饮食、菌群与抗PD-1反应

Fig. 1 | Dietary composition influences metabolic status and peripheral immunity after 15 weeks. a, Proportion of kilocalories from dietary fat, carbohydrates and protein for the experimental diets. b, Schematic of the 15-week diet model corresponding to c–p. c, Body weight over 15 weeks. Japanese: n = 14 mice; all other diets: n = 15 mice; 2 independent cohorts; mean ± s.e.m. d–f, Correlations between average body weight (n = 5 mice per diet) and metabolic assays at baseline, including percentage of fat mass (d; Japanese: n = 9 mice; all other diets: n = 10 mice), glucose tolerance test (GTT) (e; Low Fat, High Fat and Western: n = 10 mice; all other diets: n = 5 mice) and serum insulin (f; Inulin: n = 6 mice; all other diets: n = 5 mice). AUC, area under the curve. g, Diet metabolic score (mean = 0.36 is indicated). Each data point represents one mouse. h, Correlation matrix of blood immune cells, metabolic parameters and nutritional content at baseline (non-tumour-bearing) and end-point (tumour-bearing). Bubble colour represents Spearman correlation coefficient; bubble size indicates the P value. i–l, Correlation between average metabolic score (n = 5 mice per diet) and baseline blood flow cytometry (n = 5 mice per diet) for CD4+ T cells (i), CD8+ T cells (j), PD-1+ CD8+ T cells (k) and monocytes (l). m, HKP1 tumour kinetics by calliper in the 15-week diet model. Pectin, Inulin, American and Vegan: n = 4 mice; all other diets: n = 5 mice; mean ± s.e.m. n–p, Correlation between average metabolic score (n = 5 mice per diet) and end-point blood flow cytometry (Pectin, American and Vegan: n = 4 mice, all other diets: n = 5 mice) for lymphoid:myeloid ratio (n), neutrophils (o) or CD3+ T cells (p). Correlations were calculated using two-tailed Spearman correlation and are presented as mean ± s.e.m. All immune cells are calculated as percentage of CD45+ cells unless otherwise indicated.

Fig. 2 | Dietary intake dictates gut microbiome composition and anti-PD-1 response. a,b, 16S rRNA gene sequencing showing taxonomic composition of fecal bacteria at phylum (a) and family (b) levels at baseline. Diets are ordered by decreasing metabolic score, from left to right. Bars represent individual mice. c, PCoA (16S rRNA gene sequencing) using Bray–Curtis distance matrix at the ASV level for fecal bacterial DNA at baseline. d, Linear discriminant analysis effect size (LEfSe) analysis for fecal bacteria associated with high or low metabolic score diets (asterisks indicate P < 0.05, false discovery rate (FDR) < 0.1, linear discriminant analysis (LDA) ≥ 2). Bubble size represents normalized relative abundance of a given family (f_) or genus (g_); bubble colour indicates phylum. Diets are ordered by decreasing metabolic score, from left to right. e, ICI trial schematic corresponding to f. f, HKP1 tumour kinetics by calliper in IgG- or anti-PD-1-treated mice. IgG (Psyllium, Ketogenic, Pectin, Low Fat, American and Aspartame) and anti-PD-1 (Mediterranean, Japanese, Western, Inulin, American, Aspartame and High Fat): n = 5 mice; IgG (Mediterranean, Japanese, Vegan, Western, Inulin and High Fat) and anti-PD-1 (Ketogenic, Vegan, Pectin and Low Fat): n = 4 mice; anti-PD-1 (Psyllium): n = 3 mice. Multiple two-tailed Mann–Whitney tests; mean ± s.e.m. g, Anti-PD-1 sensitivity index calculated from end-point tumour volumes of IgG and anti- PD-1-treated groups as shown in f. Data are mean ± s.e.m.

