Differential intratumoral distributions of CD8 and CD163 immune cells as prognostic biomarkers in breast cancer

Abstract

Background: Tumor immune cell infiltrates are essential in hindering cancer progression and may complement the TNM classification. CD8+ and CD163+ cells have prognostic impact in breast cancer but their spatial heterogeneity has not been extensively explored in this type of cancer. Here, their potential as prognostic biomarkers was evaluated, depending on their combined densities in the tumor center (TC) and the tumor invasive margin (IM).

Methods: CD8+ and CD163+ cells were quantified by immunohistochemistry of formalin-fixed, paraffin-embedded (FFPE) tumor tissue samples from a cohort totaling 162 patients with histologically-confirmed primary invasive non-metastatic ductal breast cancer diagnosed between 2000 and 2015. Clinical follow-up (median 6.9 years) was available for 97 of these patients.

Results: Differential densities of CD8+ and CD163+ cells in the combined TC and IM compartments (i.e., high(H)/low(L), respectively for CD8+ cells and the reverse L/H combination for CD163+ cells) were found to have significant prognostic value for survival, and allowed better patient stratification than TNM stage, tumor size, lymph node invasion and histological grade. The combined evaluation of CD8+ and CD163+ cell densities jointly in TC and IM further improves prediction of clinical outcomes based on disease-free and overall survival. Patients having the favorable immune signatures had favorable clinical outcomes despite poor clinicopathological parameters.

Conclusions: Given the important roles of CD8+ and CD163+ cells in regulating opposing immune circuits, adding an assessment of their differential densities to the prognostic biomarker armamentarium in breast cancer would be valuable. Larger validation studies are necessary to confirm these findings.

Trial registrations: Study code: IRB-ID 6079/448/10-6-13 Date of approval: 10/06/2013 Retrospective study (2000-2010) First patient prospectively enrolled 14/2/2014.

Keywords: Breast cancer; CD163; CD8; Immunoscoring; Prognostic signature; Tumor infiltration.

Fig. 1

a Schematic representation of tumor center (TC) and invasive margin (IM). Representative images of low and high CD8+ and CD163+ cell infiltration densities in TC (b) and IM (c). d CD8+ and CD163+ counts in the tumor center and invasive margin for all patients analyzed (n = 162). Horizontal bars, median values

Fig. 2

Whisker plots (Tukey) of CD8+ (a, c, e, g) and CD163+ (i, k, m, o) counts in the tumor center (TC) and CD8+ (b, d, f, h) and CD163+ (j, l, n, p) counts in the invasive margin (IM) according to clinicopathological variables: Grade (a, b, i, j), T status (c, d, k, l), LN status (e, f, m, n) and TNM stage (g, h, o, p). G = grade; T = tumor size; LN = lymph node; TNM stage: early = I&IIA and advanced = IIB&III

Fig. 3

Kaplan-Meier curves illustrating DFS (a, c) and OS (b, d) for all patients analyzed according to the density (high = H or low = L) of CD8+ (a, b) or CD163+ (c, d) cells in TC or IM. Statistically significant differences or trends and hazard ratios between specific groups are given

Fig. 4

Kaplan-Meier curves illustrating DFS and OS for all patients analyzed according to the density of CD8+ or CD163+ cells in the combined tumor regions (TC and IM) (a-d). e, f shows DFS and OS for all patients analyzed according to the combined density of CD8+ and CD163+ cells in the combined tumor regions (TC and IM). For explanation of favorable or unfavorable combined immune signatures (FCIS and UCIS, respectively) see “Results”. HH/HH: high densities for both CD8+ and CD163+ cells jointly analyzed in the combined tumor regions (CT/IM); Rest: LL/LL, HH/LL, and LL/HH. Statistically significant differences and hazard ratios between specific groups are given

Fig. 5

Kaplan-Meier curves comparing DFS and OS for patients with favorable (FCIS) or unfavorable (UCIS) infiltration signatures stratified according to grade (a, b), T status (c, d), LN status (e, f) and TNM stage (g, h)

Fig. 6

Percentages of total population of patients (all; n = 162) or of patients at each histological grade (G1; n = 4, G2; n = 86,G3; n = 72); T status (T1; n = 70, T2; n = 81,T3; n = 10); node status (N0; n = 64, N1-3; n = 56, N > 3; n = 42) and pathological stage (stage I; n = 45, stage II; n = 72, stage III; n = 45) expressing FCIS or UCIS or rest (“rest” includes HH/HH; LL/LL; HH/LL and LL/HH)