Background: Triple negative breast cancer (TNBC) represents approximately 15-20% of all breast cancers and 35% of breast cancer death. TNBC has a highly aggressive behavior with the increased risk of early metastasis and relapse and poor survival rate. Given that TNBC is associated with higher levels of tumor-infiltrating lymphocytes and programmed cell death ligand 1 (PD-L1), immune checkpoint inhibitors (ICI) are considered as the appropriate treatment of patients with TNBC. However, it has been reported that ICI monotherapy is less effective compared with combination therapy. Recent several studies suggest that gut microbiome plays a critical role in the efficacy of ICI therapy. Thus, we investigated the effects of Bifidobacterium longum RAPO supplementation on anti-PD-1 therapy against TNBC and the underlying mechanisms in vivo.
Methods: The 4T1 murine metastatic breast cancer cells were injected s.c into the flank of female BALB/c mice. When tumors reached 50-150 mm3, mice were randomly assigned to one of four groups; tumor control (TC), Anti-PD-1, B. longum RAPO (RAPO), or Anti-PD-1+RAPO (n=10 per group). Anti-PD-1 antibody was injected i.p. 5 times at 3 day intervals. B. longum RAPO was orally administered daily from 2 days before the first injection of anti-PD-1. Spleen and tumor tissues were analyzed by flow cytometry, qRT-PCR, and immunohistochemistry, and fecal samples were collected for 16S rRNA gene sequencing.
Results: On day 12, tumor volume was significantly inhibited in Anti-PD-1+RAPO group compared with TC and Anti-PD-1 groups. Higher levels of PD-L1 IHC score and PD-L1 mRNA expression were observed in tumor tissues of Anti-PD-1+RAPO group. Anti-PD-1+RAPO group was also associated with the increased levels of spleen NKT cells and tumor NK cells. The proportion of pro-tumor M2 macrophages and M2/M1 ratio were significantly decreased in tumor tissues of Anti-PD-1+RAPO group than those of Anti-PD-1 group. Similarly, combination therapy of Anti-PD-1 with RAPO significantly reduced mRNA expression of M2 markers such as IL10 and Arg1, while it augmented that of anti-tumor cytokines including IFNγ and TNFα compared with Anti-PD-1 treatment alone. In addition, the relative abundance of the genus Bifidobacterium was remarkably enriched in Anti-PD-1+RAPO group compared with other groups. In Anti-PD-1+RAPO group, the relative abundance of the genus Akkermansia and Lachnospiraceae NK4A136 group were significantly increased after administration for 2 weeks.
Conclusions: Our data suggests that combination therapy of B. longum RAPO with anti-PD-1 modulates anti-tumor immunity and gut microbiota, contributing to the better response to anti-PD-1 immunotherapy in mice with TNBC. This study provides evidence that B. longum RAPO might be a potential candidate for immunomodulatory pharmabiotics in cancer immunotherapy.
Citation Format: Hyeyoon Kim, Rira Oh, Sangjun Park, Geun Eog Ji, Myeong Soo Park, Sung-Eun Kim. Bifidobacterium longum RAPO enhances efficacy of anti-PD-1 immunotherapy in a mouse model of triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 72.