Circulation Half-life Research Articles

Abstract 5740: Peptide-functionalized dendrimers for enhanced binding to immune checkpoint proteins

Abstract The human immune system has complex and dynamic mechanisms to distinguish and eliminate malignant cells and defend the body against cancer. Various therapeutic approaches to stimulate the immune system have thus been investigated, demonstrating strong anti-tumor immune responses capable of controlling metastasis and preventing recurrence. Nevertheless, existing cancer immunotherapies suffer from several limitations, including inconsistent benefits across cancer types and patients. In recent years, the utilization of peptides for immunotherapy has attracted a great deal of interest due to their ability to mimic protein functionalities while maintaining a high degree of modularity and synthetic reproducibility in their molecular design. However, the clinical translation of otherwise promising peptide-based immunotherapeutics has been hindered due to their significantly weaker binding to target proteins and short half-life in plasma circulation in comparison to their monoclonal antibody counterparts. Nevertheless, these issues can potentially be addressed by conjugation to nanoparticles, which enhances cell binding/uptake and prolongs circulation. To this end, our group has extensively investigated the use of poly(amidoamine) (PAMAM) dendrimers to improve the therapeutic utility of peptides. These dendrimers can facilitate the simultaneous binding of conjugated peptides to multiple cell receptors, resulting in a significantly enhanced binding interaction through the multivalent binding effect. Here, we report an efficient de novo nanoparticle design strategy that combines organic synthesis, phage display, and molecular dynamics to identify a novel immuno-oncology therapeutic modality. We first employed phage display to identify several highly selective sequences that bind to the mouse ortholog of TIGIT—an inhibitory receptor expressed on lymphocytes that has been identified as a potential therapeutic target to enhance anti-tumor immune responses. Subsequent computational modeling (NAMD with CHARMM force field) revealed several conserved regions of interaction that we hypothesize drive favorable binding interactions and enhance the binding affinity of the peptide inhibitors. We next incorporated experimental kinetic measurements of the identified sequences with adaptive evolution computational modeling to identify optimized mutant analogs for dendrimer conjugation. It was found that the TIGIT targeting DPCs (dendrimer-peptide conjugates) demonstrated increased binding avidity in vitro and anti-tumor efficacy in vivo, most notably when administered in combination with PD-L1 targeting DPCs to a syngeneic mouse model bearing mouse oral carcinoma (MOC1) tumors. Overall, this work demonstrates the potential of rational peptide design and our multi-functional DPC platform for the development of both effective and novel immuno-oncology therapeutics. Citation Format: Piper Anne Rawding, Francesco Coppola, DaWon Kim, Michael Poellmann, Deric Wheeler, Petr Král, Seungpyo Hong. Peptide-functionalized dendrimers for enhanced binding to immune checkpoint proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5740.

Abstract 5118: Neuroblastoma extracellular vesicles present GPC2 and activate GPC2 CAR T cells in an antigen-dependent manner

