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Molecularly Engineered Nanobodies for Tunable Pharmacokinetics and Drug Delivery
journal contribution
posted on 2020-03-19, 19:04 authored by Patrick
M. Glassman, Landis R. Walsh, Carlos H. Villa, Oscar A. Marcos-Contreras, Elizabeth D. Hood, Vladimir R. Muzykantov, Colin F. GreinederThe use of single-domain
antibody fragments, or nanobodies, has
gained popularity in recent years as an alternative to traditional
monoclonal antibody-based approaches. Relatively little is known,
however, about the utility of nanobodies as targeting agents for delivery
of therapeutic cargoes, particularly to vascular epitopes or in the
setting of acute inflammatory conditions. We used a nanobody (VCAMelid)
directed against mouse vascular cell adhesion molecule 1 (VCAM-1)
and techniques for site-specific radiolabeling and bioconjugation
to measure targeting to sites of constitutive and inducible antigen
expression and investigate the impact of various characteristics (affinity,
valence, circulation time) on nanobody biodistribution and pharmacokinetics.
Engineering of VCAMelid for bivalent binding (BiVCAMelid) increased
affinity by an order of magnitude and provided 2.8- and 3.6-fold enhancements
in splenic and brain targeting in naive mice, with a further 2.6-fold
increase in brain uptake in the setting of focal CNS inflammation.
In contrast, introduction of an albumin-binding arm (VCAM/ALB8) did
not affect binding affinity, but its prolonged circulation time resulted
in 3.5-fold and 17.4-fold increases in splenic and brain uptake at
20 min post-dose and remarkable 40-, 25-, and 15-fold enhancements
in overall exposure of blood, spleen, and brain, respectively, relative
to both VCAMelid and BiVCAMelid. Both therapeutic protein (superoxide
dismutase, SOD-1) and nanocarrier (liposome) delivery were enhanced
by conjugation to VCAM-1 targeted nanobodies. The bispecific VCAM/ALB8
maintained its superiority over VCAMelid in enhancing both circulation
time and organ targeting of SOD-1, but its advantages were largely
blunted by conjugation to liposomes.