Eric M. Pridgen

Nanoparticle technologies for cancer therapy.

Nanoparticles as drug delivery systems enable unique approaches for cancer treatment. Over the last two decades, a large number of nanoparticle delivery systems have been developed for cancer therapy, including organic and inorganic materials. Many liposomal, polymer-drug conjugates, and micellar formulations are part of the state of the art in the clinics, and an even greater number of nanoparticle platforms are currently in the preclinical stages of development. More recently developed nanoparticles are demonstrating the potential sophistication of these delivery systems by incorporating multifunctional capabilities and targeting strategies in an effort to increase the efficacy of these systems against the most difficult cancer challenges, including drug resistance and metastatic disease. In this chapter, we will review the available preclinical and clinical nanoparticle technology platforms and their impact for cancer therapy.

DNA Self‐Assembly of Targeted Near‐Infrared‐Responsive Gold Nanoparticles for Cancer Thermo‐Chemotherapy

The development of external stimulus-responsive nanoparticle (NP) systems for cancer therapy has received considerable attention in recent years, as these systems can differentially increase drug accumulation at target cancer cells/tissues, drastically decrease systemic toxicity, and potentially avoid underor over-dosing. External stimuli that have been exploited for such applications include light, magnetic field, ultrasound, and electricity. Among them, nearinfrared (NIR) light (650–900 nm) has recently become an attractive stimulus because of its minimal absorbance by skin and tissue, thus allowing for noninvasive and deep tissue penetration. In particular, NIR light can be effectively converted into heat by using photothermal NPs, such as gold nanorods (NRs), gold nanoshells, hollow gold nanospheres, and carbon nanotubes. As such, NIR-responsive NP platforms offer several important benefits for cancer therapy. For example, NIR-induced local heating can be used for cancer thermotherapy. In addition, NIR-responsive NP delivery systems enable on-demand release of drugs for cancer chemotherapy, presumably by heat-induced disruption of the delivery vehicles. Furthermore, the combination of NIR-based thermotherapy and triggered chemotherapy (thermo-chemotherapy) could provide higher therapeutic efficacy than respective monotherapies. In addition to these advantages, investigators are exploring the possibility of integrating active targeting ligands in NIR-responsive NP platforms for targeted cancer thermochemotherapy. This triple combination of thermotherapy, triggered drug release, and targeted delivery, would achieve optimal therapeutic efficacy in cancer treatment, relative to pairwise combinations. For example, Lee et al. have designed folate-conjugated, doxorubicin (Dox) loaded poly(lactic-co-glycolic acid) (PLGA)–gold half-shell NPs, and this combination led to effective tumor elimination in target tissues in a NIR-responsive manner. A current strategy in formulating this targeted NIR-responsive NP requires multiple steps, including 1) the synthesis of drug-loaded NPs, 2) deposition of gold compositions on NPs, and 3) postconjugation with targeting ligands followed by purification. However, these complex processes could increase the difficulty of adjusting bio-physicochemical properties of NPs in a reproducible manner, and could contribute to unintended drug release from NPs, thereby resulting in unfavorable batch-to-batch variability in the characteristics of drug loading. Alternatively, using pre-functionalized components to self-assemble into targeted NPs would eliminate the need for post-modification of NPs and is amenable to being scaledup with little batch-to-batch variability. This self-assembly strategy has led to the clinical translation of first-in-man targeted cyclodextrin-based NPs for small interfering RNA (siRNA) delivery, and targeted PLGA-based NPs for docetaxel delivery. Nevertheless, use of such a self-assembly strategy in the design of targeted NIR-responsive NPs has not been reported to date. Inspired by nature and the ability of complimentary strands of DNA to hybridize, we designed a DNA-based platform that can self-assemble into targeted NIR-responsive NPs for cancer therapy. As illustrated in Figure 1, this platform comprises three distinct functional components: complementary DNA strands, the gold NR (50 nm 10 nm), and a polyethylene glycol (PEG) layer. The DNA strands, which consist of sequential CG base pairs, provide loading sites for Dox, a model chemotherapeutic drug. By changing the number of CG base pairs, drug loading can be precisely tuned. In addition to serving as drug-loading scaffold, one strand of the DNA (termed capture strand) is thiolated for gold NR capture, and the complementary strand (termed targeting strand) is pre-conjugated with ligands for cellspecific targeting. Gold NRs serve as the model NIR light-to[*] Dr. Z. Xiao, Dr. C. Ji, Dr. J. Shi, J. Frieder, Dr. J. Wu, Prof. O. C. Farokhzad Laboratory of Nanomedicine and Biomaterials Department of Anesthesiology, Brigham and Women’s Hospital Harvard Medical School, Boston, MA, 02115 (USA) E-mail: ofarokhzad@zeus.bwh.harvard.edu

