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.

Matrix Metalloproteinase- and pH-Sensitive Nanoparticle System Enhances Drug Retention and Penetration in Glioblastoma.

Glioblastoma (GBM) is a primary malignant brain tumor with limited therapeutic options. One promising approach is local drug delivery, but the efficacy is hindered by limited diffusion and retention. To address this, we synthesized and developed a dual-sensitive nanoparticle (Dual-NP) system, formed between a dendrimer and dextran NPs, bound by a dual-sensitive [matrix metalloproteinase (MMP) and pH] linker designed to disassemble rapidly in the tumor microenvironment. The disassembly prompts the in situ formation of nanogels via a Schiff base reaction, prolonging Dual-NP retention and releasing small doxorubicin (Dox)-conjugated dendrimer NPs over time. The Dual-NPs were able to penetrate deep into 3D spheroid models and detected at the tumor site up to 6 days after a single intratumoral injection in an orthotopic mouse model of GBM. The prolonged presence of Dual-NPs in the tumor tissue resulted in a significant delay in tumor growth and an overall increase in survival compared to untreated or Dox-conjugated dendrimer NPs alone. This Dual-NP system has the potential to deliver a range of therapeutics for efficiently treating GBM and other solid tumors.

Microneedle-mediated Delivery of Immunomodulators Restores Immune Privilege in Hair Follicles and Reverses Immune-Mediated Alopecia.

Disorders in the regulatory arm of the adaptive immune system result in autoimmune-mediated diseases. While systemic immunosuppression is the prevailing approach to manage them, it fails to achieve long-lasting remission due to concomitant suppression of the regulatory arm and carries the risk of heightened susceptibility to infections and malignancies. Alopecia Areata is a condition characterized by localized hair loss due to autoimmunity. The accessibility of the skin provides an opportunity for local rather than systemic intervention to avoid broad immunosuppression. We hypothesized that expansion of endogenous regulatory T cells (Tregs) at the site of antigen encounter can restore the immune balance and generate a long-lasting tolerogenic response. We therefore utilized a hydrogel microneedle (MN) patch for delivery of CCL22, a chemoattractant for Tregs, and IL-2, a Treg survival factor to amplify them. In an immune-mediated murine model of alopecia, we showed local bolstering of Treg numbers leading to sustained hair regrowth and attenuation of inflammatory pathways. In a humanized skin transplant mouse model, we confirmed expansion of Tregs within human skin without engendering peripheral immunosuppression. The MN patch offered high-loading capacity and shelf-life stability for prospective clinical translation. By harmonizing immune responses locally, we aspire to reshape the landscape of autoimmune skin disease management. This article is protected by copyright. All rights reserved.

Multimodal neuro-nanotechnology: Challenging the existing paradigm in glioblastoma therapy.

Integrating multimodal neuro- and nanotechnology-enabled precision immunotherapies with extant systemic immunotherapies may finally provide a significant breakthrough for combatting glioblastoma (GBM). The potency of this approach lies in its ability to train the immune system to efficiently identify and eradicate cancer cells, thereby creating anti-tumor immune memory while minimizing multi-mechanistic immune suppression. A critical aspect of these therapies is the controlled, spatiotemporal delivery of structurally defined nanotherapeutics into the GBM tumor microenvironment (TME). Architectures such as spherical nucleic acids or poly(beta-amino ester)/dendrimer-based nanoparticles have shown promising results in preclinical models due to their multivalency and abilities to activate antigen-presenting cells and prime antigen-specific T cells. These nanostructures also permit systematic variation to optimize their distribution, TME accumulation, cellular uptake, and overall immunostimulatory effects. Delving deeper into the relationships between nanotherapeutic structures and their performance will accelerate nano-drug development and pave the way for the rapid clinical translation of advanced nanomedicines. In addition, the efficacy of nanotechnology-based immunotherapies may be enhanced when integrated with emerging precision surgical techniques, such as laser interstitial thermal therapy, and when combined with systemic immunotherapies, particularly inhibitors of immune-mediated checkpoints and immunosuppressive adenosine signaling. In this perspective, we highlight the potential of emerging treatment modalities, combining advances in biomedical engineering and neurotechnology development with existing immunotherapies to overcome treatment resistance and transform the management of GBM. We conclude with a call to action for researchers to leverage these technologies and accelerate their translation into the clinic.

Sprayable Hydrogel Sealant for Gastrointestinal Wound Shielding.

