Discovery of NO Donor-Aurovertin Hybrids as Dual Ferroptosis and Apoptosis Inducers for Treating Triple Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is a highly lethal malignancy, and its clinical management encounters severe challenges due to its high metastatic propensity and the absence of effective therapeutic targets. To improve druggability of aurovertin B (AVB), a natural polyketide with a significant antiproliferative effect on TNBC, a series of NO donor/AVB hybrids were synthesized and tested for bioactivities. Among them, compound 4d significantly inhibited the proliferation and metastasis of TNBC in vitro and in vivo with better safety than that of AVB. The structure-activity relationship analysis suggested that the types of NO donor and the linkers had considerable effects on the activities. Mechanistic investigations unveiled that 4d induced apoptosis and ferroptosis by the reduction of mitochondrial membrane potential and the down-regulation of GPX4, respectively. The antimetastatic effect of 4d was associated with the upregulation of DUSP1. Overall, these compelling results underscore the tremendous potential of 4d for treating TNBC.

Pleiotropic effects of nitric oxide sustained-release system for peripheral nerve repair.

The regenerative microenvironment after peripheral nerve injury is imbalanced and difficult to rebalance, which is mainly affected by inflammation, oxidative stress, and inadequate blood supply. The difficulty in remodeling the nerve regeneration microenvironment is the main reason for slow nerve regeneration. Traditional drug treatments have certain limitations, such as difficulty in penetrating the blood-nerve barrier and lack of pleiotropic effects. Therefore, there is an urgent need to build multifunctional nerve grafts that can effectively regulate the regenerative microenvironment and promote nerve regeneration. Nitric oxide (NO), a highly effective gas transmitter with diatomic radicals, is an important regulator of axonal growth and migration, synaptic plasticity, proliferation of neural precursor cells, and neuronal survival. Moreover, NO provides potential anti-inflammation, anti-oxidation, and blood vessel promotion applications. However, excess NO may cause cell death and neuroinflammatory cell damage. The prerequisite for NO treatment of peripheral nerve injury is that it is gradually released over time. In this study, we constructed an injectable NO slow-release system with two main components, including macromolecular NO donor nanoparticles (mPEG-P(MSNO-EG) nanoparticles, NO-NPs) and a carrier for the nanoparticles, mPEG-PA-PP injectable temperature-sensitive hydrogel. Due to the multiple physiological regulation of NO and better physiological barrier penetration, the conduit effectively regulates the inflammatory response and oxidative stress of damaged peripheral nerves, promotes nerve vascularization, and nerve regeneration and docking, accelerating the nerve regeneration process. STATEMENT OF SIGNIFICANCE: The slow regeneration speed of peripheral nerves is mainly due to the destruction of the regeneration microenvironment. Neural conduits with drug delivery capabilities have the potential to improve the microenvironment of nerve regeneration. However, traditional drugs are hindered by the blood nerve barrier and cannot effectively target the injured area. NO, an endogenous gas signaling molecule, can freely cross the blood nerve barrier and act on target cells. However, excessive NO can lead to cell apoptosis. In this study, a NO sustained-release system was constructed to regulate the microenvironment of nerve regeneration through various pathways and promote nerve regeneration.

A Randomized, Controlled Comparison of NCX 470, a Nitric Oxide-Donating Bimatoprost, and Latanoprost in Subjects with Open-Angle Glaucoma or Ocular Hypertension: The MONT BLANC Study.

