ABSTRACT
INTRODUCTION: This review explores the innovative intersection of ferroptosis, a form of iron-dependent cell death, with cancer immunotherapy. Traditional cancer treatments face limitations in efficacy and specificity. Ferroptosis as a new paradigm in cancer biology, targets metabolic peculiarities of cancer cells and may potentially overcome such limitations, enhancing immunotherapy. AREA COVERED: This review centers on the regulation of ferroptosis by nanotechnology to augment immunotherapy. It explores how nanoparticle-modulated ferroptotic cancer cells impact the TME and immune responses. The dual role of nanoparticles in modulating immune response through ferroptosis are also discussed. Additionally, it investigates how nanoparticles can be integrated with various immunotherapeutic strategies, to optimize ferroptosis induction and cancer treatment efficacy. The literature search was conducted using PubMed and Google Scholar, covering articles published up to March 2024. EXPERT OPINION: The manuscript underscores the promising yet intricate landscape of ferroptosis in immunotherapy. It emphasizes the need for a nuanced understanding of ferroptosis' impact on immune cells and the TME to develop more effective cancer treatments, highlighting the potential of nanoparticles in enhancing the efficacy of ferroptosis and immunotherapy. It calls for deeper exploration into the molecular mechanisms and clinical potential of ferroptosis to fully harness its therapeutic benefits in immunotherapy.
Subject(s)
Ferroptosis , Immunotherapy , Nanoparticles , Neoplasms , Ferroptosis/drug effects , Humans , Neoplasms/therapy , Neoplasms/immunology , Neoplasms/drug therapy , Immunotherapy/methods , Animals , Nanotechnology , Tumor MicroenvironmentABSTRACT
Cytosolic delivery of proteins and peptides provides opportunities for effective disease treatment, as they can specifically modulate intracellular processes. However, most of protein-based therapeutics only have extracellular targets and are cell-membrane impermeable due to relatively large size and hydrophilicity. The use of organelle-targeting strategy offers great potential to overcome extracellular and cell membrane barriers, and enables localization of protein and peptide therapeutics in the organelles. Although progresses have been made in the recent years, organelle-targeted protein and peptide delivery is still challenging and under exploration. We reviewed recent advances in subcellular targeted delivery of proteins/peptides with a focus on targeting mechanisms and strategies, and highlight recent examples of active and passive organelle-specific protein and peptide delivery systems. This emerging platform could open a new avenue to develop more effective protein and peptide therapeutics.
ABSTRACT
The design of intracellular delivery systems for protein drugs remains a challenge due to limited delivery efficacy and serum stability. Herein, we propose a reversible assembly strategy to assemble cargo proteins and phenolic polymers into stable nanoparticles for this purpose using a heterobifunctional adaptor (2-formylbenzeneboronic acid). The adaptor is easily decorated on cargo proteins via iminoboronate chemistry and further conjugates with catechol-bearing polymers to form nanoparticles via boronate diester linkages. The nanoparticles exhibit excellent serum stability in culture media but rapidly release the cargo proteins triggered by lysosomal acidity and GSH after endocytosis. In a proof-of-concept animal model, the strategy successfully transports superoxide dismutase to retina via intravitreal injection and efficiently ameliorates the oxidative stress and cellular damage in the retina induced by ischemia-reperfusion (I/R) with minimal adverse effects. The reversible assembly strategy represents a robust and efficient method to develop serum-stable systems for the intracellular delivery of biomacromolecules.