Abstract Glypican 2 (GPC2) chimeric antigen receptor (CAR) T cells are safe and efficacious in neuroblastoma and are currently being tested in a first-in-human Phase 1 clinical trial at the Children’s Hospital of Philadelphia. Tumor-derived extracellular vesicles (TEVs) are nanosized vesicles secreted by cancer cells, often enriched with glypicans. However, the presence of GPC2 on neuroblastoma TEVs and any interaction with GPC2 CAR T cells has not been explored. High-levels of GPC2 were found on TEVs isolated from neuroblastoma preclinical models [7 cell lines, circulating EVs from 6 patient-derived xenograft (PDX)-bearing mice] and from the peripheral blood of 16 neuroblastoma patients, but not on circulating EVs from 7 non-tumor bearing mice or 12 healthy human donors. The level of circulating GPC2+ EVs isolated from the peripheral blood of PDX-bearing mice positively correlated with PDX size (R=0.96; P<0.0001) and parent PDX GPC2 expression. To evaluate how GPC2 on EVs modulates CAR T cell functionality, we generated EVs with varied levels of GPC2 (Hi/Lo/Neg) and co-incubated each EV subset with GPC2 CAR T cells. Here using immunofluorescence and flow assays we found that EVs bind GPC2 CAR T cells proportional to the amount of GPC2 on their surface. Further, GPC2Hi EV-CAR T cell synapses induced potent T cell activation shown by CD69/Granzyme B expression and release of IL-2/IFN-γ. GPC2Hi EV pre-incubated CAR T cells also displayed enhanced target neuroblastoma cell cytotoxicity, inducing 4-fold more specific cytotoxicity than CAR T cells incubated with control GPC2Neg EVs. In vivo we observed that injection of GPC2Hi EVs into GPC2Lo neuroblastoma SK-N-AS xenografts genetically altered to have significantly decreased endogenous EV production (via RAB27A knock-out) greatly enhanced the efficacy of co-infused GPC2 CAR T cells (p<0.05). Similarly, in an isogenic SK-N-AS-GPC2 xenograft murine model treated with a limiting number of GPC2 CAR T cells concurrently with the EV-inhibiting drug GW4869, GPC2 CAR efficacy was dependent on the presence of GPC2Hi EVs. Here, pharmacologic EV inhibition in mice decreased GPC2 CAR efficacy which could be rescued by intratumoral injection of GPC2Hi EVs. In both in vivo models, the presence of intratumoral GPC2Hi EVs resulted in a significantly enhanced tumor infiltration of activated (CD69+/CD25+) GPC2 CAR T cells (p<0.05). Taken together, GPC2Hi EVs are selectively secreted from neuroblastomas and bind and activate GPC2 CAR T cells enhancing their anti-tumor cytotoxicity. To further capitalize on these findings, we engineered GPC2Hi EVs to have an extended circulation half-life through albumin binding or tumor targeting via GD2 binding that are currently being tested in vivo and results will be reported. EVs offer a versatile platform for CAR antigen presentation and should be further validated as a strategy to enhance CAR T cell efficacy for solid tumors. Citation Format: Anna M. Giudice, Stephanie Matlaga, Guillem Pascual-Pasto, Patrick M. Schuerch, Geoffrey Rouin, Brendan McIntyre, Vincent P. Zecchino, Kristopher R. Bosse. Neuroblastoma extracellular vesicles present GPC2 and activate GPC2 CAR T cells in an antigen-dependent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5118.

Abstract 4058: Nanoparticle delivery of interleukin-2 induces anti-tumor immune response and therapeutic efficacy in mouse tumor models

Abstract Immune checkpoint inhibitors (ICIs) have demonstrated robust responses in some solid tumor types. However, the low response rate, which is associated with lack of pre-existing tumor-infiltrating lymphocytes (TILs), remains a significant challenge. Interleukin-2 (IL-2) can effectively expand TILs and has been FDA-approved for metastatic renal cancer and melanoma. However, IL-2 has several drawbacks—including short circulation half-life, dose-limiting toxicities, and counterproductive activation of regulatory T cells (Tregs)—resulting in less than 10% complete response rate. Numerous efforts have been made to enhance IL-2 formulations, but none have yet received FDA approval. In this study, we utilized our proprietary nanoparticle platform (Pdx-NP™) to improve IL-2 formulation. Pdx-NP™ can load IL-2 with near-complete binding, resulting in a 160-nm nanoconstruct (IL2-NP), suitable for both local and intravenous administrations. To evaluate the efficacy, we utilized bilateral melanoma (B16F10) and colon (MC38) tumor models, in which the local tumor was treated intratumorally for 3 doses and the distal tumor was left untreated to assess the systemic anti-tumor immune response. In the B16F10 model, IL2-NP treatment demonstrated favorable immune profiles in local and distal tumors and the draining lymph nodes, such as increased CD8+ T cell proliferation, CD8+ T cells/Treg ratio, and recruitment of dendritic cells and macrophages, compared to saline or free IL-2 counterparts. These positive immune responses translated into significant growth reduction of IL2-NP treated tumors and improvement in survival of the mice compared to free IL-2. In the MC38 tumor model, the IL2-NP treatment resulted in complete regression of both local and distal tumors in 43% of the mice. When combined with ICIs (αPD-1 and αCTLA-4), the treatment achieved a cure rate of 100%, whereas ICIs alone yielded only a 29% cure rate. In conclusion, IL2-NP induces robust proliferation and activation of TILs and is efficacious both as a monotherapy and in combination with ICIs. This is due to the ability of Pdx-NP™ to protect and retain therapeutics within tumors, with an 8-fold increase in tumor accumulation over the free drug counterpart. Our ongoing efforts are focused on the targeted delivery of IL-2 to tumors upon systemic administration, using αPD-L1 antibody-conjugated Pdx-NP™. This antibody-conjugated nanoparticle has an elimination half-life (t1/2) of about 25 hrs in non-human primates. Paired with αPD-L1 for tumor homing, this nanoconstruct is hypothesized to enhance IL-2 accumulation in tumors compared to free IL-2, which has t1/2 of only 1.5 hrs in human. The current clinical landscape of cancer immunotherapy focuses on combinational approaches, and our versatile Pdx-NP™ platform, which is capable of co-delivering different classes of therapeutics, is well suited for this endeavor. Citation Format: Ruijie Wang, Alyssa Wallstrum, Natalie White, Pramod Kumar, Tongchatra Watcharawittayakul, Noah A. Crumrine, Cole Baker, Jeremy Saito, Moataz Reda, Worapol Ngamcherdtrakul, Wassana Yantasee. Nanoparticle delivery of interleukin-2 induces anti-tumor immune response and therapeutic efficacy in mouse tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4058.

