Bio-orthogonal Phosphatidylserine Conjugates for Delivery and Imaging Applications

Abstract

The syntheses of phosphatidylserine (PS) conjugates are described, including fluorescent derivatives for potential cellular delivery and bioimaging applications. Installation of terminal functional groups (amine, thiol, or alkyne) onto the sn-2 chain provides reactive sites for bio-orthogonal conjugation of cargo with suitably protected PS derivatives. An amine-containing PS forms amide bonds with peptidic cargo, a thiol derivative is designed for conjugation to cargo that contain α-halo carbonyls or Michael acceptors, and the terminal alkyne PS analog permits ‘click’ conjugation with any azide-tagged molecule. This latter conjugation method is quite versatile as it can be performed without PS headgroup protection, in aqueous media, and with acid-labile cargo.

Lipid binding domains: more than simple lipid effectors

“…Lipid sensors such as green fluorescent protein (GFP)-tagged lipid-binding domains have provided qualitative data on PI and DAG levels, and a recent DAG reporter shows promise as a more quantitative assessment of DAG levels in real-time (41). Certainly, even more quantitative lipid sensors should be possible such as those engineered to detect phosphatidylserine (42). Synthetic sensors may hold promise, as they can be engineered to harbor imaging capabilities such as those of Ca 21 indicators developed by Tsien et al (43).…”

Synthesis of Trifunctional Phosphatidylserine Probes for Identification of Lipid‐Binding Proteins

“…[43,44] Derivatization of PS lipid mimics with affinity and fluorescence tags may also be useful in target enrichment and diagnostic applications. Intermediate 13b was subjected to Cbz hydrogenolysis and directly treated with fluorescein isothiocyanate (FITC) to yield fluorescein derivative 7b (Scheme 6); this compound may be useful to follow PSR cellular localization [32] or as a reagent to screen chemical libraries for PS lipid binding elements.…”

“…While [2+3] azide-alkyne click cycloaddition has been used extensively for this purpose, [32,[45][46][47][48][49] the propargyloxybenzophenone probe module has not been previously validated as an efficient substrate. To address this issue, we carried out the desired readout and isolation reactions on soluble probe 1b (Scheme 7).…”

“…[26,31] Phosphatidylserine lipid derivatives for delivery and imaging applications have also been synthesized through enzymatic and synthetic manipulation of lysophosphatidylcholine lipid, an expensive starting material that restricts modification to the sn-2 position. [32] Glycidyl derivatives have also been used as a starting point to ether phospholipids through Lewis acid catalyzed epoxide ring opening; [12][13][14] addition to the epoxide with a long-chain carboxylic acid may yield phospholipid diesters. [33] To our surprise, regioselective stepwise acylation [34,35] of monoprotected glycerol, a robust approach that has been widely used to prepare mixed 1,2-O-diacylglycerol phospholipids, has not been previously used to prepare phosphatidylserine lipid derivatives.…”

Fluorescent labeling of alkyl chains of phosphocholine lipids by one-pot TMS cleavage-click reaction

“…The alkynefunctionalized lipid was then permeated into cells and treated with an azido-coumarin fluorogenic reactant. In another example [13] phosphatidylserine was tagged at the sn-2 acyl chain with an alkyne group, using also de novo synthesis of the glycerolipid. Scheme 1.…”