Tumor necrosis factor alpha enhanced cryosurgery: in vitro and In vivo mechanisms.

University of Minnesota Ph.D. dissertation. July 2010. Major: Biomedical Engineering. Advisor: John C. Bischof. 1 computer file (PDF); xi, 152 pages, appendices I-II. Ill. (some col.)Cryosurgery has shown potential as a minimally invasive technology for tumor treatment. However, the partial destruction of tissue at the iceball edge not only potentiates the later recurrence of the tumor, but also limits the ability of imaging techniques to predict outcomes. A variety of adjuvants have been investigated to improve the efficacy of cryosurgery and the correspondence between the imaged iceball and the destroyed tissue. The most effective of these adjuvants is tumor necrosis factor alpha (TNF-α), however, the precise role of TNF-α on cryosurgery enhancement is not well understood. Prostate cancer was selected as the tumor model to investigate the mechanisms of the combinatorial treatment both in vitro and in vivo. In the in vitro system, increased cryosensitivity in human microvascular endothelial cells (MVEC) was noted compared to human prostate cancer cells (LNCaP Pro 5) with or without TNF-α pre-treatment. This suggests that injury to the endothelium may help govern the extent and enhancement of cryoinjury in vivo. In the in vivo model system, prostate tumor (LNCaP Pro 5) was grown in a dorsal skin fold chamber implanted in a male nude mouse. Four hours pre-treatment of TNF-α was observed to induce vascular pre-conditioning, including inflammation (NF-κB) and apoptotic (caspase) pathways upregulation. This acted to convert tumor vascular endothelium from a non-thrombotic non-adhesive barrier to a pro-adhesive surface that encouraged inflammatory cell infiltrate from the blood. After TNF-α pre-conditioning a large increase in host inflammatory infiltrate within the cryolesion was observed over cryosurgery alone. Apoptosis, subsequent to inflammatory infiltrate, was also enhanced by TNF-α through leukocyte-endothelium interaction at the periphery of the cryolesion at day 1. This contributed to vascular injury and microvascular shutdown (perfusion defect) from day 1 to 7. However, caspase and NF-κB inhibition studies show that the inflammation (NF-κB) instead of apoptotic (caspase) pathway played a dominant role in TNF-α cryosurgical enhancement in vivo. Finally, this combinatorial approach was tested in ELT-3 uterine leiomyoma hindlimb tumor system allowing timing, delivery, sex and tumor type to vary. This work shows the potential of this combinatorial approach for general tumor treatment with cryosurgery