Mirkka Sarparanta

Biocompatibility of Thermally Hydrocarbonized Porous Silicon Nanoparticles and their Biodistribution in Rats

Porous silicon (PSi) particles have been studied for the effects they elicit in Caco-2 and RAW 264.7 macrophage cells in terms of toxicity, oxidative stress, and inflammatory response. The most suitable particles were then functionalized with a novel (18)F label to assess their biodistribution after enteral and parenteral administration in a rat model. The results show that thermally hydrocarbonized porous silicon (THCPSi) nanoparticles did not induce any significant toxicity, oxidative stress, or inflammatory response in Caco-2 and RAW 264.7 macrophage cells. Fluorescently labeled nanoparticles were associated with the cells surface but were not extensively internalized. Biodistribution studies in rats using novel (18)F-labeled THCPSi nanoparticles demonstrated that the particles passed intact through the gastrointestinal tract after oral administration and were also not absorbed from a subcutaneous deposit. After intravenous administration, the particles were found mainly in the liver and spleen, indicating rapid removal from the circulation. Overall, these silicon-based nanosystems exhibit excellent in vivo stability, low cytotoxicity, and nonimmunogenic profiles, ideal for oral drug delivery purposes.

Intravenous Delivery of Hydrophobin-Functionalized Porous Silicon Nanoparticles: Stability, Plasma Protein Adsorption and Biodistribution

Rapid immune recognition and subsequent elimination from the circulation hampers the use of many nanomaterials as carriers to targeted drug delivery and controlled release in the intravenous route. Here, we report the effect of a functional self-assembled protein coating on the intravenous biodistribution of (18)F-labeled thermally hydrocarbonized porous silicon (THCPSi) nanoparticles in rats. (18)F-Radiolabeling enables the sensitive and easy quantification of nanoparticles in tissues using radiometric methods and allows imaging of the nanoparticle biodistribution with positron emission tomography. Coating with Trichoderma reesei HFBII altered the hydrophobicity of (18)F-THCPSi nanoparticles and resulted in a pronounced change in the degree of plasma protein adsorption to the nanoparticle surface in vitro. The HFBII-THCPSi nanoparticles were biocompatible in RAW 264.7 macrophages and HepG2 liver cells making their intravenous administration feasible. In vivo, the distribution of the nanoparticles between the liver and spleen, the major mononuclear phagocyte system organs in the body, was altered compared to that of uncoated (18)F-THCPSi. Identification of the adsorbed proteins revealed that certain opsonins and apolipoproteins are enriched in HFBII-functionalized nanoparticles, whereas the adsorption of abundant plasma components such as serum albumin and fibrinogen is decreased.

The mucoadhesive and gastroretentive properties of hydrophobin-coated porous silicon nanoparticle oral drug delivery systems

Impediments to intestinal absorption, such as poor solubility and instability in the variable conditions of the gastrointestinal (GI) tract plague many of the current drugs restricting their oral bioavailability. Particulate drug delivery systems hold great promise in solving these problems, but their effectiveness might be limited by their often rapid transit through the GI tract. Here we describe a bioadhesive oral drug delivery system based on thermally-hydrocarbonized porous silicon (THCPSi) functionalized with a self-assembled amphiphilic protein coating consisting of a class II hydrophobin (HFBII) from Trichoderma reesei. The HFBII-THCPSi nanoparticles were found to be non-cytotoxic and mucoadhesive in AGS cells, prompting their use in a biodistribution study in rats after oral administration. The passage of HFBII-THCPSi nanoparticles in the rat GI tract was significantly slower than that of uncoated THCPSi, and the nanoparticles were retained in stomach by gastric mucoadhesion up to 3 h after administration. Upon entry to the small intestine, the mucoadhesive properties were lost, resulting in the rapid transit of the nanoparticles through the remainder of the GI tract. The gastroretentive drug delivery system with a dual function presented here is a viable alternative for improving drug bioavailability in the oral route.

Multifunctional porous silicon nanoparticles for cancer theranostics

Nanomaterials provide a unique platform for the development of theranostic systems that combine diagnostic imaging modalities with a therapeutic payload in a single probe. In this work, dual-labeled iRGD-modified multifunctional porous silicon nanoparticles (PSi NPs) were prepared from dibenzocyclooctyl (DBCO) modified PSi NPs by strain-promoted azide-alkyne cycloaddition (SPAAC) click chemistry. Hydrophobic antiangiogenic drug, sorafenib, was loaded into the modified PSi NPs to enhance the drug dissolution rate and improve cancer therapy. Radiolabeling of the developed system with (111)In enabled the monitoring of the in vivo biodistribution of the nanocarrier by single photon emission computed tomography (SPECT) in an ectopic PC3-MM2 mouse xenograft model. Fluorescent labeling with Alexa Fluor 488 was used to determine the long-term biodistribution of the nanocarrier by immunofluorescence at the tissue level ex vivo. Modification of the PSi NPs with an iRGD peptide enhanced the tumor uptake of the NPs when administered intravenously. After intratumoral delivery the NPs were retained in the tumor, resulting in efficient tumor growth suppression with particle-loaded sorafenib compared to the free drug. The presented multifunctional PSi NPs highlight the utility of constructing a theranostic nanosystems for simultaneous investigations of the in vivo behavior of the nanocarriers and their drug delivery efficiency, facilitating the selection of the most promising materials for further NP development.

