ObjectivesSeveral reports suggesting that the intestinal microbiome plays a key role in the development of inflammatory bowel disease (IBD) or colorectal cancer (CRC), but the changes of intestinal bacteria in healthy people, patients with IBD and CRC are not fully explained. The study aimed to investigate changes of intestinal bacteria in healthy subjects, patients with IBD, and patients with CRC.MaterialsWe collected data from the European Nucleotide Archive on healthy people and patients with colorectal cancer with the study accession number PRJEB6070, PRJEB7774, PRJEB27928, PRJEB12449, and PRJEB10878, collected IBD patient data from the Integrated Human Microbiome Project from the Human Microbiome Project Data Portal. We performed metagenome-wide association studies on the fecal samples from 290 healthy subjects, 512 IBD patients, and 285 CRC patients. We used the metagenomics dataset to study bacterial community structure, relative abundance, functional prediction, differentially abundant bacteria, and co-occurrence networks.ResultsThe bacterial community structure in both IBD and CRC was significantly different from healthy subjects. Our results showed that IBD patients had low intestinal bacterial diversity and CRC patients had high intestinal bacterial diversity compared to healthy subjects. At the phylum level, the relative abundance of Firmicutes in IBD decreased significantly, while the relative abundance of Bacteroidetes increased significantly. At the genus level, the relative abundance of Bacteroides in IBD was higher than in healthy people and CRC. Compared with healthy people and CRC, the main difference of intestinal bacteria in IBD patients was Bacteroidetes, and compared with healthy people and IBD, the main difference of intestinal bacteria in CRC patients was in Fusobacteria, Verrucomicrobia, and Proteobacteria. The main differences in the functional composition of intestinal bacteria in healthy people, IBD and CRC patients were L-homoserine and L-methionine biosynthesis, 5-aminoimidazole ribonucleotide biosynthesis II, L-methionine biosynthesis I, and superpathway of L-lysine, L-threonine, and L-methionine biosynthesis I. The results of stratified showed that the abundance of Firmicutes, Bacteroidetes, and Actinobacteria involved in metabolic pathways has significantly changed. Besides, the association network of intestinal bacteria in healthy people, IBD, and CRC patients has also changed.ConclusionsIn conclusion, compared with healthy people, the taxonomic and functional composition of intestinal bacteria in IBD and CRC patients was significantly changed.

As immune checkpoint inhibitors (ICIs) continue to advance, more evidence has emerged that anti-PD-1/PD-L1 immunotherapy is an effective treatment against cancers. Known as the programmed death ligand-1 (PD-L1), this co-inhibitory ligand contributes to T cell exhaustion by interacting with programmed death-1 (PD-1) receptor. However, cancer-intrinsic signaling pathways of the PD-L1 molecule are not well elucidated. Therefore, the present study aimed to evaluate the regulatory network of PD-L1 and lay the basis of successful use of anti-PD-L1 immunotherapy in acute myeloid leukemia (AML). Data for AML patients were extracted from TCGA and GTEx databases. The downstream signaling pathways of PD-L1 were identified via Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. The key PD-L1 related genes were selected by weighted gene co-expression network analysis (WGCNA), MCC algorithm and Molecular Complex Detection (MCODE). The CCK-8 assay was used to assess cell proliferation. Flow cytometry was used to determine cell apoptosis and cell cycle. Western blotting was used to identify the expression of the PI3K-AKT signaling pathway. PD-L1 was shown to be elevated in AML patients when compared with the control group, and high PD-L1 expression was associated with poor overall survival rate. The ECM-receptor interaction, as well as the PI3K-AKT signaling pathway, were important PD-L1 downstream pathways. All three analyses found eight genes (ITGA2B, ITGB3, COL6A5, COL6A6, PF4, NMU, AGTR1, F2RL3) to be significantly associated with PD-L1. Knockdown of PD-L1 inhibited AML cell proliferation, induced cell apoptosis and G2/M cell cycle arrest. Importantly, PD-L1 knockdown reduced the expression of PI3K and p-AKT, but PD-L1 overexpression increased their expression. The current study elucidates the main regulatory network and downstream targets of PD-L1 in AML, assisting in the understanding of the underlying mechanism of anti-PD-1/PD-L1 immunotherapy and paving the way for clinical application of ICIs in AML.

The tumor microenvironment is complicated and continuously evolving. This study was devoted to the identification of potential prognostic biomarkers based on the tumor microenvironment associated with immunotherapy for melanoma. This study integrates a couple of melanoma single cell and transcriptome sequencing datasets and performs a series of silico analyses as nicely as validation of molecular biology techniques. A core set of immune escape related genes was identified through Lawson et al. and the ImmPort portal. The differential proteins were identified through the cBioPortal database. Regression analysis was used to profile independent prognostic factors. Correlation with the level of immune cell infiltration was evaluated by multiple algorithms. The capacity of LCK to predict response was assessed in two independent immunotherapy cohorts. High LCK expression is associated with better prognosis, high levels of TILs and better clinical staging. Pathway analysis showed that high expression of LCK was significantly associated with activation of multiple tumor pathways as well as immune-related pathways. LCK expression tends to be higher in immunotherapy-responsive patients and those with lower IC50s treated with chemotherapeutic agents. RT-qPCR detected that LCK expression was significantly upregulated in melanoma cell lines. Single-cell transcriptome analysis showed that LCK was specifically highly expressed on T cells. CellChat analysis confirmed that LCK in C2 subpopulations and T cell subpopulations exerted immune promotion between cells by binding to CD8 receptors. In conclusion, LCK is a reliable biomarker for melanoma and will contribute to its immunotherapy.