Image-Guided Deployment and Monitoring of a Novel Tungsten Nanoparticle-Infused Radiopaque Absorbable Inferior Vena Cava Filter in a Swine Model.

PURPOSE: To improve radiopacity of radiolucent absorbable poly-p-dioxanone (PPDO) inferior vena cava filters (IVCFs) and demostrate their effectiveness in clot-trapping ability. MATERIALS AND METHODS: Tungsten nanoparticles (WNPs) were incorporated along with polyhydroxybutyrate (PHB), polycaprolactone (PCL), and polyvinylpyrrolidone (PVP) polymers to increase the surface adsorption of WNPs. The physicochemical and in vitro and in vivo imaging properties of PPDO IVCFs with WNPs with single-polymer PHB (W-P) were compared with those of WNPs with polymer blends consisting of PHB, PCL, and PVP (W-PB). RESULTS: In vitro analyses using PPDO sutures showed enhanced radiopacity with either W-P or W-PB coating, without compromising the inherent physicomechanical properties of the PPDO sutures. W-P- and W-PB-coated IVCFs were deployed successfully into the inferior vena cava of pig models with monitoring by fluoroscopy. At the time of deployment, W-PB-coated IVCFs showed a 2-fold increase in radiopacity compared to W-P-coated IVCFs. Longitudinal monitoring of in vivo IVCFs over a 12-week period showed a drastic decrease in radiopacity at Week 3 for both filters. CONCLUSIONS: The results highlight the utility of nanoparticles (NPs) and polymers for enhancing radiopacity of medical devices. Different methods of incorporating NPs and polymers can still be explored to improve the effectiveness, safety, and quality of absorbable IVCFs.

Multiple breast cancer liver metastases response to macrophage-delivered nanotherapy evaluated via a 3D continuum model.

Breast cancer liver metastases (BCLM) are usually unresectable and difficult to treat with systemic chemotherapy. A major reason for chemotherapy failure is that BCLM are typically small, avascular nodules, with poor transport and fast washout of therapeutics from surrounding capillaries. We have previously shown that nanoalbumin-bound paclitaxel (nab-PTX) encapsulated in porous silicon multistage nanovectors (MSV) is preferentially taken up by tumour-associated macrophages (TAM) in the BCLM microenvironment. The TAM alter therapeutic transport characteristics and retain it in the tumour vicinity, increasing cytotoxicity. Computational modeling has shown that therapeutic regimens could be designed to eliminate single lesions. To evaluate clinically-relevant scenarios, this study develops a modeling framework to evaluate MSV-nab-PTX therapy targeting multiple BCLM. An experimental model of BCLM, splenic injection of breast cancer 4 T1 cells was established in BALB/C mice. Livers were analyzed histologically to determine size and density of BCLM. The data were used to calibrate a 3D continuum mixture model solved via distributed computing to enable simulation of multiple BCLM. Overall tumour burden was analyzed as a function of metastases number and potential therapeutic regimens. The computational model enables realistic 3D representation of metastatic tumour burden in the liver, with the capability to evaluate BCLM growth and therapy response for hundreds of lesions. With the given parameter set, the model projects that repeated MSV-nab-PTX treatment in intervals <7 days would control the tumour burden. We conclude that nanotherapy targeting TAM associated with BCLM may be evaluated and fine-tuned via 3D computational modeling that realistically simulates multiple metastases.

The Proteolytic Landscape of Ovarian Cancer: Applications in Nanomedicine.