¹⁸F-labeled modified porous silicon particles for investigation of drug delivery carrier distribution in vivo with positron emission tomography.

Because of its biocompatibility and ability to accommodate a variety of payloads from poorly soluble drugs to biomolecules, porous silicon (PSi) is a lucrative material for the development of carriers for particle-mediated drug delivery. We report a successful direct one-step (18)F-radiolabeling of three types of PSi microparticles, thermally hydrocarbonized THCPSi, thermally oxidized TOPSi, and thermally carbonized TCPSi for the investigation of their biodistribution in vivo with positron emission tomography as part of their evaluation as carriers for particle-mediated drug delivery. FTIR and XPS characterization of the PSi materials after carrier-added (18)F/(19)F-radiolabeling reveals that depending on the material the (18)F-labeling is likely to be accomplished either by substitution for surface silyl hydrogen or silyl fluoride or by nucleophilic attack of (18)F(-) to Si-O-Si bridges. With the selected (18)F-radiolabeling method, good to excellent in vitro radiolabel stability in simulated gastric and intestinal fluids and in plasma is achieved for all the particle types studied. Finally, a preliminary evaluation of (18)F-THCPSi microparticle biodistribution in the rat gastrointestinal tract after oral administration is reported, illustrating the utility of using (18)F-radiolabeled PSi as imaging probes for PSi-based drug delivery carrier distribution in vivo.

Targeted Radiotherapy for Prostate Cancer with an Oncolytic Adenovirus Coding for Human Sodium Iodide Symporter

Purpose: Oncolytic adenoviruses are promising tools for cancer therapy. Although several clinical reports have indicated both safety and promising antitumor capabilities for these viruses, there are only a few examples of complete tumor eradication. Thus, the antitumor efficacy of oncolytic adenoviruses needs to be improved. One potentially useful approach is combination with radiotherapy. Experimental Design: To target systemically administered radioiodide to tumors, we created Ad5/3-Δ24-human sodium iodide symporter (hNIS), a Rb-p16 pathway selective infectivity enhanced oncolytic adenovirus encoding hNIS. Results: Ad5/3-Δ24-hNIS replication effectively killed prostate cancer cells in vitro and in vivo. Also, the virus-mediated radioiodide uptake into prostate cancer cells in vitro and into tumors in vivo. Furthermore, Ad5/3-Δ24-hNIS with radioiodide was significantly more effective than virus alone in mice with prostate cancer xenografts. Conclusions: These results suggest that oncolytic adenovirus-mediated targeted radiotherapy might be a potentially useful option for enhancing the efficacy or adenoviral virotherapy. (Clin Cancer Res 2009;15(17):5396–403)

A New Highly Reactive and Low Lipophilicity Fluorine-18 Labeled Tetrazine Derivative for Pretargeted PET Imaging.

A new (18)F-labeled tetrazine derivative was developed aiming at optimal radiochemistry, fast reaction kinetics in inverse electron-demand Diels-Alder cycloaddition (IEDDA), and favorable pharmacokinetics for in vivo bioorthogonal chemistry. The radiolabeling of the tetrazine was achieved in high yield, purity, and specific activity under mild reaction conditions via conjugation with 5-[(18)F]fluoro-5-deoxyribose, providing a glycosylated tetrazine derivative with low lipophilicity. The (18)F-tetrazine showed fast reaction kinetics toward the most commonly used dienophiles in IEDDA reactions. It exhibited excellent chemical and enzymatic stability in mouse plasma and in phosphate-buffered saline (pH 7.41). Biodistribution in mice revealed favorable pharmacokinetics with major elimination via urinary excretion. The results indicate that the glycosylated (18)F-labeled tetrazine is an excellent candidate for in vivo bioorthogonal chemistry applications in pretargeted PET imaging approaches.

Synthesis, in vitro and in vivo evaluation of 1,3,5-triazines as cannabinoid CB2 receptor agonists.

