the samples via IHC, and the differences and correlations in GC at different microsatellite statuses were then analyzed. PDL1 expression in tumor cells was labeled as PDL1[T], while expression of PD1 and PDL1 in tumor-infiltrating immune cells was labeled as PD1 and PDL1, respectively. Kaplan-Meier analysis was used to evaluate the significance of PD1/PDL1 expression in determining overall survival. Multivariate Cox regression analysis was performed using SPSS software. P-values were determined using the log-rank test. Results: Our results indicated that PD1, PDL1[T], and PDL1 positivity rates were 59%, 35%, and 57% in 46 microsatellite unstable (MSI) GCs and 45%, 22%, and 40% in 169 microsatellite stable (MSS) GCs, respectively. Compared with MSS GC, PD1, PDL1[T], and PDL1 expression was higher in MSI GC (P = 0.109, 0.090, and 0.044, respectively). Additionally, CD8+ T cell and CD68+ M densities were higher in MSI GC than in MSS GC (P = 0.537 and <0.001, respectively). Additionally, CD8+ T cell/CD68+ M densities were evaluated according to tumor center and invasion front. We found that PD1 expression was significantly correlated with CD8+ T cell density at the invasion front of the MSI GC (P = 0.031), whereas PDL1 expression was significantly correlated to high CD68+ M density in the tumor center and invasion front of MSS GC (P = 0.001 and 0.014, respectively). Survival analysis showed that patients with PD1-positive and PDL1[T]/PDL1-negative GC had better prognosis (P = 0.012, 0.005, and 0.022, respectively). Multivariate Cox survival analysis showed that PDL1[T] was an independent prognostic factor for GC. Conclusion: The results suggested that PD1/PDL1 expression and immune response varied at different microsatellite statuses in GC. PD1/PDL1 expression was correlated with CD8+ T cell/CD68+ M densities in GC at different microsatellite statuses, especially at the invasion front. The patients exhibiting high PD1/PDL1 expression or high CD8+ T cell/CD68+ M densities MSI GC might be potential beneficiaries of PD1/PDL1 immunotherapy.

Keywords: CD68; CD8; PD1; PDL1; gastric cancer; tumor immune microenvironment.

Figure 1

Representative immunohistochemical images of MSI/MSS GC (×200). MLH1, PMS2, MSH2 and MSH6 were all localized in GC cell nucleus. The brown yellow was considered positive, while the absence of staining was considered negative. MSI GC: MLH1 and PMS2 were negative(A,B), MSH2 and MSH6 were positive(C,D); MSS GC: MLH1, PMS2, MSH2 and MSH6 were all positive(E-H).

Figure 2

Representative immunohistochemical images of PD1, PDL1[T], PDL1, CD8 and CD68 in MSI and MSS GC (×200). PD1, PDL1[T], PDL1, CD8 and CD68 were all localized in the membrane and/or cytoplasm. PD1, CD8 and CD68 was all stained in tumor infiltrating immune cells, while PDL1 was stained in tumor cells (labeled as PDL1[T]) and tumor infiltrating immune cells (labeled as PDL1). PD1, PDL1[T] and PDL1 expression in MSI GC was greater than those in MSS GC (P=0.109,0.090 and 0.044, respectively). CD8+T cells/CD68+M density in MSI GC was higher than those in MSS GC (P=0.537 and <0.001, respectively).

Figure 3

The correlations between PD1/PDL1 expression and CD8+T cells/CD68+M density at different locations in MSI and MSS GC. PD1 expression was significantly related to CD8+T cells high density at invasion front of MSI GC (P=0.031). PDL1 expression was significantly related to CD68+M high density in tumor center and invasion front of MSS GC (P=0.001 and 0.014, respectively).

Figure 4

Kaplan-Meier overall survival curves of PD1/PDL1 expression or CD8+T cells/CD68+M density in GC. GC patients with MSI, PD1 positive, and PDL1[T]/PDL1 negative had better prognosis (P=0.006, 0.012, 0.005 and 0.022, respectively). There are no correlations between CD8+T cells/CD68+M density and prognosis in GC (P=0.870 and 0.985, respectively).