Toolbox of Biodegradable Dendritic (Poly glycerol sulfate)-SS-poly(ester) Micelles for Cancer Treatment: Stability, Drug Release, and Tumor Targeting.

In this paper, we present well-defined dPGS-SS-PCL/PLGA/PLA micellar systems demonstrating excellent capabilities as a drug delivery platform in light of high stability and precise in vitro and in vivo drug release combined with active targetability to tumors. These six amphiphilic block copolymers were each targeted in two different molecular weights (8 or 16 kDa) and characterized using 1H NMR, gel permeation chromatography (GPC), and elemental analysis. The block copolymer micelles showed monodispersed size distributions of 81-187 nm, strong negative charges between -52 and -41 mV, and low critical micelle concentrations (CMCs) of up to 1.13-3.58 mg/L (134-527 nM). The serum stability was determined as 94% after 24 h. The drug-loading efficiency for Sunitinib ranges from 38 to 83% (8-17 wt %). The release was selectively triggered by glutathione (GSH) and lipase, reaching 85% after 5 days, while only 20% leaching was observed under physiological conditions. Both the in vitro and in vivo studies showed sustained release of Sunitinib over 1 week. CCK-8 assays on HeLa lines demonstrated the high cell compatibility (1 mg/mL, 94% cell viability, 48 h) and the high cancer cell toxicity of Sunitinib-loaded micelles (IC50 2.5 µg/mL). By in vivo fluorescence imaging studies on HT-29 tumor-bearing mice, the targetability of dPGS7.8-SS-PCL7.8 enabled substantial accumulation in tumor tissue compared to nonsulfated dPG3.9-SS-PCL7.8. As a proof of concept, Sunitinib-loaded dPGS-SS-poly(ester) micelles improved the antitumor efficacy of the chemotherapeutic. A tenfold lower dosage of loaded Sunitinib led to an even higher tumor growth inhibition compared to the free drug, as demonstrated in a HeLa human cervical tumor-bearing mice model. No toxicity for the organism was observed, confirming the good biocompatibility of the system.

Recent Advances of Polycationic siRNA Vectors for Cancer Therapy.

Small interfering RNAs (siRNAs) have recently emerged as a new class of biopharmaceuticals for the treatment of various diseases, including genetic diseases, viral infections, heritable disorders, and most prominently, cancer. However, clinical applications of siRNA-based therapeutics through intravenous administration have been limited due to their rapid degradation and renal clearance, poor cellular uptake, low cytoplasmic release by escaping endocytic uptake, and off-target effects. The success of siRNA-based therapeutics depends upon the design and creation of efficient delivery vectors that should be able to protect siRNA from in vivo degradation and specifically deliver siRNA to cytosol of target cells. Over the past decade, myriad types of carrier systems composed of cationic polymers have been designed for delivery of siRNA to tumor cells. In this review, we overview recent advances in siRNA delivery by using these promising nonviral carrier systems in diverse approaches to overcome the delivery hindrances and provide valuable understanding to direct the future design of siRNA delivery carriers.

Ligand-directed active tumor-targeting polymeric nanoparticles for cancer chemotherapy.

In recent years, polymeric nanoparticles have appeared as a most viable and versatile delivery system for targeted cancer therapy. Various in vivo studies have demonstrated that virus-sized stealth particles are able to circulate for a prolonged time and preferentially accumulate in the tumor site via the enhanced permeability and retention (EPR) effect (so-called "passive tumor-targeting"). The surface decoration of stealth nanoparticles by a specific tumor-homing ligand, such as antibody, antibody fragment, peptide, aptamer, polysaccharide, saccharide, folic acid, and so on, might further lead to increased retention and accumulation of nanoparticles in the tumor vasculature as well as selective and efficient internalization by target tumor cells (termed as "active tumor-targeting"). Notably, these active targeting nanoparticulate drug formulations have shown improved, though to varying degrees, therapeutic performances in different tumor models as compared to their passive targeting counterparts. In addition to type of ligands, several other factors such as in vivo stability of nanoparticles, particle shape and size, and ligand density also play an important role in targeted cancer chemotherapy. In this review, concept and recent development of polymeric nanoparticles conjugated with specific targeting ligands, ranging from proteins (e.g., antibodies, antibody fragments, growth factors, and transferrin), peptides (e.g., cyclic RGD, octreotide, AP peptide, and tLyp-1 peptide), aptamers (e.g., A10 and AS1411), polysaccharides (e.g., hyaluronic acid), to small biomolecules (e.g., folic acid, galactose, bisphosphonates, and biotin), for active tumor-targeting drug delivery in vitro and in vivo are highlighted and discussed. With promise to maximize therapeutic efficacy while minimizing systemic side effects, ligand-mediated active tumor-targeting treatment modality has become an emerging and indispensable platform for safe and efficient cancer therapy.

