Prebiotic-Induced Anti-tumor Immunity Attenuates Tumor Growth - PubMed (original) (raw)

. 2020 Feb 11;30(6):1753-1766.e6.

doi: 10.1016/j.celrep.2020.01.035.

Lisa Elmén 1, Igor Segota 1, Yibo Xian 2, Roberto Tinoco 1, Yongmei Feng 1, Yu Fujita 1, Rafael R Segura Muñoz 2, Robert Schmaltz 2, Linda M Bradley 1, Amanda Ramer-Tait 2, Raphy Zarecki 3, Tao Long 1, Scott N Peterson 4, Ze'ev A Ronai 5

Affiliations

Prebiotic-Induced Anti-tumor Immunity Attenuates Tumor Growth

Yan Li et al. Cell Rep. 2020.

Abstract

Growing evidence supports the importance of gut microbiota in the control of tumor growth and response to therapy. Here, we select prebiotics that can enrich bacterial taxa that promote anti-tumor immunity. Addition of the prebiotics inulin or mucin to the diet of C57BL/6 mice induces anti-tumor immune responses and inhibition of BRAF mutant melanoma growth in a subcutaneously implanted syngeneic mouse model. Mucin fails to inhibit tumor growth in germ-free mice, indicating that the gut microbiota is required for the activation of the anti-tumor immune response. Inulin and mucin drive distinct changes in the microbiota, as inulin, but not mucin, limits tumor growth in syngeneic mouse models of colon cancer and NRAS mutant melanoma and enhances the efficacy of a MEK inhibitor against melanoma while delaying the emergence of drug resistance. We highlight the importance of gut microbiota in anti-tumor immunity and the potential therapeutic role for prebiotics in this process.

Keywords: MEK inhibitor; anti-tumor immunity; colon cancer; gut microbiota; inulin; melanoma; mucin; prebiotics; therapy resistance.

Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.

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Conflict of interest statement

Declaration of Interests Z.A.R. is a co-founder and serves as a scientific advisor to Pangea Therapeutics. All other authors declare no competing interests. A patent application detailing the findings disclosed in this manuscript has been filed by SBP Discovery.

Figures

Figure 1.

Figure 1.. Administration of Mucin or Inulin Reduces Tumor Growth and Induces Anti-tumor Immunity

(A) Growth of YUMM1.5 tumors that were subcutaneously transplanted in syngeneic C57BL/6 mice. Mice were provided with a control diet, 3% mucin in drinking water, or 15% inulin-supplemented chow starting 14 days before tumor inoculation (control, n = 12; mucin, n = 15; inulin, n = 15). (B) Quantification of tumor-infiltrating total CD45+ cells and effector (CD44hi) CD4+ or CD8+ T cells from mice treated as in (A) (control, n = 9; mucin, n = 10; inulin, n = 10). (C) Quantification of tumor-infiltrating, IFN-γ-producing CD4+ T cells from mice treated as in (A) (n = 10). (D) Quantification of tumor-infiltrating total DCs and DC subsets in mice treated as in (A) (control, n = 9; mucin, n = 10; inulin, n = 10). (E) MFI of MHC class I and MHC class II on tumor-infiltrating DCs in mice treated as in (A) (control, n = 9; mucin, n = 10; inulin, n = 10). (F) Wild-type C57BL/6 mice (n = 12) were fed control or 15% inulin-supplemented chow starting 14 days before subcutaneous (s.c.) injection of YUMM1.5 melanoma cells (1 × 106). Anti-mouse Thy1.2 or control immunoglobulin G (IgG, 400 μg) were injected two times a week starting 3 days after tumor inoculation (n = 12). Tumor volume was assessed two times a week. FACS analysis revealed >90% depletion of blood CD4+ and CD8+ T cells on day 8 after tumor inoculation. Data are representative of three independent experiments (A–E) or one experiment (F). Graphs show the mean ± SEM. *p < 0.05, **p < 0.005, ***p < 0.001, ****p < 0.0001 by two-way ANOVA with Bonferroni’s correction (A and F) or by one-way ANOVA with Bonferroni’s correction (B–E).

