Andrew Satterlee

In Vivo Gene Delivery by Nonviral Vectors: Overcoming Hurdles?

The promise of cancer gene therapeutics is hampered by difficulties in the in vivo delivery to the targeted tumor cells, and systemic delivery remains to be the biggest challenge to be overcome. Here, we concentrate on systemic in vivo gene delivery for cancer therapy using nonviral vectors. In this review, we summarize the existing delivery barriers together with the requirements and strategies to overcome these problems. We will also introduce the current progress in the design of nonviral vectors, and briefly discuss their safety issues.

Delivery of Oligonucleotides with Lipid Nanoparticles

Since their inception in the 1980s, oligonucleotide-based (ON-based) therapeutics have been recognized as powerful tools that can treat a broad spectrum of diseases. The discoveries of novel regulatory methods of gene expression with diverse mechanisms of action are still driving the development of novel ON-based therapeutics. Difficulties in the delivery of this class of therapeutics hinder their in vivo applications, which forces drug delivery systems to be a prerequisite for clinical translation. This review discusses the strategy of using lipid nanoparticles as carriers to deliver therapeutic ONs to target cells in vitro and in vivo. A discourse on how chemical and physical properties of the lipid materials could be utilized during formulation and the resulting effects on delivery efficiency constitutes the major part of this review.

Synergistic anti-tumor effects of combined gemcitabine and cisplatin nanoparticles in a stroma-rich bladder carcinoma model

Tumors grown in a stroma-rich mouse model resembling clinically advanced bladder carcinoma with UMUC3 and NIH 3T3 cells have high levels of fibroblasts and an accelerated tumor growth rate. We used this model to investigate the synergistic effect of combined gemcitabine monophosphate (GMP) nanoparticles and Cisplatin nanoparticles (Combo NP) on tumor-associated fibroblasts (TAFs). A single injection of Combo NP had synergistic anti-tumor effects while the same molar ratio of combined GMP and Cisplatin delivered as free drug (Combo Free) fell outside of the synergistic range. Combo NP nearly halted tumor growth with little evidence of general toxicity while Combo Free had only a modest inhibitory effect at 16mg/kg GMP and 1.6mg/kg Cisplatin. Combo NP increased levels of apoptosis within the tumor by approximately 1.3 folds (TUNEL analysis) and decreased α-SMA-positive fibroblast recruitment by more than 87% (immunofluorescence) after multiple injections compared with Combo Free, GMP NP or Cisplatin NP alone. The TAF-targeting capability of Combo NP was evaluated by double staining for TUNEL and α-SMA at various time points after a single injection. On day one after injection, 57% of the TUNEL-positive cells were identified as α-SMA-positive fibroblasts. By day four, tumor stroma was 85% depleted and 87% of the remaining TAFs were TUNEL-positive. Combo NP-treated tumors became 2.75 folds more permeable than those treated with Combo Free as measured by Evans Blue. We conclude that the antineoplastic effect of Combo NP works by first targeting TAFs and is more effective as an anti-tumor therapy than Combo Free, GMP NP or Cisplatin NP alone.

Tumor-targeted delivery of sunitinib base enhances vaccine therapy for advanced melanoma by remodeling the tumor microenvironment

&NA; Development of an effective treatment against advanced tumors remains a major challenge for cancer immunotherapy. We have previously developed a potent mannose‐modified lipid calcium phosphate (LCP) nanoparticle (NP)‐based Trp2 vaccine for melanoma therapy, but because this vaccine can induce a potent anti‐tumor immune response only during the early stages of melanoma, poor tumor growth inhibition has been observed in more advanced melanoma models, likely due to the development of an immune‐suppressive tumor microenvironment (TME). To effectively treat this aggressive tumor, a multi‐target receptor tyrosine kinase inhibitor, sunitinib base, was efficiently encapsulated into a targeted polymeric micelle nano‐delivery system (SUNb‐PM), working in a synergistic manner with vaccine therapy in an advanced mouse melanoma model. SUNb‐PM not only increased cytotoxic T‐cell infiltration and decreased the number and percentage of MDSCs and Tregs in the TME, but also induced a shift in cytokine expression from Th2 to Th1 type while remodeling the tumor‐associated fibroblasts, collagen, and blood vessels in the tumor. Additionally, inhibition of the Stat3 and AKT signaling pathways by SUNb‐PM may induce tumor cell apoptosis or decrease tumor immune evasion. Our findings indicated that targeted delivery of a tyrosine kinase inhibitor to tumors can be used in a novel synergistic way to enhance the therapeutic efficacy of existing immune‐based therapies for advanced melanoma. Graphical abstract Figure. No caption available.

