Immunocolloidal targeting of the endocytotic siglec-7 receptor using peripheral attachment of siglec-7 antibodies to poly(lactide-co-glycolide) nanoparticles.
OBJECTIVE
To prepare a nanoparticulate formulation expressing variable peripheral carboxyl density using non-endcapped and endcapped poly(lactide-co-glycolide), conjugated to antibodies recognising the siglec-7 receptor, which is expressed on most acute myeloid leukaemias. The aim is to exploit this receptor as a therapeutic target by constructing an internalising drug-loaded nanoparticle able to translocate into cytoplasm by siglec receptor-mediated internalisation.


METHODS
Antibodies to the siglec-7 (CD33-like) receptor were conjugated to dye-loaded nanoparticles using carbodiimide chemistry, giving 32.6 microg protein per mg of nanoparticles using 100% of the non-endcapped PLGA. Binding studies using cognate antigen were used to verify preservation of antibody function following conjugation.


RESULTS
Mouse embryonic fibroblasts expressing recombinant siglec-7 receptor and exposed to Nile-Red-loaded nanoparticles conjugated to antibody accumulated intracellular fluorescence, which was not observed if either antibody or siglec-7 receptor was absent. Confocal microscopy revealed internalised perinuclear cytoplasmic staining, with an Acridine Orange-based analysis showing red staining in localised foci, indicating localisation within acidic endocytic compartments.


CONCLUSIONS
Results show antibody-NP constructs are internalised via siglec-7 receptor-mediated internalisation. If loaded with a therapeutic agent, antibody-NP constructs can cross into cytoplasmic space and delivery drugs intracellularly to cells expressing CD33-like receptors, such as natural killer cells and monocytes.
School of Pharmacy, Queens University Belfast, Medical Biology Centre, Belfast, UK.
Christopher J
CJ
Scott
Waleed M
WM
Marouf
Derek J
DJ
Quinn
Richard J
RJ
Buick
Selinda J
SJ
Orr
Ryan F
RF
Donnelly
Paul A
PA
McCarron
D000165:Acridine Orange; D000818:Animals; D000906:Antibodies / Q000737:chemistry*; D002478:Cells, Cultured; D003102:Colloids; D004705:Endocytosis / Q000187:drug effects*; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005347:Fibroblasts; D005456:Fluorescent Dyes; D019344:Lactic Acid; D037102:Lectins / Q000276:immunology*; D051379:Mice; D053758:Nanoparticles; D010078:Oxazines; D010316:Particle Size; D011100:Polyglycolic Acid; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011108:Polymers; D063265:Sialic Acid Binding Immunoglobulin-like Lectins
D000906:Antibodies; D003102:Colloids; D005456:Fluorescent Dyes; D037102:Lectins; D010078:Oxazines; D011108:Polymers; D063265:Sialic Acid Binding Immunoglobulin-like Lectins; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid; D000165:Acridine Orange; C044808:nile red