Figure 2.

Figure 2.. Mucin and Inulin Induce Enhanced Expression of Immunoregulatory Genes in Tumors

(A) qPCR analysis of immune-related genes in subcutaneously transplanted melanoma grown in syngeneic C57BL/6 mice that received a control diet, 3% mucin in drinking water, or 15% inulin-supplemented chow starting 14 days before tumor inoculation (n = 6). (B) Quantification of CD45.1+ OT-I CD8+ T cells in the tumor-draining lymph nodes (TdLN) and non-draining lymph nodes (ndLN) of C57BL/6 mice (CD45.2+) treated with or without mucin and injected with B16-OVA melanoma cells (TdLN, n = 7; ndLN, n = 8). Right dot plots show gating of CD45.1+ CD8+ cells. (C) Serum cytokine and chemokine levels in naive WT mice treated with or without mucin (n = 10). (D) Serum cytokines in WT mice treated with or without mucin on day 10 after tumor inoculation (n = 10). Data are representative of three independent experiments (A and B) or one experiment (C and D). Graphs show the mean ± SEM. *p < 0.05, **p < 0.005, ***p < 0.001, ****p < 0.0001 by one-way ANOVA with Tukey’s correction (A) or by two-tailed t test or Mann-Whitney U test (B–D).

Figure 3.

Figure 3.. Mucin and Inulin Modulate the Composition and Diversity of Gut Microbiota

(A and B) Principal-component analysis of all taxa enumerated in fecal microbiota of control and mucin-treated (A) or inulin-treated (B) C57BL/6 mice, examined before prebiotic treatment, before subcutaneous injection of syngeneic YUMM1.5 tumor cells, and before tumor collection (control, n = 12; mucin, n = 15; inulin, n = 15). (C) Time course of the relative abundance of the six taxa enriched in inulin-treated mice that negatively correlate with YUMM1.5 tumor size (control, n = 12; inulin, n = 15). Time points are before inulin treatment, before tumor injection, and before tumor collection. Data are representative of two independent experiments.

Figure 4.

Figure 4.. Inulin Controls Colon Cancer Growth and Induces Anti-tumor Immunity

(A) Growth of MC-38 mouse colorectal cancer cells (1 × 106) injected subcutaneously into syngeneic C57BL/6 mice that received control diet, 3% mucin in drinking water, or 15% inulin-supplemented chow starting 14 days before tumor inoculation (control, n = 9; mucin, n = 15; inulin, n = 15). (B) MFI of MHC class II and MHC class I in MC-38 tumor-infiltrating DCs (n = 8). (C) Boxplot of the relative abundance of the taxa enriched in inulin-treated mice and positively correlated with tumor size (n = 10). Data are representative of two independent experiments. Graphs show the mean ± SEM. *p < 0.05, **p < 0.005, ***p < 0.001, ****p < 0.0001 by one-way ANOVA with Bonferroni’s correction (B) or by two-way ANOVA with Tukey’s correction (A).

Figure 5.

Figure 5.. Inulin Supplementation Overcomes Melanoma Resistance to MEKi

(A) Growth of NRASQ61K mouse melanoma cells (1 × 106) (control, n = 11; mucin, n = 9; inulin, n = 9; MEKi, n = 10; MEKi+mucin, n = 8; MEKi+inulin, n = 10) that were subcutaneously transplanted in syngeneic C57BL/6 mice that received control diet, 3% mucin in drinking water, or 15% inulin-supplemented chow starting 14 days before tumor inoculation. When tumors reached a volume of 10–20 mm2, mice were administered MEKi (PD325901, 10 mg/kg) once daily by gavage. Tumor volume was assessed every 4 days. (B) Number of tumor-infiltrating effector (CD44hi) CD4+ and CD8+ T cells and total CD45+ cells per tumor weight (in grams) in mice treated as in (A) (MEKi, n = 7; MEKi+mucin, n = 8; MEKi+inulin, n = 8). (C) Number of tumor-infiltrating DCs and DC subsets per tumor weight (in grams) and expression of MHC class I on DCs in mice treated as in (A) (MEKi, n = 8; MEKi+mucin, n = 7; MEKi+inulin, n = 8). (D) Pie chart of taxa enriched in inulin-treated mice and negatively correlated with MaN-RAS tumor size (n = 10). (E) Pie chart of taxa enriched in mucin-treated mice microbiota that negatively correlate with MaN-RAS tumor size (n = 10). (F) Relative abundance of taxa enriched in inulin+MEKi-treated mice and negatively correlated with MaN-RAS tumor size (n = 10). (G) Relative abundance of taxa enriched in mucin+MEKi-treated mice and negatively correlated with MaN-RAS tumor size (n = 10). Data are representative of two independent experiments. Graphs show the mean ± SEM. *p < 0.05, **p < 0.005, ***p < 0.001, ****p < 0.0001 by one-way ANOVA with Tukey’s correction (B and C) or by two-way ANOVA with Bonferroni’s correction (A).

