An Antigen-Free, Plasmacytoid Dendritic Cell–Targeting Immunotherapy To Bolster Memory CD8+ T Cells in Nonhuman Primates

Abstract

The priming, boosting, and restoration of memory cytotoxic CD8 T lymphocytes by vaccination or immunotherapy in vivo is an area of active research. Particularly, nucleic acid-based compounds have attracted attention due to their ability to elicit strong Ag-specific CTL responses as a vaccine adjuvant. Nucleic acid-based compounds have been shown to act as anticancer monotherapeutic agents even without coadministration of cancer Ag(s); however, so far they have lacked efficacy in clinical trials. We recently developed a second-generation TLR9 agonist, a humanized CpG DNA (K3) complexed with schizophyllan (SPG), K3-SPG, a nonagonistic Dectin-1 ligand. K3-SPG was previously shown to act as a potent monoimmunotherapeutic agent against established tumors in mice in vivo. In this study we extend the monoimmunotherapeutic potential of K3-SPG to a nonhuman primate model. K3-SPG activated monkey plasmacytoid dendritic cells to produce both IFN-α and IL-12/23 p40 in vitro and in vivo. A single injection s.c. or i.v. with K3-SPG significantly increased the frequencies of activated memory CD8 T cells in circulation, including Ag-specific memory CTLs, in cynomolgus macaques. This increase did not occur in macaques injected with free CpG K3 or polyinosinic-polycytidylic acid. Injection of 2 mg K3-SPG induced mild systemic inflammation, however, levels of proinflammatory serum cytokines and circulating neutrophil influx were lower than those induced by the same dose of polyinosinic-polycytidylic acid. Therefore, even in the absence of specific Ags, we show that K3-SPG has potent Ag-specific memory CTL response-boosting capabilities, highlighting its potential as a monoimmunotherapeutic agent for chronic infectious diseases and cancer.

STING agonists activate latently infected cells and enhance SIV-specific responses ex vivo in naturally SIV controlled cynomolgus macaques

“…The flow cytometric analysis of monkey PBMCs using a modified LSRII flow cytometer (BD Biosciences, MA) was described previously 67,68 . Briefly, frozen PBMCs were thawed and then incubated in the presence of AZT (1 μM) and adjuvant (Table 1) in R-10 medium for 12 hours.…”

“…Following permeabilization (Cytofix/Cytoperm kit; BD Biosciences, MA), cells were stained with CD3-APC-Cy7 (SP34-2), IFN-γ–Alexa Fluor 488 (B27), TNF-PE-Cy7 (Mab11), and IL-2-APC (MQ1-17H12; all from BD Biosciences). For T cell analysis, cells were analysed using a modified LSRII flow cytometer (BD Biosciences, MA) as described previously 67 . Data were analysed using FlowJo software version 9.9.6 (Tree Star, OR).…”

A unique nanoparticulate TLR9 agonist enables a HA split vaccine to confer FcγR-mediated protection against heterologous lethal influenza virus infection

“…SPG, aluminum hydroxide gel (alum), and AddaVax were purchased from Invivogen (San Diego, CA, USA). The method for K3-SPG conjugation was described in a previous report (15).…”

“…To characterize the immune response in K3-SPG HA-immunized mice, we first evaluated the frequency of NC HA-and PR8 HA-specific CD8 + T cells in splenocytes of vaccinated mice because we previously reported that K3-SPG is a strong adjuvant for eliciting antigen-specific CD8 T cells (15). Seven days after the second vaccination, splenocytes were collected and stimulated with overlapping peptides corresponding to NC or PR8 HA for 6 h. We evaluated the frequency of all functional NC-and PR8-specific CD8 + T cells, on the basis of cell positivity for IFN-γ and/or TNF-α by flow cytometry (Fig.…”

Leveraging Endogenous Dendritic Cells to Enhance the Therapeutic Efficacy of Adoptive T-Cell Therapy and Checkpoint Blockade

“…In contrast to cDCs, pDCs develop fully in the bone marrow where after they traffic to secondary lymphoid organs ( 67 ). Activated pDCs are known to augment CD8+ T cell responses, even in the absence of specific antigen stimulation ( 68 ). In addition, studies indicate that TLR-activated pDCs can function as cross-presenting cells ( 69 – 71 ) and even have direct cytotoxic effects ( 72 , 73 ).…”