Hepatocellular carcinoma (HCC) shows the highest morbidity among liver cancers which is characterized by genetic mutations in hepatocytes, leading to uncontrolled cell growth and proliferation. Current treatment include surgery, chemotherapy and immunotherapy; however, chemotherapeutics, which focus on single-targeted drug therapy, are still associated with certain limitations and may affect the treatment outcomes. Natural products also show the anticancer effect of HCC and hypotoxicity, but overall low activity of natural products limits their further application. miRNAs can modulate post-transcriptional functions of target genes. An increasing body of evidence has demonstrated that miRNAs are the key regulators in HCC by targeting different molecules in different signaling pathways. However, miRNAs are fragile and liable to catabolism by RNases in serum and other body fluids, and small molecules separated from natural products may have limited bio-availability. A chitosan based, targeted, sustained-release nanoparticle delivery miR-128-3p agomir (NA-miR-128-3p) was developed in this work. This nanoparticle was prepared by pentasodium tripolyphosphate (TPP), chitosan hydrochloride and miR-128-3p agomir with target aptamer which was loaded into the chitosan nanoparticle by self-assembly. It can intervene in HCC progress by affecting AKT1 expression. Based on this, a novel, efficient, long-acting, multi-mechanism and low-dosage combination drug delivery strategy was proposed in this work and showed a prominent anti-tumor effect. NA-miR-128-3p combined with natural product Oroxin B significantly affected HCC progression by the interference with VEGF and PI3K-AKT pathways, better than using NA-miR-128-3p and Oroxin B alone. Taken together, this nanoparticle and combinative administration compensate for the shortcomings of the fragile RNA drugs and the low activity of natural products, with high prospects in HCC treatment.

Efficient siRNA delivery is highly desirable for disease treatment. However, the application of conventional nanoparticles is limited by the inability to escape from endo-lysosomes. Herein, we report a strategy using small-molecule drugs to enhance siRNA endo‐lysosomal release,addressing this challenge. We encapsulated gentamicin(GM) into the marketed Onpattro® formulation to establish LNP-siRNA/GM nanoparticles that promote siRNA endo‐lysosomal escape through endosomal disruption, mechanistically exhibiting unique functionality and synergistic effects of LNP-siRNA/GM to improve cancer therapy. Besides, GM induced reactive oxygen species (ROS) and phospholipids accumulation in endo‐lysosomes, as well as the physical characteristics of lipid nanoparticles (LNPs) were preserved. We also revealed that GM causes endo‐lysosomal swelling and disrupts the endosomal membrane to enable siRNA release, as confirmed by Galectin 3 recruitment and acridine orange release. This approach achieved ∼81% mRNA-EGFR silencing, which is more than LNP-siEGFR (∼56.23%) by enhancing siRNA endo‐lysosomal escape efficiency. Meanwhile, LNP-siEGFR/GM exhibited significant biological activities in HepG2 cells, driven by the synergistic effects of siEGFR and GM with the VEGF and CXCL12 downregulation of, and ROS and phospholipids upregulation. Furthermore, tumor growth was notably suppressed after intravenous injection of LNP-siEGFR/GM in tumor-bearing nude mice. The combination of EGFR-siRNA and GM could also greatly inhibit angiogenesis, be antiproliferative, and induce tumor cells apoptosis. Therefore, this GM and siRNA co-delivery system would provide an efficient strategy for siRNA endosomal escape, significantly improving knockdown in various LNPs based siRNA delivery systems and efficiently enhancing cancer therapy.