Figure 6.

Figure 6.. A Cladogram Representation of Taxa Enriched in Fecal Microbiota of Mice Administered Mucin or Inulin

Cladogram representation of taxa enriched in fecal microbiota of mice (control, n = 12; mucin, n = 15; inulin, n = 15) administered mucin (red) or inulin (blue). Data are representative of two independent experiments.

Figure 7.

Figure 7.. Combination Therapy with Prebiotics and Anti-PD-1 Modulates Tumor Growth in a Context-Dependent Manner

(A) Growth of YUMM1.5 tumors that were subcutaneously transplanted in syngeneic C57BL/6 mice that were fed control chow or chow supplemented with 15% inulin starting 14 days before tumor inoculation (control, n = 7; inulin, n = 7; PD-1, n = 10; PD-1+inulin, n = 9). Mice were injected with control IgG or anti-PD-1 blocking antibody on days 7, 10, 13, and 16 after tumor inoculation. (B) Growth of YUMM1.5 tumors in C57BL/6 mice that received 0% or 3% mucin in drinking water starting 14 days before tumor inoculation (control, n = 7; mucin, n = 7; PD-1, n = 10; PD-1+mucin, n = 9). Mice were injected with antibodies as described in (A). (C) Growth of SW1 mouse melanoma cells in C3H/ HeOuJ mice that received a control diet, 3% mucin in drinking water, or 15% inulin-supplemented chow starting 14 days before tumor inoculation (control, n = 8; mucin, n = 8; inulin, n = 7; mucin+inulin, n = 9). Data are representative of two independent experiments. Graphs show the mean ± SEM. *p < 0.05, **p < 0.005, ***p < 0.001, ****p < 0.0001 by two-way ANOVA with Bonferroni’s correction.

References

    1. Adolph TE, Tomczak MF, Niederreiter L, Ko HJ, Böck J, Martinez-Naves E, Glickman JN, Tschurtschenthaler M, Hartwig J, Hosomi S, et al. (2013). Paneth cells as a site of origin for intestinal inflammation. Nature 503, 272–276. -PMC -PubMed
    1. Alexander JL, Wilson ID, Teare J, Marchesi JR, Nicholson JK, and Kinross JM (2017). Gut microbiota modulation of chemotherapy efficacy and toxicity. Nat. Rev. Gastroenterol. Hepatol 14, 356–365. -PubMed
    1. Apte RN, Dotan S, Elkabets M, White MR, Reich E, Carmi Y, Song X, Dvozkin T, Krelin Y, and Voronov E (2006). The involvement of IL-1 in tumorigenesis, tumor invasiveness, metastasis and tumor-host interactions. Cancer Metastasis Rev. 25, 387–408. -PubMed
    1. Arthur JC, Perez-Chanona E, Mühlbauer M, Tomkovich S, Uronis JM, Fan TJ, Campbell BJ, Abujamel T, Dogan B, Rogers AB, et al. (2012). Intestinal inflammation targets cancer-inducing activity of the microbiota. Science 338, 120–123. -PMC -PubMed
    1. Bäckhed F, Ley RE, Sonnenburg JL, Peterson DA, and Gordon JI (2005). Host-bacterial mutualism in the human intestine. Science 307, 1915–1920. -PubMed

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