Ovarian cancer (OvCa) is one of the leading causes of mortality globally with an overall 5-year survival of 47%. The predominant subtype of OvCa is epithelial carcinoma, which can be highly aggressive. This review launches with a summary of the clinical features of OvCa, including staging and current techniques for diagnosis and therapy. Further, the important role of proteases in OvCa progression and dissemination is described. Proteases contribute to tumor angiogenesis, remodeling of extracellular matrix, migration and invasion, major processes in OvCa pathology. Multiple proteases, such as metalloproteinases, trypsin, cathepsin and others, are overexpressed in the tumor tissue. Presence of these catabolic enzymes in OvCa tissue can be exploited for improving early diagnosis and therapeutic options in advanced cases. Nanomedicine, being on the interface of molecular and cellular scales, can be designed to be activated by proteases in the OvCa microenvironment. Various types of protease-enabled nanomedicines are described and the studies that focus on their diagnostic, therapeutic and theranostic potential are reviewed.

Nonlinear response to cancer nanotherapy due to macrophage interactions revealed by mathematical modeling and evaluated in a murine model via CRISPR-modulated macrophage polarization.

Tumor-associated macrophages (TAMs) have been shown to both aid and hinder tumor growth, with patient outcomes potentially hinging on the proportion of M1, pro-inflammatory/growth-inhibiting, to M2, growth-supporting, phenotypes. Strategies to stimulate tumor regression by promoting polarization to M1 are a novel approach that harnesses the immune system to enhance therapeutic outcomes, including chemotherapy. We recently found that nanotherapy with mesoporous particles loaded with albumin-bound paclitaxel (MSV-nab-PTX) promotes macrophage polarization towards M1 in breast cancer liver metastases (BCLM). However, it remains unclear to what extent tumor regression can be maximized based on modulation of the macrophage phenotype, especially for poorly perfused tumors such as BCLM. Here, for the first time, a CRISPR system is employed to permanently modulate macrophage polarization in a controlled in vitro setting. This enables the design of 3D co-culture experiments mimicking the BCLM hypovascularized environment with various ratios of polarized macrophages. We implement a mathematical framework to evaluate nanoparticle-mediated chemotherapy in conjunction with TAM polarization. The response is predicted to be not linearly dependent on the M1:M2 ratio. To investigate this phenomenon, the response is simulated via the model for a variety of M1:M2 ratios. The modeling indicates that polarization to an all-M1 population may be less effective than a combination of both M1 and M2. Experimental results with the CRISPR system confirm this model-driven hypothesis. Altogether, this study indicates that response to nanoparticle-mediated chemotherapy targeting poorly perfused tumors may benefit from a fine-tuned M1:M2 ratio that maintains both phenotypes in the tumor microenvironment during treatment.

Liposomal encapsulation masks genotoxicity of a chemotherapeutic agent in regulatory toxicology assessments.

The burgeoning application of nanotechnology to a variety of industries including cosmetics, food, medicine and materials has led to the exploration of nanotoxicology as a trending subject of research. However the role of a nanovector, in affecting the mutagenicity of its therapeutic payload has not yet been investigated. In this study, we compare the mutagenicity of the free drug - doxorubicin hydrochloride with its nanoencapsulated form - doxorubicin loaded liposome, using conventional methods required for regulatory approval. Contrary to free doxorubicin, doxorubicin encapsulated liposome expressed a significantly lower mutant frequency in the Ames assay, and was non-genotoxic in the in vitro micronucleus assay. Further investigation of the systems' cytotoxicity and their interaction with the bacterial cell envelope, suggests that the modification of the test parameters and release of the encapsulated drug prior to the Ames test show comparable mutagenic potential of the nanotherapeutic system to a free drug.

Magnetically Bioprinted Human Myometrial 3D Cell Rings as A Model for Uterine Contractility.

Deregulation in uterine contractility can cause common pathological disorders of the female reproductive system, including preterm labor, infertility, inappropriate implantation, and irregular menstrual cycle. A better understanding of human myometrium contractility is essential to designing and testing interventions for these important clinical problems. Robust studies on the physiology of human uterine contractions require in vitro models, utilizing a human source. Importantly, uterine contractility is a three-dimensionally (3D)-coordinated phenomenon and should be studied in a 3D environment. Here, we propose and assess for the first time a 3D in vitro model for the evaluation of human uterine contractility. Magnetic 3D bioprinting is applied to pattern human myometrium cells into rings, which are then monitored for contractility over time and as a function of various clinically relevant agents. Commercially available and patient-derived myometrium cells were magnetically bioprinted into rings in 384-well formats for throughput uterine contractility analysis. The bioprinted uterine rings from various cell origins and patients show different patterns of contractility and respond differently to clinically relevant uterine contractility inhibitors, indomethacin and nifedipine. We believe that the novel system will serve as a useful tool to evaluate the physiology of human parturition while enabling high-throughput testing of multiple agents and conditions.