PURPOSE: To compare intraocular pressure (IOP)-lowering efficacy and safety of NCX 470, a nitric oxide (NO)-donating bimatoprost, to latanoprost in subjects with open-angle glaucoma (OAG) or ocular hypertension (OHT). DESIGN: Prospective, phase 3, randomized, adaptive dose-selection, double-masked, parallel-group trial. METHODS: 691 subjects with OAG or OHT and unmedicated IOP ≥26 mmHg at 8AM, ≥24 mmHg at 10AM, and ≥22 mmHg at 4PM in the study eye were randomized to NCX 470 0.065%, NCX 470 0.1%, or latanoprost 0.005%. An interim analysis was performed to select the final dose of NCX 470. We evaluated noninferiority of NCX 470 versus latanoprost, based on IOP reduction from baseline at 8AM and 4PM at 2 weeks, 6 weeks, and 3 months. RESULTS: 661 subjects were analyzed; IOP was significantly reduced at all on-treatment time points, with reductions ranging from 8.0 to 9.7 mmHg (P < .0001 at each time point) in the NCX 470 0.1% group. Mean IOP reductions were greater with NCX 470 0.1% than latanoprost 0.005% at all 6 time points and significantly greater (P < .05) at 4 of the 6 time points. The most common adverse event was conjunctival/ocular hyperemia. CONCLUSION: The NO-donating prostaglandin analogue NCX 470 0.1% was well-tolerated and lowered IOP more than latanoprost in subjects with OAG or OHT at all 6 time points. With a dual mechanism of action that enhances both uveoscleral and trabecular outflow, NCX 470 could become an important first-line therapy for IOP reduction in glaucoma.

A coumarin-furoxan hybrid as novel nitric oxide donor induced cell apoptosis and ferroptosis in NSCLC by promoting S-nitrosylation of STAT3 and negative regulation of JAK2-STAT3 pathway.

Non-small cell lung cancer (NSCLC) still lacks effective treatment because of its extensive mutation diversity and frequent drug resistance. Therefore, it is urgent to develop new therapeutic strategies for NSCLC. In this study, we evaluated the inhibitory effect of a new coumarin-furoxan hybrid compound 9, a nitric oxide (NO) donor drug, on NSCLC proliferation and its mechanism. Our results show that compound 9 can inhibit the growth of four NSCLC cell lines and H1975 xenograft model in a dose-dependent manner. Compound 9 effectively releases high concentrations of NO within the mitochondria, leading to cellular oxidative stress, mitochondrial dysfunction, and apoptosis. Moreover, compound 9 inhibits JAK2/STAT3 protein phosphorylation and induces S-nitrosylation modification of STAT3, ultimately resulting in endogenous apoptosis in NSCLC. Additionally, compound 9 significantly induces NSCLC ferroptosis by depleting intracellular GSH, elevating MDA levels, inhibiting SLC7A11/GSH protein expression, and negatively regulating the JAK2/STAT3 pathway. In summary, this study elucidates the inhibitory effects of compound 9 on NSCLC proliferation and provides insights into the underlying mechanisms, offering new possibilities for NSCLC treatment strategies.

Development of Integrated Bioorthogonal Self-Catalyzed NO Donor/Platinum(IV) Prodrugs for Synergistical Intervention against Triple-Negative Breast Cancer.

The platinum(IV) prodrug strategy is attractive for the synergistic antitumor effect. High levels (>400 nM) of nitric oxide (NO) exert promising cancer inhibition effects via multiple mechanisms. Herein, we designed and synthesized a new group of integrated bioorthogonal self-catalyzed NO donor/Pt(IV) prodrugs bearing long alkyl chains to enhance the stability in circulation, while the cytoplasmic reductants trigger cascade activation to release Pt and NO in tumor cells. Specifically, compound 10c exhibited an improved stability, favorable pharmacokinetic properties (AUC(0-t) of 2210.10 h*ng/mL), potent anti-triple-negative breast cancer (TNBC) effects (71.08% tumor growth inhibition (TGI) against the MDA-MB-231 xenograft model), potent in vivo anti-TNBC lung metastasis activity, and acceptable low toxicity. Importantly, NO released from 10c leads to the S-nitrosation of metal transporters Atox1&ATP7a in TNBC cells, which increases the Pt retention and inhibits lysyl oxidase, generating synergistic tumoricidal and antimetastatic activity. These results may inspire further study on the synergistical therapy of Pt and NO for the treatment of TNBC.

