ANTIBODY-MEDIATED P53 PROTEIN THERAPY PREVENTS LIVER METASTASIS IN VIVO.

Abstract

Purpose To test the clinical efficacy of antibody-mediated protein transduction of p53 into cancer cells in vivo in a mouse model of colon cancer metastasis to liver. Methods An Fv fragment of mAb 3E10 was previously shown to penetrate into cancer cells and localize in the nucleus. A His6-tagged Fv-p53 fusion protein was produced in Pichia pastoris X-33 yeast cells transfected with pPICZA-Fv-p53. Fv-p53 was isolated on NiNTA-agarose and assayed for killing CT26.CL25 colon cancer cells in vitro and CT26.CL25 cells injected into the portal vein of syngeneic Balb/c mice. Results Experiments in vitro demonstrated killing of CT26.CL25 cells by Fv-p53 but not Fv or p53 alone, and immunohistochemical staining confirmed that Fv was required for transport of p53 into cells. Prevention of liver metastasis in vivo was tested by portal vein injections of 100 nM Fv-p53 10 minutes and 1 week after injection of cancer cells. Mice were sacrificed 2 weeks after the second injection of Fv-p53 and assigned metastasis scores based on the following criteria: 0 - no liver metastasis; 1 - minimal infiltration (less than 1 cm2 area of metastasis); 2 - mild to moderate infiltration (1-2 cm2 area of metastasis); 3 - severe infiltration (greater than 2 cm2 area of metastasis but the liver was not entirely infiltrated); 4 - complete infiltration. As shown in Table 1, Fv-p53 prevented liver metastasis. Conclusion Fv-p53 treatment had a profound impact on liver metastasis. These results demonstrate the clinical potential of an antibody Fv fragment as a protein transduction domain for intracellular/intranuclear delivery of p53 for prevention of cancer metastasis in vivo.