Subject(s)
Nanoparticles , Polymers , Animals , Polymers/chemistry , Nanoparticles/chemistry , Humans , Superoxide Dismutase/metabolism , Superoxide Dismutase/chemistry , Drug Delivery Systems , Phenols/chemistry , Oxidative Stress/drug effects , Boronic Acids/chemistry , Retina/metabolism , MiceABSTRACT
Developing protein drugs that can target intracellular sites remains a challenge due to their inadequate membrane permeability. Efficient carriers for cytosolic protein delivery are required for protein-based drugs, cancer vaccines, and CRISPR-Cas9 gene therapies. Here, we report a screening process to identify highly efficient materials for cytosolic protein delivery from a library of dual-functionalized polymers bearing both boronate and lipoic acid moieties. Both ligands were found to be crucial for protein binding, endosomal escape, and intracellular protein release. Polymers with higher grafting ratios exhibit remarkable efficacies in cytosolic protein delivery including enzymes, monoclonal antibodies, and Cas9 ribonucleoprotein while preserving their activity. Optimal polymer successfully delivered Cas9 ribonucleoprotein targeting NLRP3 to disrupt NLRP3 inflammasomes in vivo and ameliorate inflammation in a mouse model of psoriasis. Our study presents a promising option for the discovery of highly efficient materials tailored for cytosolic delivery of specific proteins and complexes such as Cas9 ribonucleoprotein.
Subject(s)
CRISPR-Cas Systems , Gene Editing , Animals , Mice , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Gene Transfer Techniques , Genetic Therapy , Polymers/chemistry , Ribonucleoproteins/geneticsABSTRACT
Cytosine base editors (CBEs) are effective tools for introducing C-to-T base conversions, but their clinical applications are limited by off-target and bystander effects. Through structure-guided engineering of human APOBEC3A (A3A) deaminase, we developed highly accurate A3A-CBE (haA3A-CBE) variants that efficiently generate C-to-T conversion with a narrow editing window and near-background level of DNA and RNA off-target activity, irrespective of methylation status and sequence context. The engineered deaminase domains are compatible with PAM-relaxed SpCas9-NG variant, enabling accurate correction of pathogenic mutations in homopolymeric cytosine sites through flexible positioning of the single-guide RNAs. Dual adeno-associated virus delivery of one haA3A-CBE variant to a mouse model of tyrosinemia induced up to 58.1% editing in liver tissues with minimal bystander editing, which was further reduced through single dose of lipid nanoparticle-based messenger RNA delivery of haA3A-CBEs. These results highlight the tremendous promise of haA3A-CBEs for precise genome editing to treat human diseases.
ABSTRACT
Osteoarthritis (OA) is a common degenerative disease worldwide and new therapeutics that target inflammation and the crosstalk between immunocytes and chondrocytes are being developed to prevent and treat OA. These attempts involve repolarizing pro-inflammatory M1 macrophages into the anti-inflammatory M2 phenotype in synovium. In this study, we found that phosphoglycerate mutase 5 (PGAM5) significantly increased in macrophages in OA synovium compared to controls based on histology of human samples and single-cell RNA sequencing results of mice models. To address the role of PGAM5 in macrophages in OA, we found conditional knockout of PGAM5 in macrophages greatly alleviated OA symptoms and promoted anabolic metabolism of chondrocytes in vitro and in vivo. Mechanistically, we found that PGAM5 enhanced M1 polarization via AKT-mTOR/p38/ERK pathways, whereas inhibited M2 polarization via STAT6-PPARγ pathway in murine bone marrow-derived macrophages. Furthermore, we found that PGAM5 directly dephosphorylated Dishevelled Segment Polarity Protein 2 (DVL2) which resulted in the inhibition of ß-catenin and repolarization of M2 macrophages into M1 macrophages. Conditional knockout of both PGAM5 and ß-catenin in macrophages significantly exacerbated osteoarthritis compared to PGAM5-deficient mice. Motivated by these findings, we successfully designed mannose modified fluoropolymers combined with siPGAM5 to inhibit PGAM5 specifically in synovial macrophages via intra-articular injection, which possessed desired targeting abilities of synovial macrophages and greatly attenuated murine osteoarthritis. Collectively, these findings defined a key role for PGAM5 in orchestrating macrophage polarization and provides insights into novel macrophage-targeted strategy for treating OA.
