Modulating Pharmacokinetics, Tumor Uptake and Biodistribution by Engineered Nanoparticles

Inorganic nanoparticles provide promising tools for biomedical applications including detection, diagnosis and therapy. While surface properties such as charge are expected to play an important role in their in vivo behavior, very little is known how the surface chemistry of nanoparticles influences their pharmacokinetics, tumor uptake, and biodistribution.Using a family of structurally homologous nanoparticles we have investigated how pharmacological properties including tumor uptake and biodistribution are influenced by surface charge using neutral (TEGOH), zwitterionic (Tzwit), negative (TCOOH) and positive (TTMA) nanoparticles. Nanoparticles were injected into mice (normal and athymic) either in the tail vein or into the peritoneum.Neutral and zwitterionic nanoparticles demonstrated longer circulation time via both i.p. and i.v. administration, whereas negatively and positively charged nanoparticles possessed relatively short half-lives. These pharmacological characteristics were reflected on the tumor uptake and biodistribution of the respective nanoparticles, with enhanced tumor uptake by neutral and zwitterionic nanoparticles via passive targeting

Identifying New Therapeutic Targets via Modulation of Protein Corona Formation by Engineered Nanoparticles

We introduce a promising methodology to identify new therapeutic targets in cancer. Proteins bind to nanoparticles to form a protein corona. We modulate this corona by using surface-engineered nanoparticles, and identify protein composition to provide insight into disease development.Using a family of structurally homologous nanoparticles we have investigated the changes in the protein corona around surface-functionalized gold nanoparticles (AuNPs) from normal and malignant ovarian cell lysates. Proteomics analysis using mass spectrometry identified hepatoma-derived growth factor (HDGF) that is found exclusively on positively charged AuNPs ((+)AuNPs) after incubation with the lysates. We confirmed expression of HDGF in various ovarian cancer cells and validated binding selectivity to (+)AuNPs by Western blot analysis. Silencing of HDGF by siRNA resulted s inhibition in proliferation of ovarian cancer cells.We investigated the modulation of protein corona around surface-functionalized gold nanoparticles as a promising approach to identify new therapeutic targets. The potential of our method for identifying therapeutic targets was demonstrated through silencing of HDGF by siRNA, which inhibited proliferation of ovarian cancer cells. This integrated proteomics, bioinformatics, and nanotechnology strategy demonstrates that protein corona identification can be used to discover novel therapeutic targets in cancer

Applying surface modified gold nanoparticles to biological systems

Gold nanoparticles hold promise as both a stable drug delivery system as well as a targeted inhibitor of protein:protein interactions. The flexibility and ease of modifying the surface of nanoparticles enables them to be tailored to specific cellular environments and tasks. Due to these characteristics, I have been able to effectively modify gold. To further investigate gold nanoparticles as drug delivery systems, it is important to understand where they localize once inside the cell. One possible way of determining the cellular fate of the nanoparticles is conduction cell fractionization studies coupled with ICP-MS. A protocol using sucrose gradients along with high-speed ultracentrifugation allows for the various cellular organelles to be separated and digested. Once each fraction is digested, I can monitor where the gold nanoparticles localize inside the cell

Enrichment and functional consequence of HDGF.

<p>a) Western blot confirming the presence of HDGF on ⁺AuNP- corona whereas Hsp90 on ⁻AuNP- corona. Also shown is Hsp70 (known to be present in all NP coronas via MS). b) Expression of HDGF in ovarian cell lines. c) Knockdown of HDGF in A2780 cell line using HDGF-siRNAs (KD-siRNA) and compared with the scrambled control (scRNA); d) Effect of silencing HDGF on proliferation of ovarian cancer cells analyzed by [3H]-thymidine incorporation assay.</p

Characterization of the AuNP and protein corona made from cell lysates of OSE and OV167.

<p>a) A cartoon showing functionalization of 5 nm gold nanoparticle to create positively charged (⁺AuNP) or negatively charged (⁻AuNP) gold nanoparticles. b) Amount of protein bound on the nanoparticle as determined by Bradford assay. The binding of protein is evident from the increase in surface size via DLS on c) ⁺AuNP and d) ⁻AuNP.</p

Venn diagram demonstrating the difference between the proteins that make up the corona for a) ⁺AuNP, b) ⁻AuNP from the cell lysates and c) the different between proteins in the OV167 and OSE cell lysates.

<p>These differences in the composition of the lysates could be exploited to identify new therapeutic targets in ovarian cancer.</p

Selectivity of the proteins bound to positively charged (⁺AuNP) vs negatively charged (⁻AuNP) particles.

<p>Venn diagrams show proteins identified in the protein corona around ⁺AuNP and ⁻AuNP from a) normal OSE cell lysates and b) malignant OV167 cell lysates. The figure clearly depicts the preferential enrichment of low abundance proteins by engineered nanoparticles that were not detectable in the lysates by proteomics analysis. These proteins, which were otherwise undetected, could potentially be new therapeutic targets.</p

Dynamic Light Scattering (DLS) to determine hydrodynamic diameter (d_H) of AuNPs before and after incubation with OV167 and OSE lysates.

<p>Dynamic Light Scattering (DLS) to determine hydrodynamic diameter (d_H) of AuNPs before and after incubation with OV167 and OSE lysates.</p

Zeta Potential in mV of ⁺AuNP and ⁻AuNP before and after incubation with the lysates of OV167 and OSE, respectively.

<p>Zeta Potential in mV of ⁺AuNP and ⁻AuNP before and after incubation with the lysates of OV167 and OSE, respectively.</p