Advanced Nitric Oxide Generating Nanomedicine for Therapeutic Applications

Abstract

Nitric oxide (NO), a gaseous transmitter extensively present in the human body, regulates vascular relaxation, immune response, inflammation, neurotransmission, and other crucial functions. Nitrite donors have been used clinically to treat angina, heart failure, pulmonary hypertension, and erectile dysfunction. Based on NO's vast biological functions, it further can treat tumors, bacteria/biofilms and other infections, wound healing, eye diseases, and osteoporosis. However, delivering NO is challenging due to uncontrolled blood circulation release and a half-life of under five seconds. With advanced biotechnology and the development of nanomedicine, NO donors packaged with multifunctional nanocarriers by physically embedding or chemically conjugating have been reported to show improved therapeutic efficacy and reduced side effects. Herein, we review and discuss recent applications of NO nanomedicines, their therapeutic mechanisms, and the challenges of NO nanomedicines for future scientific studies and clinical applications. As NO enables the inhibition of the replication of DNA and RNA in infectious microbes, including COVID-19 coronaviruses and malaria parasites, we highlight the potential of NO nanomedicines for antipandemic efforts. This review aims to provide deep insights and practical hints into design strategies and applications of NO nanomedicines.

Keywords: Bacterial; Biofilm; Biotechnology; Cancer; Coronavirus; Glaucoma; Nitric oxide nanomedicine; Osteoporosis; Wound healing.

Figure 1.

NO inhibits P-gp expression to reverse MDR. A) Schematic mechanisms of nitric oxide (NO) against multidrug resistance (MDR). Reprinted with permission from ref . Copyright 2017 Elsevier. B) pH-responsive NO generating to reverse P-gp-mediated MDR. (I) Schematic composition of HMs and NO generating mechanism to responsive acid tumors environment. (II) Confocal images of P-gp expression levels after different treatments. Reprinted with permission from ref . Copyright 2015 WILEY-VCH. C) Polyprodrug nanoparticles (CPNs) using cisplatin and ONOO⁻ to overwhelm cisplatin-resistant cancers. (I) Mechanism of CPNs to overcome cisplatin-resistant cancers. (II) P-gp expression level for cells imaged after CPN and different treatments. Reprinted with permission from ref . Copyright 2020 Elsevier. D) Combined NO generation, mild photothermal, and chemotherapy to overcome MDR by PNOC-PDA/DOX. (I) Schematic mechanism. (II) Western blot detection of P-gp expression in MCF-7/ADR upon different treatments. Reprinted with permission from ref . Copyright 2019 American Chemical Society. P-glycoprotein 1 (permeability glycoprotein), P-gp; multidrug resistance protein 1, MDR1; Hypoxia-inducible factor, HIF-1; Photothermal Therapy, PTT.

Figure 2.

NO promotes tumour environment normalization and enhances tumor chemotherapy. A) Schematic of the mechanism NanoNO to suppress hepatocellular carcinoma (HCC) progression in mice. The perivascular gradient produces nitric oxide (NO) and promotes the normalization of vessels and cancer suppression via apoptotic and programmed cell death ligand 1 (PD-L1) pathways in the tumor microenvironment (TME). Reprinted with permission from ref . Copyright 2019 Nature. B) NO inhibited collagen expression and improved chemotherapeutic penetration in tumors. (I) Mechanism of matrix metalloproteinase aka matricin (MMP) production and collagen degradation induced by ONOO⁻. (II) NO-generating nanoparticles increased the production of MMP-1 and MMP-2 shown by Western blots. (III) Confocal imaging for detecting collagenase activity under various treatments. (IV) immunofluorescent staining of Collagen I in tumors. Reprinted with permission from ref . Copyright 2019 American Chemical Society.

Figure 3.

NO enhances cancer radiotherapy. A) X-Ray-inducible ONOO⁻ from nanosized scintillators of LiLuF₄:Ce³⁺for radiosensitization. (I) Schematic illustration of RBS-T-SCNPs and the X-ray-controlled ONOO⁻ generation for improving radiotherapy. (II) Detection of cell viability of A549 cells, expression of γ-H2AX, and Nitro-Tyrosine with various treatments (scale bar= 20 μm). Reprinted with permission from ref . Copyright 2018 WILEY-VCH. B) X-ray-triggered depth-independent on-demand NO-release for hypoxic radiosensitization. (I) Construction of PEG-USMSs-SNO. (II) X-ray-triggered NO release in zebrafish larvae from PEG-USMSs-SNO. (III) X-ray dose-dependent NO release from PEG-USMSs-SNO in one hour. (IV) Cumulative NO release during the first day from PEG-USMSs-SNO after dose-dependent X-ray irradiation. (V) Relative tumor growth and (VI) weight change curve of mice with 4T1 tumors after indicated treatments. Reprinted with permission from ref . Copyright 2015 WILEY-VCH.

Figure 4.