PVA-based bulk microneedles capable of high insulin loading and pH-triggered degradation for multi-responsive and sustained hypoglycemic therapy.

"Closed-loop" insulin-loaded microneedle patche shows great promise for improving therapeutic outcomes and life quality for diabetes patients. However, it is typically hampered by limited insulin loading capacity, random degradation, and intricate preparation procedures for the independence of the "closed-loop" bulk microneedles. In this study, we combined the solubility of microneedles and "closed-loop" systems and designed poly(vinyl alcohol)-based bulk microneedles (MNs@GI) through in situ photopolymerization for multi-responsive and sustained hypoglycemic therapy, which significantly simplified the preparation process and improved insulin loading. GOx/insulin co-encapsulated MNs@GI with a phenylboronic ester structure improved glycemic responsiveness to control the insulin release under high glucose conditions and reduced inflammation risk in the normal skin. MNs@GI could further degrade to increase insulin release due to the crosslinked acetal-linkage hydrolysis in the presence of gluconic acid, which was caused by GOx-mediated glucose-oxidation in a hyperglycemic environment. The in vivo results showed that MNs@GI effectively regulated glycemic levels within the normal range for approximately 10 h compared to that of only insulin-loaded microneedles (MNs@INS). Consequently, the highly insulin-loaded, multi-responsive, and pH-triggered MN system has tremendous potential for diabetes treatment.

Bioresponsive Nanoparticles Boost Starvation Therapy and Prevent Premetastatic Niche Formation for Pulmonary Metastasis Treatment.

In the process of tumor metastasis, tumor cells can acquire invasion by excessive uptake of nutrients and energy and interact with the host microenvironment to shape a premetastatic niche (PMN) that facilitates their colonization and progression in the distal sites. Pyruvate is an essential nutrient that engages in both energy metabolism and remodeling of the extracellular matrix (ECM) in the lungs for PMN formation, thus providing a target for tumor metastasis treatment. There is a paucity of strategies focusing on PMN prevention, which is key to metastasis inhibition. Here, we design a bioresponsive nanoparticle (HP/GU) based on a disulfide-cross-linked hyperbranched polyethylenimine (D-PEI) core and a hyaluronic acid (HA) shell with a reactive oxygen species (ROS)-sensitive cross-linker between them to encapsulate glucose oxidase (GOX) and a mitochondrial pyruvate carrier (MPC) inhibitor via electrostatic interaction, which reinforces starvation therapy and reduces PMN formation in the lungs via inhibiting pyruvate metabolism. In tumor cells, GOX and MPC inhibitors can be rapidly released and synergistically reduce the energy supply of tumor cells by consuming glucose and inhibiting pyruvate uptake to decrease tumor cell invasion. MPC inhibitors can also reduce ECM remodeling by blocking cellular pyruvate metabolism to prevent PMN formation. Consequently, HP/GU achieves an efficient inhibition of both primary and metastatic tumors and provides an innovative strategy for the treatment of tumor metastases.

Ursodeoxycholic Acid-Decorated Zwitterionic Nanoparticles for Orally Liver-Targeted Close-Looped Insulin Delivery.

Oral insulin therapies targeting the liver and further simulating close-looped secretion face significant challenges due to multiple trans-epithelial barriers. Herein, ursodeoxycholic acid (UDCA)-decorated zwitterionic nanoparticles (NPs) (UC-CMs@ins) are designed to overcome these barriers, target the liver, and respond to glycemia, thereby achieving oral one-time-per-day therapy. UC-CMs@ins show excellent mucus permeability through the introduction of zwitterion (carboxy betaine, CB). Furthermore, UC-CMs@ins possess superior cellular internalization via proton-assisted amino acid transporter 1 (PAT1, CB-receptor) and apical sodium-dependent bile acid transporter (ASBT, UDCA-receptor) pathways. Moreover, UC-CMs@ins exhibit excellent endolysosomal escape ability and improve the basolateral release of insulin into the bloodstream via the ileal bile acid-binding protein and the heteromeric organic solute transporter (OSTα- OSTß) routes compared with non-UDCA-decorated C-CMs@ins. Therefore, CB and UDCA jointly overcome mucus and intestinal barriers. Additionally, UC-CMs@ins prevent insulin degradation in the gastrointestinal tract for crosslinked structure, improve insulin accumulation in the liver for UDCA introduction, and effectively regulate glycemia for "closed-loop" glucose control. Surprisingly, oral ingestion of UC-CMs@ins shows a superior effect on glycemia (≈22 h, normoglycemia) and improves postprandial glycemic levels in diabetic mice, illustrating the enormous potential of the prepared NPs as a platform for oral insulin administration in diabetes treatment.

