We have previously reported the results of a clinical trial in which patients with biochemically recurrent prostate cancer were immunized 6 times at 2-week intervals with a DNA vaccine encoding PAP.21
Ultimately, the goal of antitumor vaccines is to elicit a sustainable immune response able to eradicate or restrain a tumor. Hence, most clinical vaccine trials have used schedules of immunization, informed by animal studies, conducted over a limited period of time with the hope of eliciting durable anti-tumor immune responses. This approach is in contrast to most current targeted molecular pharmacologic approaches to the treatment of cancer in which it is generally assumed that chronic therapy is necessary. In addition, as most preclinical immunization studies are performed in inbred rodent strains, estimations of appropriate vaccine schedules may not be uniformly applicable to human trials. Moreover, the means of immunization may greatly affect optimal schedules of administration. To date, despite anti-tumor vaccines having entered phase 3 clinical trials, there has been very little evaluation of whether the number and schedule of immunizations is sufficient to elicit an immune response that is potentially therapeutic and durable. This leaves questions about whether prolonged schedules of administration should be considered, or booster immunizations should be used. In addition, there have been concerns that repetitive immunization might lead to counterproductive tolerant immune responses.23
In an earlier vaccine trial, patients were immunized 6 times at 2-week intervals with a DNA vaccine encoding PAP, a schedule that we had investigated earlier in rats.16
The current analysis was conducted to answer several questions aimed at guiding the schedule of vaccine administration to be used in future clinical trials with this particular DNA vaccine. Although the number of samples evaluable was relatively small, we sought to determine the following:
- At what point in the series of 6 vaccinations were immune responses detectable? This was aimed primarily at answering whether 6 vaccinations were sufficient, or whether ongoing immunization should be considered.
- Were immune responses, once elicited, durable, and/or were immune responses only detectable months later?
- Was the development of a durable immune response associated with beneficial changes in PSA kinetics?
- Once immunized, could booster immunizations with the same DNA vaccine subsequently augment an antigen-specific T-cell response.
Using IFNγ ELISPOT as a measure of PAP-specific T cells, we found that in those individuals in whom PAP-specific CD4+
T-cell responses were elicited over the course of immunization, the majority of responses were elicited late in the course of DNA immunization. This was similar to what we observed earlier in rats in which although immune responses to the human PAP xenoantigen were elicited after as few as 2 vaccinations, more vaccinations were necessary with a vaccine encoding the rat PAP self-antigen to elicit responses.17
This finding suggests that it could be advantageous to continue immunization beyond 6 initial vaccinations, using DNA immunization alone without a heterologous immunization strategy. Alternatively, it is possible that some individuals were tolerant to PAP at baseline and might not develop detectable PAP-specific T cells. Future studies will explore continued immunization, beyond 6 initial vaccinations, to evaluate these possibilities. However, as suggested earlier, these types of studies must be performed with some caution given that genetic vaccines delivered at frequent intervals over prolonged periods may elicit tolerant responses, and have in fact been investigated by other groups as a means of inducing T-cell tolerance.24
We have identified earlier that rats immunized multiple times with the same DNA vaccine encoding human PAP developed PAP-specific memory CD4+
T cells detectable by antigen-specific T-cell proliferation for at least 1 year after immunization (data not shown). Although in that system the human PAP is a xenoantigen, we similarly identified that several patients had long-lived responses detectable many months after immunization (). Although such findings complicate clinical trial evaluation, as it is difficult to define an appropriate time to measure immune responses, they suggest that immune monitoring at static time points may be insufficient to characterize what is likely a more dynamic process. This may be one reason why many vaccine trials to date have failed to demonstrate an association between the development of an immunologic response and a clinical response. In our previous trial, patients were enrolled with biochemical (serum PSA) recurrence only, and we identified that 8 patients experienced a ≥200% increase in PSA doubling time from pretreatment to the 1-year period of time after treatment.21
We also identified that 10 of 22 patients had a PAP-specific response detectable immediately after immunization, although this was not associated with changes in PSA doubling time.21
Although changes in PSA doubling time have to be interpreted with caution, because this has not been validated prospectively as a clinical trial endpoint, changes in PSA kinetics remain the only marker of possible clinical response in this population of patients with early recurrent disease. In this study, we identified that changes in PSA doubling time were highly associated with the development of late/durable immune responses. Future clinical trials will evaluate whether the durability of PAP-specific T cells, and effector and memory T-cell immune responses in particular, are positively associated with clinical measures such as time to disease progression.
Finally, our studies demonstrate that immune responses to PAP can be subsequently augmented with the same DNA vaccine encoding PAP. In one individual, PAP-specific responses were detected after as few as 2 booster immunizations in terms of proliferative T-cell responses and specific CD8+
T-cell populations. Of note, this particular individual was treated with only 100 μg of plasmid DNA for each immunization. Although the immunization of a single patient is anecdotal, it does provide evidence that comparable doses of plasmid DNA as used in rodent studies, not dosed per weight, can be immunologically effective in human immunization. A similar observation that higher plasmid DNA doses are not necessarily immunologically superior to lower doses was recently reported in a phase 2 DNA vaccine trial for patients with multiple sclerosis.25
Our results suggest that ongoing boosters may have led to decreased antigen-specific proliferative responses, however, this again remains anecdotal. In any case, our findings demonstrate that antigen-specific T-cell immune responses, having been elicited earlier, can be augmented with a DNA vaccine, although optimal schedules of vaccination may need to be evaluated. The specific finding that this DNA vaccine encoding PAP can augment PAP-specific T-cell immunity may be of particular relevance given that a vaccine being evaluated by Dendreon Corporation, sipuleucel-T, is targeting the same PAP antigen and has demonstrated clinical benefit in terms of improving patients’ overall survival.7,26
An “off-the-shelf ” DNA vaccine targeting the PAP antigen could potentially be investigated in combination with that vaccine to maintain or augment long-term PAP-specific T-cell immunity.