Solid lipid nanoparticles (SLN) could enhance the oral bioavailability of loaded protein and peptide drugs through lymphatic transport. Natural oligopeptides regulate nearly all vital processes and serve as a nitrogen source for nourishment. They are mainly transported by oligopeptide transporter-1 (PepT-1) which are primarily expressed in the intestine with the characteristics of high-capacity and low energy consumption. Our preliminary research discovered the transmembrane transport of SLN could be improved by stimulating the oligopeptide absorption pathway. This implied the potential of combining the advantages of SLN with oligopeptide transporter mediated transportation. Herein, two kinds of dipeptide modified SLN were designed with insulin and glucagon like peptide-1 (GLP-1) analogue exenatide as model drugs. These drugs loaded SLN showed enhanced oral bioavailability and hypoglycemic effect in both type I diabetic C57BL/6 mice and type II diabetic KKAy mice. Compared with un-modified SLN, dipeptide-modified SLN could be internalized by intestinal epithelial cells via PepT-1-mediated endocytosis with higher uptake. Interestingly, after internalization, more SLN could access the systemic circulation via lymphatic transport pathway, highlighting the potential to combine the oligopeptide-absorption route with SLN for oral drug delivery.

Temozolomide (TMZ) is considered as a standard-of-care DNA alkylating agent for treating glioblastoma multiforme. Despite being a highly potent molecule, TMZ poses several limitations, including short half-life, rapid metabolism, low brain bioavailability and dose-dependent toxicities. Attempts have been made to improve the delivery of TMZ that mainly exhibited nominal therapeutic outcomes. In the current study, we have conjugated TMZ to mPEG-b-P(CB-{g-COOH}) copolymer to obtain mPEG-b-P(CB-{g-COOH; g-TMZ_n}) that demonstrated improvement in stability and efficacy. Further, a hybrid TMZ nanoconjugate formulation was developed using mPEG-b-P(CB-{g-COOH; g-TMZ₄₀}) and mPEG-polylactic acid (mPEG-PLA) showed an average size of 105.7 nm with narrow PDI of <0.2 and TMZ loading of 21.6%. Stability was assessed under physiological conditions wherein TMZ was found to be stable with a half-life of ∼194 h compared to 1.8 h for free TMZ. The Hybrid TMZ nanoconjugates showed improved intracellular uptake and reduced IC₅₀ values in C6 and U87MG glioma cells. Furthermore, they exhibited better in vivo therapeutic outcome, i.e., reduced brain weight, hemispherical width ratio and improved survival rate in C6-cell induced orthotropic glioma model in Sprague Dawley rats compared to the free TMZ-treated and positive control animals. Histopathological evaluation also revealed reduced cell infiltration in the lungs and reduced toxicity in major organs. Overall, the hybrid nanoconjugates of TMZ significantly improved its stability and efficacy in the GBM model, thereby opening newer avenues for treatment.

The effective intracellular accumulation of doxorubicin (DOX) is crucial for improving antitumor efficacy, which is severely impeded by limited drug penetration, uncontrollable drug release, and drug resistance. In this study, a thermal-deformative polymer embedding ultrasmall ceria (CeO₂) was rationally designed for deep tumor drug shuttling and hypoxia reversal to improve chemotherapy. Structurally, the CeO₂ nanozyme was covalently grafted with a polymer of p(NIPAM-co-AM) that could sharply shrink for DOX loading, which was consolidated with polydopamine (PDA) film encapsulation. Thereafter, a tumor penetration guide of apolipoprotein A-I (apoA-I) conjugated iRGD peptide (apoA-I-iRGD) was further decorated onto the PDA shell via Michael addition for preparing CeO₂P/DOX@iAPDA. With the aid of apoA-I-iRGD, CeO₂P/DOX@iAPDA penetrated both the tumor spheroids (∼78 µm) and the tumors of the mouse model deeply. After internalization by tumor cells and triggering by low pH in lysosomes, rapid DOX release was achieved by peeling off the PDA shell and thermosensitive deformation of p(NIPAM-co-AM). CeO₂P/DOX@iAPDA provided 66.4% tumor suppression in 4T1-derived tumor spheroids and 63.2% in 4T1-tumor-bearing mice, respectively. Preliminary mechanistic research involving western blotting and immunohistochemistry revealed that CeO₂P/DOX@iAPDA reversed resistance through the through HIF-1α-P-gp/lipid axis. Collectively, this study intelligently integrated CeO₂ nanozymes, temperature-sensitive polymers, and imitated biochemical modifications to improve chemotherapy for breast cancer.

