Higher intratumoral infiltrated Foxp3+ Treg numbers and Foxp3+/CD8+ ratio are associated with adverse prognosis in resectable gastric cancer

Abstract

Purpose: The aim of the present study was to investigate the prognostic value of tumor-infiltrated lymphocytes (TILs), especially the prognostic value of Foxp3+ regulatory T cells (Tregs), CD8+ CTLs and Tregs/CD8+ ratios in gastric cancer patients after R0 resection.

Patients and methods: From 133 patients, CD4+, CD8+ and Foxp3+ TILs were assessed by immunohistochemistry in tissue microarrays and N1 regional lymph nodes sections containing gastric cancer. The prognostic effects of low- or high-density TIL subsets were evaluated by Cox regression and Kaplan-Meier analysis using median values as cutoff, while the effects of Foxp3+/CD8+ ratios were evaluated using the value determined by ROC cure analysis as cutoff.

Results: It was found that CD4+ and CD8+ TILs were not associated with overall survival (OS). In the tumor sites, higher Foxp3+ Tregs/CD8+ ratio was an independent factor for worse OS (multivariate analysis HR = 2.827, P = 0.037). The 1-year, 2-year and 3-year OS rates were 90, 77.5 and 70% for the group with intratumoral high Tregs/CD8+ ratio, compared with 100, 94.3 and 90.5% for the group with intratumoral low ratio. At the same time, the presence of intratumoral high Foxp3+ Tregs was also associated with worse OS (log rank test, P = 0.025); however, it was not an independent predictor and correlated with intratumoral Foxp3+ Tregs/CD8+ ratio (chi(2) test, P < 0.001). Although the infiltration of Foxp3+ Tregs in N1 regional lymph nodes was associated with lymph node metastasis (P = 0.028), it was not associated with prognosis (P = 0.458).

Conclusions: Intratumoral high Foxp3+ Tregs/CD8+ ratio was an independent predictor for the prognosis of gastric cancer. It can be inferred that a combination of deletion of Tregs and stimulation of CD8+ effector T cells may be an effective immunotherapy to prolong survival after surgery.

Fig. 1

Immunohistochemical staining of different tissue microarrays in gastric cancer pathological specimens. Specimens were collected from peritumor, intratumor and tumor-draining lymph nodes followed by immunohistochemical staining for CD4+, CD8+ and Foxp3+. The slides were viewed with light microscopy (magnification ×400). CD4+ and CD8+ staining were noted on the cell surface, whereas Foxp3+ staining was confined to the nucleus

Fig. 2

Different frequency of TILs in different tissues. Low power showing core from tissue microarray (magnification ×100). The frequency of intratumoral TILs was significantly higher than in peritumor (paired-sample t test, **P < 0.001); while the frequency of TILs in lymph nodes was significantly higher than in intratumor (paired-sample t test, **P < 0.001)

Fig. 3

K–M analysis of overall survival for intratumoral Foxp3+ and Foxp3+/CD8. a Low Foxp3+ was associated with prolonged survival (*P = 0.025); b low rate of Foxp3+ and CD8 was also associated with improved survival (**P = 0.002)

Fig. 4

K–M analysis for the direct correlation between the infiltration of TILs and clinical overall survival (OS) after surgery. a, b The correlation of introtumor infiltrated CD4+, CD8+ T cells with OS. c, d The correlation of infiltrated CD4+, CD8+ T cells in L.N with OS. e, f The correlation of infiltrated Foxp3+ Treg cells with OS