ROS-responsive core-shell nano-inhibitor impedes pyruvate metabolism for reinforced photodynamic therapy and interrupted pre-metastatic niche formation.

The formation of pre-metastatic niche (PMN) in a hospitable organ derived from the primary tumor requires the communication between the tumor cells and the host environment. Pyruvate is a fundamental nutrient by which the tumor cells metabolically reshape the extracellular matrix in the lung to facilitate their own metastatic development. Here we report a combination regimen by integrating the photo-sensitizer and the mitochondrial pyruvate carrier (MPC) inhibitor in a dendritic polycarbonate core-hyaluronic acid shell nano-platform with multivalent reversible crosslinker embedded in it (DOH-NI+L) to reinforce photodynamic therapy (PDT) toward the primary tumor and interrupt PMN formation in the lung via impeding pyruvate uptake. We show that DOH-NI+L mediates tumor-specific MPC inhibitor liberation, inhibiting the aerobic respiration for facilitated PDT and restraining ATP generation for paralyzing cell invasion. Remarkably, DOH-NI+L is demonstrated to block the metabolic crosstalk of tumor cell-host environment by dampening pyruvate metabolism, provoking a series of metabolic responses and resulting in the pulmonary PMN interruption. Consequently, DOH-NI+L realizes a significant primary tumor inhibition and an efficient pulmonary metastasis prevention. Our research extends nano-based anti-metastatic strategies aiming at PMN intervention and such a dendritic core-shell nano-inhibitor provides an innovative paradigm to inhibit tumor growth and prevent metastasis efficiently. STATEMENT OF SIGNIFICANCE: In the progression of cancer metastasis, the formation of a pre-metastatic niche (PMN) in a hospitable organ derived from the primary tumor is one of the rate-limiting stages. The current nano-based anti-metastatic modalities mainly focus on targeted killing of tumor cells and specific inhibition of tumor cell invasion, while nanomedicine-mediated interruption of PMN formation has been rarely reported. Here we report a combination regimen by integrating a photo-sensitizer and an inhibitor of mitochondrial pyruvate carrier in a dendritic core-shell nano-platform with a reversible crosslinker embedded in it to reinforce PDT toward the primary tumor and interrupt PMN formation via impeding the uptake of pyruvate that is a fundamental nutrient facilitating aerobic respiration and PMN formation. Our research proposed a nano-based anti-metastatic strategy aiming at PMN intervention.

Photothermal-controlled NO-releasing Nanogels reverse epithelial-mesenchymal transition and restore immune surveillance against cancer metastasis.

Metastasis leads to high mortality among cancer patients. It is a complex, multi-step biological process that involves the dissemination of cancer cells from the primary tumor and their systemic spread throughout the body, primarily through the epithelial-mesenchymal transition (EMT) program and immune evasion mechanisms. It presents a challenge in how to comprehensively treat metastatic cancer cells throughout the entire stage of the metastatic cascade using a simple system. Here, we fabricate a nanogel (HNO-NG) by covalently crosslinking a macromolecular nitric oxide (NO) donor with a photothermal IR780 iodide-containing hyaluronic acid derivative via a click reaction. This enables stable storage and tumor-targeted, photothermia-triggered release of NO to combat tumor metastasis throughout all stages. Upon laser irradiation (HNO-NG+L), the surge in NO production within tumor cells impairs the NF-κB/Snail/RKIP signaling loop that promotes the EMT program through S-nitrosylation, thus inhibiting cell dissemination from the primary tumor. On the other hand, it induces immunogenic cell death (ICD) and thereby augments anti-tumor immunity, which is crucial for killing both the primary tumor and systemically distributed tumor cells. Therefore, HNO-NG+L, by fully leveraging EMT reversal, ICD induction, and the lethal effect of NO, achieved impressive eradication of the primary tumor and significant prevention of lung metastasis in a mouse model of orthotropic 4T1 breast tumor that spontaneously metastasizes to the lungs, extending the NO-based therapeutic approach against tumor metastasis.

