Introduction: Recurrent urinary tract infections have been linked to increased risk of bladder cancer, suggesting a potential role of the urinary microbiome in bladder cancer pathogenesis.Objective: Compare the urinary microbiomes in mice with and without bladder. Methods:Longitudinal study of mice exposed to a dilute bladder-specific carcinogen (0.05% nbutyl-n-(4-hydroxybutyl) nitrosamine, BBN mice, n=10), and control mice (n=10). Urine was sampled monthly from individual mice for 4 months. Microbial DNA was extracted from the urine, and the V4 region of the 16S rRNA gene sequenced. Animals were sacrificed and their bladders harvested for histopathology. Bladder sections were graded by a blinded pathologist. The composition and diversity of the urinary microbiome were compared between the BBN and control mice. Metabolic pathway analysis was completed using PICRUST.Results: Bladder histology in the BBN group showed normal tissue with inflammation (BBNnormal, n=5), precancerous pathologies, (BBN-precancerous, n=3), and invasive cancer (BBNcancer, n=2). Alpha diversity did not differ between the mice exposed to BBN and the control mice at any timepoint. There were no differences in the urinary microbiomes between the BBN and control mice at baseline. At month 4, mice exposed to BBN had higher proportion of both Gardnerella and Bifidobacterium compared to control mice. There were no differences in proportions of specific bacteria between either the BBN-precancer or BBN-cancer and controls at month 4. However, the BBN-normal mice had higher proportions of Gardnerella, Haemophilus, Bifidobacterium, and Ureaplasma Actinobaculum, and lower proportions of Actinomyces, compared to control mice at month 4. Functional pathway analysis demonstrated increases in genes related to purine metabolism, phosphotransferase systems, peptidases, protein folding, and bacterial toxins in the BBN-mice compared to control mice at month 4.Conclusion: Mice exposed to 4 months of BBN, a bladder-specific carcinogen, have distinct urine microbial profiles compared to control mice.

Herpes simplex virus type 1 (HSV-1) mutants lacking the γ134.5 neurovirulence loci are promising agents for treating malignant glioma. Arming oncolytic HSV-1 (oHSV) to express immunostimulatory genes may potentiate therapeutic efficacy. We have previously demonstrated improved pre-clinical efficacy, biodistribution, and safety of M002, a γ134.5-deleted HSV-1 engineered to express murine IL-12. Herein, we describe the safety and biodistribution of M032, a γ134.5-deleted HSV-1 virus that expresses human IL-12 after intracerebral administration to non-human primates (NHPs), Aotus nancymae. Cohorts were administered vehicle, 106, or 108 pfu of M032 on Day 1 and subjected to detailed clinical observations performed serially over a 92-day trial. Animals were sacrificed on Days 3, 31, and 91 for detailed histopathologic assessments of all organs and to isolate and quantify virus in all organs. With the possible exception of one animal euthanized on Day 16, neither adverse clinical signs nor sex- or dose-related differences were attributed to M032. Elevated white blood cell and neutrophil counts were observed in virus-injected groups on Day 3, but no other significant changes were noted in clinical chemistry or coagulation parameters. Minimal to mild inflammation and fibrosis detected, primarily in meningeal tissues, in M032-injected animals on Days 3 and 31had mostly resolved by Day 91. The highest viral DNA levels were detected at the injection site and motor cortex on Day 3 but decreased in CNS tissues over time. These data demonstrate the requisite safety of intracerebral M032 administration for consideration as a therapeutic for treating malignant brain tumors.