Multiscale benchmarking of drug delivery vectors.

Cross-system comparisons of drug delivery vectors are essential to ensure optimal design. An in-vitro experimental protocol is presented that separates the role of the delivery vector from that of its cargo in determining the cell response, thus allowing quantitative comparison of different systems. The technique is validated through benchmarking of the dose-response of human fibroblast cells exposed to the cationic molecule, polyethylene imine (PEI); delivered as a free molecule and as a cargo on the surface of CdSe nanoparticles and Silica microparticles. The exposure metrics are converted to a delivered dose with the transport properties of the different scale systems characterized by a delivery time, τ. The benchmarking highlights an agglomeration of the free PEI molecules into micron sized clusters and identifies the metric determining cell death as the total number of PEI molecules presented to cells, determined by the delivery vector dose and the surface density of the cargo.

Liposomes: a nanoscale drug carrying system to prevent indomethacin passage to the fetus in a pregnant mouse model.

OBJECTIVE: Indomethacin (IND) is a prostaglandin production inhibitor that reduces uterine contractions, but crosses the placenta leading to adverse fetal effects. Liposomes (LIP) are nanoscale systems clinically used to preferentially deliver a drug to the tissue of interest and simultaneously prevent distribution to unwanted locations. Our objective was to determine whether LIP could prevent the transfer of IND across the placenta to the fetus while preserving its pharmacological activity. STUDY DESIGN: Multilamellar LIP were designed with a 150- to 200-nm size, fluorescently labeled, and loaded with IND. Timed pregnant CD1 mice (n = 6/group) on gestational day 18 were administered LIP, LIP-IND (1 mg IND/kg), or saline (SAL) via tail vein injection, or IND (1 mg/kg) via oral gavage. After 4 hours, the uterus, placenta, and fetuses were retrieved. LIP levels were visualized using fluorescent microscopy and quantitatively assessed by National Institutes of Health image processing software. LIP brightness values (mean ± SEM) in arbitrary units (AU) were normalized to the autofluorescence of the same tissue (as measured in SAL group). IND and prostaglandin E2 levels were assessed using liquid chromatography-tandem mass spectrometry and enzyme-linked immunosorbent assay, respectively. RESULTS: The qualitative analysis of LIP distribution revealed that the system was primarily confined within the uterus, minimally detected within the placenta, and absent in the fetus. LIP fluorescence was greater in the uterus compared to placenta and fetus (uterus 15.3 ± 5.4 AU vs placenta 3.0 ± 3.5 AU vs fetus 4.4 ± 2.5 AU; P = .009). LIP-IND resulted in a 7.6-fold reduction in the IND levels in the fetus compared to IND alone (LIP-IND 10.7 ± 17.1 ng/g vs IND 81.3 ± 24.7 ng/g; P = .041). Prostaglandin E2 levels were significantly reduced in the uterus of animals given LIP-IND and IND compared to LIP and SAL. CONCLUSION: LIP localized within the uterus and did not cross the placenta to the fetus. IND within the fetus was reduced 7.6-fold while encapsulated within the LIP and the pharmacologic effects of IND were maintained. Thus, LIP provide a novel therapeutic approach to correct the primary clinical limitation of IND by reducing placental passage to the fetus.

Macrophage Checkpoint Nanoimmunotherapy Has the Potential to Reduce Malignant Progression in Bioengineered In Vitro Models of Ovarian Cancer.

