Cancer vaccines using synthetic long peptides (SLP) targeting tumor antigens have been tested in the clinic, but the outcomes have been unimpressive. In this study, Arbelaez et al. investigated the impact of delivering neoantigen peptides in liposomes on tumor-infiltrating CD8+ T cells in C57BL/6 mice bearing MC38-G12D tumors.

Study Description

The mutated neoantigen Adpgk peptide derived from the antigen G12D, and the adjuvant CpG were formulated into liposomes (Neo-Lpx) to immunize mice with MC38-G12D tumors five days after subcutaneous tumor implantation in C57BL/6 mice. Mice were administered Neo-Lpx (50µg CpG/peptide) after randomization on day 14. Tumor-infiltrating leukocytes (TIL) were prepared from harvested MC38-G12D tumors to detect Adpgk-specific CD8+ T cells by flow cytometry using custom Adpgk/H2Db Dextramer® reagents.


CD8+ TILs specific for the neoantigen peptide Adpgkwere enriched in tumors after treatment with Neo-Lpx (Fig. 1a). Due to heterogeneity in antigen specificity and activity of TILs, it was important to analyze the tumor-infiltrating T cells responding to the tumor antigens. Thus, researchers focused on markers of chronic antigen stimulation that may lead to functional T-cell exhaustion. High levels of PD-1 and Tim3 on all the TILs from Neo-Lpxtreated tumors were observed. Using the AdpgkDextramer®reagents, researchers compared the lipoplex vaccine-expanded TILs to bulk TILs and found that Neo-Lpxtreated TILs exhibited a higher dysfunctional signature (Fig. 1b).

Fig. 1: Detection and checkpoint signature of tumor-infiltrating, Adpgk-specific CD8+ T cells. a) Detection of mutant Adpgk-specific CD8+TILs in mice immunized with Neo-Lpx using an MHC I Dextramer® specific for the mutated neoantigen peptide Adgk derived from the antigen G12D. b) TILs specific or not for Adpgk using MHC I Dextramer® staining were analyzed for PD-1 and Tim3 expression.


  • Researchers successfully detected Adpgk-specific CD8+ T cells in MC38 tumors from Neo-Lpx immunized C57BL/6 mice using MHC I Dextramer® reagents customized for the purpose
  • The higher dysfunctional signature compared to untreated bulk TILs, as measured by PD-1 and Tim3 expression led to further studies combining the Neo-Lpx with checkpoint inhibitor anti-PD-1


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