Tumors require ongoing angiogenesis to support their growth. Inhibition of angiogenesis by production of angiostatic factors should be a viable approach for cancer gene therapy. Endostatin, a potent angiostatic factor, was expressed in mouse muscle and secreted into the bloodstream for up to 2 weeks after a single intramuscular administration of the endostatin gene. The biological activity of the expressed endostatin was demonstrated by its ability to inhibit systemic angiogenesis. Moreover, the sustained production of endostatin by intramuscular gene therapy inhibited both the growth of primary tumors and the development of metastatic lesions. These results demonstrate the potential utility of intramuscular delivery of an antiangiogenic gene for treatment of disseminated cancers.

No abstract

We have previously shown that talactoferrin-alfa (TLF), a recombinant human lactoferrin, is an immunomodulatory protein that is active against implanted tumors, both as a single agent and in combination with chemotherapy. In this study, we show that talactoferrin is active against autochthonous tumors in a transgenic mouse line, which is more analogous to human cancers, and identify key mechanistic steps involved in the anticancer activity of oral TLF. BALB/c mice transgenic for the rat neu (ErbB2) oncogene (BALBneuT) treated with oral TLF showed a significant delay in carcinogenesis, with 60% tumor protection relative to vehicletreated mice at week 21. Oral TLF also showed tumor growth inhibition in wild-type BALB/c mice implanted with neu + mammary adenocarcinoma, with one third displaying a longlasting or complete response. Oral TLF induces an increase in intestinal mucosal IFN-; production and an increase in Peyer's patch cellularity, including expansion of CD8 + T lymphocytes and NKT cells, and the enhancement of CD8 + Tcell cytotoxicity. In IFN-; knockout mice, there is an absence of the TLF-induced Peyer's patch cellularity, no expansion of CD8 + T lymphocytes and NKT cells, and loss of TLF anticancer activity. TLF antitumor activity is also lost in mice depleted of CD8 + T cells and in CD1 knockout mice, which lack NKT activity. Thus, the inhibition of distant tumors by oral TLF seems to be mediated by an IFN-;-dependent enhancement of CD8 + T-and NKT cell activity initiated within the intestinal mucosa. [Cancer Res 2007;67(13):6425-32]

Administration of plasmid/lipid complexes to the lung airways for the treatment of metastatic pulmonary diseases represents a new strategy of gene therapy. In this study we present evidence that intratracheal administration of a plasmid encoding murine IL-12 complexed with N-[1-(2,3-dioleyloxy)propyl)-N,N,N-trimethylammonium chloride:cholesterol inhibits the growth of lung metastases, using a renal cell carcinoma model. Instillation of pIL-12/lipid complexes resulted in expression of biologically active IL-12 (170-240 pg/ml) and IFN-gamma (100-190 pg/ml) in the bronchoalveolar lavage fluid. A significantly reduced number of lung metastases (26+/-24) was observed in mice instilled with IL-12/lipid complexes 24 hr after tumor challenge, whereas more than 250 metastatic foci were counted in lungs of untreated mice. Moreover, IL-12/lipid inhibited the growth of 3-day-old established metastases when compared with empty plasmid/lipid or IL-12 plasmid in saline. Mice receiving IL-12 gene therapy survived significantly longer (median survival of 43 days) than untreated mice (median survival of 31 days) or mice treated with control plasmid/lipid complexes (median survival of 35 days). These data demonstrate that a nonviral IL-12 gene therapy employing synthetic cationic lipids as a delivery system can be used to inhibit the development of lung metastases. Thus, this method provides support for the use of IL-12/lipid complexes to control the growth of pulmonary metastases and represents a potentially safer alternative to IL-12 protein immunotherapy.

Fluorescence microscopy analysis indicated that the instilled formulation reaches the lower airways and alveolar region. Data also suggests cationic lipid-mediated gene expression is primarily localized in the lung parenchyma and not infiltrating cells isolated from the BALF.