Cancer stem cells (CSCs) are a major challenge in cancer therapy. Stem cell-like cells form a unique subpopulation within many tumors, which govern the degree of malignancy by promoting metastasis, recurrence, heterogeneity, and resistance to drug and radiation. Furthermore, these cells can persist in patients even after undergoing multiple cycles of conventional cancer therapy via dormancy, where they no longer dividing but remain active. These may cause cancer recurrence at any time, even years after a supposed cure, and remain invisible to the immune system. Targeting specific surface markers, signaling pathways and tumor microenvironment, which all have a significant effect on CSC function and maintenance, could help to eradicate CSCs and improve patient survival. Combinations of traditional therapies with nano-based drug delivery systems can efficiently target CSCs. Considering the biology and properties of CSCs, we classify recent approaches involving nanoparticle engineering, extracellular matrix modulation, cocktail strategies, multi-stage therapy, CSC defanging, Trojan horse systems, targeted therapy and organelle targeting. We highlight the most recent advances in nanocarrier design and drug delivery technologies to target CSCs, combined with conventional treatment in preclinical and clinical trials. The prospects of these approaches for CSCs elimination and recurrent cancer treatment are discussed.

Abnormal activation of macrophages and osteoclasts (OCs) contributes significantly to rheumatoid arthritis (RA) development by secretion of numerous inflammatory factors. Notably, these cells exhibit significant upregulation of folate receptor proteins on their surfaces. Unfortunately, there is a current lack of safe and effective therapeutic drugs for RA. Xuetongsu (XTS), a triterpenoid compound extracted from Kadsura heteroclita Roxb Craib, has demonstrated the ability to significantly inhibit the proliferation of RA fibroblast-like synoviocytes (RAFLS). However, its clinical application is hampered by poor targeting and short half-life. To address these drawbacks, we previously developed a nano-drug system named HRPS nanoparticles (NPs), which effectively targets RAFLS and inhibits synovial hyperplasia. However, this system overlooked the essential role of OCs in RA-related bone destruction. Therefore, we designed a novel folate-modified biomimetic Prussian blue (PB)-XTS NP (FMPX NP) for the selective delivery of XTS into inflammatory macrophages and OCs. The NP exhibits an excellent photothermal effect when assisted by laser irradiation, facilitating targeted release of XTS within inflammatory macrophages and OCs. The synergistic anti-inflammatory and reactive oxygen species scavenging effects of PB NPs and XTS are mediated by the inhibition of the NF-κB signaling pathway in inflammatory macrophages and RANK/RANKL/NFATc1 signaling pathway in OCs. In vivo experiments showed that FMPX NPs extended the half-life of XTS by 2.32 times, decreased hind foot swelling from 12.10 ± 0.49 mm to 8.24 ± 0.09 mm in the model group, and prevented bone damage. In conclusion, this study introduces a novel dual-targeted nano-based therapy for RA joints and highlights its potential for biochemical photothermal triple therapy for RA. FMPX NPs inhibit arthritis-related inflammation and bone destruction through a dual-target strategy, providing new insights for targeted drug therapies in clinical RA treatment.

Mimicking the hierarchical structure of the skin is one of the most important strategies in skin tissue engineering. Monolayer wound dressings are usually not able to provide several functions at the same time and cannot meet all clinical needs. In order to maximize therapeutic efficiency, herein, we fabricated a Tri-layer wound dressing, where the middle layer was fabricated via 3D-printing and composed of alginate, tragacanth and zinc oxide nanoparticles (ZnO NPs). Both upper and bottom layers were constructed using electrospinning technique; the upper layer was made of hydrophobic polycaprolactone to mimic epidermis, while the bottom layer consisted of Soluplus® and insulin-like growth factor-1 (IGF-1) to promote cell behavior. Swelling, water vapor permeability and tensile properties of the dressings were evaluated and the Tri-layer dressing exhibited impressive antibacterial activity and cell stimulation following by the release of ZnO NPs and IGF-1. Additionally, the Tri-layer dressing led to faster healing of full-thickness wound in rat model compared to monolayer and Bilayer dressings. Overall, the evidence confirmed that the Tri-layer wound dressing is extremely effective for full-thickness wound healing.

