By Dean A. Haycock

Special To BioWorld Today

Gene therapy really could use a shot in the arm.

The snag with traditional therapeutic approaches is not in locating and harnessing the best cancer fighting genes, but in delivering them efficiently to their targets, which are often hidden deep in the body. Repeated, local injections aren¿t practical for deep tumors and widely scattered metastases. And ex vivo gene therapy, in which a therapeutic gene is inserted into a patient¿s cells and re-injected, takes a lot of time and effort.

The needed ¿shot in the arm¿ might some day come in the form of a literal shot in the arm, or other muscle, of patients with cancer and other diseases. At least, that might happen if the results from animal experiments described by researchers at Vical Inc., of San Diego, can be extended to human patients. In the Feb. 16 issue of the Proceedings of the National Academy of Sciences (PNAS), first author Holly Horton and her colleagues describe their success at treating three types of cancer in mice with intramuscular injections of plasmid DNA (pDNA) encoding murine interferon-a (mIFN-a). Their paper, ¿A gene therapy for cancer using intramuscular injection of plasmid DNA encoding interferon-a,¿ shows that it is possible to obtain potent anti-tumor effects on both primary and metastatic tumors in mice using this technique.

A plasmid is a kind of ¿paragene,¿ a genetic particle that exists separately from chromosomes. Found mostly in bacteria, plasmids can function and replicate as chromosome-bound genes, but they are not essential to basic cell function. Fitted with therapeutic genes, plasmids can serve as vectors for gene therapy. In this case, they were fitted with a mouse gene for interferon mIFN-a, a cytokine. Cytokines are low-molecular-weight proteins that act like hormones. Many different types of cells use them to signal other cells. They interest cancer researchers, because they regulate the duration and intensity of immune responses. In this way, they be used as anticancer agents.

Injections Worked In Three Types of Mouse Melanoma

Intramuscular injections of mIFN-a pDNA significantly slowed tumor growth and increased survival rates in three mouse tumor models: B16F10 melanoma, Cloudman melanoma, and Glioma 261. Mice received mIFN-a pDNA or control pDNA on the fourth day after tumor cells were implanted. Injections continued twice a week for three weeks. The finding was that mIFN-a pDNA treatment reduced B16F10 melanoma tumor volume by 89 percent after 28 days, compared to control pDNA. And 90 percent of the mice treated with the cytokine gene survived 39 days, compared to only 10 percent of controls. Similar results were found in the other cancer models. The treatment also significantly reduced the number of metastatic lung tumors in the B16F10 melanoma cancer model.

In mice with melanoma tumors, efficacy of the treatment appears to be due to the effects of mIFN-a on a particular type of immune cell, CD8+ T cells. Depletion of this cell type in mice, but not of CD4+ T cells, prevented the gene therapy from working. The idea of delivering genes in the form of pDNA is not new, of course, nor is the use of cytokines like interferon-a to fight cancer.

¿Traditionally in cancer gene therapy, either a plasmid or a viral vector that encodes a cytokine gene is injected directly into a tumor or around the tumor site,¿ first author Horton, a research investigator at Vical, told BioWorld Today. ¿The other way is to transfect cells in culture and transplant the cells that contain the plasmid.¿ Previously, intramuscular injections have also been used to deliver plasmids containing cytokine genes. Previous studies did not measure effects on growth of both solid and metastatic tumors, however.

The approach described by Horton involves injection of pDNA ¿into the muscle at a site distant from the tumor, a completely different place. In fact, you could treat metastatic disease when it is difficult to access an internal organ, by just a simple intramuscular injection,¿ Horton said.

Recombinant IFN-a protein ¿ not the gene ¿ is currently used as an adjuvant therapy for advanced melanoma. While showing signs of clinical benefit, this therapy still requires frequent injections and high doses that approach toxic levels.

The significance of the recent PNAS report lies in its expansion of previous pDNA intramuscular injection research. As few as three injections over six weeks produced noticeably beneficial effects against melanoma in mice. ¿Just a single injection produces pretty stable levels over a two week period,¿ Horton said. ¿It suggests that maybe you would just have to inject a few times to get stable serum levels.¿ This approach could avoid the peaks and valleys of serum cytokine levels that accompany injection of a protein with a short half-life.

Method Light Years Away¿ From Treating Humans

¿We did not see any side effects in the mice, even when we were giving them plasmids twice a week for three weeks,¿ Horton said.

Dawn Willis, scientific program director in the research department of the Atlanta-based American Cancer Society, said the finding is ¿an interesting observation but light years away from application in humans. The protein [interferon] is toxic so they can¿t give much [by injection]. This might be an alternative way to get it in, but we need a lot more work to find what cells it is going into.¿

Vical has no immediate plans to develop this particular approach independently, said Alan Engbring, director of investor relations at the company. ¿This paper was the result of a demonstration project,¿ he said. ¿We are very excited about the results, because it certainly demonstrates the potential of naked DNA as a delivery vehicle for a therapeutic protein. In this instance, that was for cancer, but [the method] can be used for a number of different indications.¿

Vical has several plasmid-based cancer products in clinical trials. Allovectin-7 (HLA-B7) is in Phase III for metastatic melanoma. Leuvectin, the IL-2 gene, is in Phase II for kidney cancer and in Phase I/II for prostate cancer. In collaboration with the National Cancer Institute, in Bethesda, Md., Vical is testing intramuscular injection of DNA encoding a melanoma antigen for use as cancer vaccine. n