Thrombotic microenvironment responsive crosslinking cyclodextrin metal-organic framework nanocarriers for precise targeting and thrombolysis

Global public health is seriously threatened by thrombotic disorders because of their high rates of mortality and disability. Most thrombolytic agents, especially protein-based pharmaceuticals, have a short half-life in circulation, reducing their effectiveness in thrombolysis. The creation of an intelligent drug delivery system that delivers medication precisely and releases it under regulated conditions at nearby thrombus sites is essential for effective thrombolysis. In this article, we present a unique medication delivery system (MCRUA) that selectively targets platelets and releases drugs by stimulation from the thrombus' microenvironment. The thrombolytic enzyme urokinase-type plasminogen-activator (uPA) and the anti-inflammatory medication Aspirin (acetylsalicylic acid, ASA) are both loaded onto pH-sensitive CaCO3/cyclodextrin crosslinking metal-organic frameworks (MC) that make up the MCRUA system. c(RGD) is functionalized on the surface of MC, which is functionalized by RGD to an esterification reaction. Additionally, the thrombus site's acidic microenvironment causes MCRUA to disintegrate to release uPA for thrombolysis and aiding in vessel recanalization. Moreover, cyclodextrin-encapsulated ASA enables the treatment of the inflammatory environment within the thrombus, enhancing the antiplatelet aggregation effects and promoting cooperative thrombolysis therapy. When used for thrombotic disorders, our drug delivery system (MCRUA) promotes thrombolysis, suppresses rethrombosis, and enhances biosafety with fewer hemorrhagic side effects.

Delivering Relaxin Plasmid by Polymeric Metformin Lipid Nanoparticles for Liver Fibrosis Treatment.

Liver fibrosis usually progresses to liver cirrhosis and even results in hepatocellular carcinoma, which accounts for one million deaths annually worldwide. To date, anti-liver fibrosis drugs for clinical treatment have not yet been approved. Nowadays, as a natural regulator, Relaxin (RLX) has received increased attention because the expression of RLX could deactivate the activation of hepatic stellate cells (aHSCs) and resolve liver fibrosis. However, its application in treatment is limited due to the short half-life in circulation and low accumulation within the target organ. To address these problems, a kind of polymeric metformin (PolyMet)-loaded relaxin plasmid (pRLX) core-membrane lipid nanoparticle (PolyMet-pRLX-LNPs, PRLNP) was prepared. Here, PolyMet was used as a carrier to replace the traditional polymer polyethylene diene (PEI), which is of higher toxicity, to prolong the circulation time of pRLX in vivo. Then, the antifibrotic ability of PRLNP to overcome liver fibrosis was carried out in C57BL/6 mice. It is worth mentioning that this is the first time to investigate the potential of PRLNP in carbon tetrachloride-induced liver fibrosis. The results showed that PRLNP effectively downregulated fibrosis-related biomarkers such as alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Meanwhile, histopathological examinations also showed low collagen accumulation, revealing that PRLNP could histologically and functionally alleviate liver fibrosis. In addition, no significant difference in serum biochemical value between the PRLNP and the normal group, suggesting the safety profile of PRLNP. This research proposed a novel non-toxic treatment method for liver fibrosis with a nanosystem to effectively treat liver fibrosis.