Historical Perspectives of the Role of NO/NO Donors in Anti-Tumor Activities: Acknowledging Dr. Keefer's Pioneering Research.

The role of nitric oxide (NO) in cancer has been a continuous challenge and particularly the contradictory findings in the literature reporting NO with either anti-cancer properties or pro-cancer properties. This dilemma was largely resolved by the level of NO/inducible nitric oxide synthase in the tumor environment as well as other cancer-associated gene activations in different cancers. The initial findings on the role of NO as an anti-cancer agent was initiated in the late 1990's in Dr. Larry Keefer's laboratory, who had been studying and synthesizing many compounds with releasing NO under different conditions. Using an experimental model with selected NO compounds they demonstrated for the first time that NO can inhibit tumor cell proliferation and sensitizes drug-resistant cancer cells to chemotherapy-induced cytotoxicity. This initial finding was the backbone and the foundation of subsequent reports by the Keefer's laboratory and followed by many others to date on NO-mediated anti-cancer activities and the clinical translation of NO donors in cancer therapy. Our laboratory initiated studies on NO-mediated anti-cancer therapy and chemo-immuno-sensitization following Keefer's findings and used one of his synthesized NO donors, namely, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETANONOate), throughout most of our studies. Many of Keefer's collaborators and other investigators have reported on the selected compound, O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl] diazen-1-ium-1,2-diolate (JS-K), and its therapeutic role in many tumor model systems. Several lines of evidence that investigated the treatment with NO donors in various cancer models revealed that a large number of gene products are modulated by NO, thus emphasizing the pleiotropic effects of NO on cancers and the identification of many targets of therapeutic significance. The present review reports historically of several examples reported in the literature that emanated on NO-mediated anti-cancer activities by the Keefer's laboratory and his collaborators and other investigators including my laboratory at the University of California at Los Angeles.

Fifty Years of Diazeniumdiolate Research: A Tribute to Dr. Larry K. Keefer.

The pioneering studies of Dr. Larry Keefer and colleagues with diazeniumdiolates or NONOates as a platform have unraveled the chemical biology of many nitric oxides and have led to the design of a variety of promising therapeutic agents in oncology, gastroenterology, antimicrobials, wound healing, and the like. This dedication to Dr. Larry Keefer briefly highlights some of his studies using the diazeniumdiolate platform in the cancer arena.

Novel Nitric Oxide Donor-Azole Conjugation Strategy for Efficient Treatment of Cryptococcus neoformans Infections.

Invasive fungal infections (IFIs) such as cryptococcal meningitis (CM) remain a serious health issue worldwide due to drug resistance closely related to biofilm formation. Unfortunately, available antifungal drugs with ideal safety and promising potency are still lacking; thus, the research of new candidate and therapeutic approach is urgently needed. As an important gas messenger molecule, nitric oxide (NO) shows vital inhibition on various microorganism biofilms. Hence, three series of novel NO-donating azole derivatives were designed and synthesized, and the in vitro antifungal activity as well as the mechanism of action was investigated. Among them, 3a and 3e displayed excellent antifungal activity against Cryptococcus neoformans and biofilm depending on the release of NO. Moreover, a more stable analogue 3h of 3a demonstrated markedly anti-CM effects via intranasal dropping, avoiding the first-pass effects and possessing a better brain permeability bypass blood-brain barrier. These results present a promising antifungal candidate and intranasal dropping approach for the treatment of CM, warranting further studies.

Nitric Oxide: Physiological Functions, Delivery, and Biomedical Applications.

