Can DNA Vaccines Get a Boost from Cytokines?

By Mark Boaz and Richard Jefferys*

Although DNA-based vaccines are beingdeveloped against many diseases, results in humans have not lived up to the initial promise shown in animal models. By themselves they are usually poor inducers of antigen-specific immunity; as the first of two vaccines in a prime-boost combination—the most common way to use DNA vaccines—the jury is still out, although Merck’s data on DNA/Adenovirus5-based vaccines in humans so far are disappointing (see Update on Merck's AIDS Vaccine Program ).

Several studies presented in Banff set out to address this problem by combining DNA vaccines with “molecular adjuvants”—molecules such as cytokines that may enhance immune responses.

One such approach was described in back-to-back talks by collaborators John Eldridge (Wyeth-Aherst) and David Weiner (University of Pennsylvania), who are analyzing the effects of cytokines IL-12 and IL-15 on the immunogenicity of a DNA vaccine against SIV. Their data in macaques show that these cytokines enhance not only cellular immune responses but also the level of antibodies, which are not usually induced by DNA vaccines. Challenge data are not yet available, but will provide crucial information on whether this enhanced immunogenicity leads to better protection, as it did in earlier studies of this approach (using IL-2) by Dan Barouch and Norman Letvin of Harvard Medical School.

IL-12 and DNA: Proof of Principle

Eldridge began by presenting a study in macaques comparing responses to an SIV-gag DNA vaccine given with and without a plasmid encoding IL-12. This cytokine is thought to be one of the most potent adjuvants for inducing cellular immune responses, and has been shown to improve vaccine protection in flu, malaria and other disease models in animals. In the Wyeth study, five groups of five macaques were immunized at weeks 0, 4 and 8 with SIV-gag (1.5 or 5mg of DNA) with or without IL-12. Immunogenicity was determined by measuring IFN-gamma-producing cells using Elispot assays.

Looking at the 10-week timepoint, the data show clear enhancement of immune responses in animals vaccinated with IL-12 plus SIV-gag, compared to those given DNA vaccine alone. The increase was seen in both the number of responders (10/10 versus 5/10 with DNA only) and the magnitude of the T-cell response, with peak responses about 5-fold higher when IL-12 is present. (Mean numbers of spot-forming cells [SFC] with vaccine plus IL-12 were 1,344 and 1,433, for low and high DNA doses, respectively, compared to 256 and 338 with DNA alone.) IL-12 also led to more persistent responses, with roughly two-thirds of peak response levels retained at week 20 (about 1,000 SFC), but only 30-50% (about 100 SFC) with DNA vaccine alone.

Next, Eldridge described findings on IL-12 in a prime-boost regimen with SIV-gag DNA and a second vaccine based on Vesicular Stomatitis Virus (VSV), made by John Rose’s group at Yale University together with Wyeth. After priming with SIV-gag and IL-12 DNA, animals boosted twice with VSV vaccine (containing SIV-gag and HIV-env) showed an 8-fold increase in Elispot responses to Gag, peaking at a mean of 3,772 SFC. Anti-Gag binding antibodies were also boosted by half a log (to a mean titer of 32,768, compared to 4,096 after DNA priming). The observed response levels are comparable to those obtained with other prime-boost regimens that protect monkeys from disease (such as Harriet Robin-son’s DNA/MVA and Merck’s DNA/Adenovirus).

IL-15: Boosting Memory

Following Eldridge’s talk, David Weiner described studies showing enhancement by IL-15, an important cytokine for the induction and maintenance of CD8 memory. Macaques were divided into three groups of six animals, with one group receiving DNA vaccine alone, the second given DNA vaccine plus IL-15 plasmid, and the third, placebo. Data after two of three planned vaccinations (the study is still ongoing) showed the highest responses in the DNA/IL-15 group (averaging 447 SFC), with all 6 animals responding. Only 3/6 animals in the DNA-only group responded, with one “outlier” showing a robust response (557 SFC) and the remaining ones averaging 50 SFC.

Weiner also presented data from mice showing that IL-15 can partially replace CD4 T-cells as helpers for CD8 responses in animals depleted of CD4 cells and vaccinated with DNA/IL-15. He speculates that this might suggest an important use for IL-15 in therapeutic vaccination, since most HIV-infected people have reduced CD4 cell numbers and/or impaired function.

Summarizing the overall approach, Weiner concluded that the use of IL-12 or IL-15 might enable DNA vaccines to achieve levels of immunogenicity comparable to those reported for viral vectors. He also emphasized that humoral as well as cellular responses appear achievable—raising the possibility that DNA might be a feasible vector to deliver future immunogens that induce neutralizing antibodies.

Clinical Development Pathway

To move the approach into clinical trials, plans call for two preventive vaccine studies with the US HIV Vaccine Trials Network, plus a therapeutic trial at the University of Pennsylvania. The preventive trials will use an HIV-gagDNA vaccine and either IL-12 or IL-15, and should start within a year; both studies may expand later to include a boost with the HIV-VSV vaccine. Discussions are also underway with the Walter Reed Army Institute of Research (WRAIR) on combining its multi-clade DNA vaccine candidate with cytokines.

In the meantime, studies are underway to investigate some regulatory questions arising from the use of cytokines as vaccine adjuvants. In a poster by Sangeeta Bhargava Periwal and the Wyeth/U. Penn. team, the researchers showed that expression of IL-12 after immunization with plasmid is relatively short-lived, peaking at six days and showing only trace levels at 30 days. Weiner says that ongoing discussions with the FDA are defining other potential issues and the studies needed to address them.

*Richard Jefferys is Basic Science Project Director at the Treatment Action Group, an AIDS advocacy organization based in New York.