Most ovarian carcinoma (OvCa) patients present with advanced disease at the time of diagnosis. Malignant, metastatic OvCa is invasive and has poor prognosis, exposing the need for improved therapeutic targeting. High CD47 (OvCa) and SIRPα (macrophage) expression has been linked to decreased survival, making this interaction a significant target for therapeutic discovery. Even so, previous attempts have fallen short, limited by CD47 antibody specificity and efficacy. Macrophages are an important component of the OvCa tumor microenvironment and are manipulated to aid in cancer progression via CD47-SIRPα signaling. Thus, we have leveraged lipid-based nanoparticles (LNPs) to design a therapy uniquely situated to home to phagocytic macrophages expressing the SIRPα protein in metastatic OvCa. CD47-SIRPα presence was evaluated in patient histological sections using immunohistochemistry. 3D tumor spheroids generated on a hanging drop array with OVCAR3 high-grade serous OvCa and THP-1-derived macrophages created a representative model of cellular interactions involved in metastatic OvCa. Microfluidic techniques were employed to generate LNPs encapsulating SIRPα siRNA (siSIRPα) to affect the CD47-SIRPα signaling between the OvCa and macrophages. siSIRPα LNPs were characterized for optimal size, charge, and encapsulation efficiency. Uptake of the siSIRPα LNPs by macrophages was assessed by Incucyte. Following 48 h of 25 nM siSIRPα treatment, OvCa/macrophage heterospheroids were evaluated for SIRPα knockdown, platinum chemoresistance, and invasiveness. OvCa patient tumors and in vitro heterospheroids expressed CD47 and SIRPα. Macrophages in OvCa spheroids increased carboplatin resistance and invasion, indicating a more malignant phenotype. We observed successful LNP uptake by macrophages causing significant reduction in SIRPα gene and protein expressions and subsequent reversal of pro-tumoral alternative activation. Disrupting CD47-SIRPα interactions resulted in sensitizing OvCa/macrophage heterospheroids to platinum chemotherapy and reversal of cellular invasion outside of heterospheroids. Ultimately, our results strongly indicate the potential of using LNP-based nanoimmunotherapy to reduce malignant progression of ovarian cancer.

Telomerase mRNA Enhances Human Skin Engraftment for Wound Healing.

Deep skin wounds represent a serious condition and frequently require split-thickness skin grafts (STSG) to heal. The application of autologous human-skin-cell-suspension (hSCS) requires less donor skin than STSG without compromising the healing capacity. Impaired function and replicative ability of senescent cutaneous cells in the aging skin affects healing with autologous hSCS. Major determinants of senescence are telomere erosion and DNA damage. Human telomerase reverse transcriptase (hTERT) adds telomeric repeats to the DNA and can protect against DNA damage. Herein, hTERT mRNA lipid nanoparticles (LNP) are proposed and evaluated for enhancing cellular engraftment and proliferation of hSCS. Transfection with optimized hTERT mRNA LNP system enables delivery and expression of mRNA in vitro in keratinocytes, fibroblasts, and in hSCS prepared from donors' skin. Telomerase activity in hSCS is significantly increased. hTERT mRNA LNP enhance the generation of a partial-thickness human skin equivalent in the mouse model, increasing hSCS engraftment (Lamin) and proliferation (Ki67), while reducing cellular senescence (p21) and DNA damage (53BP1).

Controlled Delivery of Rosuvastatin or Rapamycin through Electrospun Bismuth Nanoparticle-Infused Perivascular Wraps Promotes Arteriovenous Fistula Maturation.