Subject(s)
Osteoarthritis , Phosphoglycerate Mutase , Humans , Animals , Mice , beta Catenin , Osteoarthritis/genetics , Inflammation , Macrophages , Phosphoprotein Phosphatases , Mitochondrial ProteinsABSTRACT
Osteoclasts are over-activated as we age, which results in bone loss. Src deficiency in mice leads to severe osteopetrosis due to a functional defect in osteoclasts, indicating that Src function is essential in osteoclasts. G-protein-coupled receptors (GPCRs) are the targets for â¼35% of approved drugs but it is still unclear how GPCRs regulate Src kinase activity. Here, we reveal that GPR54 activation by its natural ligand Kisspeptin-10 (Kp-10) causes Dusp18 to dephosphorylate Src at Tyr 416. Mechanistically, Gpr54 recruits both active Src and the Dusp18 phosphatase at its proline/arginine-rich motif in its C terminus. We show that Kp-10 binding to Gpr54 leads to the up-regulation of Dusp18. Kiss1, Gpr54 and Dusp18 knockout mice all exhibit osteoclast hyperactivation and bone loss, and Kp-10 abrogated bone loss by suppressing osteoclast activity in vivo. Therefore, Kp-10/Gpr54 is a promising therapeutic target to abrogate bone resorption by Dusp18-mediated Src dephosphorylation.
Subject(s)
Bone Resorption , Osteoclasts , Animals , Mice , Osteoclasts/metabolism , Kisspeptins/genetics , Kisspeptins/metabolism , Receptors, G-Protein-Coupled/metabolism , src-Family Kinases/genetics , src-Family Kinases/metabolism , Mice, Knockout , Bone Resorption/genetics , Receptors, Kisspeptin-1ABSTRACT
The rising prevalence of global antibiotic resistance evokes the urgent need for novel antimicrobial candidates. Cationic lipopeptides have attracted much attention due to their strong antimicrobial activity, broad-spectrum and low resistance tendency. Herein, a library of fluoro-lipopeptide amphiphiles was synthesized by tagging a series of cationic oligopeptides with a fluoroalkyl tail via a disulfide spacer. Among the lipopeptide candidates, R6F bearing six arginine moieties and a fluorous tag shows the highest antibacterial activity, and it exhibits an interesting fluorine effect as compared to the non-fluorinated lipopeptides. The high antibacterial activity of R6F is attributed to its excellent bacterial membrane permeability, which further disrupts the respiratory chain redox stress and cell wall biosynthesis of the bacteria. By co-assembling with lipid nanoparticles, R6F showed high therapeutic efficacy and minimal adverse effects in the treatment of MRSA-induced sepsis and chronic wound infection. This work provides a novel strategy to design highly potent antibacterial peptide amphiphiles for the treatment of drug-resistant bacterial infections.
Subject(s)
Anti-Bacterial Agents , Methicillin-Resistant Staphylococcus aureus , Microbial Sensitivity Tests , Sepsis , Methicillin-Resistant Staphylococcus aureus/drug effects , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Sepsis/drug therapy , Sepsis/microbiology , Wound Infection/drug therapy , Wound Infection/microbiology , Animals , Mice , Staphylococcal Infections/drug therapy , Surface-Active Agents/chemistry , Surface-Active Agents/pharmacology , Surface-Active Agents/chemical synthesis , Lipopeptides/pharmacology , Lipopeptides/chemistry , Lipopeptides/therapeutic useABSTRACT
Ferroptosis as programmed cell death received considerable attention in cancer research. Recently, studies have associated ferroptosis with photodynamic therapy (PDT) because PDT promotes glutathione (GSH) deletion, glutathione peroxidase 4 (GPX4) degradation, and lipid peroxide accumulation. However, PDT-induced ferroptosis may be potentially prevented by ferroptosis suppressor protein 1 (FSP1). To address this limitation, herein, a novel strategy is developed to trigger ferroptosis by PDT and FSP1 inhibition. For enhancement of this strategy, a photoresponsive nanocomplex, self-assembled by BODIPY-modified poly(amidoamine) (BMP), is utilized to stably encapsulate the inhibitor of FSP1 (iFSP1) and chlorin e6 (Ce6). The nanosystem promotes intracellular delivery, penetration, and accumulation of ferroptosis inducers in tumors with light irradiation. The nanosystem presents high-performance triggering of ferroptosis and immunogenic cell death (ICD) in vitro and in vivo. Importantly, the nanoparticles increase tumor infiltration of CD8+ T cells and further enhance the efficacy of anti-PD-L1 immunotherapy. The study suggests the potential of photo-enhanced synergistic induction of ferroptosis by the photoresponsive nanocomplexes in cancer immunotherapy.