Predicting the spontaneous vaporisation of superheated perfluorocarbon droplets

Superheated perfluorocarbon droplets have been widely explored as agents for ultrasound imaging and therapy, as well as for other applications such as radiation dosimetry. Submicrometre, or “nano” droplets offer a number of potential advantages over microbubbles, e.g., longer circulation half-lives, higher surface area-to-volume ratio and the ability to perfuse the microvasculature more easily. A key challenge in the use of nanodroplets, however, is the need to avoid spontaneous vaporisation whilst keeping the energy required for acoustic activation within the range of pressures that can be used safely in humans. This is especially important for imaging applications. Perfluoropropane (C;3F;8) microbubbles can be condensed to form liquid nanodroplets that offer a good trade-off between thermal stability and acoustic vaporisation threshold. Anecdotal reports, however, suggest that C;3F;8 droplets can spontaneously vaporise, and may therefore pose a potential safety risk, especially if bubble coalescence occurs. The aim of this study was to build on recent theoretical models of droplet vaporisation and investigate the probability of vaporisation as a function of temperature and interfacial tension. The results are compared with experimental measurements of vaporisation rates for different droplet formulations.

Liver sinusoidal cells eliminate blood-borne phage K1F.

Phage treatment has regained attention due to an increase in multiresistant bacteria. For phage therapy to be successful, phages must reach their target bacteria in sufficiently high numbers. Blood-borne phages are believed to be captured by macrophages in the liver and spleen. Since liver sinusoids also consist of specialized scavenger liver sinusoidal endothelial cells (LSECs) and Kupffer cells (KCs), this study investigated the contribution of both cell types in the elimination of Escherichia coli phage K1Fg10b::gfp (K1Fgfp) in mice. Circulatory half-life, organ, and hepatocellular distribution of K1Fgfp were determined following intravenous administration. Internalization of K1Fgfp and effects of phage opsonization on uptake were explored using primary mouse and human LSEC and KC cultures. When inoculated with 107 virions, >95% of the total K1Fgfp load was eliminated from the blood within 20 min, and 94% of the total retrieved K1Fgfp was localized to the liver. Higher doses resulted in slower elimination, possibly reflecting temporary saturation of liver scavenging capacity. Phage DNA was detected in both cell types, with a KC:LSEC ratio of 12:1 per population following cell isolation. Opsonization with plasma proteins increased time-dependent cellular uptake in both LSECs and KCs in vitro. Internalized phages were rapidly transported along the endocytic pathway to lysosomal compartments. Reduced viability of intracellular K1Fgfp corroborated inactivation following endocytosis. This study is the first to identify phage distribution in the liver at the hepatocellular level, confirming clearance of K1Fgfp performed mostly by KCs with a significant uptake also in LSECs.IMPORTANCEFaced with the increasing amounts of bacteria with multidrug antimicrobial resistance, phage therapy has regained attention as a possible treatment option. The phage field has recently experienced an emergence in commercial interest as research has identified new and more efficient ways of identifying and matching phages against resistant superbugs. Currently, phages are unapproved drugs in most parts of the world. For phages to reach broad clinical use, they must be shown to be clinically safe and useful. The results presented herein contribute to increased knowledge about the pharmacokinetics of the T7-like phage K1F in the mammalian system. The cell types of the liver that are responsible for rapid phage blood clearance are identified. Our results highlight the need for more research about appropriate dose regimens when phage therapy is delivered intravenously and advise essential knowledge about cell systems that should be investigated further for detailed phage pharmacodynamics.

Cholesterol-Modified DNA Nanostructures Serve as Effective Non-Viral Carriers for Delivering siRNA to the Kidneys to Prevent Acute Kidney Injury.