In the context of arteriovenous fistula (AVF) failure, local delivery enables the release of higher concentrations of drugs that can suppress neointimal hyperplasia (NIH) while reducing systemic adverse effects. However, the radiolucency of polymeric delivery systems hinders long-term in vivo surveillance of safety and efficacy. We hypothesize that using a radiopaque perivascular wrap to deliver anti-NIH drugs could enhance AVF maturation. Through electrospinning, we fabricated multifunctional perivascular polycaprolactone (PCL) wraps loaded with bismuth nanoparticles (BiNPs) for enhanced radiologic visibility and drugs that can attenuate NIH─rosuvastatin (Rosu) and rapamycin (Rapa). The following groups were tested on the AVFs of a total of 24 Sprague-Dawley rats with induced chronic kidney disease: control (i.e., without wrap), PCL-Bi (i.e., wrap with BiNPs), PCL-Bi-Rosu, and PCL-Bi-Rapa. We found that BiNPs significantly improved the wraps' radiopacity without affecting biocompatibility. The drug release profiles of Rosu (hydrophilic drug) and Rapa (hydrophobic drug) differed significantly. Rosu demonstrated a burst release followed by gradual tapering over 8 weeks, while Rapa demonstrated a gradual release similar to that of the hydrophobic BiNPs. In vivo investigations revealed that both drug-loaded wraps can reduce vascular stenosis on ultrasonography and histomorphometry, as well as reduce [18F]Fluorodeoxyglucose uptake on positron emission tomography. Immunohistochemical studies revealed that PCL-Bi-Rosu primarily attenuated endothelial dysfunction and hypoxia in the neointimal layer, while PCL-Bi-Rapa modulated hypoxia, inflammation, and cellular proliferation across the whole outflow vein. In summary, the controlled delivery of drugs with different properties and mechanisms of action against NIH through a multifunctional, radiopaque perivascular wrap can improve imaging and histologic parameters of AVF maturation.

Mechanisms and Barriers in Nanomedicine: Progress in the Field and Future Directions.

In recent years, steady progress has been made in synthesizing and characterizing engineered nanoparticles, resulting in several approved drugs and multiple promising candidates in clinical trials. Regulatory agencies such as the Food and Drug Administration and the European Medicines Agency released important guidance documents facilitating nanoparticle-based drug product development, particularly in the context of liposomes and lipid-based carriers. Even with the progress achieved, it is clear that many barriers must still be overcome to accelerate translation into the clinic. At the recent conference workshop "Mechanisms and Barriers in Nanomedicine" in May 2023 in Colorado, U.S.A., leading experts discussed the formulation, physiological, immunological, regulatory, clinical, and educational barriers. This position paper invites open, unrestricted, nonproprietary discussion among senior faculty, young investigators, and students to trigger ideas and concepts to move the field forward.

Sex-dependent improvement in traumatic brain injury outcomes after liposomal delivery of dexamethasone in mice.

Traumatic Brain Injury (TBI) can have long-lasting physical, emotional, and cognitive consequences due to the neurodegeneration caused by its robust inflammatory response. Despite advances in rehabilitation care, effective neuroprotective treatments for TBI patients are lacking. Furthermore, current drug delivery methods for TBI treatment are inefficient in targeting inflamed brain areas. To address this issue, we have developed a liposomal nanocarrier (Lipo) encapsulating dexamethasone (Dex), an agonist for the glucocorticoid receptor utilized to alleviate inflammation and swelling in various conditions. In vitro studies show that Lipo-Dex were well tolerated in human and murine neural cells. Lipo-Dex showed significant suppression of inflammatory cytokines, IL-6 and TNF-α, release after induction of neural inflammation with lipopolysaccharide. Further, the Lipo-Dex were administered to young adult male and female C57BL/6 mice immediately after a controlled cortical impact injury. Our findings demonstrate that Lipo-Dex can selectively target the injured brain, thereby reducing lesion volume, cell death, astrogliosis, the release of proinflammatory cytokines, and microglial activation compared to Lipo-treated mice in a sex-dependent manner, showing a major impact only in male mice. This highlights the importance of considering sex as a crucial variable in developing and evaluating new nano-therapies for brain injury. These results suggest that Lipo-Dex administration may effectively treat acute TBI.

Modeling critical interaction for metastasis between circulating tumor cells (CTCs) and platelets adhered to the capillary wall.

