Systemic TLR7/8 micelles trigger a novel and potent anti-tumor response by strong recruitment of neutrophils leading to massive tumor cell killing.
Authors: Jensen Simon Skjøde , Christensen Esben , Petersen Morten Just , Munther Rasmus , Hansen Anders , Bak Martin , Stavnsbjerg Camilla , Panina Svetlana , Andresen Thomas 
Published: June 2022

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Background: Clinical use of TLR7/8 agonists is currently restricted to topical application of Imiquimod for treatment of superficial basal cell carcinoma since systemic administration is prevented due to dose-limiting toxicity. Here, we present a novel micelle-based drug delivery technology containing a lipid-anchored TLR7/8 agonist for intravenous administration. The MBS8 formulation shows good efficacy in mouse cancer models, is well tolerated in rodents and non-human primates and is currently being tested in clinical studies (NCT04855435). Methods: MBS8 was tested in 12 syngeneic mouse cancer models as monotherapy or in combination anti-PD-1 and anti-PD-L1 antibodies. MBS8 was administered IV as slow bolus. Tumors and spleens were analysed using histology, immunohistochemistry, ELISPOT and flow cytometry. Phenotyping of tumor microenvironment was done using flow cytometry and Nanostring analyses. Safety and tolerability were tested in mice, rats and monkeys. Safety-related and PD biomarkers were identified in monkeys using a multiplex MSD technology and validated in whole human blood stimulated with MBS8 ex-vivo. Results: In several syngeneic mouse tumor models, MBS8 led to complete eradication of established tumors. The complete responders showed tumor rejection upon re-challenge. In these mice, a CD8+-dependent tumor-specific immune memory response was evident. Tumors demonstrated a massive tissue necrosis ( > 80%) within 24-48h after the first drug administration and showed massive neutrophil infiltration 6h after dosing. Antigen specific CD8+ T-cells were detected in tumors 24 hours after treatment with increased CD8+:Treg cell ratio and enhanced antigen presentation in tdLN. MBS8 showed both additive and synergistic effect when combined with anti-PD-1 or anti-PD-L1 antibody treatment. Moreover, in tumors being resistant to anti-PD-1 treatment, co-administration of MBS8 reverted sensitivity to anti-PD-1 treatment in a synergistic manner. A panel of cytokines induced by MBS8 upon repeat dose administration in monkeys was identified in plasma as potential biomarkers. Ten of them were further evaluated in whole human blood treated with MBS8 ex vivo. Currently, these cytokines are being used in Phase I clinical studies. Conclusions: MBS8 showed significant anti-cancer activity in multiple in vivo solid tumor models when administered either as monotherapy or in combination with ICIs. MBS8 demonstrated a novel mode of action with neutrophils playing a central role as primary effector cells causing a rapid killing of tumors. Further, adaptive immune response was initiated including generation of tumor specific CD8+ cells and establishment of the immune memory. MBS8 was well tolerated in rodents and cynomolgus monkeys at the dose levels above therapeutic effective doses identified in mice, thus providing a good therapeutic window.