Thiolated polyglycerol sulfate as potential mucolytic for muco-obstructive lung diseases.

Increased disulfide crosslinking of secreted mucins causes elevated viscoelasticity of mucus and is a key determinant of mucus dysfunction in patients with cystic fibrosis (CF) and other muco-obstructive lung diseases. In this study, we describe the synthesis of a novel thiol-containing, sulfated dendritic polyglycerol (dPGS-SH), designed to chemically reduce these abnormal crosslinks, which we demonstrate with mucolytic activity assays in sputum from patients with CF. This mucolytic polymer, which is based on a reportedly anti-inflammatory polysulfate scaffold, additionally carries multiple thiol groups for mucolytic activity and can be produced on a gram-scale. After a physicochemical compound characterization, we compare the mucolytic activity of dPGS-SH to the clinically approved N-acetylcysteine (NAC) using western blot studies and investigate the effect of dPGS-SH on the viscoelastic properties of sputum samples from CF patients by oscillatory rheology. We show that dPGS-SH is more effective than NAC in reducing multimer intensity of the secreted mucins MUC5B and MUC5AC and demonstrate significant mucolytic activity by rheology. In addition, we provide data for dPGS-SH demonstrating a high compound stability, low cytotoxicity, and superior reaction kinetics over NAC at different pH levels. Our data support further development of the novel reducing polymer system dPGS-SH as a potential mucolytic to improve mucus function and clearance in patients with CF as well as other muco-obstructive lung diseases.

Acetal-linked paclitaxel prodrug micellar nanoparticles as a versatile and potent platform for cancer therapy.

Endosomal pH-activatable paclitaxel (PTX) prodrug micellar nanoparticles were designed and prepared by conjugating PTX onto water-soluble poly(ethylene glycol)-b-poly(acrylic acid) (PEG-PAA) block copolymers via an acid-labile acetal bond to the PAA block and investigated for potent growth inhibition of human cancer cells in vitro. PTX was readily conjugated to PEG-PAA with high drug contents of 21.6, 27.0, and 42.8 wt % (denoted as PTX prodrugs 1, 2, and 3, respectively) using ethyl glycol vinyl ether (EGVE) as a linker. The resulting PTX conjugates had defined molecular weights and self-assembled in phosphate buffer (PB, pH 7.4, 10 mM) into monodisperse micellar nanoparticles with average sizes of 158.3-180.3 nm depending on PTX contents. The in vitro release studies showed that drug release from PTX prodrug nanoparticles was highly pH-dependent, in which ca. 86.9%, 66.4% and 29.0% of PTX was released from PTX prodrug 3 at 37 °C in 48 h at pH 5.0, 6.0, and pH 7.4, respectively. MTT assays showed that these pH-sensitive PTX prodrug nanoparticles exhibited high antitumor effect to KB and HeLa cells (IC(50) = 0.18 and 0.9 µg PTX equiv/mL, respectively) as well as PTX-resistant A549 cells. Notably, folate-decorated PTX prodrug micellar nanoparticles based on PTX prodrug 3 and 20 wt % folate-poly(ethylene glycol)-b-poly(D,L-lactide) (FA-PEG-PLA) displayed apparent targetability to folate receptor-overexpressing KB cells with IC(50) over 12 times lower than nontargeting PTX prodrug 3 under otherwise the same conditions. Furthermore, PTX prodrug nanoparticles could also load doxorubicin (DOX) to simultaneously release PTX and DOX under mildly acidic pH. These acetal-linked PTX prodrug micellar nanoparticles have appeared as a highly versatile and potent platform for cancer therapy.

Gold nanorod-cored biodegradable micelles as a robust and remotely controllable doxorubicin release system for potent inhibition of drug-sensitive and -resistant cancer cells.