The cannabinoid receptors type 2 (CBR2) are attractive therapeutic targets of the endocannabinoid signaling system (ECS) as they are not displaying the undesired psychotropic and cardiovascular side-effects seen with cannabinoid receptor type 1 (CB1R) agonists. In continuation of our previous work on 2,4,6-trisubstituted 1,3,5-triazines as potent CB2 agonists, we synthesized an additional series of more polar analogues (1-10), which were found to possess high CB2R agonist activity with enhanced water solubility. The most potent compound in the series was N-(adamantan-1-yl)-4-ethoxy-6-(4-(2-fluoroethyl)piperazin-1-yl)-1,3,5-triazin-2-amine (9) with EC50 value of 0.60nM. To further evaluate the biological effects of the compounds, the selected compounds were tested in vitro against four different cell lines. A human retinal pigment epithelial cell line (ARPE-19) was used to evaluate the cytotoxicity of the compounds whereas an androgen-sensitive human prostate adenocarcinoma cell line (LNCaP), a Jurkat leukemia cell line and a C8161 melanoma cell line were used to assess the antiproliferative activity of the compounds. The most interesting results were obtained for N-(adamantan-1-yl)-4-ethoxy-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine (6), which induced cell viability decrease in prostate and leukemia cell lines, and diminished proliferation of C8161 melanoma cells. The results could be reversed in leukemia cells with the selective CB2R antagonist AM630, whereas in prostate cells the AM630 induced a significant cell viability decrease with a mechanism probably unlinked to CB2 cannabinoid receptor. The antiproliferative effect of 6 on the melanoma cells seemed not to be mediated via the CB1R or CB2R. No cytotoxicity was detected against ARPE-19 cell line at concentrations of 1 and 10μM for compound 6. However, at 30μM concentration the compound 6 decreased the cell viability. Finally, in order to estimate in vivo behavior of these compounds, (18)F labeled PET ligand, N-cyclopentyl-4-ethoxy-6-(4-(2-fluoro-18-ethyl)piperazin-1-yl)-1,3,5-triazin-2-amine ([(18)F]5), was synthesized and its biodistribution was determined in healthy male Sprague-Dawley rats. As a result, the tracer showed a rapid (<15min) elimination in urine accompanied by a slower excretion via the hepatobiliary route. In conclusion, we further demonstrated that 1,3,5-triazine scaffold serves as a suitable template for the design of highly potent CB2R agonists with reasonable water solubility properties. The compounds may be useful when studying the role of the endocannabinoid system in different diseases. The triazine scaffold is also a promising candidate for the development of new CB2R PET ligands.

In vivo evaluation of porous silicon and porous silicon solid lipid nanocomposites for passive targeting and imaging.

The use of nanoparticle carriers for the sustained release of cytotoxic drugs in cancer therapy can result in fewer adverse effects and can thus be of great benefit for the patient. Recently, a novel nanocomposite, prepared by the encapsulation of THCPSi nanoparticles within solid lipids (SLN), was developed and characterized as a promising drug delivery carrier in vitro. The present study describes the in vivo evaluation of unmodified THCPSi nanoparticles and THCPSi-solid lipid nanocomposites (THCPSi-SLNCs) as potential drug delivery carriers for cancer therapy by using (18)F radiolabeling for the detection of the particle biodistribution in mice. Passive tumor targeting of (18)F-THCPSis and (18)F-THCPSi-SLNCs by the enhanced permeation and retention effect was investigated in a murine breast cancer model. Encapsulation of THCPSi nanoparticles with solid lipids improved their accumulation in tumors at a 7 week time point (tumor-to-liver ratio 0.10 ± 0.08 and 0.24 ± 0.09% for (18)F-THCPSis and (18)F-THCPSi-SLNCs, respectively).

SPECT/CT Imaging of hNIS -Expression after Intravenous Delivery of an Oncolytic Adenovirus and 131I

Oncolytic adenoviruses can be engineered for better tumor selectivity, gene delivery and be armed for imaging and concentrating radionuclides into tumors for synergistic oncolysis. We constructed Ad5/3-hTERT-hNIS where replication is controlled by hTERT-promoter. Ad5/3-hTERT-hNIS expresses hNIS for imaging of transgene expression and for treatment of infected tumors by radioiodine. Ad5/3-hTERT-hNIS efficiently killed prostate cancer cells and induced iodine uptake in vitro and in vivo after intratumoral virus administration. Survival of mice treated with intravenous Ad5/3-hTERT-hNIS significantly prolonged survival over mock or radioiodine only but the combination of virus with radioiodine was not more effective than virus alone. Temporal and spatial changes in hNIS-expression during therapy were detected with SPECT, demonstrating feasibility of evaluation of the combination therapy with hNIS-expressing adenoviruses and radioiodide.