Subject(s)
Ferroptosis , Photochemotherapy , Photosensitizing Agents/pharmacology , Cell Line, Tumor , CD8-Positive T-Lymphocytes , ImmunotherapyABSTRACT
BACKGROUND: Autophagy related protease 4B (ATG4B) is a protease required for autophagy processing, which is strongly implicated in cancer progression. Phosphorylation of ATG4B is crucial for activation of its protease activity. However, little is known about the relationship of ATG4B and its phosphorylated form at Ser 383 and 392 sites (pS383/392-ATG4B), with clinical outcomes, particularly in colorectal cancer (CRC). METHODS: The ATG4B gene expression in CRC patients was obtained from The Cancer Genome Atlas (TCGA) database to analyze its clinical relevance. Tissue microarrays composed of 118 CRC patient specimens were used to determine the associations of ATG4B and pS383/392-ATG4B protein levels with prognosis. The biological functions of ATG4B in CRC cells were inspected with cell proliferation, mobility and spheroid culture assays. RESULTS: ATG4B gene expression was elevated in tumor tissues of CRC patients compared to that in adjacent normal tissues and high level of ATG4B expression was associated with poor survival. Similarly, protein levels of ATG4B and pS383/392-ATG4B were highly correlated with worse overall survival and disease-free survival. Stratification analysis results showed that high level of ATG4B had significantly higher risk of mortality in males and elderly patients compared to those female patients and patients 60 years or younger. In contrast, multivariate Cox's regression analysis indicated that high level of pS383/392-ATG4B was significantly linked to unfavorable overall survival and disease-free survival of males and elderly patients, whereas, it had no correlation with female patients and patients 60 years or younger. Moreover, high level of ATG4B was positively associated with increased mortality risk in patients with advanced AJCC stages (III and IV) and lymph node invasion (N1 and N2) for both overall survival and disease-free survival. Nevertheless, high level of pS383/392-ATG4B was positively correlated with increased mortality risk in patients with early AJCC stages (I and II) and without lymph node invasion (N0). In addition, silencing ATG4B attenuated migration, invasion, and further enhanced the cytotoxic effects of chemotherapeutic drugs in two and three-dimensional cultures of CRC cells. CONCLUSIONS: Our results suggest that ATG4B and pS383/392-ATG4B might be suitable biomarkers and therapeutic targets for CRC.
ABSTRACT
Fluorous tagged peptides have shown promising features for biomedical applications such as drug delivery and multimodal imaging. The bioconjugation of fluoroalkyl ligands onto cargo peptides greatly enhances their proteolytic stability and membrane penetration via a proposed "fluorine effect". The tagged peptides also efficiently deliver other biomolecules such as DNA and siRNA into cells via a co-assembly strategy. The fluoroalkyl chains on peptides with antifouling properties enable efficient gene delivery in the presence of serum proteins. Besides intracellular biomolecule delivery, the amphiphilic peptides can be used to stabilized perfluorocarbon-filled microbubbles for ultrasound imaging. The fluorine nucleus on fluoroalkyls provides intrinsic probes for background-free magnetic resonance imaging. Labeling of fluorous tags with radionuclide 18 F also allows tracing the biodistribution of peptides via positron emission tomography imaging. This mini-review will discuss properties and mechanism of the fluorous tagged peptides in these applications.