Naturally occurring internal bleeding, such as in stomach ulcers, and complications following interventions, such as polyp resection post-colonoscopy, may result in delayed (5-7 days) post-operative adverse events-such as bleeding, intestinal wall perforation, and leakage. Current solutions for controlling intra- and post-procedural complications are limited in effectiveness. Hemostatic powders only provide a temporary solution due to their short-term adhesion to GI mucosal tissues (less than 48 h). In this study, a sprayable adhesive hydrogel for facile application and sustained adhesion to GI lesions is developed using clinically available endoscopes. Upon spraying, the biomaterial (based on polyethyleneimine-modified Pluronic micelles precursor and oxidized dextran) instantly gels upon contact with the tissue, forming an adhesive shield. In vitro and in vivo studies in guinea pigs, rabbits, and pig models confirm the safety and efficacy of this biomaterial in colonic and acidic stomach lesions. The authors' findings highlight that this family of hydrogels ensures prolonged tissue protection (3-7 days), facilitates wound healing, and minimizes the risk of delayed complications. Overall, this technology offers a readily adoptable approach for gastrointestinal wound management.

Investigation of the enhanced antitumour potency of STING agonist after conjugation to polymer nanoparticles.

Intravenously administered cyclic dinucleotides and other STING agonists are hampered by low cellular uptake and poor circulatory half-life. Here we report the covalent conjugation of cyclic dinucleotides to poly(ß-amino ester) nanoparticles through a cathepsin-sensitive linker. This is shown to increase stability and loading, thereby expanding the therapeutic window in multiple syngeneic tumour models, enabling the study of how the long-term fate of the nanoparticles affects the immune response. In a melanoma mouse model, primary tumour clearance depends on the STING signalling by host cells-rather than cancer cells-and immune memory depends on the spleen. The cancer cells act as a depot for the nanoparticles, releasing them over time to activate nearby immune cells to control tumour growth. Collectively, this work highlights the importance of nanoparticle structure and nano-biointeractions in controlling immunotherapy efficacy.

Poly(ß-amino ester)s-Based Delivery Systems for Targeted Transdermal Vaccination.

Nucleic acid vaccines have become a transformative technology to fight emerging infectious diseases and cancer. Delivery of such via the transdermal route could boost their efficacy given the complex immune cell reservoir present in the skin that is capable of engendering robust immune responses. We have generated a novel library of vectors derived from poly(ß-amino ester)s (PBAEs) including oligopeptide-termini and a natural ligand, mannose, for targeted transfection of antigen presenting cells (APCs) such as Langerhans cells and macrophages in the dermal milieu. Our results reaffirmed terminal decoration of PBAEs with oligopeptide chains as a powerful tool to induce cell-specific transfection, identifying an outstanding candidate with a ten-fold increased transfection efficiency over commercial controls in vitro. The inclusion of mannose in the PBAE backbone rendered an additive effect and increased transfection levels, achieving superior gene expression in human monocyte-derived dendritic cells and other accessory antigen presenting cells. Moreover, top performing candidates were capable of mediating surface gene transfer when deposited as polyelectrolyte films onto transdermal devices such as microneedles, offering alternatives to conventional hypodermic administration. We predict that the use of highly efficient delivery vectors derived from PBAEs could advance clinical translation of nucleic acid vaccination over protein- and peptide-based strategies.

Localized Nanoparticle-Mediated Delivery of miR-29b Normalizes the Dysregulation of Bone Homeostasis Caused by Osteosarcoma whilst Simultaneously Inhibiting Tumor Growth.

Patients diagnosed with osteosarcoma undergo extensive surgical intervention and chemotherapy resulting in dismal prognosis and compromised quality of life owing to poor bone regeneration, which is further compromised with chemotherapy delivery. This study aims to investigate if localized delivery of miR-29b-which is shown to promote bone formation by inducing osteoblast differentiation and also to suppress prostate and cervical tumor growth-can suppress osteosarcoma tumors whilst simultaneously normalizing the dysregulation of bone homeostasis caused by osteosarcoma. Thus, the therapeutic potential of microRNA (miR)-29b is studied to promote bone remodeling in an orthotopic model of osteosarcoma (rather than in bone defect models using healthy mice), and in the context of chemotherapy, that is clinically relevant. A formulation of miR-29b:nanoparticles are developed that are delivered via a hyaluronic-based hydrogel to enable local and sustained release of the therapy and to study the potential of attenuating tumor growth whilst normalizing bone homeostasis. It is found that when miR-29b is delivered along with systemic chemotherapy, compared to chemotherapy alone, the therapy provided a significant decrease in tumor burden, an increase in mouse survival, and a significant decrease in osteolysis thereby normalizing the dysregulation of bone lysis activity caused by the tumor.

Polymeric microneedles enable simultaneous delivery of cancer immunomodulatory drugs and detection of skin biomarkers.