CRISPR-Cas system permanently deletes any harmful gene-of-interest to combat cancer growth. Chitosan (CS) is a potential cancer therapeutic that mediates via PI3K/Akt/mTOR, MAPK and NF-kβ signaling pathway modulation. CS and its covalent derivatives have been designed as nanocarrier of CRISPR-Cas9 alone (plasmid or ribonucleoprotein) or in combination with chemical drug for cancer treatment. The nanocarrier was functionalized with polyethylene glycol (PEG), targeting ligand, cell penetrating ligand and its inherent positive zeta potential to mitigate premature clearance and particulate aggregation, and promote cancer cell/nucleus targeting and permeabilization to enable CRISPR-Cas9 acting on the host DNA. Different physicochemical attributes are required for the CS-based nanocarrier to survive from the administration site, through the systemic circulation-extracellular matrix-mucus-mucosa axis, to the nucleus target. CRISPR-Cas9 delivery is met with heterogeneous uptake by the cancer cells. Choice of excipients such as targeting ligand and PEG may be inappropriate due to lacking overexpressed cancer receptor or availability of excessive metabolizing enzyme and immunoglobulin that defies the survival and action of these excipients rendering nanocarrier fails to reach the target site. Cancer omics analysis should be implied to select excipients which meet the pathophysiological needs, and chitosan nanocarrier with a “transformative physicochemical behavior” is essential to succeed CRISPR-Cas9 delivery.

Current treatments for glioblastoma face challenges such as the blood-brain barrier and lack of targeted therapy, compounded by the aggressive nature, high invasiveness, and heterogeneity of the disease. Exosomes, a subtype of extracellular vesicles are emerging as promising nanocarrier drug delivery systems to address these limitations. Exosomes released by all cell types can be easily obtained and modified as delivery vehicles or therapeutic agents. A systematic review was conducted to evaluate various methods for exosome isolation, characterization, engineering or modification, drug loading and delivery efficiency, including exosome biodistribution and treatment efficacy. A search of four databases for in vitro and in vivo studies (2000-,2023) identified 6165 records, of which 23 articles were found eligible and included for analyses. Most studies applied ultracentrifugation (UC) for exosomes isolation. Cancer cell lines being the most frequently used source of exosomes, followed by stem cells. The incubation approach was predominantly utilized to modify exosomes for drug loading. In vivo analysis showed that exosome biodistribution was primarily concentrated in the brain region, peaking in the first 6 h and remained moderately high. Compared to native exosomes and untreated control groups, utilizing modified native exosomes (cargo loaded) for treating glioblastoma disease models led to more pronounced suppression of tumor growth and proliferation, enhanced stimulation of immune response and apoptosis, effective restoration of drug chemosensitivity, increased anti-tumor effect and prolonged survival rates. Modified exosomes whether through incubation, sonication, transfection, freeze-thawing or their combination, improve targeted delivery and therapeutic efficacy against glioblastoma.

Microneedle-mediated drug delivery systems (MDDS) have experienced robust growth in recent years, with designers leveraging their creativity to apply these systems for direct drug delivery to the skin, mucous membranes, blood vessel walls and even internal organs. In order to achieve precise drug delivery, various delicately conceived drug release modes based on MDDS have been developed. Herein, to elucidate the design concepts of numerous reported MDDS, we have categorized them into two levels (Level-Ⅰ MDDS and Level-Ⅱ MDDS) depending on whether nanoscale and microscale carriers are integrated within the microneedles. In this work, the design strategies of MDDS, as well as the current status of their applications in targeted and intelligent drug delivery were reviewed, while their prospects and challenges for future industrialization and clinical applications were also discussed.