Mesenchymal stem cell homing to advanced and metastatic prostate cancer

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Prostate cancer (PCa) is the most common cancer affecting men worldwide. Current treatment strategies to combat advanced and metastatic disease are ineffective and this has created a need to explore novel therapies, such as cell and gene therapies. A promising strategy involves capitalising on the innate ability of bone marrow-derived mesenchymal stem cells (BMSCs) to home to sites of cancer and release a genetic payload. BMSCs have the added benefit of being immune evasive, which is a major problem for other cell and gene therapy protocols. BMSCs used in this study had been previously stably nucleofected to express a cell tracking reporter gene firefly luciferase (fl) and the yeast fusion suicide gene cytosine deaminase and uracil phosphoribosyltransferase (BMSC-Fcy:Fur). RM1 murine PCa cells were gene modified with the cell tracking reporter gene renilla luciferase (rl). An immune intact B6 albino mouse model was developed to investigate BMSC-Fcy:Fur homing to RM1 lung pseudometastases, and therapeutic effect. Using bioluminescence imaging (BLI) it was discovered that BMSC-Fcy:Fur showed greater persistence in the lungs of mice with RM1 tumours at 3 hours post-injection compared to their cancer free counterparts, however the BMSCs did not persist for longer than 24 hours in the lungs likely because of their advanced passage. By delivering prodrug when BMSC-Fcy:Fur were largely present in the lungs, a significant decrease lung RM1 colonies and a 25% improvement in survival was achieved. Importantly, BMSC-Fcy:Fur treatment was associated with no adverse events and did not promote PCa growth, confirming their safety for allogenic use in the treatment of PCa. Previous findings have led to investigations into why BMSCs are attracted to PCa and how to best capitalise BMSC-PCa tropism for the development of novel therapies for metastatic PCa. It was anticipated that a greater understanding of the molecular events governing BMSC tropism for cancer may permit improved therapeutic BMSC targeting to metastatic PCa. Towards this aim, I isolated and characterised a subpopulation of BMSCs from B6 albino mouse BM. Cells were sorted from early passage following dual colour staining with antibodies against a stem cell antigen-1 (Sca-1) and a haematopoietic marker (CD45). Specifically, when compared to unsorted BM cells (UBMCs), the subpopulation showed: typical BMSC phenotype: Sca-1⁺, CD44⁺, CD90⁺, CD106⁺, CD31⁻, CD34⁻ and CD45; enhanced adipogenic ability; similar osteogenic ability; and similar colony formation ability. Subsequently, these BMSCs were used to understand the signals mediating migration towards PCa. BMSC migration to RM1 derived conditioned medium (CM) was assessed using Transwell migration assays. These assays revealed that migration was due to soluble chemokines. Cytokine arrays and genome wide microarrays identified candidate chemokine receptors CCR2, CXCR2, CCR1, and CCR5 as potential mediators of BMSC migration towards RM1 CM. Inhibition of individual chemokine receptors led to a reduction in migration of BMSCs to RM1 CM, however not to level observed with pertussis toxin alone suggesting multiple or other chemoattractant receptors (chemokine receptors or growth factor receptors) may be involved in migration. Importantly, it was found that overexpression of CCR2 and CXCR2 on the surface of BMSC-Fcy:Fur significantly improved their migration to RM1 PCa CM. In this thesis, I have demonstrated that systemic delivery of allogenic BMSCs followed by GDPET activation at a time when BMSCs are most localised to the tumour, is a safe and efficacious method to combat late stage PCa. Moreover, I showed for the first time that BMSCs can be enriched from B6 albino mice using Sca-1 and CD45 cell sorting. Lastly, I showed that BMSCs migration is facilitated by various chemokine receptors and overexpression of CCR2 and CXCR2 promotes migration to RM1 CM. These findings provide new insights into the signals mediating migration to PCa and may be exploited in future to improve on migration of BMSCs to PCa.
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