The bacterial neurometabolic signature of the gut microbiota of young children with autism spectrum disorders.

Introduction. The human gut microbiota is currently seen as an important factor that can promote autism spectrum disorder (ASD) development in children.Aim. This study aimed to detect differences in the taxonomic composition and content of bacterial genes encoding key enzymes involved in the metabolism of neuroactive biomarker compounds in the metagenomes of gut microbiota of children with ASD and neurotypical children.Methodology. A whole metagenome sequencing approach was used to obtain metagenomic data on faecal specimens of 36 children with ASD and 21 healthy neurotypical children of 3-5 years old. Taxonomic analysis was conducted using MetaPhlAn2. The developed bioinformatics algorithm and created catalogue of the orthologues were applied to identify bacterial genes of neuroactive compounds in the metagenomes. For the identification of metagenomic signatures of children with ASD, Wilcoxon's test and adjustment for multiple comparisons were used.Results. Statistically significant differences with decreases in average abundance in the microbiota of ASD children were found for the genera Barnesiella and Parabacteroides and species Alistipes putredinis, B. caccae, Bacteroides intestinihominis, Eubacterium rectale, Parabacteroides distasonis and Ruminococcus lactaris. Average relative abundances of the detected genes and neurometabolic signature approach did not reveal many significant differences in the metagenomes of the groups that were compared. We noted decreases in the abundance of genes linked to production of GABA, melatonine and butyric acid in the ASD metagenomes.Conclusion. For the first time, the neurometabolic signature of the gut microbiota of young children with ASD is presented. The data can help to provide a comparative assessment of the transcriptional and metabolomic activity of the identified genes.

Targeting TLR-4 with a novel pharmaceutical grade plant derived agonist, Immunomax®, as a therapeutic strategy for metastatic breast cancer.

BACKGROUND: Previously we demonstrated that the resection of primary 4T1 tumors only slightly prolongs mouse survival, but importantly, creates a "window of opportunity" with attenuated suppressor cell and increased activated T cell populations. This suggests that additional activation of the immune system by immunostimulatory agents during this period may enhance anti-tumor immunity and potentially eradicate micro-metastatic disease in this stringent model. We hypothesized that the immunostimulator Immunomax®, which is comprised of a plant-derived polysaccharide, is non-toxic in humans and stimulates immune defense during the infectious diseases treatment, may have also anti-tumor activity and be beneficial in the adjuvant setting when endogenous anti-tumor responses are present and during the "window of opportunity" in post-resection metastatic breast cancer model. Here we provide the initial report that Immunomax® demonstrates the capacity to eliminate micro-metastatic disease in the post-resection, 4T1 mouse model of breast cancer. METHODS: The efficacy of Immunomax® was evaluated by analyzing survival rate and the number of spontaneous clonogenic tumor cells in the lung homogenates of mice. The frequencies of activated NK, CD4(+) and CD8(+) cells as well as myeloid-derived suppressor cells and Treg cells were evaluated using flow cytometry. Highly purified mouse and human dendritic and NK cells were sorted and the effect of Immunomax® on activation status of these cells was assessed by flow cytometry. The property of Immunomax® as TLR-4 agonist was determined by NF-κB/SEAP reporter gene assay, WB, RT-PCR. RESULTS: Immunomax® injections significantly prolonged overall survival and cured 31% of mice. This immunostimulator activates DCs via the TLR-4, which in turn stimulates tumoricidal NK cells and in vitro, completely inhibits growth of 4T1 cells. Incubation of PBMC from healthy donors with Immunomax® activates NK cells via activation of plasmacytoid DC leading significantly higher efficacy in killing of human NK-target cells K562 compared with non-treated cells. CONCLUSION: This is the first demonstration that Immunomax® is a TLR-4 agonist and the first report of a documented role for this pharmaceutical grade immunostimulator in augmenting anti-tumor activity, suggesting that incorporation of Immunomax® into developing breast cancer therapeutic strategies may be beneficial and with less potential toxicity than checkpoint inhibitors.