Interferon alpha and antisense k-ras rna combination gene therapy

This application is a continuation of U.S. application Ser. No. 11/114,088, filed Apr. 26, 2005, which claims benefit of priority of U.S. provisional application No. 60/565,526, filed Apr. 27, 2004, the entire disclosures of each of which are incorporated by reference herein in their entireties.

1. Field of the Invention

The invention relates to a combination therapy comprising the use of interferon alpha (IFN-α) and antisense K-ras RNA against cancer. The invention also relates to vector-mediated co-expression of antisense K-ras RNA and IFN-α gene for the treatment of cancer. In addition, the invention relates to a synergistic cytotoxic effect of the combination therapy of antisense K-ras RNA and IFN-α against pancreatic cancer.

2. Discussion of Background Information

Adenocarcinoma of the pancreas is highly aggressive and is one of the most difficult cancers to treat at present (1, 2). One major reason for the poor prognosis is that the cancer has a high potential to infiltrate to the surrounding tissues and metastasize even in the early stage. Despite advances in chemo- and radiotherapies and surgical techniques, patients with pancreatic cancer often succumb to local recurrence or metastatic spread (1-3), and new therapeutic strategies against this cancer are an urgent need.

Interferon alpha (IFN-α) protein is a cytokine with multiple biological activities that include antiviral activity, regulation of cell proliferation and differentiation and immunomodulation (4), and it is used worldwide for the treatment of more than 14 types of cancers including some hematological malignancies (hairy cell leukemia, chronic myeloid leukemia, some B- and T-cell lymphoma) and certain solid tumors such as melanoma, renal carcinoma, and Kaposi\'s sarcoma (4, 5). The growth inhibitory effect of IFN-α protein has been documented in pancreatic cancer cells (6, 7). Recent clinical trials using this cytokine in combination with standard chemotherapeutic drugs also showed some potential anti-tumor activity against pancreatic cancer, but the current clinical protocols failed to document sufficient evidence to enlist pancreatic cancer as a clinically effective target of IFN-α (8-11).

An improved therapeutic effect and safety are possibly achieved by IFN-α in a gene therapy context, because it may allow increased and sustained local concentrations of this cytokine in the target sites while preventing unnecessary systemic distribution (12). Actually, in recent years, IFN-α delivered by adenovirus (12-15), retrovirus (16, 17) or nonviral vectors (18, 19), has been shown to suppress growth of various cancers such as breast cancer, renal cell cancer, basal cell cancer and leukemia in mice. However, the study of IFN-α gene transfer against pancreatic cancer has not been reported. Thus, the antiproliferative effect of IFN-alpha gene transduction into pancreatic cancer cells has been investigated. IFN-alpha expression significantly suppressed cell growth and induced cell death in pancreatic cancer.

It has previously been reported that the expression of antisense K-ras RNA suppressed the growth of pancreatic cells with K-ras point mutation (20-22). The interferon-inducible 2′,5′-oligoadenylate synthetase (2-5AS)/RNase L pathway has been shown to activate the apoptotic pathway (23-26). 2-5AS is a double-stranded-RNA-dependent synthetase that generates the activators for Rnase L (23-26) (FIG. 1). The use of IFN-α in combination with antisense K-ras RNA for suppressing growth or inducing apoptosis of pancreatic cancer cells has not been previously reported.

The invention relates to a method of treating pancreatic cancer. The invention also relates to combination therapy for treating pancreatic cancer comprising administering therapeutically effective amounts of IFN and antisense K-ras to a mammal in need thereof.

The invention provides a method of treating pancreatic cancer cells in a mammal, comprising:

administering therapeutically effective amounts of AxCA-IFN adenoviral vector and antisense K-ras expressing vector to pancreatic cancer cells,

wherein the AxCA-IFN adenoviral vector comprises a DNA sequence encoding a human IFN-alpha gene, wherein the human IFN-alpha gene is operably linked to a promoter, and the administered cancer cells express the human IFN-alpha gene; and the antisense K-ras expressing vector comprises a DNA sequence encoding an antisense K-ras gene, wherein the antisense K-ras gene is operably linked to a promoter, and the administered cancer cells express the antisense K-ras gene; and

obtaining regression of pancreatic cancer cells in the mammal by expression of the human IFN-alpha gene and the antisense K-ras gene.

The invention may further provide the above method wherein the promoter of the AxCA-IFN adenoviral vector is CAG promoter, wherein the CAG promoter is a beta-actin promoter fused to a cytomegalovirus enhancer element.

The invention may also provide that the promoter of the antisense K-ras expressing vector may be a CAG promoter, wherein the CAG promoter is a beta-actin promoter fused to a cytomegalovirus enhancer element.

The invention may also comprise AxCA-IFN adenoviral vector administered in a dose, for example, ranging from about 4×10⁹ to 20×10⁹ pfu (plaque forming unit)/day/kg body weight.

The invention may also comprise antisense K-ras expressing vector administered in a dose, for example, ranging from about 4×10⁹ to 20×10⁹ pfu (plaque forming unit)/day/kg body weight.

The invention may include administering both vectors to pancreatic cancer cells at the pancreas, administering both vectors to pancreatic cancer cells anywhere in the mammal, or a combination thereof.