Nanoparticle (NP)-based mRNA cancer vaccines hold great promise to realize personalized cancer treatments. To advance this technology requires delivery formulations for efficient intracellular delivery to antigen-presenting cells. We developed a class of bioreducible lipophilic poly(beta-amino ester) nanocarriers with quadpolymer architecture. The platform is agnostic to the mRNA sequence, with one-step self-assembly allowing for delivery of multiple antigen-encoding mRNAs as well as codelivery of nucleic acid?based adjuvants. We examined structure?function relationships for NP-mediated mRNA delivery to dendritic cells (DCs) and identified that a lipid subunit of the polymer structure was critical. Following intravenous administration, the engineered NP design facilitated targeted delivery to the spleen and preferential transfection of DCs without the need for surface functionalization with targeting ligands. Treatment with engineered NPs codelivering antigen-encoding mRNA and toll-like receptor agonist adjuvants led to robust antigen-specific CD8+ T cell responses, resulting in efficient antitumor therapy in in vivo models of murine melanoma and colon adenocarcinoma.

In recent years, the immune system has increasingly been recognized as a critical component to understanding cancer progression and cellular microenvironments. This has led to massive growth in the field of cancer immunotherapy, in which the patient's natural defense mechanisms are harnessed and enhanced to fight cancer. In this field, T-cell engineering has been the most widely studied and developed approach, with checkpoint inhibitors like anti-PD-1 antibody recently showing impressive clinical success for solid tumors and ex vivo engineering of chimeric antigen receptor T-cells making a significant impact on liquid tumors. There has been continued basic and translational research interest in developing engineering technologies for delivery of therapeutic agents to induce T-cell reprogramming both ex vivo and in vivo for cancer immunotherapy. In this chapter, delivery technologies that provide surface stimulation to T-cells (outside-in) as well as technologies that deliver intracellular mediators to T-cells (inside-out) for reprogramming to enhance anti-cancer activity are discussed.

Glioblastoma (GBM) is an aggressive central nervous system cancer with a dismal prognosis. The standard of care involves surgical resection followed by radiotherapy and chemotherapy, but five-year survival is only 5.6% despite these measures. Novel therapeutic approaches, such as immunotherapies, targeted therapies, and gene therapies, have been explored to attempt to extend survival for patients. Nanoparticles have been receiving increasing attention as promising vehicles for non-viral nucleic acid delivery in the context of GBM, though delivery is often limited by low blood-brain barrier permeability, particle instability, and low trafficking to target brain structures and cells. In this review, nanoparticle design considerations and new advances to overcome nucleic acid delivery challenges to treat brain cancer are summarized and discussed.

Clinical translation of polymer-based nanocarriers for systemic delivery of RNA has been limited due to poor colloidal stability in the blood stream and intracellular delivery of the RNA to the cytosol. To address these limitations, this study reports a new strategy incorporating photocrosslinking of bioreducible nanoparticles for improved stability extracellularly and rapid release of RNA intracellularly. In this design, the polymeric nanocarriers contain ester bonds for hydrolytic degradation and disulfide bonds for environmentally triggered small interfering RNA (siRNA) release in the cytosol. These photocrosslinked bioreducible nanoparticles (XbNPs) have a shielded surface charge, reduced adsorption of serum proteins, and enable superior siRNA-mediated knockdown in both glioma and melanoma cells in high-serum conditions compared to non-crosslinked formulations. Mechanistically, XbNPs promote cellular uptake and the presence of secondary and tertiary amines enables efficient endosomal escape. Following systemic administration, XbNPs facilitate targeting of cancer cells and tissue-mediated siRNA delivery beyond the liver, unlike conventional nanoparticle-based delivery. These attributes of XbNPs facilitate robust siRNA-mediated knockdown in vivo in melanoma tumors colonized in the lungs following systemic administration. Thus, biodegradable polymeric nanoparticles, via photocrosslinking, demonstrate extended colloidal stability and efficient delivery of RNA therapeutics under physiological conditions, and thereby potentially advance systemic delivery technologies for nucleic acid-based therapeutics.

INTRODUCTION: Gene delivery technologies are being developed for an increasing number of biomedical applications, with delivery vehicles including viruses and non-viral materials. Among biomaterials used for non-viral gene delivery, poly(beta-amino ester)s (PBAEs), a class of synthetic, biodegradable polymers, have risen as a leading gene delivery vehicle that has been used for multiple applications in vitro and in vivo.

AREAS COVERED: This review summarizes the key properties of PBAEs and their development, including a discussion of the advantages and disadvantages of PBAEs for gene delivery applications. The use of PBAEs to improve the properties of other drug delivery vehicles is also summarized.

EXPERT OPINION: PBAEs are designed to have multiple characteristics that are ideal for gene delivery, including their reversible positive charge, which promotes binding to nucleic acids as well as imparting high buffering capacity, and their rapid degradability under mild conditions. Simultaneously, some of their properties also lead to nanoparticle instability and low transfection efficiency in physiological environments. The ease with which PBAEs can be chemically modified as well as non-covalently blended with other materials, however, allows them to be customized specifically to overcome delivery barriers for varied applications.