Fig. 3 | Microbial and immune signatures of diet-associated anti-PD-1 efficacy. a, LEfSe analysis for fecal bacteria associated with the top 4 responsive and non-responsive diets (asterisks indicate P < 0.05, FDR < 0.1, LDA ≥ 2). Bubble size indicates normalized relative abundance of a given family (f_) or genus (g_); bubble colour represents phylum. b,c, Bacteroides and Lactobacillus from 16S rRNA gene sequencing on fecal samples before tumour injection (b) and at the end-point (anti-PD-1-treated mice only) (c). Non-responder (NR): n = 27 pre-tumour mice, n = 15 end-point mice; responder (R): n = 29 pre-tumour mice, n = 16 end-point mice. d–h, Flow cytometry analysis of anti-PD-1-treated mice, quantifying intratumoural T cells (d), neutrophils (e), neutrophil:T cell ratio (f) and PD-1+Ki-67− T cells (g), and blood PD-1+Ki-67− T cells (h). Non-responder tumour: n = 15 mice; responder tumour: n = 19 mice; non-responder blood: n = 18 mice; responder blood: n = 20 mice. i, PCoA (16S rRNA gene sequencing) using Bray–Curtis distance matrix at the ASV level for fecal bacterial DNA after 3 and 15 weeks on diet. Homogeneity of group dispersions (PERMDISP) test. j, Body weight increase between 3 and 15 weeks of diet; n = 15 mice per group, 2 independent cohorts. k, Schematic of the 3-week diet model corresponding to l,n,o. l, HKP1 tumour kinetics by calliper comparing 3-week and 15-week models. 3-week, High Fat: n = 4 mice; 3-week, all other groups:, n = 5 mice; 15-week data are from Fig. 1m. m, HKP1 tumour kinetics by calliper in IgG- or anti-PD-1-treated mice in the 3-week diet model. Left, Psyllium diet; right, High Fat diet. n = 5 mice per group. n,o, Intratumoural CD3+ T cells (n) and PD-1+Ki-67−CD8+ T cells (o) in the 3-week model. Non-responders: n = 10 mice; responders: n = 18 mice. Data are mean ± s.e.m. Two-tailed Mann–Whitney test (b,c,e–h,l–n); two-tailed unpaired t-test (d,o).

Fig. 4 | Modulation of the food–gut axis to elicit anti-PD-1 sensitivity. a, Psyllium to High Fat (Psy to HF) diet switch trial schematic corresponding to b,c,g–j. b,c, HKP1 tumour kinetics by calliper (b) and end-point tumour volume (c) following a switch from the Psy to HF diet. n = 10 mice per group, 2 independent cohorts. d, High Fat to Psyllium (HF to Psy) diet switch trial schematic corresponding to e–i,k. e,f, HKP1 tumour kinetics by calliper (e) and end-point tumour volume (f) following a switch from HF to Psy diet. IgG: n = 8 mice; anti-PD-1: n = 10 mice; 2 independent cohorts. g–i, Flow cytometry analysis of intratumoural CD8+ T cells from anti-PD-1-treated mice, quantifying production of IFNγ (g) and TNF (h), and expression of CD44, CD11a, GZMB and PD-1 (i). Psy to HF: n = 4 mice; HF to Psy: n = 5 mice. j,k, Quantitative PCR (qPCR) to detect Lactobacillus spp. and Bacteroides spp. in fecal samples from anti-PD-1-treated mice in the Psy to HF diet switch model (j) or the HF to Psy diet switch model (k). n = 6 mice, each with paired stool samples that include one before and one 48 h after the diet switch. l, Psy donor to HF recipient FMT model schematic corresponding to m–o. Abx, antibiotic treatment. m,n, HKP1 tumour kinetics by calliper (m) and end-point tumour volume (n) in the Psy donor to HF recipient FMT model. IgG: n = 10 mice; anti-PD-1: n = 13 mice; 2 independent cohorts. o, PCoA (16S rRNA gene sequencing) using Bray–Curtis distance matrix at the ASV level for fecal bacterial DNA from the FMT trial. permutational multivariate ANOVA (PERMANOVA) test, n = 5 mice per group. Tx, treatment. Data are mean ± s.e.m. Two-tailed Mann–Whitney test (b,c,e–i,m,n), two-tailed Wilcoxon test (j,k).