Background: Immune-modulating therapies impart positive outcomes in a subpopulation of cancer patients. Improved delivery strategies and non-invasive monitoring of anti-tumor effects can help enhance those outcomes and understand the mechanisms associated with the generation of anti-tumor immune responses following immunotherapy. Methods: We report on the design of a microneedle (MN) platform capable of simultaneous delivery of immune activators and collection of interstitial skin fluid (ISF) to monitor therapeutic responses. While either approach has shown promise, the integration of the therapy and diagnostic arms into one MN platform has hardly been explored before. MNs were synthesized out of crosslinked hyaluronic acid (HA) and loaded with a model immunomodulatory nanoparticle-containing drug, CpG oligodinucleotides (TLR9 agonist), for cancer therapy in melanoma and colon cancer models. The therapeutic response was monitored by longitudinal analysis of entrapped immune cells in the MNs following patch retrieval and digestion. Results: Transdermal delivery of CpG-containing NPs with MNs induced anti-tumor immune responses in multiple syngeneic mouse cancer models. CpG-loaded MNs stimulated innate immune cells and reduced tumor growth. Intravital microscopy showed deposition and spatiotemporal co-localization of CpG-NPs within the tumor microenvironment when delivered with MNs. Analysis of MN-sampled ISF revealed similar immune signatures to those seen in the bulk tumor homogenate, such as increased populations of macrophages and effector T cells following treatment. Conclusions: Our hydrogel-based MNs enable effective transdermal drug delivery into immune cells in the tumor microenvironment, and upon retrieval, enable studying the immune response to the therapy over time. This platform has the theranostic potential to deliver a range of combination therapies while detecting biomarkers.

Sprayable Hydrogel for Instant Sealing of Vascular Anastomosis.

Bleeding-related complications following vascular surgeries occur in up to half of the patients-500 000 cases annually in the United States alone. This results in additional procedures, increased mortality rate, and prolonged hospitalization, posing a burden on the healthcare system. Commercially available materials rely, in large, on forming covalent bonds between the tissue and the biomaterial to achieve adhesion. Here, it is shown that a biomaterial based on oxidized alginate and oxidized dextran together with polyamidoamine (PAMAM) dendrimer amine provides simultaneous electrostatic and covalent interactions between the biomaterial and the tissue, maximizing adhesion. This study finds that the material withstands supraphysiological pressures (≈300 mmHg) and prevents bleeding in a rabbit aortic puncture model and in a pig carotid bilateral poly(tetrafluoroethylene) graft model-achieving superior performance to commercially available materials such as Tisseel and BioGlue. Material biocompatibility is validated in comprehensive in vitro and in vivo studies in accordance with the US Food and Drug Administration (FDA) guidelines, including in vitro neutral red uptake test, subcutaneous implantation in rabbits, ames genotoxicity, and guinea pig maximization test. This material has the potential to provide with adequate seal and reduced complications following complex vascular surgeries, including hard-to-seal tissue-graft interfaces.

The landscape of receptor-mediated precision cancer combination therapy via a single-cell perspective.

Mining a large cohort of single-cell transcriptomics data, here we employ combinatorial optimization techniques to chart the landscape of optimal combination therapies in cancer. We assume that each individual therapy can target any one of 1269 genes encoding cell surface receptors, which may be targets of CAR-T, conjugated antibodies or coated nanoparticle therapies. We find that in most cancer types, personalized combinations composed of at most four targets are then sufficient for killing at least 80% of tumor cells while sparing at least 90% of nontumor cells in the tumor microenvironment. However, as more stringent and selective killing is required, the number of targets needed rises rapidly. Emerging individual targets include PTPRZ1 for brain and head and neck cancers and EGFR in multiple tumor types. In sum, this study provides a computational estimate of the identity and number of targets needed in combination to target cancers selectively and precisely.

A Spheroid Model of Early and Late-Stage Osteosarcoma Mimicking the Divergent Relationship between Tumor Elimination and Bone Regeneration.

Osteosarcoma is the most diagnosed bone tumor in children. The use of tissue engineering strategies after malignant tumor resection remains a subject of scientific controversy. As a result, there is limited research that focuses on bone regeneration postresection, which is further compromised following chemotherapy. This study aims to develop the first co-culture spheroid model for osteosarcoma, to understand the divergent relationship between tumor elimination and bone regeneration. By manipulating the ratio of stromal to osteosarcoma cells the modelled cancer state (early/late) is modified, as is evident by the increased tumor growth rates and an upregulation of a panel of well-established osteosarcoma prognostic genes. Validation of the authors' model is conducted by analyzing its ability to mimic the cytotoxic effects of the FDA-approved chemotherapeutic Doxorubicin. Next, the model is used to investigate what effect osteogenic supplements have, if any, on tumor growth. When their model is treated with osteogenic supplements, there is a stimulatory effect on the surrounding stromal cells. However, when treated with chemotherapeutics this stimulatory effect is significantly diminished. Together, the results of this study present a novel multicellular model of osteosarcoma and provide a unique platform for screening potential therapeutic options for osteosarcoma before conducting in vivo experiments.