Efficient cytosolic protein delivery is necessary to fully realize the potential of protein therapeutics. Current methods of protein delivery often suffer from low serum tolerance and limited in vivo efficacy. Here, we report the synthesis and validation of a previously unreported class of carboxylated branched poly(β-amino ester)s that can self-assemble into nanoparticles for efficient intracellular delivery of a variety of different proteins. In vitro, nanoparticles enabled rapid cellular uptake, efficient endosomal escape, and functional cytosolic protein release into cells in media containing 10% serum. Moreover, nanoparticles encapsulating CRISPR-Cas9 ribonucleoproteins (RNPs) induced robust levels of gene knock-in (4%) and gene knockout (>75%) in several cell types. A single intracranial administration of nanoparticles delivering a low RNP dose (3.5 pmol) induced robust gene editing in mice bearing engineered orthotopic murine glioma tumors. This self-assembled polymeric nanocarrier system enables a versatile protein delivery and gene editing platform for biological research and therapeutic applications.

Improved delivery materials are needed to enable siRNA transport across biological barriers, including the blood-brain barrier (BBB), to treat diseases like brain cancer. We engineered bioreducible nanoparticles for systemic siRNA delivery to patient-derived glioblastoma cells in an orthotopic mouse tumor model. We first utilized a newly developed biomimetic in vitro model to evaluate and optimize the performance of the engineered bioreducible nanoparticles at crossing the brain microvascular endothelium. We performed transmission electron microscopy imaging which indicated that the engineered nanoparticles are able to cross the BBB endothelium via a vesicular mechanism. The nanoparticle formulation engineered to best cross the BBB model in vitro led to safe delivery across the BBB to the brain in vivo. The nanoparticles were internalized by human brain cancer cells, released siRNA to the cytosol via environmentally-triggered degradation, and gene silencing was obtained both in vitro and in vivo. This study opens new frontiers for the in vitro evaluation and engineering of nanomedicines for delivery to the brain, and reports a systemically administered biodegradable nanocarrier for oligonucleotide delivery to treat glioma.

Polymeric nanoparticles have tremendous potential to improve the efficacy of therapeutic cancer treatments by facilitating targeted delivery to a desired site. The physical and chemical properties of polymers can be tuned to accomplish delivery across the multiple biological barriers required to reach diverse subsets of cells. The use of biodegradable polymers as nanocarriers is especially attractive, as these materials can be designed to break down in physiological conditions and engineered to exhibit triggered functionality when at a particular location or activated by an external source. We present how biodegradable polymers can be engineered as drug delivery systems to target the tumor microenvironment in multiple ways. These nanomedicines can target cancer cells directly, the blood vessels that supply the nutrients and oxygen that support tumor growth, and immune cells to promote anticancer immunotherapy.

OBJECTIVES: The release of magnesium ions (Mg²⁺ ) from titanium surfaces has been shown to boost the initial biological response of peri-implant bone and to increase the biomechanical strength of osseointegration. The objective of the present paper was to investigate if the initial improvement in osseointegration would influence the bone remodeling also during the maturation stage of bone healing.

METHODS: Titanium implants were coated with mesoporous titania layers and either loaded with Mg²⁺ (test group) or left untreated (control group). The implants were inserted in the tibiae of 10 New Zealand White rabbits. Osseointegration was assessed after 6 weeks by means of biomechanical testing (RTQ), non-decalcified histology and histomorphometry (BIC%, BA%, NBA%). The expression of genes involved in the bone formation and remodeling was quantified using qPCR.

RESULTS: Mg²⁺ releasing mesoporous titania coatings showed, on average, higher removal torques and histomorphometrical outcomes (RTQ: 17.2 Ncm vs. 15 Ncm; BIC: 38.8% vs. 32.1%; BA%: 71.6% vs. 64%; NBA% 62.5% vs. 54% for the tests vs the controls); however, the differences were not statistically significant. Three osteogenic markers, osteocalcin (OC), collagen 1 alpha 1 (COL1A1), and alkalin phosphatase (ALPL), were respectively 2-fold, 1.53-fold, and 1.13-fold up-regulated in the control group compared to the test. The expression of COL1A1 was particularly high in both groups, while the biomarkers for remodeling and inflammation showed a low expression in both groups.

SIGNIFICANCE: The results suggested that the initial enhancement in osseointegration induced by magnesium release from mesoporous titania coatings has no detrimental effects during bone maturation. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 2118-2125, 2017.

UNLABELLED: Magnesium has a key role in osteoporosis and could enhance implant osseointegration in osteoporotic patients. Titanium implants impregnated with Mg ions were installed in the tibia of ovariectomized rats. The release of Mg induced a significant increase of bone formation and the expression of anabolic markers in the peri-implant bone.

INTRODUCTION: The success of endosseous implants is highly predictable in patients possessing normal bone status, but it may be impaired in patients with osteoporosis. Thus, the application of strategies that adjuvate implant healing in compromized sites is of great interest. Magnesium has a key role in osteoporosis prevention and it is an interesting candidate for this purpose. In this study, the cellular and molecular effects of magnesium release from implants were investigated at the early healing stages of implant integration.

METHODS: Osteoporosis was induced in 24 female rats by means of ovariectomy and low-calcium diet. Titanium mini-screws were coated with mesoporous titania films and were loaded with magnesium (test group) or left as native (control group). The implants were inserted in the tibia and femur of the rats. One, 2 and 7 days after implantation, the implants were retrieved and histologically examined. In addition, expression of genes was evaluated in the peri-implant bone tissue at day 7 by means of quantitative polymerase chain reactions with pathway-oriented arrays.

RESULTS: The histological evaluation revealed that new bone formation started already during the first week of healing for both groups. However, around the test implants, new bone was significantly more abundant and spread along a larger surface of the implants. In addition, the release of magnesium induced a significantly higher expression of BMP6.

CONCLUSIONS: These results provide evidence that the release of magnesium promoted rapid bone formation and the activation of osteogenic signals in the vicinity of implants placed in osteoporotic bone.