BACKGROUND AND OBJECTIVE: We used a 2D fluid-solid interaction (FSI) model to investigate the critical conditions for the arrest of the CTCs traveling through the narrowed capillary with a platelet attached to the capillary wall. This computational model allows us to determine the deformations and the progression of the passage of the CTC through different types of microvessels with platelet included. METHODS: The modeling process is obtained using the strong coupling approach following the remeshing procedure. Also, the 1D FE rope element for simulating active ligand-receptor bonds is implemented in our computational tool (described below). RESULTS: A relationship between the CTCs properties (size and stiffness), the platelet size and stiffness, and the ligand-receptor interaction intensity, on one side, and the time in contact between the CTCs and platelet and conditions for the cell arrest, on the other side, are determined. The model is further validated in vitro by using a microfluidic device with metastatic breast tumor cells. CONCLUSIONS: The computational framework that is presented, with accompanying results, can be used as a powerful tool to study biomechanical conditions for CTCs arrest in interaction with platelets, giving a prognosis of disease progression.

Rosuvastatin-Eluting Gold Nanoparticle-Loaded Perivascular Implantable Wrap for Enhanced Arteriovenous Fistula Maturation in a Murine Model.

Background: Arteriovenous fistulas (AVFs) are a vital intervention for patients requiring hemodialysis, but they also contribute to overall mortality due to access malfunction. The most common cause of both AVF non-maturation and secondary failure is neointimal hyperplasia (NIH). Absorbable polycaprolactone (PCL) perivascular wraps can address these complications by incorporating drugs to attenuate NIH, such as rosuvastatin (ROSU), and metallic nanoparticles for visualization and device monitoring. Objectives: This study aimed to assess the impacts of gold nanoparticle (AuNP) and ROSU-loaded perivascular wraps on vasculature NIH and AVF maturation and patency in a chronic kidney disease rat model. Methods: Electrospun wraps containing combinations of PCL, AuNP, and ROSU were monitored for in vitro drug elution, nanoparticle release, tensile strength, and cell viability. Perivascular wraps were implanted in chronic kidney disease rats for in vivo ultrasound (US) and micro-computed tomography (mCT) imaging. AVF specimens were collected for histological analyses. Results: No difference in cell line viability was observed in ROSU-containing grafts. In vitro release studies of ROSU and AuNPs correlated with decreasing radiopacity over time on in vivo mCT analysis. The mCT study also demonstrated increased radiopacity in AuNP-loaded wraps compared with PCL and control. The addition of ROSU demonstrated decreased US and histologic measurements of NIH. Conclusions: The reduced NIH seen with ROSU-loading of perivascular wraps suggests a synergistic effect between mechanical support and anti-hyperplasia medication. Furthermore, the addition of AuNPs increased wrap radiopacity. Together, our results show that radiopaque, AuNP-, and ROSU-loaded PCL grafts induce AVF maturation and suppress NIH while facilitating optimal implanted device visualization.

Image-guided deployment and monitoring of a novel tungsten nanoparticleâ€"infused radiopaque absorbable inferior vena cava filter in pigs.

The use of absorbable inferior vena cava filters (IVCFs) constructed with poly-p-dioxanone (PPDO) eliminates risks and complications associated with the use of retrievable metallic filters. Radiopacity of radiolucent PPDO IVCFs can be improved with the incorporation of nanoparticles (NPs) made of high-atomic number materials such as gold and bismuth. In this study, we focused on incorporating tungsten NPs (WNPs), along with polyhydroxybutyrate (PHB), polycaprolactone (PCL), and polyvinylpyrrolidone (PVP) polymers to increase the surface adsorption of the WNPs. We compared the imaging properties of WNPs with single-polymer PHB (W-P) and WNPs with polymer blends consisting of PHB, PCL, and PVP (W-PB). Our in vitro analyses using PPDO sutures showed enhanced radiopacity with either W-P or W-PB coating, without compromising the inherent physico-mechanical properties of the PPDO sutures. We observed a more sustained release of WNPs from W-PB-coated sutures than W-P-coated sutures. We successfully deployed W-P- and W-PB-coated IVCFs into the inferior vena cava of pig models, with monitoring by fluoroscopy. At the time of deployment, W-PB-coated IVCFs showed a 2-fold increase in radiopacity compared to W-P-coated IVCFs. Longitudinal monitoring of in vivo IVCFs over a 12-week period showed a drastic decrease in radiopacity at week 3 for both filters. Results of this study highlight the utility of NPs and polymers for enhancing radiopacity of medical devices; however, different methods of incorporating NPs and polymers can still be explored to improve the efficacy, safety, and quality of absorbable IVCFs.