Biodegradable, polymeric nanoparticle delivery systems for cancer therapy

Nanotechnology has the potential to impact the treatment of cancer significantly. This review will explore how this potential is beginning to be realized through the design of polymeric nanoparticle delivery systems. Current research is focused on developing biocompatible nanoparticles capable of targeting specific cancer markers and delivering imaging and therapeutic agents for the detection and treatment of cancer, resulting in a number of preclinical and clinical applications. More sophisticated nanoparticle designs are now in development, including particles able to release multiple drugs for enhanced treatment efficacy and targeted, multifunctional particles capable of combining imaging and drug release.

Transepithelial Transport of Fc-Targeted Nanoparticles by the Neonatal Fc Receptor for Oral Delivery

Nanoparticles targeted to the neonatal Fc receptor cross the intestinal epithelium and reach systemic circulation after oral administration. A Spoonful of Nanomedicine Oral delivery of drug-loaded nanoparticles is, to some, the Holy Grail of nanomedicine. Patients can easily pop a pill, which makes them more compliant with a therapeutic regimen. The difficulty with ingesting these tiny particles is that they are not readily absorbed in the intestine, thus eliminating most of the particles from the body and, in turn, limiting efficacy. In response, Pridgen et al. designed polymeric nanoparticles targeting a receptor expressed on the surface of the intestine to actively transport the particle across the cell into the patient’s circulation. The nanoparticles were decorated with Fc fragments that readily bind to the neonatal Fc receptor (FcRn) in the intestinal epithelium. The authors observed that the Fc-targeted nanoparticles crossed the intestinal barrier both in vitro, using human epithelial cells, and in vivo in mice (who also express FcRn), ending up in high concentrations in several organs of the body. By contrast, nontargeted nanoparticles were barely visible. To demonstrate the therapeutic benefits of these Fc-targeted nanoparticles, Pridgen et al. administered insulin-laden targeted and nontargeted particles orally to mice. Free insulin given orally did not generate a glucose response in the animals, similar to the nontargeted, insulin-containing particles. However, Fc-targeted nanoparticles containing insulin produced a significant hypoglycemic response in the mice. To confirm that the targeting and epithelial transport is important for this mode of delivery, the authors showed that animals lacking FcRn did not respond to the insulin-filled Fc-targeted nanoparticles. The ability to deliver nanomedicine orally would open doors to treating many chronic diseases that require daily therapy, such as diabetes and cancer. This study by Pridgen et al. is an exciting proof of concept but will require longer periods of testing in disease models to confirm that FcRn targeting is essential and safe for human use. Nanoparticles are poised to have a tremendous impact on the treatment of many diseases, but their broad application is limited because currently they can only be administered by parenteral methods. Oral administration of nanoparticles is preferred but remains a challenge because transport across the intestinal epithelium is limited. We show that nanoparticles targeted to the neonatal Fc receptor (FcRn), which mediates the transport of immunoglobulin G antibodies across epithelial barriers, are efficiently transported across the intestinal epithelium using both in vitro and in vivo models. In mice, orally administered FcRn-targeted nanoparticles crossed the intestinal epithelium and reached systemic circulation with a mean absorption efficiency of 13.7%*hour compared with only 1.2%*hour for nontargeted nanoparticles. In addition, targeted nanoparticles containing insulin as a model nanoparticle-based therapy for diabetes were orally administered at a clinically relevant insulin dose of 1.1 U/kg and elicited a prolonged hypoglycemic response in wild-type mice. This effect was abolished in FcRn knockout mice, indicating that the enhanced nanoparticle transport was specifically due to FcRn. FcRn-targeted nanoparticles may have a major impact on the treatment of many diseases by enabling drugs currently limited by low bioavailability to be efficiently delivered though oral administration.