Gold nanorod-cored biodegradable micelles were prepared by coating gold nanorods (AuNRs) with lipoylated poly(ethylene glycol)-b-poly(ε-caprolactone) (PEG-PCL-LA) block copolymer and investigated for remotely triggered release of doxorubicin (DOX) and effective inhibition of drug-sensitive and multidrug-resistant (MDR) cancer cells. The micelles had uniform sizes and excellent colloidal stability. The in vitro release studies showed that drug release from DOX-loaded AuNR-cored micelles (AuNR-M-DOX) was minimal under physiological conditions but markedly enhanced upon NIR irradiation at a low power density of 0.2 W/cm2, most likely due to photothermally induced phase transition of PCL regime. As revealed by confocal microscopy and flow cytometry, NIR could also trigger effective DOX release in drug-sensitive as well as drug-resistant MCF-7 cells. MTT assays showed that antitumor activity of AuNR-M-DOX to drug-sensitive MCF-7 cells was significantly boosted by mild NIR irradiation, reaching a comparable level to free DOX. Most remarkably, AuNR-M-DOX combined with NIR irradiation could also effectively kill drug-resistant MCF-7 cells, in which a cell viability of 38% was observed at a DOX dosage of 10 µg equiv/mL, whereas 100% cell viability was maintained for cells treated with free DOX under otherwise the same conditions. These AuNR-cored biodegradable micelles with high stability, photo-triggered drug release, and effective reversal of multidrug resistance in cancer cells have appeared as a novel platform for targeted cancer therapy.

Ligand-directed reduction-sensitive shell-sheddable biodegradable micelles actively deliver doxorubicin into the nuclei of target cancer cells.

The therapeutic performance of biodegradable micellar drugs is far from optimal due to existing challenges like poor tumor cell uptake and intracellular drug release. Here, we report on ligand-directed reduction-sensitive shell-sheddable biodegradable micelles based on poly(ethylene glycol)-poly(ε-caprolactone) (PEG-PCL) copolymer actively delivering doxorubicin (DOX) into the nuclei of target cancer cells, inducing superb in vitro antitumor effects. The micelles were constructed from PEG-SS-PCL and galactose-PEG-PCL (Gal-PEG-PCL) block copolymers, in which Gal-PEG-PCL was designed with a longer PEG than that in PEG-SS-PCL (6.0 vs 5.0 kDa) to fully expose Gal ligands onto the surface of micelles for effective targeting to hepatocellular carcinoma cells. PEG-SS-PCL combining with 10 or 20 wt % of Gal-PEG-PCL formed uniform micelles with average sizes of 56.1 and 58.2 nm (denoted as PEG-SS-PCL/Gal10 and PEG-SS-PCL/Gal20, respectively). The in vitro release studies showed that about 81.1 and 75.0% DOX was released in 12 h from PEG-SS-PCL/Gal10 and PEG-SS-PCL/Gal20 micelles under a reducing condition containing 10 mM dithiothreitol (DTT). In contrast, minimal DOX release (<12%) was observed for PEG-SS-PCL/Gal10 and PEG-SS-PCL/Gal20 micelles under nonreducing conditions as well as for reduction-insensitive Gal-PEG-PCL and PEG-PCL/Gal20 micelles in the presence of 10 mM DTT. MTT assays in HeLa and HepG2 cells showed that DOX-loaded PEG-SS-PCL/Gal20 micelles exhibited apparent targetability and significantly enhanced antitumor efficacy toward asialoglycoprotein receptor (ASGP-R)-overexpressing HepG2 cells with a particularly low half maximal inhibitory concentration (IC50) of 1.58 µg DOX equiv/mL, which was comparable to free DOX and approximately six times lower than that for nontargeting PEG-SS-PCL counterparts under otherwise the same conditions. Interestingly, confocal microscopy observations using FITC-labeled PEG-SS-PCL/Gal20 micelles showed that DOX was efficiently delivered and released into the nuclei of HepG2 cells in 8 h. Flow cytometry revealed that cellular DOX level in HepG2 cells treated with DOX-loaded PEG-SS-PCL/Gal20 micelles was much greater than that with reduction-insensitive PEG-PCL/Gal20 and nontargeting PEG-SS-PCL controls, signifying the importance of combining shell-shedding and active targeting. Ligand-directed, reduction-sensitive, shell-sheddable, and biodegradable micelles have emerged as a versatile and potent platform for targeted cancer chemotherapy.

Super-small zwitterionic micelles enable the improvement of blood-brain barrier crossing for efficient orthotopic glioblastoma combinational therapy.