Lysicamine Reduces Protein Kinase B (AKT) Activation and Promotes Necrosis in Anaplastic Thyroid Cancer.

Anaplastic thyroid cancer (ATC) is an aggressive form of thyroid cancer (TC), accounting for 50% of total TC-related deaths. Although therapeutic approaches against TC have improved in recent years, the survival rate remains low, and severe adverse effects are commonly reported. However, unexplored alternatives based on natural compounds, such as lysicamine, an alkaloid found in plants with established cytotoxicity against breast and liver cancers, offer promise. Therefore, this study aimed to explore the antineoplastic effects of lysicamine in papillary TC (BCPAP) and ATC (HTH83 and KTC-2) cells. Lysicamine treatment reduced cell viability, motility, colony formation, and AKT activation while increasing the percentage of necrotic cells. The absence of caspase activity confirmed apoptosis-independent cell death. Necrostatin-1 (NEC-1)-mediated necrosome inhibition reduced lysicamine-induced necrosis in KTC-2, suggesting necroptosis induction via a reactive oxygen species (ROS)-independent mechanism. Additionally, in silico analysis predicted lysicamine target proteins, particularly those related to MAPK and TGF-ß signaling. Our study demonstrated lysicamine's potential as an antineoplastic compound in ATC cells with a proposed mechanism related to inhibiting AKT activation and inducing cell death.

Multi-stage delivery nano-particle systems for therapeutic applications.

BACKGROUND: The daunting task for drug molecules to reach pathological lesions has fueled rapid advances in Nanomedicine. The progressive evolution of nanovectors has led to the development of multi-stage delivery systems aimed at overcoming the numerous obstacles encountered by nanovectors on their journey to the target site. SCOPE OF REVIEW: This review summarizes major findings with respect to silicon-based drug delivery vectors for cancer therapeutics and imaging. Based on rational design, well-established silicon technologies have been adapted for the fabrication of nanovectors with specific shapes, sizes, and porosities. These vectors are part of a multi-stage delivery system that contains multiple nano-components, each designed to achieve a specific task with the common goal of site-directed delivery of therapeutics. MAJOR CONCLUSIONS: Quasi-hemispherical and discoidal silicon microparticles are superior to spherical particles with respect to margination in the blood, with particles of different shapes and sizes having unique distributions in vivo. Cellular adhesion and internalization of silicon microparticles is influenced by microparticle shape and surface charge, with the latter dictating binding of serum opsonins. Based on in vitro cell studies, the internalization of porous silicon microparticles by endothelial cells and macrophages is compatible with cellular morphology, intracellular trafficking, mitosis, cell cycle progression, cytokine release, and cell viability. In vivo studies support superior therapeutic efficacy of liposomal encapsulated siRNA when delivered in multi-stage systems compared to free nanoparticles. This article is part of a Special Issue entitled Nanotechnologies - Emerging Applications in Biomedicine.

Discoidal Porous Silicon Particles: Fabrication and Biodistribution in Breast Cancer Bearing Mice.