With the emergence of gene therapy utilizing viral vectors, the potential risks associated with these vectors have prompted increased attention toward non-viral alternatives. DNA nanotechnology enables the assembly of specific oligonucleotide chains into nanostructures possessing defined spatial configurations. Due to their inherent characteristics, DNA nanostructures possess natural advantages as carriers for regulating gene expression in a non-viral manner. Cholesterol modification can convert DNA nanostructures from hydrophilic materials to amphiphilic materials, thereby extending their systemic circulation time. In this study, the high-dimensional design and cholesterol modification are shown to prolong the systemic circulation half-life of DNA nanostructures in mice. Specifically, the tetrahedron structure modified with three cholesterol molecules (TDN-3Chol) exhibit excellent circulation time and demonstrate a preference for renal uptake. The unique characteristics of TDN-3Chol can effectively deliver p53 siRNA to the mouse renal tubular tissue, resulting in successful knockdown of p53 and demonstrating its potential for preventing acute kidney injury. Furthermore, TDN-3Chol is not exhibited significant toxicity in mice, highlighting its promising role as a non-viral vector for targeted gene expression regulation in the kidneys. The designed non-viral vector as a prophylactic medication shows potential in addressing the current clinical challenges associated with nephrotoxic drugs.

Mono-amino acid linkers enable highly potent small molecule-drug conjugates by conditional release

The endosome cleavable linkers have been widely employed by antibody-drug conjugates and small molecule-drug conjugates (SMDCs) to control the accurate release of payloads. An effective linker should provide stability in systemic circulation but efficient payload release at its targeted tumor sites. This conflicting requirement always leads to linker design with increasing structural complexity. Balance of the effectiveness and structural complexity presents a linker design challenge. Here, we explored the possibility of mono-amino acid as so far the simplest cleavable linker (X-linker) for SMDC-based auristatin delivery. Within a diverse set of X-linkers, the SMDCs differed widely in bioactivity, with one (Asn-linker) having significantly improved potency (IC50= 0.1nM) and fast response to endosomal cathepsin B cleavage. Notably, this SMDC, once grafted with effector protein fragment crystallizable (Fc), demonstrated a profound invivo therapeutic effect in aspects of targetability, circulation half-life (t1/2= 73 h), stability, and anti-tumor efficacy. On the basis of these results, we believe that this mono-amino acid linker, together with the new SMDC-Fc scaffold, has significant potential in targeted delivery application.

Curcumin Nanocarriers with a Focus on Cellular Uptake Studies – An Innovative Cancer Therapy

Once a dye, curcumin (CUR) has transformed into a versatile therapeutic agent with antioxidative, anti-inflammatory, and anticancer properties. Despite its potent anticancer effects, CUR encounters challenges such as poor solubility and a short circulation half-life. To address this, researchers utilize nanocarriers like nanoparticles, liposomes, and micelles for efficient CUR delivery. With its multifaceted anticancer activity, CUR holds promise as a cancer therapeutic. Recent studies concentrate on crafting nanocarriers tailored for size, charge, and functionalization, offering adaptable tools for combinational cancer therapy. The synergistic combination of CUR with chemotherapy, magnetic nano hyperthermia, or photodynamic therapy amplifies the efficacy of malignancy treatment. The investigation into CUR-loaded nanocarriers, whether used alone or in combination with other modalities, aims to enhance cancer treatment outcomes, highlighting the diverse potential of curcumin in contemporary therapeutic strategies. This research underscores the importance of combinational drug delivery therapies, providing a renewed perspective on the versatile applications of curcumin in modern medicine.

An Albumin-Holliday Junction Biomolecular Modular Design for Programmable Multifunctionality and Prolonged Circulation.

Combinatorial properties such as long-circulation and site- and cell-specific engagement need to be built into the design of advanced drug delivery systems to maximize drug payload efficacy. This work introduces a four-stranded oligonucleotide Holliday Junction (HJ) motif bearing functional moieties covalently conjugated to recombinant human albumin (rHA) to give a "plug-and-play" rHA-HJ multifunctional biomolecular assembly with extended circulation. Electrophoretic gel-shift assays show successful functionalization and purity of the individual high-performance liquid chromatography-purified modules as well as efficient assembly of the rHA-HJ construct. Inclusion of an epidermal growth factor receptor (EGFR)-targeting nanobody module facilitates specific binding to EGFR-expressing cells resulting in approximately 150-fold increased fluorescence intensity determined by flow cytometric analysis compared to assemblies absent of nanobody inclusion. A cellular recycling assay demonstrated retained albumin-neonatal Fc receptor (FcRn) binding affinity and accompanying FcRn-driven cellular recycling. This translated to a 4-fold circulatory half-life extension (2.2 and 0.55 h, for the rHA-HJ and HJ, respectively) in a double transgenic humanized FcRn/albumin mouse. This work introduces a novel biomolecular albumin-nucleic acid construct with extended circulatory half-life and programmable multifunctionality due to its modular design.

