By David N. Leff

Synergism is the name of the game being played at McMaster University, in Hamilton, Ontario.

The players are virologists and immunologists, who have synergized two molecules of the immune system and scored much greater anticancer effects than with either one alone. The first is a cytokine, interleukin-12 (IL-12); the second, an antigen-fingering protein called B7.

The synergizing force that brings these two together, and sends them into action against tumors, is an adenoviral vector, much used in gene therapy despite its drawbacks.

A paper in the Sept. 30 Proceedings of the National Academy of Sciences (PNAS) gives a play-by-play account of the game. Its title: "Interleukin 12 and B7-1 costimulatory molecule expressed by an adenovirus vector act synergistically to facilitate tumor regression."

Its senior author is molecular biologist and pathologist Frank Graham, at McMaster. Molecular virologist Mary Hitt is a co-author.

"Previously," she told BioWorld Today, "we used an adenovirus (AV) that expressed IL-12 only. With it, we got about 30 percent tumor regression in mice."

IL-12 is a cytokine that recruits armies of cell-killing T cells to attack malignant tumors. B7 bullies those tumor cells into running their antigenic immune-system targets up the flagpole, where T cells will notice them and mow those cells down.

By doubling up those two anticancer co-conspirators into a single AV-driven gene delivery package, that 3 percent regression score more than doubled, to 70 percent.

"With our new virus," Hitt observed, "the dose of IL-12 and B7 we needed to get really good tumor regression was below the level that gives toxicity. So the mice weren't getting sick, but they were regressing the tumors."

Hitt pointed out: "You can stimulate the immune system in two different ways. One, by saying: 'Here's the antigen that you need to react to.' And also by saying, 'Send out the IL-12 cytokine, and tell it it's time to make killer T cells differentiate and proliferate.'"

IL-12 does both of those things at the same time. "And we put the B7 in because it makes the tumor cell itself look — to a T lymphocyte — like an antigen-presenting cell. That's how it stimulates the T cell response."

The McMaster team constructed a transgenic mouse programmed to contract massive murine mammary tumors by the time it was four months old.

"So we took out tumor cells from the transgenic animals," Hitt recounted, "explanted them ex vivo for a couple of days, then implanted one million cells in the flanks of other, normal mice, where they grew into large murine breast cancer tumors."

Into 10 mice carrying these tumors, the co-authors injected the adenoviral vector containing their combined IL-12/B7 plasmid, and recorded total regression in nine animals.

Tumor-Challenged Mice Win, 9:0

Two to three months later, they challenged these tumor-free rodents with an inoculation of fresh mammary cancer cells in the opposite flank. "No tumors were induced in any of the mice," the PNAS paper reported, "whereas 100 percent of age-matched syngeneic control animals developed tumors within 21 days after injection. These results," it concluded, "indicate that [the combined vector] administration is associated with long-term antitumor immunity, which might inhibit tumor recurrence and prevent metastasis."

Hitt volunteered that "because this is really effective in mice, we have a desire to go into human trials with a similar construct. It's now in the works here, extremely preliminary; our plan is for clinical trials, probably in months."

Meanwhile, at the neighboring University of Toronto, a close collaborator of the McMaster group has just completed a Phase I trial of that same adenovirus vector, but delivering interleukin-2 only, not IL-12.

Molecular oncologist Keith Stewart told BioWorld Today: "It's Frank Graham's mouse model and vector. We just made human clinical-grade material, and tested it in 21 patients with breast cancer or melanoma."

That trial ran from November 1996 to September 1997. It was sponsored, Stewart said, by the gene therapy division of Baxter Healthcare Corp., in Deerfield, Ill.

Besides finding "no toxicity to speak of," he observed that preliminary efficacy results indicated "some local tumor responses only."

Additional Clinical Trials Planned

Stewart is now "in the process of repeating the same study with the new combined IL-12-B7 McMaster vector. We're sort of their clinical branch," he remarked. "Now we're going for production of clinical material, and expect to start the study in the spring of next year, because it takes so long to make the stuff."

This upcoming Phase I protocol will enroll the same types of patients. "Those with melanoma," Stewart said, "will require no prior therapy to enter; those with breast cancer have to have failed at least one round of chemotherapy."

He added that "Genetics Institute [of Cambridge, Mass.], which owns the patents to both IL-12 and B7, has expressed interest in this planned trial."

Assuming that the synergistic combination vector passes all three clinical trial phases, Stewart envisages a scenario in clinical oncology practice "that it would be used as a minimal disease strategy. So for breast cancer, for example, this would probably mean injecting a primary lesion prior to surgical resection. In melanoma, it's not quite so clear. It would probably be what we're doing now, multiple injections."

One drawback of the adenovirus as a gene therapy vector is the antiviral immune response it provokes in a recipient patient. Stewart foresees this as happening in the course of future therapy. "But it's not clear," he commented, "that that would abrogate any antitumor effect. In fact," he added, "it might be beneficial. You're generating an immune response against virally infected cells.

"In this circumstance," he concluded, "enhanced antibody production against the virally infected cells may actually be of benefit, rather than the reverse." *