Sigma receptor-mediated targeted delivery of anti-angiogenic multifunctional nanodrugs for combination tumor therapy.

The potential of low molecular weight heparin (LMWH) in anti-angiogenic therapy has been tempered by poor in vivo delivery to the tumor cell and potentially harmful side effects, such as the risk of bleeding due to heparins anticoagulant activity. In order to overcome these limitations and further improve the therapeutic effect of LMWH, we designed a novel combination nanosystem of LMWH and ursolic acid (UA), which is also an angiogenesis inhibitor for tumor therapy. In this system, an amphiphilic LMWH-UA (LHU) conjugate was synthesized and self-assembled into core/shell nanodrugs with combined anti-angiogenic activity and significantly reduced anticoagulant activity. Furthermore, DSPE-PEG-AA-modified LHU nanodrugs (A-LHU) were developed to facilitate the delivery of nanodrugs to the tumor. The anti-angiogenic activity of A-LHU was investigated both in vitro and in vivo. It was found that A-LHU significantly inhibited the tubular formation of human umbilical vein endothelial cells (HUVECs) (p<0.01) and the angiogenesis induced by basic fibroblast growth factor (bFGF) in a Matrigel plug assay (p<0.001). More importantly, A-LHU displayed significant inhibition on the tumor growth in B16F10-bearing mice in vivo. The level of CD31 and p-VEGFR-2 expression has demonstrated that the excellent efficacy of antitumor was associated with a decrease in angiogenesis. In conclusion, A-LHU nanodrugs are a promising multifunctional antitumor drug delivery system.

A radio-theranostic nanoparticle with high specific drug loading for cancer therapy and imaging.

We have developed a theranostic nanoparticle delivering the model radionuclide (177)Lu based on the versatile lipid-calcium-phosphate (LCP) nanoparticle delivery platform. Characterization of (177)Lu-LCP has shown that radionuclide loading can be increased by several orders of magnitude without affecting the encapsulation efficiency or the morphology of (177)Lu-LCP, allowing consistency during fabrication and overcoming scale-up barriers typical of nanotherapeutics. The choice of (177)Lu as a model radionuclide has allowed in vivo anticancer therapy in addition to radiographic imaging via the dual decay modes of (177)Lu. Tumor accumulation of (177)Lu-LCP was measured using both SPECT and Cerenkov imaging modalities in live mice, and treatment with just one dose of (177)Lu-LCP showed significant in vivo tumor inhibition in two subcutaneous xenograft tumor models. Microenvironment and cytotoxicity studies suggest that (177)Lu-LCP inhibits tumor growth by causing apoptotic cell death via double-stranded DNA breaks while causing a remodeling of the tumor microenvironment to a more disordered and less malignant phenotype.

Current and Future Theranostic Applications of the Lipid-Calcium-Phosphate Nanoparticle Platform

Over the last four years, the Lipid-Calcium-Phosphate (LCP) nanoparticle platform has shown success in a wide range of treatment strategies, recently including theranostics. The high specific drug loading of radiometals into LCP, coupled with its ability to efficiently encapsulate many types of cytotoxic agents, allows a broad range of theranostic applications, many of which are yet unexplored. In addition to providing an overview of current medical imaging modalities, this review highlights the current theranostic applications for LCP using SPECT and PET, and discusses potential future uses of the platform by comparing it with both systemically and locally delivered clinical radiotherapy options as well as introducing its applications as an MRI contrast agent. Strengths and weaknesses of LCP and of nanoparticles in general are discussed, as well as caveats regarding the use of fluorescence to determine the accumulation or biodistribution of a probe.