A Modular Albumin-Oligonucleotide Biomolecular Assembly for Delivery of Antisense Therapeutics.

Antisense nucleic acid drugs are susceptible to nuclease degradation, rapid renal clearance, and short circulatory half-life. In this work, we introduce a modular-based recombinant human albumin-oligonucleotide (rHA-cODN) biomolecular assembly that allows incorporation of a chemically stabilized therapeutic gapmer antisense oligonucleotide (ASO) and FcRn-driven endothelial cellular recycling. A phosphodiester ODN linker (cODN) was conjugated to recombinant human albumin (rHA) using maleimide chemistry, after which a complementary gapmer ASO, targeting ADAMTS5 involved in osteoarthritis pathogenesis, was annealed. The rHA-cODN/ASO biomolecular assembly production, fluorescence labeling, and purity were confirmed using polyacrylamide gel electrophoresis. ASO release was triggered by DNase-mediated degradation of the linker strand, reaching 40% in serum after 72 h, with complete release observed following 30 min of incubation with DNase. Cellular internalization and trafficking of the biomolecular assembly using confocal microscopy in C28/I2 cells showed higher uptake and endosomal localization by increasing incubation time from 4 to 24 h. FcRn-mediated cellular recycling of the assembly was demonstrated in FcRn-expressing human microvascular endothelial cells. ADAMTS5 in vitro silencing efficiency reached 40%, which was comparable to free gapmer after 72 h incubation with human osteoarthritis patients' chondrocytes. This work introduces a versatile biomolecular modular-based "Plug-and-Play" platform potentially applicable for albumin-mediated half-life extension for a range of different types of ODN therapeutics.

Bionanoengineered 2D monoelemental selenene for piezothrombolysis

Thrombosis-related diseases represent the leading causes of disability or death worldwide. However, conventional thrombolytic therapies are subjected to narrow therapeutic window, short circulation half-life and bleeding. Herein, we rationally design and develop a safe and efficient nonpharmaceutical thrombolysis strategy based on a specific piezocatalytic effect arising from platelet membrane (PM)-conjugated two-dimensional (2D) piezoelectric selenene, Se-PM nanosheets (NSs). The 2D selenene is fabricated from nonlayered bulk selenium powder by a facile liquid-phase exfoliation method, and the PM conjugation confers selenene with the distinct thrombus-homing feature. Under ultrasonic activation, the piezoelectric characteristic of selenene triggers electrons and holes separation, resulting in generation of reactive oxygen species (ROS) by reacting with surrounding H2O and O2 in the thrombosis microenvironment for thrombolysis. Both systematic in vitro and in vivo assessments demonstrate that the biocompatible Se-PM NSs efficiently degrade erythrocytes, fibrin and artificial blood clots under ultrasound irradiation. Compared to the clinical thrombolytic drug urokinase plasminogen activator, the engineered Se-PM NSs possess excellent thrombolytic efficacy by single treatment in the tail thrombosis animal model without bleeding risk. The engineered Se-PM nanoplatform marks an exciting jumping-off point for research into the application of piezocatalysis in clinical treatment of thrombosis.

Magnetic Resonance Angiography with Hour-Scale Duration after Single Low-Dose Administration of Biocompatible Gadolinium Oxide Nanoprobe.

Long-term contrast-enhanced angiography offers significant advantages in theranostics for diverse vascular diseases, particularly in terms of real-time dynamic monitoring during acute vascular events; However, achieving vascular imaging with a duration of hours through a single administration of low-dose contrast agent remains challenging. Herein, a hyaluronic acid-templated gadolinium oxide (HA@Gd2O3) nanoprobe-enhanced magnetic resonance angiography (MRA) is proposed to address this bottleneck issue for the first time. The HA@Gd2O3 nanoprobe synthesized from a facile one-pot biomineralization method owns ultrasmall size, good biocompatibility, optimal circulation half-life (≈149 min), and a relatively high T1 relaxivity (r1) under both clinical 3 T (8.215 mM-1s-1) and preclinical 9.4 T (4.023 mM-1s-1) equipment. The HA@Gd2O3 nanoprobe-enhanced MRA highlights major vessels readily with significantly improved contrast, extended imaging duration for at least 2 h, and ultrahigh resolution of 0.15mm under 9.4 T, while only requiring half clinical dosage of Gd. This technique can enable rapid diagnosis and real-time dynamic monitoring of vascular changes in a model of acute superior mesenteric vein thrombosis with only a single injection of nanoprobe. The HA@Gd2O3 nanoprobe-enhanced MRA provides a sophisticated approach for long-term (hour scale) vascular imaging with ultrahigh resolution and high contrast through single administration of low-dose contrast agent.