Nitric oxide (NO) is a gaseous molecule that has a central role in signaling pathways involved in numerous physiological processes (e.g., vasodilation, neurotransmission, inflammation, apoptosis, and tumor growth). Due to its gaseous form, NO has a short half-life, and its physiology role is concentration dependent, often restricting its function to a target site. Providing NO from an external source is beneficial in promoting cellular functions and treatment of different pathological conditions. Hence, the multifaceted role of NO in physiology and pathology has garnered massive interest in developing strategies to deliver exogenous NO for the treatment of various regenerative and biomedical complexities. NO-releasing platforms or donors capable of delivering NO in a controlled and sustained manner to target tissues or organs have advanced in the past few decades. This review article discusses in detail the generation of NO via the enzymatic functions of NO synthase as well as from NO donors and the multiple biological and pathological processes that NO modulates. The methods for incorporating of NO donors into diverse biomaterials including physical, chemical, or supramolecular techniques are summarized. Then, these NO-releasing platforms are highlighted in terms of advancing treatment strategies for various medical problems.

Design, Synthesis, and Biological Evaluation of Esculetin-Furoxan-DEAC Ternary Hybrids for Anti-Triple Negative Breast Cancer.

Twelve new hybrid compounds of Esculetin with nitric oxide (NO) donors and/or mitochondrial targeting groups were designed, synthesized, and evaluated for their anti-tumor activity and mechanism in vitro and in vivo. Notably, the most potent compound A11 exhibited nanomolar antiproliferative activity on triple-negative breast cancer (TNBC) MDA-MB-231 cells (IC50 = 8 nM) with a strikingly selective inhibitory effect. The mechanism of A11 involves targeting MDA-MB-231 cells' mitochondria, releasing a high NO concentration, and increasing the expression of cyclophilin D (CypD), leading to increased reactive oxygen species (ROS) and triggering cancer cell apoptosis. Additionally, A11 could arrest the cell cycle at the G2/M phase to achieve anti-tumor effects. Moreover, A11 demonstrated a superior TNBC inhibition rate and diminished toxicity relative to doxorubicin (DOX) in vivo. In summary, A11 serves as a noteworthy contender for TNBC treatment with high potency and minimal toxicity.

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.

A Multifunctional Nanozyme with NADH Dehydrogenase-Like Activity and Nitric Oxide Release under Near-Infrared Light Irradiation as an Efficient Therapeutic for Antimicrobial Resistance Infection and Wound Healing.

In recent years, antimicrobial resistance (AMR) has become one of the greatest threats to human health. There is an urgent need to develop new antibacterial agents to effectively treat AMR infection. Herein, a novel nanozyme platform (Cu,N-GQDs@Ru-NO) is prepared, where Cu,N-doped graphene quantum dots (Cu,N-GQDs) are covalently functionalized with a nitric oxide (NO) donor, ruthenium nitrosyl (Ru-NO). Under 808 nm near-infrared (NIR) light irradiation, Cu,N-GQDs@Ru-NO demonstrates nicotinamide adenine dinucleotide (NADH) dehydrogenase-like activity for photo-oxidizing NADH to NAD+ , thus disrupting the redox balance in bacterial cells and resulting in bacterial death; meanwhile, the onsite NIR light-delivered NO effectively eradicates the methicillin-resistant Staphylococcus aureus (MRSA) bacterial and biofilms, and promotes wound healing; furthermore, the nanozyme shows excellent photothermal effect that enhances the antibacterial efficacy as well. With the combination of NADH dehydrogenase activity, photothermal therapy, and NO gas therapy, the Cu,N-GQDs@Ru-NO nanozyme displays both in vitro and in vivo excellent efficacy for MRSA infection and biofilm eradication, which provides a new therapeutic modality for effectively treating MRSA inflammatory wounds.

Protective role of nitric oxide donors on endothelium in ischemia-reperfusion injury: a meta-analysis of randomized controlled trials.