Ultrasonic Thermal Monitoring of the Brain Using Golay-Coded Excitations-Feasibility Study.

Thermal monitoring during focused ultrasound (FUS) transcranial procedures is mandatory and commonly performed by MRI. Transcranial ultrasonic thermal monitoring is an attractive alternative. Furthermore, using the therapeutic FUS transducer itself for this task is highly desirable. Nonetheless, such application is challenged by massive skull-induced signal attenuation and aberrations. This study examined the feasibility of implementing the Golay-coded excitations (CoE) for temperature monitoring in bovine brain samples in the range of 35 °C-43 °C (hyperthermia). Feasibility was assessed using computer simulations, water-based phantoms, and ex vivo bovine brain white-matter samples. The samples were gradually heated to about 45 °C and sonicated during cool down with a 1-MHz therapeutic FUS implementing Golay CoE. Initially, a calibration curve correlating the normalized time-of-flight (TOF) changes and the temperature was generated. Next, a bovine bone was positioned between the FUS and the brain samples, and the scanning process was repeated for different fresh samples. The calibration curve was then used as a mean for estimating the temperature, which was compared to thermocouple measurements. The simulations demonstrated a substantial improvement in signal-to-noise ratio (SNR) and suggested that the implementation of 4-bit sequences is advantageous. The experimental measurements with bone demonstrated good temperature estimation with an average absolute error for the water phantoms and brains of 1.46 °C ± 1.22 °C and 1.23 °C ± 0.99 °C, respectively. In conclusion, a novel noninvasive method utilizing the Golay CoE for ultrasonic thermal monitoring using a therapeutic FUS transducer is introduced. This method can lead to the development of an acoustic tool for brain thermal monitoring.

Scale-up manufacturing of gelatin-based microcarriers for cell therapy.

Microcarriers, including crosslinked porous gelatin beads (Cultispher G) are widely used as cell carriers for cell therapy applications. Microcarriers can support a range of adherent cell types in stirred tank bioreactor culture, which is scalable up to several thousands of liters. Cultispher G in particular is advantageous for cell therapy applications because it can be dissolved enzymatically, and thus cells can be harvested without the need to perform a large-scale cell-bead filtration step. This enzymatic dissolution, however, is challenged by the slow degradation of the carriers in the presence of enzymes as new extracellular matrix is being deposited by the proliferating cells. This extended dissolution timelimits the yield of cell recovery while compromising cellular viability. We report herein the development of crosslinked porous gelatin beads that afford rapid, stimuli-triggered dissolution for facile cell removal using human mesenchymal stem cells (hMSC) as a model system. We successfully fabricated redox-sensitive beads (RS beads) and studied their cell growth, dissolution time and cell yield, compared to regular gelatin-based beads (Reg beads). We have shown that RS beads allow for much faster dissolution compared to Reg beads, supporting better hMSC detachment and recovery following 8 days of culture in spinner flasks, or in 3L bioreactors. These newly synthesized RS beads show promise as cellular microcarriers and can be used for scale-up manufacturing of different cell types while providing on-demand degradation for facile cell retrieval.

Prolonged Local In Vivo Delivery of Stimuli-Responsive Nanogels That Rapidly Release Doxorubicin in Triple-Negative Breast Cancer Cells.

Triple negative breast cancer patients remain with chemotherapy as their only viable therapeutic option. However, the toxicity of available anticancer drugs and their inefficient delivery have limited the development of effective chemotherapy administration protocols and combination therapies. Drug delivery devices that can properly target chemotherapy to the right cells with efficient cancer-cell killing may play a vital role in eliminating triple-negative breast cancer. While systemic delivery results in low drug accumulation at the tumor site and for a short period of time, local delivery enables sustained drug release. However, a system that is able to provide rapid, yet prolonged action, would enable efficient tumor elimination. Herein, the development of dual-sensitive nanogels is described that are designed to rapidly dislodge the chemotherapy drug, doxorubicin, inside cancer cells through dual-sensitive action-pH and redox sensitivities-enabling efficient cancer-cell killing while eliminating systemic side effects. Their embedding within a hydrogel injected next to a tumor in a triple-negative breast-cancer mouse model enables prolonged release of the drug with instantaneous action when inside the cells resulting in efficacious tumor elimination compared to sustained local delivery only. This technology can be used for the delivery of combination therapies and for the treatment of other solid tumors.