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. 2019 Dec 1;11(12):1909.
doi: 10.3390/cancers11121909.

A Novel Anti-PD-L1 Vaccine for Cancer Immunotherapy and Immunoprevention

Affiliations
Free PMC article

A Novel Anti-PD-L1 Vaccine for Cancer Immunotherapy and Immunoprevention

Jie Chen et al. Cancers (Basel). .
Free PMC article

Abstract

Dendritic cells (DCs) are potent antigen-presenting cells that play a critical role in activating cellular and humoral immune responses. DC-based tumor vaccines targeting tumor-associated antigens (TAAs) have been extensively tested and demonstrated to be safe and potent in inducing anti-TAA immune responses in cancer patients. Sipuleucel-T (Provenge), a cancer vaccine of autologous DCs loaded with TAA, was approved by the United States Food and Drug Administration (FDA) for the treatment of castration-resistant prostate cancer. Sipuleucel-T prolongs patient survival, but has little or no effect on clinical disease progression or biomarker kinetics. Due to the overall limited clinical efficacy of tumor vaccines, there is a need to enhance their potency. PD-L1 is a key immune checkpoint molecule and is frequently overexpressed on tumor cells to evade antitumor immune destruction. Repeated administrations of PD-L1 or PD-1 antibodies have induced sustained tumor regression in a fraction of cancer patients. In this study, we tested whether vaccinations with DCs, loaded with a PD-L1 immunogen (PDL1-Vax), are able to induce anti-PD-L1 immune responses. We found that DCs loaded with PDL1-Vax induced anti-PD-L1 antibody and T cell responses in immunized mice and that PD-L1-specific CTLs had cytolytic activities against PD-L1+ tumor cells. We demonstrated that vaccination with PDL1-Vax DCs potently inhibited the growth of PD-L1+ tumor cells. In summary, this study demonstrates for the first time the principle and feasibility of DC vaccination (PDL1-Vax) to actively induce anti-PD-L1 antibody and T cell responses capable of inhibiting PD-L1+ tumor growth. This novel anti-PD-L1 vaccination strategy could be used for cancer treatment and prevention.

Keywords: PD-L1; dendritic cells; immune checkpoint; immunotherapy; tumor vaccine.

Conflict of interest statement

J.C., Y.L., Z.C., and X.F.H. are employees of Pomona Biotechnology Corp.

Figures

Figure 1
Figure 1
Expression and purification of the recombinant protein PDL1-Vax. (A) Protein fractions after lysis of bacterial cells (pET-PDL1-vax) were electrophoresed on 12% SDS–PAGE gels and then stained with Coomassie brilliant blue R250. Lane M, prestained protein molecular weight marker; lane 1, the cell pellets before IPTG induction; lanes 2, the cell pellets after isopropyl β-D-1-thiogalactopyranoside (IPTG) induction; lane 3, the soluble fraction after IPTG induction; and lanes 4, the insoluble protein fraction after IPTG induction. The recombinant protein PD-L1-Vax (arrow) is indicated. (B) Purified recombinant proteins PDL1-Vax were electrophoresed on 12% SDS–PAGE gels and then stained with Coomassie brilliant blue R250. (C) Western blot analysis of the purified recombinant protein PDL1-Vax using antibodies against human PD-L1 and IgG Fc. An irrelevant protein bovine serum albumin (BSA) was used as a negative control.
Figure 2
Figure 2
Induction of anti-PD-L1 antibody responses. Groups of C57BL/6 mice were immunized with various antigen-loaded BM-derived DCs (1 × 106 cells/mouse) twice at a weekly interval, and sera were collected from each group of mice 14 d later. (A) PD-L1-specific IgG levels from the pooled sera of each group (4–6 mice/group) were determined by ELISA. (B) PD-L1-specific IgG subclass levels from the pooled sera of PDL1-Vax-DCs-immunized mice (4–6 mice/group) were also determined by ELISA. p < 0.01, PDL1-Vax-DCs versus PDL1-DCs or IgG Fc-DCs.
Figure 3
Figure 3
Activation of PD-L1-specific B cells. Groups of C57BL/6 mice were immunized with various antigen-loaded BM-derived DCs (1 × 106 cells/mouse) twice at a weekly interval, and splenocytes were prepared from each group of mice (5 per group) 14 d later. Frequencies of anti-PD-L1 antibody-secreting B cells (ASC) in different groups of mice were determined and presented as the number of cells secreting PD-L1-specific IgG per 2 × 105 B cells. p < 0.01, PDL1-Vax-DCs versus PDL1-DCs or IgG Fc-DCs.
Figure 4
Figure 4
Inhibition of PD-1 and PD-L1 interaction. Sera were collected from each group of mice (immunized with various antigen-loaded BM-DCs. Inhibition of PD-1 and PD-L1 interaction by the addition of different amounts of the sera of the mice (5 per group), immunized with various antigen-loaded DCs, was performed using a competitive ELISA. The percentages of inhibition were determined and presented. p < 0.01, PDL1-Vax-DCs versus PDL1-DCs or IgG Fc-DCs.
Figure 5
Figure 5
Induction of PD-L1-specific CD4+ Th and CD8+ CTL responses. Groups of C57BL/6 mice were immunized with various antigen-loaded BM-derived DCs (1 × 106 cells/mouse) twice at a weekly interval, and, 14 d later, CD3+ T cells, CD4+ T cells, and CD8+ T cells were isolated from splenocytes of immunized mice (5 per group) were used for IFNγ ELISPOT assays. (A) Frequencies of PD-L1-specific CD3+ T cells are presented as the number of IFNγ-producing cells per 1 × 105 or 2.5 × 104 splenocytes. (B) Frequencies of PD-L1-specific CD4+ T cells. (C) Frequencies of PD-L1-specific CD8+ T cells. p < 0.01, PDL1-Vax-DCs versus PDL1-DCs or IgG Fc-DCs.
Figure 6
Figure 6
CTL assays. Groups of C57BL/6 mice were immunized with various antigen-loaded BM-derived DCs (1 × 106 cells/mouse) twice at a weekly interval, and, 14 d later, splenocytes taken from different group of mice were restimulated in vitro with recombinant PD-L1 protein. The restimulated splenocytes (E) were cocultured with 51Cr-labeled target cells Panc02-PD-L1 (T) at various ratios. The percentages of target cell killing by the splenocytes from different immunized mice were determined and shown. p < 0.01, PDL1-Vax-DCs versus PDL1-DCs or IgG Fc-DCs. The data represent the means of triplicate samples from one representative experiment of three (six mice/group); bars, SE.
Figure 7
Figure 7
Induction of antitumor immunity. Groups of mice (5 per group) were inoculated with 3 × 105 exponentially growing PD-L1+ tumor cells (murine pancreatic adenocarcinoma cells Panc02-PD-L1 or murine colon carcinoma cells MC38-PD-L1). Seven days later, mice were immunized with various antigen-loaded BM-derived DCs (1 × 106 cells/mouse) twice at a weekly interval. Tumor sizes were measured every 3–4 days. (A). Tumor volumes in each group are presented. p < 0.01, PDL1-Vax-DCs versus PDL1-DCs or IgG Fc-DCs. Data represent the means of one of three independent experiments; bars, SE. (B). Mouse death was recorded for evaluation of mouse survival percentage.

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