Subject(s)
Fluorine , Peptides , Fluorine/chemistry , Tissue Distribution , Peptides/chemistry , Positron-Emission Tomography , Drug Delivery SystemsABSTRACT
The design of efficient materials for intracellular protein delivery has attracted great interest in recent years; however, most current materials for this purpose are limited by poor serum stability due to the early release of cargoes triggered by abundant serum proteins. Here, we propose a light-activated crosslinking (LAC) strategy to prepare efficient polymers with excellent serum tolerance for intracellular protein delivery. A cationic dendrimer engineered with photoactivatable O-nitrobenzene moieties co-assembles with cargo proteins via ionic interactions, followed by light activation to yield aldehyde groups on the dendrimer and the formation of imine bonds with cargo proteins. The light-activated complexes show high stability in buffer and serum solutions, but dis-assemble under low pH conditions. As a result, the polymer successfully delivers cargo proteins green fluorescent protein and ß-galactosidase into cells with maintained bioactivity even in the presence of 50% serum. The LAC strategy proposed in this study provides a new insight to improve the serum stability of polymers for intracellular protein delivery.
ABSTRACT
Peptide drugs have been successfully used for the treatment of various diseases. However, it is still challenging to develop therapeutic peptides working on intracellular targets due to their poor membrane permeability. Here, we proposed a type of dual-responsive bioconjugates bearing a heterobifunctional adaptor containing both aldehyde and catechol moieties for efficient cytosolic peptide delivery. Hydrazine-terminated cargo peptides were tagged to a boronated dendrimer with the help of the adaptor via dynamic acylhydrazone and catecholboronate linkages. The bioconjugates efficiently delivered peptides with distinct physicochemical properties into various cells, and could release the cargo peptides triggered by intracellular reactive oxygen species and endolysosomal acidity, restoring the biofunctions of delivered peptides. In addition, the designed complexes efficiently delivered a pro-apoptotic peptide into osteosarcoma cancer cells and successfully inhibited the tumor growth both in vitro and in vivo. This study provides a universal and efficient platform for cytosolic therapeutic peptide delivery to intracellular targets for treating various diseases.
Subject(s)
Neoplasms , Peptides , Humans , Peptides/chemistry , Neoplasms/drug therapyABSTRACT
Rational design of efficient cytosolic protein delivery carriers holds enormous promise for biotherapeutics development. Several delivery systems have been developed during the past decades, while tailoring the balance between extracellular protein binding and intracellular cargo release is still challenging. In this study, we synthesized a series of oxygen-sensitive reactive polymers, rich in boron, by radical polymerization and post-modification for cytosolic protein delivery in vitro and in vivo. The introduction of boronate building blocks into the polymer scaffold significantly enhanced its protein binding affinity, and the polymer/protein complexes with high stability were obtained by tailoring the molecular ratios between the boronate ligands and the amine groups. The lead material screened from the polymer library exhibited efficient protein delivery efficacy that can release cargo proteins in cytosol in a reactive oxygen species responsive manner, which enables intracellular delivery of proteins with maintained bioactivity. In addition, the polymer-based nanoformulations efficiently delivered saporin, a toxin protein, into osteosarcoma cells and tumor tissues, and exhibited high therapeutic efficacy in an osteosarcoma mouse model. The synthesized polymer in this study can be developed as a promising nanocarrier for cytosolic delivery of protein therapeutics to treat a variety of diseases.
Subject(s)
Osteosarcoma , Polymers , Animals , Mice , Polymers/chemistry , Drug Carriers/chemistry , Reactive Oxygen Species , ProteinsABSTRACT
All-small-molecule dynamic hydrogels have shown great promise in cell culture, tissue engineering, and controlled drug release. The further development of more kinds of all-small-molecule dynamic hydrogels is severely hindered by the lack of enough commensurate building blocks from nature and on the market. Inspired by the widely developed metal-organic framework structures, herein we report a facile fabrication of metallogels by direct gelation of small molecular compounds including aminoglycosides (AGs), 2,2'-bipyridine-4,4'-dicarboxaldehyde (BIPY), and metal ions via coordination interactions and Schiff base reactions. These prepared metallogels exhibited good biodegradability and biosafety, excellent conductivity, tunable mechanical properties and potent antibacterial activities both in vitro and in vivo. This study provides a new strategy for expanding the scope of all-small-molecule dynamic metallogels for various biomedical applications.
