Make It Simple: (SR-A1+TLR7) Macrophage Targeted NANOarchaeosomes

PARRA, FEDERICO LEONEL; CAIMI, AYELEN TATIANA; ALTUBE, MARIA JULIA; CARGNELUTTI, DIEGO ESTEBAN; VERMEULEN, MÓNICA ELBA; DE FARIAS, MARCELO ALEXANDRE; PORTUGAL, RODRIGO VILLARES; MORILLA, MARIA JOSE; ROMERO, EDER LILIA

Frontiers in Bioengineering and Biotechnology

Frontiers Media S.A.

Año: 2018 vol. 6

Hyperhalophilic archaebacteria exclusively produce sn2,3 diphytanylglycerol dietherarchaeolipids, unique structures absent in bacteria and eukaryotes. Nanovesiclesmade of archaeolipids known as nanoarchaeosomes (nanoARC), possess highly stablebilayers, some of them displaying specific targeting ability. Here we hypothesize thatnanoARC made from Halorubrum tebenquichense archaebacteria, may constituteefficient carriers for the TLR7 agonist imiquimod (IMQ). NanoARC-IMQ takesadvantage of the intense interaction between IMQ and the highly disordered, poorlyfluid branched archaeolipid bilayers, rich in archaeol analog of methyl ester ofphosphatidylglycerophosphate (PGP-Me), a natural ligand of scavenger receptor A1(SR-A1). This approach lacks complex manufacture steps required for bilayers labeling,enabling future analytical characterization, batch reproducibility, and adaptation tohigher scale production. SR-A1 mediated internalization of particulate material is mostlytargeted to macrophages and is extensive because it is not submitted to a negativefeedback. A massive and selective intracellular delivery of IMQ may concentrate itseffect specifically into the endosomes, where the TLR7 is expressed, magnifying itsimmunogenicity, at the same time reducing its systemic bioavailability, and therefore it?sin vivo adverse effects. NanoARC-IMQ (600?900 nm diameter oligolamellar vesicles of∼−43 mV Z potential) were heavily loaded with IMQ at ∼44 µg IMQ/mg phospholipids[∼20 folds higher than the non-SR-A1 ligand soyPC liposomes loaded with IMQ(LIPO-IMQ)]. In vitro, nanoARC-IMQ induced higher TNF-α and IL-6 secretion by J774A1macrophages compared to same dose of IMQ and same lipid dose of LIPO-IMQ. In vivo,3 subcutaneous doses of nanoARC-IMQ+ 10 µg total leishmania antigens (TLA) at50 µg IMQ per Balb/C mice, induced more pronounced DTH response, accompaniedby a nearly 2 orders higher antigen-specific systemic IgG titers than IMQ+TLA andLIPO-IMQ. The isotype ratio of nanoARC-IMQ+TLA remained ∼0.5 indicating, thesame as IMQ+TLA, a Th2 biased response distinguished by a pronounced increasein antibody titers, without negative effects on splenocytes lymphoproliferation, with a potential CD8+LT induction 10 days after the last dose. Overall, this first approachshowed that highly SR-A1 mediated internalization of heavily loaded nanoARC-IMQ,magnified the effect of IMQ on TLR7 expressing macrophages, leading to a more intensein vivo immune response.