Mini-intronic plasmid vaccination elicits tolerant LAG3⁺CD8⁺T cells and inferior antitumor responses

Abstract

Increasing transgene expression has been a major focus of attempts to improve DNA vaccine-induced immunity in both preclinical studies and clinical trials. Novel mini-intronic plasmids (MIPs) have been shown to cause elevated and sustained transgene expression in vivo. We sought to test the antitumor activity of a MIP, compared to standard DNA plasmid immunization, using the tumor-specific antigen SSX2 in an HLA-A2-restricted tumor model. We found that MIP vaccination elicited a greater frequency of antigen-specific CD8C T cells when compared to conventional plasmid, and protected animals from subsequent tumor challenge. However, therapeutic vaccination with the MIP resulted in an inferior antitumor effect, and CD8C tumor-infiltrating lymphocytes from these mice expressed higher levels of surface LAG3. Antitumor efficacy of MIP vaccination could be recovered upon antibody blockade of LAG3. In non-tumor bearing mice, MIP immunization led to a loss of epitope dominance, attenuated CD8 C cytokine responses to the dominant p103 epitope, and increased LAG3 expression on p103-specific CD8 C T cells. Further, LAG3 expression on CD8 C T cells was associated with antigen dose and persistence in spite of DNA-induced innate immunity. These data suggest that for antitumor immunization, approaches leading to increased antigen expression following vaccination might optimally be combined with LAG3 inhibition in human trials. On the other hand, mini-intronic vector approaches may be a superior means to elicit LAG3-dependent tolerance in the treatment of autoimmune diseases.

Engineering DNA vaccines against infectious diseases

“…It has been reported that the FDA recommends integration studies, regardless of the delivery method, if there are greater than 10,000 copies of foreign DNA per microgram of host DNA [71]. Interestingly, minicircle DNA [51,53,54,72,73] and ministring DNA [74,75] have been reported to have less risk of insertional mutagenesis because the major bacterial DNA (i.e., the unmethylated CpG repeats functioning as PAMPs) is removed.…”

DNA vaccines for prostate cancer

“…We found that increasing antigen dose or duration of antigen expression by DNA immunization increased the expression of LAG3 on CD8+ T cells and rendered them ineffective against an SSX2-expressing tumor. These data suggest that increased gene expression per se may not confer additional benefit in the absence of methods to block tumor mechanisms of resistance (Colluru et al, 2016b). However, other approaches to increase gene expression are being explored.…”

“…Similarly, we have demonstrated that modifications to DNA vaccines that led to increased antigen expression resulted in increased expression of another checkpoint inhibitor, LAG-3, on antigen-specific CD8+ T cells. Combining LAG-3 blockade with DNA vaccination similarly elicited greater anti-tumor activity than either treatment alone (Colluru et al, 2016b). We have also found that patients with prostate cancer, treated with a DNA vaccine, developed PD-1-regulated antigen-specific T-cell responses, and increases in PD-L1 on circulating tumor cells (Rekoske et al, 2016).…”

“…Finally, ongoing studies evaluating the means by which tumors avoid immune detection, and whether some of these mechanisms are used preferentially by prostate cancer, will provide the most rational combination treatments using DNA vaccines in combination with agents targeting specific mechanisms of resistance. Specifically, we believe that trials combing DNA vaccines with checkpoint inhibitors, including agents targeting CTLA-4, PD-1/PD-L1, and/or LAG3, will be most effective given recent results in murine studies (Rekoske et al, 2015; Colluru et al, 2016b). Moreover, a combination of a DNA vaccine with PD-1 blockade has recently demonstrated encouraging findings with objective clinical responses observed in patients with advanced stage prostate cancer, NCT 02499835 (McNeel et al, 2016).…”

Trial watch: DNA-based vaccines for oncological indications

“…77 However, most malignancies arise from normal tissues and only express endogenous TAAs, which are normally covered by central or peripheral tolerance (and hence are unable to drive an immune response). 78,79 Hence, several strategies have been developed to overcome tolerance and improve the immunogenicity of TAAs, including the co-expression (in cis or trans) of adjuvant-like molecules. 27,79 Of note, neoplastic cells also express a large panel of so-called "tumor neoantigens", which result from the mutational processes that accompany tumor progression.…”