Abstract LB150: Metagenomics and metabolomics identify differential biomarkers in MASLD and hepatocellular carcinoma in South Texas Hispanics

Background: The Rio Grande Valley (RGV) is a major hotspot for hepatocellular carcinoma (HCC) and liver diseases. HCC often develops from metabolic dysfunction-associated steatotic liver disease (MASLD), progressing through stages like simple steatosis, MASH, and cirrhosis. However, the mechanisms driving progression remain poorly understood. In this study, we examined liver microbiome signatures to identify those that differentiate MASLD patients at risk for HCC. We also explored microbiome-metabolome associations, uncovering microbe-metabolite links in RGV patients. These findings demonstrate microbiome's role in HCC progression, potentially guiding future diagnostic and therapeutic strategies. Methods: FFPE tissue blocks from MASLD, MASH, HCC, and cirrhosis samples were selected. DNA extraction was carried out using the QIAamp DNA FFPE Advanced Kit with protocol modifications, and quality was assessed with NanoDrop, Qubit, Bioanalyzer, and TapeStation. 16S rRNA metagenomics was performed on Illumina MiSeq, and raw sequences were processed using Trim Galore to ensure high-quality reads. Taxonomic classification was conducted with Kraken 2, comparing against the United Human Gastrointestinal Genome (UHGG) v2.0.2 reference database, retaining species-level classified reads for further analysis. Microbial community diversity and comparative analyses were carried out using R packages in Microbiome Analyst, employing Welch t-tests, Bray-Curtis ordination, and LEfSe to identify differential abundance. Additionally, metabolomics was performed using Bruker and Thermo instruments to explore the correlation between microbial populations and metabolites. Results: Proteobacteria was most abundant in the liver of MASLD patients, followed by Firmicutes, Actinobacteria, Bacteroidetes, and Fusobacteria, which decreased in MASH, Cirrhosis, and HCC. Elevated Beijerinckiaceae, Moraxellaceae, Streptococcaceae, and Enterobacteriaceae were found in MASLD compared to the other stages. Alpha and beta diversity analyses revealed significant microbial differences, particularly between MASLD and HCC (p = 0.001). MASH showed increased Ruminococcus, Enterobacter, and Enterococcus, while Cirrhosis had more Collinsella, Veillonella, and Prevotella. Linear discriminant analysis identified key taxa, including Ruminococcus, Enterobacter, and Staphylococcus in MASH, and Bifidobacterium_thermophilum, Lacticaseibacillus_rhamnosus, and Lactobacillus spp. in MASLD. Metabolomics revealed altered metabolic pathways linked to these microbial shifts. Combined microbiome and metabolomics analyses showed correlations with metabolic activity, and random forest models validated these taxa as potential biomarkers, with Spearman correlation confirming the link between microbiome diversity and liver disease progression. Conclusion: This study is crucial for identifying the etiology of HCC and developing microbial species-based diagnostic and prognostic biomarkers. It is particularly relevant for the Hispanic population in the RGV, addressing a significant regional health disparity. Citation Format: Shweta Singh, Sierra Vidaurri, Anupam Dhasmana, Swati Dhasmana, Bablu Kumar, Jacob Galan, Natasha S. Garcia-Rodriguez, Murali Mohan Yallapu, Subhash Chauhan, Sheema Khan. Metagenomics and metabolomics identify differential biomarkers in MASLD and hepatocellular carcinoma in South Texas Hispanics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited s); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2): nr LB150.