A Squalene-Based Nanoemulsion for Therapeutic Delivery of Resiquimod

Abstract

Agonists for toll-like receptors (TLRs) have shown promising activities against cancer. In the present study, a squalene-based nanoemulsion (NE) was loaded with resiquimod, a TLR7/8 agonist for therapeutic delivery. R848 NE was developed and characterized for long-term stability. In vitro and in vivo antitumor immunity of R848 NE were also evaluated in combination with SD-101, a CpG-containing TLR9 agonist. In vitro studies demonstrated strong long-term stability and immune responses to R848 NE. When combined with SD-101, strong antitumor activity was observed in MC38 murine colon carcinoma model with over 80% tumor growth inhibition. The combination treatment showed a 4-fold increase in systemic TNFa production and a 2.6-fold increase in Cd8a expression in tumor tissues, suggesting strong cell-mediated immune responses against the tumor. The treatment not only demonstrated a strong antitumor immunity by TLR7/8 and TLR9 activations but also induced PD-L1 upregulation in tumors, suggesting a potential therapeutic synergy with immune checkpoint inhibitors.

Lipid metabolism in tumor immunology and immunotherapy

“…Lipid nanoparticles are widely used to enhance antitumor responses by increasing the immunogenic effects of cancer immunotherapy. One example is resiquimod, a hydrophobic agonist of the Toll-like receptor (TLR) -7/8 used as an adjuvant in topical preparations for skin carcinomas; it has also been made more water-soluble by combining it with lipid particles and administering the compound systemically, to increase the efficacy of immunotherapy ( 106 ). Another group also showed that a liposomal preparation of resiquimod improved the adjuvant’s pharmacokinetics and prolonged its retention time in the blood, which also improved the effectiveness of immunotherapy ( 107 ).…”

Application of lipid-based nanoparticles in cancer immunotherapy

“…The viscosity, refractive index, density, phase behavior, and interfacial tension of the oils greatly determine the stability and functional properties of NEs ( 61 ). Other function of oils in NE-based drug delivery is to solubilize hydrophobic agents such as poor water-soluble chemotherapies and immunomodulatory agents ( 62 , 63 ). It has been reported that some of the fusogenic oils (such as medium-chain oils, long-chain oils, and squalene) may stimulate innate immune system by activating toll-like receptors (TLRs) ( 64 – 66 ).…”

“…Imidaziquinoline-like TLR 7/8 agonists, including imiquimod and resiquimod (R848), was loaded into the liposomal formulation to prolong its retention time in blood circulation ( 96 ). When considering the poor water solubility of TLR7/8 agonists, NEs formulation is also a suitable delivery platform to improve drug solubility thus the drug loading ( 63 ). Synthetic TLR9 agonists are oligonucleotides (ODNs) with cytosine-phosphate-guanine (CpG) motifs.…”

mRNA delivery in cancer immunotherapy

“…Albeit the accomplishment in optimizing mRNA moieties, targeted cellular uptake is an urgent need for safe, precise, and efficient delivery materials. In recent decades, multiple drug delivery nanomaterials have been proposed and developed, including lipids 28 , 29 , 30 , 31 , 32 , polymers 33 , lipid‒polymer hybrid 34 , and protein/peptide-mediated carriers ( Fig. 2 ) 35 , 36 .…”