Enhancing Nanoparticle Accumulation and Retention in Desmoplastic Tumors via Vascular Disruption for Internal Radiation Therapy.

Aggressive, desmoplastic tumors are notoriously difficult to treat because of their extensive stroma, high interstitial pressure, and resistant tumor microenvironment. We have developed a combination therapy that can significantly slow the growth of large, stroma-rich tumors by causing massive apoptosis in the tumor center while simultaneously increasing nanoparticle uptake through a treatment-induced increase in the accumulation and retention of nanoparticles in the tumor. The vascular disrupting agent Combretastatin A-4 Phosphate (CA4P) is able to increase the accumulation of radiation-containing nanoparticles for internal radiation therapy, and the retention of these delivered radioisotopes is maintained over several days. We use ultrasound to measure the effect of CA4P in live tumor-bearing mice, and we encapsulate the radio-theranostic isotope 177Lutetium as a therapeutic agent as well as a means to measure nanoparticle accumulation and retention in the tumor. This combination therapy induces prolonged apoptosis in the tumor, decreasing both the fibroblast and total cell density and allowing further tumor growth inhibition using a cisplatin-containing nanoparticle.

Theranostic etoposide phosphate/indium nanoparticles for cancer therapy and imaging

Etoposide phosphate (EP), a water-soluble anticancer prodrug, is widely used for treatment of many cancers. After administration it is rapidly converted to etoposide, its parent compound, which exhibits anticancer activity. Difficulty in parenteral administration necessitates the development of a suitable nanoparticle delivery system for EP. Here we have used indium both as a carrier to deliver etoposide phosphate to tumor cells and as a SPECT imaging agent through incorporation of (111)In. Etoposide phosphate was successfully encapsulated together with indium in nanoparticles, and exhibited dose dependent cytotoxicity and induction of apoptosis in cultured H460 cancer cells via G2/M cell cycle arrest. In a mouse xenograft lung cancer model, etoposide phosphate/indium nanoparticles induce tumor cell apoptosis, leading to significant enhancement of tumor growth inhibition compared to the free drug.

Quantitative Evaluation of the Mechanism Underlying the Biotransportation of the Active Ingredients in Puerariae lobatae Radix and Chuanxiong rhizoma

The objective of this study was to establish a quantitative method to evaluate the biotransportation of a drug across the cell membrane. Through the application of the law of mass conservation, the drug transportation rate was calculated based on Ficks law of passive diffusion and the Michaelis–Menten equation. The overall membrane‐transportation rate was the sum of the passive diffusion rate and the carrier‐mediated diffusion rate, which were calculated as the transportation mass divided by the respective rate. The active ingredients of Puerariae lobatae Radix and Chuanxiong rhizoma, namely, puerarin and ferulic acid, respectively, were used as two model drugs. The transportation rates of puerarin and ferulic acid were obtained by fitting a model that includes both Ficks law of diffusion and the Michaelis–Menten equation. Compared with the overall transportation, the carrier‐mediated transport and passive diffusion of 1.59 mmol/L puerarin were −35.07% and 64.93%, respectively, whereas the respective transportation modes of 0.1 mmol/L ferulic acid were −35.40% and 64.60%, respectively. Verapamil and MK‐571 increased the overall transport rate and ratio, and MK‐571 treatment changed the carrier‐mediated transport from negative to positive. However, the transport rate and ratio of ferulic acid did not change significantly. The cell transportation mechanisms of puerarin and ferulic acid primarily involve simple passive diffusion and carrier‐mediated transportation. Moreover, P‐glycoprotein and multidrug resistance‐associated protein efflux proteins, and other transportation proteins were found to be involved in the transportation of puerarin. Copyright