Glioblastoma multiforme (GBM) remains incurable in clinical, nanotechnology-based drug delivery strategies show promising perspective in alleviating GBM, while limited blood-brain-barrier (BBB) permeation, short blood half-live accompanied by the poor tumor accumulation and penetration, significantly restrict the therapeutic outcomes. Herein, a versatile super-small zwitterionic nano-system (MCB(S)) based on carboxybetaine (CB) zwitterion functionalized hyperbranched polycarbonate (HPCB) is developed to overcome the brain delivery challenges. After grafting with amino-functionalized IR780 (free IR780), the ultimate paclitaxel (PTX)-encapsulated micelles (MCB(S)-IR@PTX) are precisely activated by near-infrared (NIR) for accelerated drug release and effective combinational GBM therapy. Importantly, MCB(S)-IR@PTX with the crosslinked structure and CB zwitterion prolongs blood-circulation, and CB-zwitterion further facilitates BBB-traversing through betaine/γ-aminobutyric acid (GABA) transporter-1 (BGT-1) pathway. Combined with the benefit of super small-size, MCB(S)-IR@PTX highly accumulates at tumor sites and penetrates deeply, thus efficiently inhibiting tumor growth and strikingly improving survival time in U87MG orthotopic GBM-bearing mouse model. The ingenious nanoplatform furnishes a versatile strategy for delivering therapeutics into the brain and realizing efficient brain cancer therapy.

Macromolecular NO-Donor Micelles for Targeted and Augmented Chemotherapy against Prostate Cancer.

Mitoxantrone (MTO) is clinically utilized for treating hormone-refractory prostate cancer (PCa), however, the therapeutic outcome is far from optimal due to the lack of proper drug carrier as well as the inherent MTO detoxification mechanisms of DNA lesion repair and anti-oxidation. Herein, a bombesin-installed nanoplatform combining the chemotherapeutic MTO and the chemotherapeutic sensitizer of nitric oxide (NO) is developed based on MTO-loaded macromolecular NO-donor-containing polymeric micelles (BN-NMMTO ) for targeted NO-sensitized chemotherapy against PCa. BN-NMMTO actively target and accumulates in PCa sites and are internalized into the tumor cells. The macromolecular NO-donor of BN-NMMTO undergoes a reductive reaction to unleash NO upon intracellular glutathione (GSH), accompanying by micelle swelling and MTO release. The targeted intracellular MTO release induces DNA lesion and reactive oxygen species (ROS) generation in tumor cells without damage to the normal cells, and MTO's cytotoxicity is further augmented by NO release via the inhibition of both DNA repair and anti-oxidation pathways as compared with traditional MTO therapies.

Reversibly "double locked" hydroxycamptothecin prodrug nanoparticles for targeted chemotherapy of lung cancer.

Prodrug assembled nanoparticles integrate the merits of both prodrug and nanoparticle, which significantly improve pharmacokinetic parameters, enhance tumorous accumulation and decrease adverse effects, while they are challenged by disassembly upon dilution in blood, masking the superiority of nanoparticles (NPs). Herein, a reversibly "double locked" hydroxycamptothecin (HCPT) prodrug nanoparticle decorated with cyclic RGD peptide (cRGD) is developed for safe and efficient chemotherapy of orthotopic lung cancer in mice. HCPT prodrug is constructed from acetal (ace)-linked cRGD-PEG-ace-HCPT-ace-acrylate polymer, which is self-assembled into the nanoparticles with "the first lock" of HCPT. Then the nanoparticles undergo the in situ UV-crosslinking of the acrylate residues for constructing "the second lock" of HCPT. The obtained "double locked" nanoparticles (T-DLHN) with simple and well-defined construction are demonstrated to possess extremely high stability against 100-fold dilution and acid-triggered "unlock" including de-crosslinking and liberation of the pristine HCPT. In an orthotopic lung tumor of mouse model, T-DLHN reveals a prolonged circulation time of about 5.0 h, superb lung tumor-homing capacity with tumorous drug uptake of about 7.15%ID/g, resulting in significantly boosted anti-tumor activity and reduced adverse effects. Hence, these nanoparticles utilizing "double lock" and acid-triggered "unlock" strategies represent a unique and promising nanoplatform for safe and efficient drug delivery. STATEMENT OF SIGNIFICANCE: Prodrug assembled nanoparticles have the unique properties of the well-defined structure, systemic stability, improved pharmacokinetics, passive targeting and decreased adverse effects. However, prodrug assembled NPs would disassemble against extensive dilution in the blood circulation when intravenously injected into the body. Herein, we have designed a cRGD-directed reversibly "double-locked" HCPT prodrug nanoparticle (T-DLHN) for safe and efficient chemotherapy of orthotopic A549 human lung tumor xenografts. Upon intravenous injection, T-DLHN can overcome the shortcoming of disassembly against extensive dilution, prolong the circulation time due to the "double locked" configuration and then mediate targeted drug delivery into the tumors. After uptaken into the cells, T-DLHN undergoes concurrent de-crosslinking and liberation of HCPT under acidic condition for enhanced chemotherapeutic efficacy with negligible adverse effects.

