Abstract 239: Establishing a murine intratumoral bacterial model
Authors: Hannon Ryan A. , Xu Dongqing , Wan Fengyi 
Published: March 2024

Abstract
Background: Long thought to be sterile spaces, current evidence now suggests that host microbiota can be found within cancerous tissue. Emerging data has illuminated that microbes are integral components of tumor tissue itself, which may correlate with tumor progression, treatment responsiveness, and disease prognosis. A recent Cell article from Fu et al. demonstrated that tumor-resident intracellular microbiota can promote metastatic colonization in breast cancer. Others within this research landscape see therapeutic opportunity - harnessing the tropism of these microbes to deliver TME modifying payloads directly to cancerous lesions.
Purpose: Before studying the mechanism underlying their tropism to solid tumors, it is necessary to first establish a murine model that would allow for an oncotropic bacteria to be detected and quantified within tissues of interest. My research entailed the design and execution of pilot experiments developing xenograft melanoma and genetic colorectal models for future research projects.
Methods: In all mouse studies, an E. coli Nissle 1917 strain called “NisLux” was used, who’s integration with a luxCDABE cassette allowed it to be selectively identified via luminescence using traditional exposure photography or endpoint luminescence detection using a 96-well plate reader. A xenograft melanoma model was developed using C57BL/6J mice and a B16-F10 murine melanoma line. A genetics model was developed using C57BL/6J-Apcmin mice which spontaneously develop colorectal adenomas as they age. Tumors, spleens, and healthy tissue were harvested and processed following tail vein injections of either NisLux or PBS solution. Harvested tissue was normalized by weight and concentration of resident NisLux was calculated from serially diluted samples plated on LB agar.
Results: In both the xenograft and genetics murine models, NisLux exhibited tropism to tumor sites, enriching at a concentration above that of healthy background tissue. NisLux dosing correlated with splenomegaly without changes to the mass of the primary lesion compared to control.
Conclusions: NisLux exhibits tropism to tumor sites in our xenograft and genetics murine models. Both models should undergo further procedural and control optimization. Two possible modifications to future melanoma xenograft experiments include the collection of non-cancerous skin negative control samples and a further delayed takedown day for tissue processing. In future C57BL/6J-Apcmin mice experiments, healthy colon sections from NisLux dosed C57BL/6J mice should be used as negative controls. Overall, the results from both models were promising and encourage future exploration of the mechanism underlying NisLux’s tropism to solid tumors. Subsequent experiments could involve comparing the tropism of engineered KO strains to WT NisLux or performing comparative sequencing of isolated tumor resident NisLux to the initial dosing population.
Citation Format: Ryan A. Hannon, Dongqing Xu, Fengyi Wan. Establishing a murine intratumoral bacterial model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 239.