BACKGROUND: Decreased bioavailability of nitric oxide (NO) under hypoxic conditions can lead to endothelial dysfunction. NO supplementation may protect endothelial function in ischemia-reperfusion (IR) injury. Therefore, a meta-analysis of randomized controlled trials (RCTs) was performed to verify the protective effect of NO donors on endothelium in IR injury. METHODS: Medline, Embase, Cochrane Library, and Web of Science databases were searched from inception to April 1, 2023. The specific inclusion criteria were as follows: (1) RCTs; (2) trials comparing NO donors with placebo control groups; and (3) trials reporting the effects of these interventions on vascular endothelial functional outcomes in IR injury. Random-effects models were used to assess pooled effect sizes, which were expressed as standardized mean differences (SMD). RESULTS: Seven studies satisfied the inclusion criteria and consisted of a total of 149 participants. NO donors were protective of endothelial function in IR injury (SMD: - 1.60; 95% confidence interval [CI]: - 2.33, - 0.88, P < 0.0001; heterogeneity [I2 = 66%, P = 0.001]). Results of the subgroup analysis showed the following: absence of protective effect of NO donor use following ischemia on endothelial function in IR injury - 1.78 (95% CI: - 2.50, - 1.07) and loss of protective effect on endothelial function after prolonged NO donor use - 0.89 (95% CI: - 2.06, 0.28). CONCLUSION: The short-period use of NO donors before the onset of ischemia can protect endothelial function in IR injury.

Liquid nitric oxide donor for adjuvant therapy of acute ischemic stroke via nasal administration.

OBJECTIVES: The aim of this study was to synthesize and characterize a novel NO donor, PEI-PO-NONOate, using propylene oxide and to investigate its biosafety and therapeutic efficacy via nasal administration in vitro and vivo. EXPERIMENTAL PROCEDURES: The PEI-PO-NONOate was synthesized based on polyethylenimine (PEI) with different molecular weights and characterized using Fourier transform infrared (FTIR), nuclear magnetic resonance (NMR), and ultraviolet (UV) spectroscopy. Cytotoxicity assays were performed on mouse fibroblast cells L929 and human nasal mucosa epithelial cells (HNEpC), and a rat middle cerebral artery occlusion (MCAO) model was established to evaluate the therapeutic efficacy of PEI-PO-NONOate via nasal administration. RESULTS: The PEI-PO-NONOate was found to be stable under dark, dry, and airproof conditions, and its release was accelerated in an aqueous phase or acidic environment, while it was slowed down in a polyethylene glycol (PEG) mixture system. The NO donor released approximately 0.4, 0.5, and 0.6 µmol of gaseous NO from 1.0 mg of the polymer based on PEI600, PEI1800, and PEI10K, respectively. Cytotoxicity assays showed that the PEI-PO-NONOates had a cryoprotective effect as compared with PEI and PEI-PO. Furthermore, nasal administration of PEI-PO-NONOates resulted in a significant reduction in overall necrotic ratio as compared with the control group (16.4% versus 24.6%, p < 0.05). CONCLUSION: The findings of this study suggest that PEI-PO-NONOates may have potential as an adjuvant therapy for acute ischemic stroke when administered via the nasal route.

The Nitric Oxide (NO) Donor Molsidomine Counteract Social Withdrawal and Cognition Deficits Induced by Blockade of the NMDA Receptor in the Rat.

The deficiency of the gaseous molecule nitric oxide (NO) seems to be critically involved in the pathogenesis of schizophrenia. Thus, molecules that can normalize NO levels, as are NO donors, might be of utility for the medication of this psychiatric disease. The aim of the present study was to detect the ability of the NO donor molsidomine to reduce schizophrenia-like impairments produced by the blockade of the N-methyl-D-aspartate (NMDA) receptor in rats. Molsidomine's ability to attenuate social withdrawal and spatial recognition memory deficits induced by the NMDA receptor antagonist ketamine were assessed using the social interaction and the object location test, respectively. Further, the efficacy of the combination of sub-effective doses of molsidomine with sub-effective doses of the atypical antipsychotic clozapine in alleviating non-spatial recognition memory deficits was evaluated utilizing the object recognition task. Molsidomine (2 and 4 mg/kg) attenuated social withdrawal and spatial recognition memory deficits induced by ketamine. Co-administration of inactive doses of molsidomine (1 mg/kg) and clozapine (0.1 mg/kg) counteracted delay-dependent and ketamine-induced non-spatial recognition memory deficits. The current findings suggest that molsidomine is sensitive to glutamate hypofunction since it attenuated behavioral impairments in animal models mimicking the negative symptoms and cognitive deficits of schizophrenia. Additionally, the present results support the potential of molsidomine as an adjunctive drug for the therapy of schizophrenia.

