Polymorphism of formyl peptide receptor 1 (FPR1) reduces the therapeutic efficiency and antitumor immunity after neoadjuvant chemoradiotherapy (CCRT) treatment in locally advanced rectal cancer

Abstract

Immunosurveillance and immunoscavenging prompted by preoperative chemoradiotherapy (CCRT) may contribute to improve local control and increase survival outcomes for patients with locally advanced rectal cancer (LARC). In this study, we investigated several genotypes of pattern recognition receptors (PRRs) and their impact on therapeutic efficacy in LARC patients treated with CCRT. We found that homozygosis of formyl peptide receptor 1 (FPR1) (E346A/rs867228) was associated with reduced 5-year overall survival (OS) by Kaplan-Meier analysis (62% vs. 81%, p = 0.014) and multivariate analysis [hazard ratio (HR) = 3.383, 95% CI = 1.374-10.239, p = 0.007]. Moreover, in an animal model, we discovered that the FPR1 antagonist, Boc-MLF (Boc-1), reduced CCRT therapeutic efficacy and decreased cytotoxic T cells and T effector memory cells after chemoradiotherapy treatment. Pharmacologic inhibition of FPR1 by Boc-1 decreased T lymphocyte migration to irradiated tumor cells. Therefore, these results revealed that the FPR1 genotype participates in CCRT-elicited anticancer immunity by reducing T lymphocytes migration and infiltration, and that the FPR1-E346A CC genotype can be considered an independent biomarker for chemo- and radiotherapy outcomes.

Keywords: CCRT; Cancer immunity; DAMP; FPR1; ICD.

Fig. 1

The association between disease-free survival (DFS), overall survival (OS) and FPR1 genotypes in LARC a Kaplan–Meier curve showed that the FPR1-E348A CC genotype is associated with 10-year DFS. b Kaplan–Meier curve showed that the FPR1-E348A CC genotype is associated with 10-year OS

Fig. 2

FPR1 signaling is required for TIL infiltration and antitumor immunity in vivo. a The graphic scheme of the concurrent chemoradiotherapy regimen on BALB/c mice. b Tumor growth of CT26-driven colon carcinoma established in BALB/c mice (n = 5 per group) that were treated with concurrent chemoradiotherapy [CCRT, 5-FU (5 mg/kg) and 5 Gy for 2 fractions or CCRT/Boc-1 (25 mg/kg FPR inhibitor one hour before 5-FU). Tumor growth is calculated as the mean tumor volume ± SD over time. ***p < 0.001. ANOVA test. c Resected tumors were extracted and examined (n = 5). ***p < 0.001. ANOVA test. d TILs were isolated from resected tumors and analyzed by flow cytometry. Dot plot of CD8a⁺ and CD44⁺ TILs was based on the gating of CD45⁺ cells. e Statistical analyses of CD8a⁺ and CD8a⁺ CD44⁺ TILs density in resected tumors (n = 3 per group). The density (%/cm³) was calculated by percentage of the cells and adjusted by each tumor weight, which 1 g tumor is approximately 1 cm3. ***p < 0.001, **p < 0.01, *p < 0.05. ANOVA test. f Immunofluorescence stain with CD8a⁺ and granzyme B in the central tumor region. g and h The number of CD8a⁺ or CD8a⁺/granzyme B TILs was counted under high-power-field microscopy (n = 5). ***p < 0.001. ANOVA test

Fig. 3

FPR1 blockade affects T cells recruitment and closed contact formation with DCs in vitro.HCT116 cells were seeded and irradiated (10 Gy) on the lower well of Transwell for overnight culture. The CMTPX–labeled (Red) THP1-iDCs and/or CSFE-labeled (Green) Jurkat T lymphocytes were seeded on the upper wells for 24 h incubation. a The number of migrated THP1-iDCs into irradiated HCT116 cells with or without 20 μM Boc-1 treated (n = 5). Unpaired t test. b The number of migrated Jurkat T cells into irradiated HCT116 cells with or without 20 μM Boc-1 treated (n = 5). **p < 0.01 and ***p < 0.001. Unpaired t test. These data were obtained from at least three independent experiments. The values represent the means ± S.D. c The closed contact between THP-1-derived immature DCs and Jurkat T lymphocytes within irradiated HCT116 cells (200X). Scale bars = 20 μm. d Immunoblotting analysis of FPR1 expression. THP1-iDCs and Jurkat T lymphocytes were transfected with siRNA against negative control (NC) and FPR1. e The number of THP1-iDCs-siNC, THP1-iDCs-siFPR1, Jurkat-siNC and Jurkat-siFRP1 migrated into irradiated HCT116 cells (n = 3). ***p < 0.001. Unpaired t test. The values represent the means ± S.D