Porous silicon (pSi) is emerging as a promising material in the development of nanovectors for the systemic delivery of therapeutic and imaging agents. The integration of photolithographic patterning, typical of the semiconductor industry, with electrochemical silicon etching provides a highly flexible strategy to fabricate monodisperse and precisely tailored nanovectors. Here, a microfabrication strategy for direct lithographic patterning of discoidal pSi particles is presented that enables precise and independent control over particle size, shape, and porous structure. Discoidal pSi nanovectors with diameters ranging from 500 to 2600 nm, heights from 200 to 700 nm, pore sizes from 5 to 150 nm, and porosities from 40 to 90% are demonstrated. The degradation in serum, interaction with immune and endothelial cells in vitro, and biodistribution in mice bearing breast tumors are assessed for two discoidal nanovectors with sizes of 600 nm × 400 nm and 1000 nm × 400 nm. It is shown that both particle types are degraded after 24 h of continuous gentle agitation in serum, do not stimulate cytokine release from macrophages or affect endothelial cell viability, and accumulate up to about 10% of the injected dose per gram tissue in orthotopic murine models of breast cancer. The accumulation of the discoidal pSi nanovectors into the breast tumor mass is found to be up to five times higher than for spherical silica beads with similar diameters.

Multistage nanovectors: from concept to novel imaging contrast agents and therapeutics.

Over the last few decades a great variety of nanotechnology based platforms have been synthesized and fabricated to improve the delivery of active compounds to a disease site. Nanoparticles currently used in the clinic, and the majority of nanotherapeutics/nanodiagnostics under investigation, accommodate single- or multiple- functionalities on the same entity. Because many heterogeneous biological barriers can prevent therapeutic and imaging agents from reaching their intended targets in sufficient concentrations, there is an emerging requirement to develop a multimodular nanoassembly, in which different components with individual specific functions act in a synergistic manner. The multistage nanovectors (MSVs) were introduced in 2008 as the first system of this type. It comprises several nanocomponents or "stages", each of which is designed to negotiate one or more biological barriers. Stage 1 mesoporous silicon particles (S1MPs) were rationally designed and fabricated in a nonspherical geometry to enable superior blood margination and to increase cell surface adhesion. The main task of S1MPs is to efficiently transport nanoparticles that are loaded into their porous structure and to protect them during transport from the administration site to the disease lesion. Semiconductor fabrication techniques including photolithography and electrochemical etching allow for the exquisite control and precise reproducibility of S1MP physical characteristics such as geometry and porosity. Furthermore, S1MPs can be chemically modified with negatively/positively charged groups, PEG and other polymers, fluorescent probes, contrast agents, and biologically active targeting moieties including antibodies, peptides, aptamers, and phage. The payload nanoparticles, termed stage 2 nanoparticles (S2NPs), can be any currently available nanoparticles such as liposomes, micelles, inorganic/metallic nanoparticles, dendrimers, and carbon structures, within the approximate size range of 5-100 nm in diameter. Depending upon the physicochemical features of the S1MP (geometry, porosity, and surface modifications), a variety of S2NPs or nanoparticle "cocktails" can be loaded and efficiently delivered to the disease site. As demonstrated in the studies reviewed here, once the S2NPs are loaded into the S1MPs, a variety of novel properties emerge, which enable the design of new and improved imaging contrast agents and therapeutics. For example, the loading of the MRI Gd-based contrast agents onto hemispherical and discoidal S1MPs significantly increased the longitudal relaxivity (r1) to values of up to 50 times larger than those of clinically available gadolinium-based agents (~4 mM(-1) s(-1)/Gd(3+) ion). Furthermore, administration of a single dose of MSVs loaded with neutral nanoliposomes containing small interfering RNA (siRNA) targeted against the EphA2 oncoprotein enabled sustained EphA2 gene silencing for at least 21 days. As a result, the tumor burden was reduced in an orthotopic mouse model of ovarian cancer. We envision that the versatility of the MSV platform and its emerging properties will enable the creation of personalized solutions with broad clinical implications within and beyond the realm of cancer theranostics.