Interactions of nanomaterials and biological systems: implications to personalized nanomedicine

The application of nanotechnology to personalized medicine provides an unprecedented opportunity to improve the treatment of many diseases. Nanomaterials offer several advantages as therapeutic and diagnostic tools due to design flexibility, small sizes, large surface-to-volume ratio, and ease of surface modification with multivalent ligands to increase avidity for target molecules. Nanomaterials can be engineered to interact with specific biological components, allowing them to benefit from the insights provided by personalized medicine techniques. To tailor these interactions, a comprehensive knowledge of how nanomaterials interact with biological systems is critical. Herein, we discuss how the interactions of nanomaterials with biological systems can guide their design for diagnostic, imaging and drug delivery purposes. A general overview of nanomaterials under investigation is provided with an emphasis on systems that have reached clinical trials. Finally, considerations for the development of personalized nanomedicines are summarized such as the potential toxicity, scientific and technical challenges in fabricating them, and regulatory and ethical issues raised by the utilization of nanomaterials.

Enhancing tumor cell response to chemotherapy through nanoparticle-mediated codelivery of siRNA and cisplatin prodrug

Significance The development of acquired chemoresistance is often a clinical problem limiting the successful treatment of cancers. RNAi is showing promising results in human clinical trials. The combination of chemotherapy with RNAi approaches to suppress the expression of proteins involved in the emergence of drug resistance represents a promising synergistic strategy to circumvent or reverse acquired chemoresistance. Such combination therapy approaches require specific delivery vehicles to encapsulate and deliver chemotherapy and siRNA therapeutics simultaneously in a controlled manner. Herein, we describe a nanoparticle technology to codeliver a DNA-damaging chemotherapeutic and siRNAs that impair the cell’s ability to repair the DNA damage. This combination therapeutic approach can sensitize cancer cells to chemotherapeutics and shows superior tumor inhibition compared with monochemotherapy. Cisplatin and other DNA-damaging chemotherapeutics are widely used to treat a broad spectrum of malignancies. However, their application is limited by both intrinsic and acquired chemoresistance. Most mutations that result from DNA damage are the consequence of error-prone translesion DNA synthesis, which could be responsible for the acquired resistance against DNA-damaging agents. Recent studies have shown that the suppression of crucial gene products (e.g., REV1, REV3L) involved in the error-prone translesion DNA synthesis pathway can sensitize intrinsically resistant tumors to chemotherapy and reduce the frequency of acquired drug resistance of relapsed tumors. In this context, combining conventional DNA-damaging chemotherapy with siRNA-based therapeutics represents a promising strategy for treating patients with malignancies. To this end, we developed a versatile nanoparticle (NP) platform to deliver a cisplatin prodrug and REV1/REV3L-specific siRNAs simultaneously to the same tumor cells. NPs are formulated through self-assembly of a biodegradable poly(lactide-coglycolide)-b-poly(ethylene glycol) diblock copolymer and a self-synthesized cationic lipid. We demonstrated the potency of the siRNA-containing NPs to knock down target genes efficiently both in vitro and in vivo. The therapeutic efficacy of NPs containing both cisplatin prodrug and REV1/REV3L-specific siRNAs was further investigated in vitro and in vivo. Quantitative real-time PCR results showed that the NPs exhibited a significant and sustained suppression of both genes in tumors for up to 3 d after a single dose. Administering these NPs revealed a synergistic effect on tumor inhibition in a human Lymph Node Carcinoma of the Prostate xenograft mouse model that was strikingly more effective than platinum monotherapy.

HER-2-targeted nanoparticle-affibody bioconjugates for cancer therapy.

Affibodies are a class of polypeptide ligands that are potential candidates for cell- or tissue-specific targeting of drug-encapsulated controlled release polymeric nanoparticles (NPs). Here we report the development of drug delivery vehicles comprised of polymeric NPs that are surface modified with Affibody ligands that bind to the extracellular domain of the trans-membrane human epidermal growth factor receptor 2 (HER-2) for targeted delivery to cells which over express the HER-2 antigen. NPs lacking the anti-HER-2 Affibody did not show significant uptake by these cells. Using paclitaxel encapsulated NP-Affibody (1 wt% drug loading), we demonstrated increased cytotoxicity of these bioconjugates in SK-BR-3 and SKOV-3 cell lines. These targeted, drug encapsulated NPAffibody bioconjugates may be efficacious in treating HER-2 expressing carcinoma.

