The impact of ketogenic diet on the gut microbiome and tumor growth in an in vivo epithelial ovarian cancer model (105)

Abstract

<b>Objectives:</b> The ketogenic diet (KD) is hypothesized to protect against cancer progression due to metabolic reprogramming. Sustained dietary modifications alter the composition of the gut microbiome, which may influence tumor growth. Here, we present the impact of KD on the gut microbiome in relation to epithelial ovarian cancer (EOC) progression. <b>Methods:</b> C57Bl/6J ID8 (luciferase-tagged) tumor-bearing mice were fed a KD (10% protein, 0% carbohydrates, 90% fat), high fat diet (HFD) (10% protein, 15% carbohydrates, 75% fat), and control diet (CD) (10% protein, 77% carbohydrates, and 13% fat) (<i>n</i>=10 mice per group). Blood ketone and glucose levels were serially measured. EOC tumor growth was monitored with <i>in Vivo</i> Imaging System (IVIS) via intraperitoneal D-luciferin. Total tumor burden was measured in total flux (photons/second). Stool was collected at baseline, midpoint (four weeks after tumor cell injection), and endpoint (tumor mass of >150 mm³, ascites development, or weight loss >20%); 16S rDNA was isolated, and sequence reads were assigned to genus-level amplicon sequence variants (ASV) in DADA2 as well as analyzed for alpha and beta microbial diversity using Phyloseq. Statistical tests included analysis of variance (ANOVA), permutational multivariate analysis (PERMANOVA), linear regression, and Wilcoxon test. A p-value of <0.05 was considered statistically significant. <b>Results:</b> KD-fed mice exhibited an increase in ketone blood levels (2.22 mmol/L) compared to both HFD- (0.7 mmol/L, p<0.0001) and CD-fed mice (0.61 mmol/L, p<0.0001) at the endpoint. There was no significant difference in overall glucose levels or body mass composition between the groups. By day 42, there was a 9.1-fold increase in tumor growth from baseline in the KD-fed group compared to HFD-and CD-fed groups (9.1 vs 2.0 vs 3.1-fold, respectively, p<0.001 and p<0.001). By study endpoint, KD-fed mice exhibited a statistically significant increase in gut microbial alpha diversity in comparison to HFD and CD (p<0.01 at the midpoint and p<0.05 at the endpoint). Brays-Curtis dissimilarity based on microbial beta diversity demonstrated that the relative abundance of gut microbial species between the three groups was similar at baseline. However, KD-fed mice had significantly altered gut microbial composition than HFD-and CD-fed mice at both midpoint and endpoint (p = 0.001); 16S data analysis demonstrated a relative decrease in <i>Lactobacillus</i> and <i>Coriobacteriaceae</i> species and a relative increase in <i>Romboutsia</i> and <i>Akkermansia</i> species in the KD-fed mice. <b>Conclusions:</b> In our EOC murine model, KD was associated with increased EOC growth and distinct gut microbial alterations in comparison to HFD and CD. KD-fed mice exhibited increased gut microbial diversity over time, a relative depletion of bacterial species <i>Lactobacillus</i> and <i>Coriobacteriaceae</i>, and a relative enrichment of bacterial species <i>Romboutsia</i> and <i>Akkermansia</i>. Further studies are needed to understand the mechanisms underlying the relationship between KD, the gut microbiome, and EOC progression.Fig. 1