In Vivo Quantitative Detection of PEGylated Macromolecules by Magnetic Resonance Spectroscopy

Large poly(ethylene) glycol (PEG) chains are often conjugated to proteins or biomolecules to inhibit proteolytic degradation, mask immunogenic response, reduce clearance rates, and improve biodistribution of therapeutics, vaccines, drug delivery systems, and gene therapy formulations. The PEG macromolecular chain can also be used as a noninvasive reporter to track biologics in vivo by magnetic resonance spectroscopy (MRS). Rapid internal dynamics of PEG render the transverse ¹H spin relaxation time to be comparable to water (~0.5 s) and amenable to imaging through traditional pulsed field gradient techniques. While water signal grossly exceeds that of PEG it is possible to filter ¹H MRS signal of PEGylated conjugates through one of two ways-(1) stimulated echo acquisition mode (STEAM) MRS, which leverages huge differences in the diffusion of water versus PEGylated constructs, and (2) ¹³C-edited ¹H MRS of fully ¹³C-enriched PEGylated constructs. Here, we compare both approaches. A 15 kDa ¹³C-enriched PEG chain was prepared alone, conjugated to bovine serum-albumin (BSA), and incorporated into a 52-nm-diameter PEG-poly(lactic acid) (PLA) nanoparticle. These three PEG constructs were then separately monitored in real time by ¹³C-edited ¹H MRS, after introducing them into rat animal models intravenously. A ¹³C-editing scheme was employed to monitor ¹H MRS PEG signal in the vasculature via a radiofrequency coil placed around the tail. An observed two-component decay of the PEG signal is attributed to perfusion and early equilibration (alpha phase) and slow clearance (beta phase). ¹³C-PEG alone, ¹³C-PEG-BSA, and ¹³C-PEG-PLA nanoparticles exhibited half-lives of 38.6 min, 23.4 h, and 11.9 h, respectively. The relatively rapid clearance rates associated with the PEG-PLA nanoparticles is expected to arise from enzymatic degradation of the PLA chain. Using STEAM-based editing schemes, we then evaluated sensitivity and water suppression in diffusion-edited ¹H MRS for (¹²C)-PEGylated BSA contrasting 2-, 20-, and 40-kDa PEG chains, in imaging phantom samples. Larger molecular weight PEG chains (i.e., 40 kDa) proved far superior to smaller PEG chain reporters due to reduced inhomogeneities and longer T₂, upon employing either a ¹³C-HQMC filter or a STEAM-based diffusion filter.