Microfluidic platform for combinatorial synthesis and optimization of targeted nanoparticles for cancer therapy.

Taking a nanoparticle (NP) from discovery to clinical translation has been slow compared to small molecules, in part by the lack of systems that enable their precise engineering and rapid optimization. In this work we have developed a microfluidic platform for the rapid, combinatorial synthesis and optimization of NPs. The system takes in a number of NP precursors from which a library of NPs with varying size, surface charge, target ligand density, and drug load is produced in a reproducible manner. We rapidly synthesized 45 different formulations of poly(lactic-co-glycolic acid)-b-poly(ethylene glycol) NPs of different size and surface composition and screened and ranked the NPs for their ability to evade macrophage uptake in vitro. Comparison of the results to pharmacokinetic studies in vivo in mice revealed a correlation between in vitro screen and in vivo behavior. Next, we selected NP synthesis parameters that resulted in longer blood half-life and used the microfluidic platform to synthesize targeted NPs with varying targeting ligand density (using a model targeting ligand against cancer cells). We screened NPs in vitro against prostate cancer cells as well as macrophages, identifying one formulation that exhibited high uptake by cancer cells yet similar macrophage uptake compared to nontargeted NPs. In vivo, the selected targeted NPs showed a 3.5-fold increase in tumor accumulation in mice compared to nontargeted NPs. The developed microfluidic platform in this work represents a tool that could potentially accelerate the discovery and clinical translation of NPs.

Nanoparticle Encapsulation of Mitaplatin and the Effect Thereof on In Vivo Properties

Nanoparticle (NP) therapeutics have the potential to significantly alter the in vivo biological properties of the pharmaceutically active agents that they carry. Here we describe the development of a polymeric NP, termed M-NP, comprising poly(D,L-lactic-co-glycolic acid)-block-poly(ethylene glycol) (PLGA-PEG), stabilized with poly(vinyl alcohol) (PVA), and loaded with a water-soluble platinum(IV) [Pt(IV)] prodrug, mitaplatin. Mitaplatin, c,c,t-[PtCl2(NH3)2(OOCCHCl2)2], is a compound designed to release cisplatin, an anticancer drug in widespread clinical use, and the orphan drug dichloroacetate following chemical reduction. An optimized preparation of M-NP by double emulsion and its physical characterization are reported, and the influence of encapsulation on the properties of the platinum agent is evaluated in vivo. Encapsulation increases the circulation time of Pt in the bloodstream of rats. The biodistribution of Pt in mice is also affected by nanoparticle encapsulation, resulting in reduced accumulation in the kidneys. Finally, the efficacy of both free mitaplatin and M-NP, measured by tumor growth inhibition in a mouse xenograft model of triple-negative breast cancer, reveals that controlled release of mitaplatin over time from the nanoparticle treatment produces long-term efficacy comparable to that of free mitaplatin, which might limit toxic side effects.

Hybrid Lipid-Polymer Nanoparticles for Sustained siRNA Delivery and Gene Silencing

UNLABELLED The development of controlled-release nanoparticle (NP) technologies has great potential to further improve the therapeutic efficacy of RNA interference (RNAi), by prolonging the release of small interfering RNA (siRNA) for sustained, long-term gene silencing. Herein, we present an NP platform with sustained siRNA-release properties, which can be self-assembled using biodegradable and biocompatible polymers and lipids. The hybrid lipid-polymer NPs showed excellent silencing efficacy, and the temporal release of siRNA from the NPs continued for over one month. When tested on luciferase-expressed HeLa cells and A549 lung carcinoma cells after short-term transfection, the siRNA NPs showed greater sustained silencing activity than lipofectamine 2000-siRNA complexes. More importantly, the NP-mediated sustained silencing of prohibitin 1 (PHB1) generates more effective tumor cell growth inhibition in vitro and in vivo than the lipofectamine complexes. We expect that this sustained-release siRNA NP platform could be of interest in both fundamental biological studies and clinical applications. FROM THE CLINICAL EDITOR Emerging gene silencing applications could be greatly enhanced by prolonging the release of siRNA for sustained gene silencing. This team of scientists presents a hybrid lipid-polymer nanoparticle platform that successfully accomplishes this goal, paving the way to future research studies and potential clinical applications.