Biomimetic Biomembrane Encapsulation and Targeted Delivery of a Nitric Oxide Release Platform for Therapy of Parkinson's Disease.

The existence of the blood-brain barrier (BBB) and the complex inflammatory environment in the brain are two major obstacles in the treatment of Parkinson's disease (PD). As a target group, we modified the red blood cell membrane (RBCM) on the surface of upconversion nanoparticles (UCNPs) in this study to effectively target the brain. Mesoporous silicon, coated with UCNPs (UCM), was loaded with S-nitrosoglutathione (GSNO) as the nitric oxide (NO) donor. Then, UCNPs were excited to emit green light (540 nm) by 980 nm near-infrared (NIR). In addition, it produced a light-responsive anti-inflammatory effect by promoting the release of NO from GSNO and lowering the brain's level of proinflammatory factors. A series of experiments demonstrated that this strategy could effectively mitigate the inflammatory response damage of neurons in the brain.

Combination of chemotherapy and gaseous signaling molecular therapy: Novel ß-elemene nitric oxide donor derivatives against leukemia.

This study aimed to design and synthesize active hybrids of ß-elemene and nitric oxide (NO) donor pharmacophore as potential agents for treating leukemia. Derivatives reported herein exerted better inhibitory effects against human chronic myeloid leukemia K562 cells compared to ß-elemene (IC50 > 100 µM). The most potent compound 18f showed an IC50 value of 0.53 µM against K562 cells, as well as a high NO release level in vitro. In the K562 xenograft tumor mice model, compound 18f effectively inhibited the growth of the tumor, with a significant inhibition rate of 73.18%. After treatment with compound 18f, the body weight of mice did not decrease, indicating that it possessed good safety profile. All these proved that compound 18f was an excellent potential agent against leukemia.

Renal replacement therapy removes a large number of nitric oxide donors responsible for the nitrate-nitrite-nitric oxide pathway.

BACKGROUNDS: Nitric oxide has a broad-spectrum antibacterial property promising as a new therapeutic agent for severe acute respiratory syndrome coronavirus-2 because nitric oxide donor (such as S-nitroso-N-acetylpenicillamine) reduces the replication of coronavirus-2. Patients with coronavirus disease 2019 undergoing dialysis generally have a higher mortality rate than the general population. Although the higher mortality rate in these patients may be related to their advanced age, it has been suggested that plasma nitrite and nitrate levels (products of nitric oxide metabolism) are significantly decreased after hemodialysis which may compromise the nitrate-nitrite-nitric oxide pathway and impair nitric oxide homeostasis. It results in increased cardiovascular mortality in patients undergoing dialysis. However, the profile of nitric oxide-producing substances is poorly understood during renal replacement therapy. METHODS: We simulated continuous hemodialysis and hemodiafiltration to measure the amount of nitric oxide (nitric oxide-producing substance) clearance in vitro. RESULTS: The results demonstrated increased nitric oxide clearance and higher clearance than creatinine (molecular weight: 113) and vitamin B12 (molecular weight: 1355) using highly efficient renal replacement therapy modes. CONCLUSION: The high nitric oxide clearance may have partly contributed to the high cardiovascular and coronavirus-2 mortality risk in patients on dialysis.

Repurposing nitric oxide donating drugs in cancer therapy through immune modulation.

