(496b) Monitoring Nanoparticle Stability and Mobility in Whole Blood and Tissues in situ

Low tumor accumulation following systemic delivery remains a key challenge for advancing many cancer nanomedicines. One obstacle in engineering nanoparticles for tumor accumulation is a lack of techniques to monitor their stability and mobility in situ, for example in whole blood or in tissue. One way to monitor the stability and mobility of nanoparticles in complex fluids is through measurements of their rotational diffusivity. We have developed a technique to monitor the rotational diffusivity of magnetic nanoparticles that is applicable to small samples, does not require optic access, and is based on the nanoparticle’s rotational response to low amplitude alternating magnetic fields, through what we call Dynamic Magnetic Susceptibility measurements.^1,2 Here we apply DMS measurements to study the stability and mobility of inorganic nanoparticles with a variety of surface coatings, in blood and in xenograft tumor explants.

Cobalt ferrite and iron oxide nanoparticles with Brownian relaxation component were synthesized by co-precipitation and thermal decomposition methods. The nanoparticles were coated with polymers such as poly(ethylene imine) (PEI), carboxymethyl dextran (CMDx), and poly(ethylene glycol) (PEG). For comparison, commercially available nanoparticles coated with hydroxyethyl starch (HES) were included in the studies. All nanoparticles were characterized according to their physicochemical, colloidal, and magnetic properties in aqueous media. Blood and xenograft tumor tissues were obtained from inoculated mice under IACUC approved protocols. The nanoparticles were dispersed in heparinized blood and their DMS spectra was acquired immediately and 24 hours after dispersion. Excised tumor xenografts were cut into pieces and nanoparticles directly injected by slow infusion. Samples were maintained immersed in phosphate buffered saline. DMS spectra were acquired immediately and 24 hours after injection.

DMS measurements show that nanoparticle stability and mobility in whole blood and tumor tissues decreases with time and is affected by the nature of the nanoparticle’s coating. Furthermore, good stability/mobility in whole blood does not translate to stability/mobility in tumor tissue. Of the various coatings, it was found that particles coated with the positively charged polymer PEI had the worst stability/mobility characteristics – their DMS signal dropped significantly immediately upon dispersion in blood or injection in tumor tissue. This was to be expected given the nanoparticle’s positive zeta potential and the prevalence of negatively-charged proteins and macromolecules in biological environments, which would lead to adsorption and aggregation of the nanoparticles. This was confirmed by transmission electron microscopy (TEM) of fixed and microtomed tumor tissue fragments containing these nanoparticles. On the opposite end of the spectrum, DMS measurements of nanoparticles coated with dense, covalently bonded brushes of PEG suggested that these particles did not significantly lose stability or mobility after dispersion in blood and injection in tumor tissue. This is consistent with the expectation that dense brushes of PEG can lead to prolonged stability in biological media.^3-6 Interestingly, prolonged stability/mobility in blood did not guarantee stability/mobility in tumor tissue. This was evident in the DMS measurements of nanoparticles coated with CMDx, where the particles appeared to retain stability/mobility in whole blood but experienced loss of mobility and increasing hydrodynamic diameter when injected in tumor tissue. Similar observations were made for the commercially obtained HES-coated nanoparticles, which quickly lost stability/mobility when injected in tumor tissue.

In conclusion, DMS measurements offer the potential of monitoring nanoparticle stability and mobility in biological milieu in situ, such as in whole blood and tumor tissue. This can provide valuable insights into the role of nanoparticle surface coatings and modification on their ability to navigate the biological environments, and can serve as a means to screen nanoparticles intended for biomedical application.

1 A. C. Bohorquez and C. Rinaldi, "In Situ Evaluation of Nanoparticle-Protein Interactions by Dynamic Magnetic Susceptibility Measurements," Particle & Particle Systems Characterization, vol. 31, pp. 561-570, 2014. http://onlinelibrary.wiley.com/doi/10.1002/ppsc.201300296/full

2 V. L. Calero-DdelC, D. I. Santiago-Quiñonez, and C. Rinaldi, "Quantitative nanoscale viscosity measurements using magnetic nanoparticles and SQUID AC susceptibility measurements," Soft Matter, vol. 7, p. 4497, 2011. http://xlink.rsc.org/?DOI=c0sm00902d

3 Q. Xu, L. M. Ensign, N. J. Boylan, A. Schon, X. Gong, J. C. Yang, N. W. Lamb, S. Cai, T. Yu, E. Freire, and J. Hanes, "Impact of Surface Polyethylene Glycol (PEG) Density on Biodegradable Nanoparticle Transport in Mucus ex Vivo and Distribution in Vivo," ACS Nano, vol. 9, pp. 9217-27, Sep 22 2015. https://www.ncbi.nlm.nih.gov/pubmed/26301576

4 D. E. Owens, 3rd and N. A. Peppas, "Opsonization, biodistribution, and pharmacokinetics of polymeric nanoparticles," Int J Pharm, vol. 307, pp. 93-102, Jan 03 2006. https://www.ncbi.nlm.nih.gov/pubmed/16303268

5 C. D. Walkey, J. B. Olsen, H. Guo, A. Emili, and W. C. Chan, "Nanoparticle size and surface chemistry determine serum protein adsorption and macrophage uptake," J Am Chem Soc, vol. 134, pp. 2139-47, Feb 01 2012. https://www.ncbi.nlm.nih.gov/pubmed/22191645

6 Q. Dai, C. Walkey, and W. C. Chan, "Polyethylene glycol backfilling mitigates the negative impact of the protein corona on nanoparticle cell targeting," Angew Chem Int Ed Engl, vol. 53, pp. 5093-6, May 12 2014. https://www.ncbi.nlm.nih.gov/pubmed/24700480