Fig. 5 | L. johnsonii underlies High Fat diet-associated ICI efficacy. a, PCoA (shotgun metagenomics) of Bray–Curtis dissimilarities in fecal microbiome after 3 weeks of obesogenic responder or non-responder diets. Shading shows 95% confidence intervals; vectors represent top 15 species by SIMPER; arrow length indicates magnitude of contribution and direction represents correlation with ordination axes. PERMANOVA test, n = 3 mice per group. b, LEfSe analysis for fecal bacteria species associated with obesogenic responder and non-responder diets (n = 3 mice per diet; top 15 taxa with P < 0.05, FDR < 0.1, LDA ≥ 2). Bubble size represents normalized relative abundance and bubble colour indicates family. Amer, American; Keto, Ketogenic; Med, Mediterranean. c, L. johnsonii abundance from metagenomic data in a,b. Ob_NR, obesogenic non-responder diet; Ob_R, obesogenic responder diet. d, Schematic for monocolonization (germ-free (GF)) and single-strain supplementation (specific pathogen-free (SPF)) models corresponding to e–h. e,f, HKP1 tumour kinetics by calliper (e) and end-point tumour volume (f) in the monocolonization model. L. johnsonii (Lj): n = 10 mice per group; PBS: n = 9 mice per group; 2 independent cohorts. g,h, HKP1 tumour kinetics by calliper (g) and end-point tumour volume (h) in the single-strain supplementation model. n = 5 mice per group. i, Schematic for human-to-mouse FMT from an ICI-refractory donor, corresponding to j–l. j,k, HKP1 tumour kinetics by calliper (j) and end-point tumour volume (k) in FMT recipient mice. High Fat diet plus IgG: n = 4 mice; all other groups: n = 5 mice. l, qPCR for L. johnsonii in faeces from FMT recipients after anti-PD-1 treatment. Psyllium: n = 5 mice; High Fat: n = 4 mice; one stool per mouse. m, Pathway enrichment (serum metabolomics) from obesogenic responder versus non-responder models. Bubble size represents pathway effect; bubble colour indicates significance. n, Normalized concentration (log2(ratio)) of tryptophan-derived (top) and tyrosine-derived (bottom) metabolites in obesogenic responders versus non-responders. HP, hydroxyphenyl. o–q, Normalized concentration (log2(ratio)) of serum DAT in anti-PD-1-treated mice from the monocolonization model (o; n = 5 mice per group), serum DAT in mice on obesogenic responder versus non-responder diets (p; n = 10 mice per group, including 5 mice per diet) and DAT in bacterial supernatants (q; n = 3 independent cultures). r, Schematic for generating diet-conditioned intratumoural T cells, corresponding to s. s, Flow cytometry for IFNγ and TNF from intratumoural CD8+ T cells after DAT stimulation, normalized to vehicle. n = 4 wells per group, with T cells from 5 mice per group. t, DAT supplementation trial schematic, corresponding to u,v. u,v, HKP1 tumour kinetics by calliper (u) and end-point tumour volume (v) in the DAT supplementation trial. n = 5 mice per group. w, Differentially abundant plasma metabolites from ICI responder versus non-responder patients with NSCLC. Fold change threshold = 1.2; P value threshold < 0.05. DHICA, 5,6-dihydroxyindole-2-carboxylic acid. x,y, Normalized concentration (log2(ratio)) of indolelactic acid (x) and 3-hydroxy-3-(3-hydroxyphenyl) propionic acid-O-sulfate (3-HPP sulfate) (y) in plasma of ICI responders (n = 32 patients) and non-responders (n = 21 patients). Data are mean ± s.e.m.; two-tailed unpaired t-test (c,l,s,x), two-tailed Mann–Whitney test (e,g,j,n,p,u,y), one-way ANOVA Kruskal–Wallis test (f,h), one-way ANOVA with Tukey’s multiple comparisons test (k,o,v) and one-way ANOVA with Dunnett’s multiple comparisons test (q). NS, not significant.