NO enhances hypoxia tumor photodynamic therapy by inhibiting HIF-α expression and generation of ONOO⁻. A) Schematic illustration of PDT-specific O₂ economizer used to inhibit cellular respiration to combat hypoxia tumor. Reprinted with permission from ref . Copyright 2019 American Chemical Society. B) A cascade reaction of NO and hydrogen radicals for anti-hypoxia PDT. (I) Schematic generation of the H^•, O₂^•–/HO₂^•, NO, ONOO^–, and ¹O₂ from DANO and GSH upon light irradiation. (II) Cell viability images with DANO, calcein-AM, and PI staining under normoxic and hypoxic conditions with LED irradiation. Scale bars, 100 μm. Reprinted with permission from ref . Copyright 2021 American Chemical Society.

Figure 5.

NO enhances photothermal therapy for cancer. A) NIR light-triggered NO release for sensitizing mild photothermal therapy (PTT). (I) Schematic illustration of the synthesis of multifunctional BNN-Bi₂S₃ and NIR triggered NO and mild PTT in cancer therapy. (II) LC3-II, LC3-I, and p62 expression in BEL-7402 cells with different treatments by Western blot. Reprinted with permission from ref . Copyright 2018 WILEY-VCH. B) Schematic illustration of NIR-II -responsive NO-release anti-angiogenesis hydrogel. (I) Construction of WB@hydrogel and NIR-II laser-triggered anti-angiogenesis therapy of cancer. (II) Anti-angiogenesis mechanism of WB@hydrogel under laser irradiation. Reprinted with permission from ref , Copyright 2021 WILEY-VCH. Vascular endothelial growth factor, VEGF; Basic fibroblast growth factor, bFGF; Thrombospondins 1, TSP-1; Prolyl 4-hydroxylase subunit alpha-2, P4HA2.

Figure 6.

NO sensitizes photoacoustic therapy by generating ONOO⁻. A) Schematic illustration of photoacoustic (PA) cavitation-triggered ONOO⁻ generation for cancer therapy. B) In vitro NO-releasing of NO-NCPs under different treatments. C) Western blotting analysis of apoptosis-related proteins (cleaved-caspase 3 and Nitro-Tyrosine proteins). D) Gel electrophoresis detected DNA fragmentation of EMT6 cells after different treatments. E) In vivo PA imaging of tumor-bearing mice following intravenous injection of NO-NCPs. F) Tumor response to various treatment processes. Reprinted with permission from ref . Copyright 2021 WILEY-VCH.

Figure 7.

NO enhances chemodynamic therapy for liver tumors. A) Illustration of Fe (III)-BNCP and Fe (II)-BNCP synthesis as nanoscale coordination polymers. B) The mechanism of NO-CDT synergistic therapy using Fe (II)-BNCP for tumor cells. C) NO release at different content of GSH. D) Tumor growth curves during various treatment processes: (a) saline, (b) Zn (II)-DNCP, (c) Fe (II)-DNCP, (d) Zn (II)-BNCP, and (e) Fe (II)-BNCP. Reprinted with permission from ref . Copyright 2019 American Chemical Society.

Figure 8.

NO for antibacterial applications. A) Illustration of visible light-triggered simultaneous release NO and formaldehyde as a broad-spectrum antibacterial. Reprinted with permission from ref . Copyright 2021 American Chemical Society. B) Synergistic photothermal and NO antibacterial research based on dendritic Fe₃O₄@Poly(dopamine)@PAMAM nanocomposite for NO-delivery. Reprinted with permission from ref . Copyright 2018 WILEY-VCH. C) Synergistic NO and carbon monoxide (CO) for combating methicillin-resistant Staphylococcus aureus (MRSA) infections. (I) Synthetic routes of PCNO diblock copolymers and their mechanism for combating bacteria. (II) SEM images of E. coli (top) and S. aureus (bottom) incubated with PCNO micelles after irradiation (410 nm light). (III) Quantitative analysis of MRSA infection wound healing after receiving various treatments. (IV) Quantitative analysis of bacteria on days 1, 3, and 5 in wound tissues of MRSA-infected mice receiving different treatments. Reprinted with permission from ref . Copyright 2021 WILEY-VCH.

Figure 9.

NO for antibiofilm eradication. A) Visible-light-triggered NO releasing to eradicate biofilm. (I) Illustration of PEO-b-PCouNO nanoparticle preparation and synergistic therapeutic mechanism of NO and antibiotic against P. aeruginosa biofilm. (II) Two-dimensional and 3D confocal laser scanning microscopy (CLSM) images for detecting P. aeruginosa biofilms eradiation. (III) Quantitative analysis of biofilm viability. (IV) ATP assay for analysis of Planktonic bacteria. Reprinted with permission from ref . Copyright 2019 American Chemical Society. B) Red-light triggered NO release for efficient antibacterial treatment. (I) Illustration of the formulation of red-light triggered micelles and the NO-releasing mechanism of CouN(NO)-R derivatives responding to red light. (II) Representative images of the abscess during the treatment process in vivo. Reprinted with permission from ref . Copyright 2021 WILEY-VCH.