Two-in-One Polymeric NO-Donor Prodrugs Mediate Precision and Synergistic Prostate Cancer Treatment.

Chemotherapy predominates in clinical treatment of prostate cancer (PCa), while irreversible resistance to chemotherapeutics and severe side effects hinder the therapeutic efficacy, especially in castration-resistant PCa (CRPC). Herein, a bombesin (BBN)-decorated two-in-one prodrug (T-NO/E2-PMs) incorporating a polymeric nitric oxide (NO) donor and acetal-linked 17ß-estradiol (E2) in one backbone is developed, aiming to inhibit androgen receptor (AR) expression, reprogram the tumor microenvironment of CRPC, and enhance estradiol-mediated hypoxic CRPC therapy. Following efficient internalization mediated by BBN, T-NO/E2-PMs releases estradiol and NO in response to the unique intracellular environments. Both in vitro and in vivo studies demonstrate that the T-NO/E2-PMs nano-prodrug along with NO release potently downregulates AR levels to reverse CRPC and further enhances the chemo-sensitization of estradiol to PCa PC-3 cell apoptosis and the inhibition of metastasis. Collectively, this two-in-one nano-prodrug strategy offers a promising platform for construction of advanced nanomedicine to boost the therapeutic efficacy.

Black Phosphorus-Synergic Nitric Oxide Nanogasholder Spatiotemporally Regulates Tumor Microenvironments for Self-Amplifying Immunotherapy.

The lack of tumor immunogenicity coupled with the presence of tumor immunosuppression severely hinders antitumor immunity, especially in the treatment of "immune cold" tumors. Here, we have developed a drug-free and NIR-enabled nitric oxide (NO)-releasing nanogasholder (NOPS@BP) composed of an outer cloak of nitrate-containing polymeric NO donor and an inner core of black phosphorus (BP) as the energy converter to spatiotemporally regulate NO-mediated tumor microenvironment remodeling and achieve multimodal therapy. Following NIR-irradiation, BP-induced photothermia and its intrinsic reducing property accelerate NO release from the outer cloak, by which the instantaneous NO burst concomitant with mild photothermia, on the one hand, induces immunogenic cell death (ICD), thereby provoking antitumor responses such as the maturation of dendritic cells (DCs) and the infiltration of cytotoxic T lymphocytes (CTLs); on the other hand, it reverses tumor immunosuppression via Treg inhibition, M2 macrophage restraint, and PD-L1 downregulation, further strengthening antitumor immunity. Therefore, this drug-free NOPS@BP by means of multimodal therapy (NO gas therapy, immune therapy, photothermal therapy) realizes extremely significant curative effects against primary and distant tumors and even metastasis in B16F10 tumor models, providing a new modality to conquer immune cold tumors by NO-potentiated ICD and immunosuppression reversal.

Dual tumor- and subcellular-targeted photodynamic therapy using glucose-functionalized MoS2 nanoflakes for multidrug-resistant tumor ablation.

Photodynamic therapy (PDT) is emerging as an efficient strategy to combat multidrug-resistant (MDR) cancer. However, the short half-life and limited diffusion of reactive oxygen species (ROS) undermine the therapeutic outcomes of this therapy. To address this issue, a tumor-targeting nanoplatform was developed to precisely deliver mitochondria- and endoplasmic reticulum (ER)-targeting PDT agents to desired sites for dual organelle-targeted PDT. The nanoplatform is constructed by functionalizing molybdenum disulfide (MoS2) nanoflakes with glucose-modified hyperbranched polyglycerol (hPG), and then loading the organelle-targeting PDT agents. The resultant nanoplatform Cy7.5-TG@GPM is demonstrated to mediate both greatly enhanced internalization within MDR cells and precise subcellular localization of PDT agents, facilitating in situ near-infrared (NIR)-triggered ROS generation for augmented PDT and reversal of MDR, causing impressive tumor shrinkage in a HeLa multidrug-resistant tumor mouse model. As revealed by mechanistic studies of the synergistic mitochondria- and ER-targeted PDT, ROS-induced ER stress not only activates the cytosine-cytosine-adenosine-adenosine thymidine/enhancer-binding protein homologous protein (CHOP) pro-apoptotic signaling pathway, but also cooperates with ROS-induced mitochondrial dysfunction to trigger cytochrome C release from the mitochondria and induce subsequent cell death. Furthermore, the mitochondrial dysfunction reduces ATP production and thereby contributes to the reversal of MDR. This nanoplatform, with its NIR-responsive properties and ability to target tumors and subcellular organelles, offers a promising strategy for effective MDR cancer therapy.