VISUALIZATION OF NANOPOROUS SILICA NANOPARTICLE DISTRIBUTION FOR IMPLANT-DIRECTED MAGNETIC DRUG TARGETING BY 68GA-LABELING AND PET/CT

In orthopedic surgery, implant infections are a serious issue and difficult to treat. The aim of this study was to use superparamagnetic nanoporous silica nanoparticles (MNPSNP) as candidates for directed drug delivery. Currently, short blood circulation half-life due to interactions with the host's immune system hinder nanoparticles in general from being clinically used. PEGylation is an approach to reduce these interactions and to enhance blood circulation time. The effect of PEGylation of the used 68Ga-labelled MNPSNP on the distribution and implant accumulation was examined by PET/CT imaging and gamma counting in an implant mouse model.Female Balb/c mice (n=24) received a magnetic implant subcutaneously on the left and a titanium implant on the right hind leg. On day one, 12 of these mice received an additional clodronate®-injection for macrophage depletion. On the second postoperative day, mice were anaesthetized and MNPSNP (native or PEGylated) injected intravenously, followed by a dynamic PET-scan over 60 minutes, a CT- and a static PET-scan at 120 min. As control, 12 mice received only 68Ga-MNPSNP (native or PEGylated). Gamma counting of inner organs, urine, blood and implant area was performed as further final analysis. Although PEGylation of the nanoparticles already resulted in lower liver uptakes, both variants of 68Ga-labeled MNPSNP accumulated in liver and spleen. Combination of PEGylation with clodronate®-injection led to a highly significant effect whereas clodronate®-injection alone could not reveal significant differences. In gamma counting, a significantly higher %I.D./g was found for the tissue surrounding the magnetic implants compared to the titanium control, although in a low range. PEGylation and/or clodronate®-injection revealed no significant differences regarding nanoparticle accumulation at the implantation site. PEGylation increases circulation time, but MNPSNP accumulation at the implant site was still insufficient for treatment of infections. Additional efforts have to further increase circulation time and local accumulation.Acknowledgements: This work is funded by the German Research Foundation (DFG, project number 280642759).

The Potential of Single-Chain Variable Fragment Antibody: Role in Future Therapeutic and Diagnostic Biologics.

The advancement of genetic engineering has revolutionized the field of immunology by allowing the utilization of intrinsic antibody structures. One of the biologics that are being produced by recombinant antibody technology is single-chain fragments variable (scFv). Genes of variable regions, the heavy and light chains that are genetically linked into a single transcript by a short flexible linker peptide, are used to generate this fragment from cellular and synthetic libraries. The specificity and affinity of these molecules are comparable to those of parental antibodies. Fusion with marker proteins and other potent molecules improves their stability, circulation half-life, activity, and efficient purification. Besides, this review comprises construction protocols, therapeutics, and diagnostic applications of scFv, as well as related challenges. Nonetheless, there are still issues with efficacy, stability, safety, intracellular administration, and production costs that need to be addressed.

Long-circulating nanoparticles as passive targeting nanocarriers for the treatment of thrombosis.

Thrombosis is the major cause of cardiovascular diseases. Only a small subset of patients could benefit from thrombolytic therapy due to the high bleeding risk brought about by the repeated administration of thrombolytic drugs. Nanoparticles with targeting ligands have been developed as nanocarriers of thrombolytic drugs to deliver the drug to the thrombus through active targeting. However, the passive targeting effect of nanoparticles on the thrombus is yet to be investigated. Herein, we prepared silica cross-linked micelles (SCLMs) with a long blood circulation half-life as drug carriers to target the thrombus through passive targeting. Compared with SCLMs modified with an active targeting ligand cRGD, the SCLMs exhibited similar targeting behavior to the thrombus in vivo. Loaded with the thrombolytic drug tirofiban, the passive targeting SCLMs showed a comparable therapeutic effect to cRGD-modified SCLMs in a mice model with pulmonary embolism and arterial thrombosis.