Subject(s)
Hydrogels , Sepsis , Humans , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Anti-Bacterial Agents/chemistry , Metals , Tissue Engineering , Sepsis/drug therapyABSTRACT
Intracellular protein delivery has attracted considerable attention in the development of protein-based therapeutics, however, the design of highly efficient materials for robust delivery of native proteins remains challenging. This study proposes a Cu+ -based coordination polymer for cytosolic protein delivery with high efficacy and robustness. The phenylthiourea grafted dendrimer is coordinated with cuprous ions to prepare the polymeric carrier, which efficiently bind cargo proteins via a combination of coordination, ionic and hydrophobic interactions. The incorporation of Cu+ ions in the polymer greatly improves its cellular uptake and endosomal escape. The cuprous-based coordination polymer successfully delivered a variety of structurally diverse proteins into various cell lines with reserved bioactivities. This study provides a new type of coordination polymers for cytosolic delivery of biomacromolecules.
Subject(s)
Dendrimers , Dendrimers/chemistry , Endosomes/metabolism , Polymers/chemistry , Proteins/metabolism , Thiourea , Copper/chemistryABSTRACT
Polymers are one of the most promising protein delivery carriers; however, their applications are hindered by low delivery efficacy owing to their undesirable performance in protein binding, cellular uptake and endosomal escape. Here, we designed a series of histidine-based coordinative polymers for efficient intracellular protein delivery. Coordination of metal ions such as Ni2+, Zn2+, and Cu2+ with histidine residues on a polymer greatly improved its performance in protein binding, complex stability, cellular uptake and endosomal escape, therefore achieving highly improved protein delivery efficacy. Among the coordinative polymers, the Zn2+-coordinated one exhibited the highest cellular uptake, while the Cu2+-coordinated one exhibited the highest endosomal escape. The Ni2+-coordinated polymer formed large-sized aggregates with cargo proteins and showed insufficient protein release after endocytosis. The results obtained in this study provided new insight into the development of coordinative polymer-based protein delivery systems.
Subject(s)
Histidine , Polymers , Polymers/chemistry , Histidine/metabolism , Protein Binding , Endosomes/metabolism , Endocytosis , Proteins/metabolismABSTRACT
Intracellular protein delivery has been a major challenge due to various physiological barriers including low proteolytic stability and poor membrane permeability of the biologics. Nanoparticles were widely proposed to deliver cargo proteins into cells by endocytosis, however, the materials and complexes with proteins are often entrapped in endosomes and subject to lysosome degradation. In this study, we report a piperazine modified dendrimer for stabilizing the complexes via a combination of electrostatic interaction and hydrophobic interactions. The complexes show rapid cell internalization and the loaded proteins are released into the cytosols as early as half an hour post incubation. Mechanism study suggests that the complexes are endocytosed into cells via caveolae-based pathways, which could be inhibited by inhibitors such as genistein, filipin III, brefeldin A and nystatin. The phenylpiperazine-modified polymer enables the delivery of cargo proteins with reserved bioactivity and show high permeability in three-dimensional cell spheroids. The results prove the beneficial roles of phenylpiperazine ligands in polymer-mediated cytosolic protein delivery systems. STATEMENT OF SIGNIFICANCE: We synthesized a list of piperazine and derivatives modified dendrimers as cytosolic protein delivery vectors via facile reactions. Phenylpiperazine modification enables the efficient protein binding through the combination of electrostatic, hydrogen bonding and hydrophobic interactions. Phenylpiperazine modified dendrimers were internalized into the cells via a caveolae-based endo/lysosome-independent path and could release the cargo proteins into the cytosols as early as half an hour post incubation. Phenylpiperazine modified dendrimers delivered cargo proteins with reserved bioactivity and showed high permeability in three-dimensional cell spheroids.