Recent advances on next generation of polyzwitterion-based nano-vectors for targeted drug delivery.

Poly (ethylene glycol) (PEG)-based nanomedicines are perplexed by the challenges of oxidation damage, immune responses after repeated injections, and limited excretion from the body. As an alternative to PEG, bioinspired zwitterions bearing an identical number of positive and negative ions, exhibit exceptional hydrophilicity, excellent biomimetic nature and chemical malleability, endowing zwitterionic nano-vectors with biocompatibility, non-fouling feature, extended blood circulation and multifunctionality. In this review, we innovatively classify zwitterionic nano-vectors into linear, hyperbranched, crosslinked, and hybrid nanoparticles according to different chemical architectures in rational design of zwitterionic nano-vectors for enhanced drug delivery with an emphasis on zwitterionic engineering innovations as alternatives of PEG-based nanomedicines. Through combination with other nanostrategies, the intelligent zwitterionic nano-vectors can orchestrate stealth and other biological functionalities together to improve the efficacy in the whole journey of drug delivery.

Lipoic acid modified low molecular weight polyethylenimine mediates nontoxic and highly potent in vitro gene transfection.

The clinical success of gene therapy intimately relies on the development of safe and efficient gene carrier systems. We found here that 1.8 kDa polyethylenimine (PEI) following hydrophobic modification with lipoic acid (LA) mediated nontoxic and highly potent in vitro gene transfection in both HeLa and 293T cells. 1.8 kDa PEI-LA conjugates were prepared with controlled degree of substitution (DS) by coupling LA to PEI using carbodiimide chemistry. Gel electrophoresis measurements showed that the DNA binding ability of 1.8 kDa PEI was impaired by lipoylation, in which an N/P ratio of 2/1 and 4-6/1 was required for 1.8 kDa PEI and 1.8 kDa PEI-LA conjugates, respectively, to completely inhibit DNA migration. Interestingly, dynamic light scattering measurements (DLS) revealed that PEI-LA conjugates condensed DNA into much smaller sizes (183-84 nm) than unmodified 1.8 kDa PEI (444-139 nm) at N/P ratios ranging from 20/1 to 60/1. These polyplexes revealed similar surface charges of ca. +22 to +30 mV. 1.8 kDa PEI-LA(2) polyplexes formed at an N/P ratio of 10/1 were stable against exchange with 12-fold excess of negatively charged dextran sodium sulfate (DSS) relative to DNA phosphate groups while 1.8 kDa PEI controls dissociated at 6-fold excess of DSS, indicating that lipoylation of 1.8 kDa PEI resulted in stronger binding with DNA. Importantly, DNA was released from 1.8 kDa PEI-LA(2) polyplexes upon addition of 10 mM dithiothreitol (DTT). Reduction-triggered unpacking of 1.8 kDa PEI-LA(2) polyplexes was also confirmed by DLS. MTT assays demonstrated that all PEI-LA conjugates and polyplexes were essentially nontoxic to HeLa and 293T cells up to a tested concentration of 50 µg/mL and an N/P ratio of 80/1, respectively. The in vitro gene transfection studies in HeLa and 293T cells showed that lipoylation of 1.8 kDa PEI markedly boosted its transfection activity. For example, 1.8 kDa PEI-LA(2) polyplexes displayed 400-fold and 500-fold higher levels of gene expression than unmodified 1.8 kDa PEI controls, which were ca. 2-fold and 3-fold higher than 25 kDa PEI controls, in serum-free and 10% serum media, respectively. The transfection efficiency decreased with increasing DS, following an order of 1.8 kDa PEI-LA(2) > 1.8 kDa PEI-LA(4) > 1.8 kDa PEI-LA(6) ≫ 1.8 kDa PEI. Confocal laser scanning microscopy (CLSM) studies corroborated that 1.8 kDa PEI-LA(2) delivered and released DNA into the nuclei of HeLa cells more efficiently than 25 kDa PEI. These nontoxic 1.8 kDa PEI-LA conjugates form a superb basis for the development of targeting, biocompatible and highly efficient carriers of gene delivery.