BACKGROUND: Nitric oxide-releasing drugs are used for cardiovascular diseases; however, their effects on the tumor immune microenvironment are less clear. Therefore, this study explored the impact of nitric oxide donors on tumor progression in immune-competent mice. METHODS: The effects of three different nitric oxide-releasing compounds (SNAP, SNP, and ISMN) on tumor growth were studied in tumor-bearing mouse models. Three mouse tumor models were used: B16F1 melanoma and LL2 lung carcinoma in C57BL/6 mice, CT26 colon cancer in BALB/c mice, and LL2 lung carcinoma in NOD/SCID mice. After nitric oxide treatment, splenic cytokines and lymphocytes were analyzed by cytokine array and flow cytometry, and tumor-infiltrating lymphocytes in the TME were analyzed using flow cytometry and single-cell RNA sequencing. RESULTS: Low doses of three exogenous nitric oxide donors inhibited tumor growth in two immunocompetent mouse models but not in NOD/SCID immunodeficient mice. Low-dose nitric oxide donors increase the levels of splenic cytokines IFN-γ and TNF-α but decrease the levels of cytokines IL-6 and IL-10, suggesting an alteration in Th2 cells. Nitric oxide donors increased the number of CD8+ T cells with activation gene signatures, as indicated by single-cell RNA sequencing. Flow cytometry analysis confirmed an increase in infiltrating CD8+ T cells and dendritic cells. The antitumor effect of nitric oxide donors was abolished by depletion of CD8+ T cells, indicating the requirement for CD8+ T cells. Tumor inhibition correlated with a decrease in a subtype of protumor macrophages and an increase in a subset of Arg1-positive macrophages expressing antitumor gene signatures. The increase in this subset of macrophages was confirmed by flow cytometry analysis. Finally, the combination of low-dose nitric oxide donor and cisplatin induced an additive cancer therapeutic effect in two immunocompetent animal models. The enhanced therapeutic effect was accompanied by an increase in the cells expressing the gene signature of NK cell. CONCLUSIONS: Low concentrations of exogenous nitric oxide donors inhibit tumor growth in vivo by regulating T cells and macrophages. CD8+ T cells are essential for antitumor effects. In addition, low-dose nitric oxide donors may be combined with chemotherapeutic drugs in cancer therapy in the future.

Metal-organic framework for biomimetic nitric oxide generation and anticancer drug delivery.

The potential therapeutic implications of nitric oxide (NO) have drawn a great deal of interest for reversing multidrug resistance (MDR) in cancer; however, previous strategies utilized unstable or toxic NO donors often oxidized by the excessive addition of reactive oxygen species, leading to unexpected side effects. Therefore, this study proposed a metal-organic framework (MOF), Porous coordination network (PCN)-223-Fe, to be loaded with a biocompatible NO donor, L-arginine (L-arg; i.e., PCN-223-Fe/L-arg). This specific MOF possesses a ligand of Fe-porphyrin, a biomimetic catalyst. Thus, with PCN-223-Fe/L-arg, L-arg was released in a sustained manner, which generated NO by a catalytic reaction between L-arg and Fe-porphyrin in PCN-223-Fe. Through this biomimetic process, PCN-223-Fe/L-arg could generate sufficient NO to reverse MDR at the expense of hydrogen peroxide already present and highly expressed in cancer environments. For treatment of MDR cancer, this study also proposed PCN-223-Fe loaded with an anticancer drug, irinotecan (CPT-11; i.e., PCN-223-Fe/CPT-11), to be formulated together with PCN-223-Fe/L-arg. Owing to the synergistic effect of reversed MDR by NO generation and sustained release of CPT-11, this combined formulation exhibited a higher anticancer effect on MDR cancer cells (MCF-7/ADR). When intratumorally injected in vivo, coadministration of PCN-223-Fe/L-arg and PCN-223-Fe/CPT-11 greatly suppressed tumor growth in nude mice bearing MDR tumors.