Revolutionizing Healthcare through Applications and Challenges of Nanomaterials in Targeted Drug Delivery and Therapeutics

This research investigates the cutting edge of healthcare advancement through the applications and challenges of nanomaterials focused on sedate conveyance and therapeutics. Gold nanoparticles (AuNPs) and liposomes were synthesized, characterized, and utilized as carriers for doxorubicin, illustrating remarkable medicate stacking capacities of 4.5% and 80.2%, individually. The discharge energy uncovered a Higuchi demonstration for AuNPs and first-order energy for liposomes, exhibiting custom-fitted medicate discharge profiles. In vitro studies illustrated critical cytotoxicity, with an IC50 of 12.3 µM for AuNPs in A549 cells and 8.7 µM for liposomes in MCF-7 cells. Cellular take-up rates of 35.6% and 50.2% advance asserted their adequacy. Moving to in vivo studies, the circulation half-life of AuNPs was 6.2 hours, whereas liposomes displayed a longer half-life of 8.5 hours. Bio distribution ponders illustrated particular aggregation within the tumor, coming to 4.8% ID/g for AuNPs and 6.2% ID/g for liposomes. Helpful results included tumor measure decrease and by and large survival rates of 75.4% for AuNPs and 82.7% for liposomes. Comparison with related works highlighted the competitive execution of the created nanomaterials, emphasizing their potential in progressing exactness medication. This research contributes to the advancing scene of nanomedicine, emphasizing personalized and maintainable healthcare arrangements.

Impacts of polyethylene glycol (PEG) dispersity on protein adsorption, pharmacokinetics, and biodistribution of PEGylated gold nanoparticles.

PEGylated gold nanoparticles (PEG-AuNPs) are widely used in drug delivery, imaging and diagnostics, therapeutics, and biosensing. However, the effect of PEG dispersity on the molecular weight (M W) distribution of PEG grafted onto AuNP surfaces has been rarely reported. This study investigates the effect of PEG dispersity on the M W distribution of PEG grafted onto AuNP surfaces and its subsequent impact on protein adsorption and pharmacokinetics, by modifying AuNPs with monodisperse PEG methyl ether thiols (mPEG n -HS, n = 36, 45) and traditional polydisperse mPEG2k-SH (M W = 1900). Polydisperse PEG-AuNPs favor the enrichment of lower M W PEG fractions on their surface due to the steric hindrance effect, which leads to increased protein adsorption. In contrast, monodisperse PEG-AuNPs have a uniform length of PEG outlayer, exhibiting markedly lower yet constant protein adsorption. Pharmacokinetics analysis in tumor-bearing mice demonstrated that monodisperse PEG-AuNPs possess a significantly prolonged blood circulation half-life and enhanced tumor accumulation compared with their polydisperse counterpart. These findings underscore the critical, yet often underestimated, impacts of PEG dispersity on the in vitro and in vivo behavior of PEG-AuNPs, highlighting the role of monodisperse PEG in enhancing therapeutic nanoparticle performance.

Novel Strategies for Spatiotemporal and Controlled BMP-2 Delivery in Bone Tissue Engineering.

Bone morphogenetic protein-2 (BMP-2) has been commercially approved by the Food and Drug Administration for use in bone defects and diseases. BMP-2 promotes osteogenic differentiation of mesenchymal stem cells. In bone tissue engineering, BMP-2 incorporated into scaffolds can be used for stimulating bone regeneration in organoid construction, drug testing platforms, and bone transplants. However, the high dosage and uncontrollable release rate of BMP-2 challenge its clinical application, mainly due to the short circulation half-life of BMP-2, microbial contamination in bone extracellular matrix hydrogel, and the delivery method. Moreover, in clinical translation, the requirement of high doses of BMP-2 for efficacy poses challenges in cost and safety. Based on these, novel strategies should ensure that BMP-2 is delivered precisely to the desired location within the body, regulating the timing of BMP-2 release to coincide with the bone healing process, as well as release BMP-2 in a controlled manner to optimize its therapeutic effect and minimize side effects. This review highlights improvements in bone tissue engineering applying spatiotemporal and controlled BMP-2 delivery, including molecular engineering, biomaterial modification, and synergistic therapy, aiming to provide references for future research and clinical trials.