Figure 10.

NO enhances phototherapy against biofilms. A) NIR-mediated NO-enhanced photodynamic therapy (PDT) and mild photothermal therapy (PTT) for biofilm elimination. (I) Schematic illustration of the mechanism by which NIR triggers NO release to enhance PDT and PTT for biofilm ablation. (II) Scanning electron microscopy (SEM) images of biofilm after various treatments. (III) Live/dead stained 3D confocal laser scanning microscopy (CLSM) of biofilms challenged with other treatments. Reprinted with permission from ref . Copyright 2020 American Chemical Society. B) supramolecular nanocarriers with a switchable surface charge for synergistic NO and PDT destruction of biofilms. Reprinted with permission from ref . Copyright 2020 American Chemical Society. C) Illustration of the mechanism of MRSA biofilm eradication via NO-triggered gene downregulation. Reprinted with permission from ref . Copyright 2020 American Chemical Society.

Figure 11.

NO accelerates wound healing. A) Keratin composite mats release NO based on S-nitrosated to accelerate wound healing. Reprinted with permission from ref . Copyright 2020 Elsevier. B) ROS-triggered NO-releasing for synergistically combat bacterial infection and accelerate wound healing based on L-Arg-enriched amphiphilic peptide. Reprinted with permission from ref . Copyright 2021 WILEY-VCH. C) Photon-mediated NO-releasing from hemin-derived colloids to promote angiogenesis, microcirculation, and collagen deposition during wound healing. (I) Illustration of a NO-carrying Prussian blue (PB-NO) nanocubes for NIR-responsive NO-release healing of incisional wounds. (II) In vivo assessment of NO release for incisional wound healing. Reprinted with permission from ref . Copyright 2019 American Chemical Society.

Figure 12.

NO for treatment of eye diseases. A) Localized and controlled delivery of NO for glaucoma therapy. Reprinted with permission from ref . Copyright 2017 WILEY-VCH. B) NO releasing in intraocular pressure (IOP) reduction pathway for precision glaucoma therapy. Reprinted with permission from ref . Copyright 2021 Elsevier. C) Smart NO delivery for corneal wound healing by light-induced acid generation from pH@MSN-CaP-NO. Reprinted with permission from ref . Copyright 2016 American Chemical Society. D) Light-triggered NO release from polymersomes for corneal wound healing. Reprinted with permission from ref . Copyright 2019 Royal Society of Chemical.

Figure 13.

Smart microparticles (MPs) release NO to reverse osteoporosis. A) After subcutaneous administration, MPs were converted into small micelles through a phase transition to generate NO to alleviate osteoporosis. (I) Illustration of fabrication, structure, and functional mechanism of MPs. (II) Micro-CT images and H&E staining images of bones from test rats under varying treatments. (III) Serum biomarker levels of alkaline phosphatase (ALP) and osteocalcin after multiple treatments. Reprinted with permission from ref . Copyright 2017 WILEY-VCH. B) NIR-induced NO therapy for osteoporosis mediated by upconversion nanoparticle (UCPA-BNN). (I) Schematic illustration of NIR-triggered NO therapy for Osteoporosis based on UCPN-BNN. (II) Alizarin red staining photos of calcium nodules after varying treatments. (III-V) The expression of Col-1, Runx2, and ALP of MC3T3-E1 cells after varying treatments. Reprinted with permission from ref . Copyright 2021 American Chemical Society.

Figure 14.

Application of NO in the therapy of COVID-19. A) The mechanisms of nitric oxide (NO) antiviral. Reprinted with permission from ref . Copyright 2021 Elsevier. B) NO can efficiently inhibit viral RNA. Reprinted with permission from ref . Copyright 2005 American Society for Microbiology. C) Mitigation of the replication of SARS-CoV-2 by NO in vitro. (I) Comparison of the cytopathic effect development between cells treated with SNAP (a NO donor) and untreated controls. (II, III) Effect of NO generation on the activity of recombinant SARS-CoV-2 protease. Reprinted with permission from ref . Copyright 2020 Elsevier.

Figure 15.

Physiological and Immunological mechanism of NO action. Reprinted with permission from ref . Copyright 2001 Elsevier. Nitric oxide synthase enzymes, NOS; lipopolysaccharide, LPS; interleukin-1, IL-1; Tumor necrosis factor alpha, TNF-α; Interferon, IFN; messenger RNA, mRNA.

Scheme 1.

The treatment mechanisms of nitric oxide (NO) for various diseases. In addition to the direct effects of NO on biomolecules, NO reacts with oxygen or other reactive oxygen species (ROS) to generate reactive nitrogen species (RNS) to act on proteins, lipids, nucleosides, and metals as well as to induce transnitration, which can cause DNA strand breaks, abasic sites, enzyme activity inhibitions, mitochondrial depolarization, mitochondrial dysfunction, and DNA/RNA replication inhibitions. High-intensity focused ultrasound, HIFU; Glutathione, GSH.