FDA-approved Iron Chelator Research Articles

Iron Chelation Therapy Elicits Innate Immune Control of Metastatic Ovarian Cancer.

Iron accumulation in tumors contributes to disease progression and chemoresistance. Although targeting this process can influence various hallmarks of cancer, the immunomodulatory effects of iron chelation in the tumor microenvironment are unknown. Here, we report that treatment with deferiprone, an FDA-approved iron chelator, unleashes innate immune responses that restrain ovarian cancer. Deferiprone reprogrammed ovarian cancer cells toward an immunostimulatory state characterized by the production of type-I IFN and overexpression of molecules that activate NK cells. Mechanistically, these effects were driven by innate sensing of mitochondrial DNA in the cytosol and concomitant activation of nuclear DNA damage responses triggered upon iron chelation. Deferiprone synergized with chemotherapy and prolonged the survival of mice with ovarian cancer by bolstering type-I IFN responses that drove NK cell-dependent control of metastatic disease. Hence, iron chelation may represent an alternative immunotherapeutic strategy for malignancies that are refractory to current T-cell-centric modalities. Significance: This study uncovers that targeting dysregulated iron accumulation in ovarian tumors represents a major therapeutic opportunity. Iron chelation therapy using an FDA-approved agent causes immunogenic stress responses in ovarian cancer cells that delay metastatic disease progression and enhance the effects of first-line chemotherapy. See related commentary by Bell and Zou, p. 1771.

A small molecular “albumin hitchhiking” deferoxamine conjugate improved iron clearance efficacy

Deferoxamine (DFO), the first FDA-approved iron chelator, showed multifaceted potential in the treatment of iron accumulation-related diseases. However, the extremely short half-life of DFO leads to low patient compliance, and required high doses result in nonspecific toxicity. To overcome these obstacles, a new conjugate 2-sulfo-9-fluorenylmethoxycarbonyl-deferoxamine (FMS-DFO) was developed, which could be leveraged to hand-in-hand associate with the albumin in plasma to bypass the rapid elimination and reduce the related toxicities. FMS-DFO was observed in comparable albumin binding capacity as the amphiphilic modular “albumin hitchhiking” DSPE-PEG2000-DFO and maintained identical iron chelating potency of DFO. As compared with DSPE-PEG2000-DFO (t1/2–1.30 h), FMS-DFO (t1/2–4.49 h) remarkably more prolonged the exposure circulation of marketed DFO (t1/2–0.08 h) in mice. FMS-DFO could spare the dosing frequency from 5 times to 3 times weekly in iron overload therapy, which can improve patient compliance and reduce the difficulty of clinical usage. Additionally, the combined utilization of 5-ALA and FMS-DFO demonstrated excellent anti-cancer efficacy in synergistic photodynamic therapy model, broadening the clinical potential of the developed iron chelators. The strategy of “albumin hitchhiking” FMS-DFO is thus of great clinical interest and merits further exploration for treating iron accumulation-related diseases.

Deferasirox Derivatives: Ligands for the Lanthanide Series.

Deferasirox is an FDA-approved iron chelator used in the treatment of iron toxicity. In this work, we report the use of several deferasirox derivatives as lanthanide chelators. Solid-state structural studies of three representative trivalent lanthanide cations, La(III), Eu(III), and Lu(III), revealed the formation of 2:2 complexes in the solid state. A 1:1 stoichiometry dominates in DMSO solution, with Ka values of 472 ± 14, 477 ± 11, and 496 ± 15 M-1 being obtained in the case of these three cations, respectively. Under the conditions of competitive precipitation in the presence of triethylamine, high selectivity (up to 80%) for lutetium(III) was observed in competition with La(III), Ce(III), and Eu(III). Theoretical calculations provided support for the observed selective crystallization.

Retinal ferroptosis as a critical mechanism for the induction of retinochoroiditis during ocular toxoplasmosis

Toxoplasmosis is a major infectious disease, affecting approximately one-third of the world's population; its main clinical manifestation, ocular toxoplasmosis (OT), is a severe sight-threatening disease. Nevertheless, the diagnosis of OT is based on clinical findings, which needs improvement, even with biochemical tests, such as polymerase chain reaction and antibody detections. Furthermore, the efficacy of OT-targeted treatment is limited; thus, additional measures for diagnosis and treatments are needed. Here, we for the first time report a significantly reduced iron concentration in the vitreous humor (VH) of human patients infected with OT. To obtain further insights into molecular mechanisms, we established a mouse model of T. gondii infection, in which intravitreally injected tracer 57Fe, was accumulated in the neurosensory retina. T. gondii-infected eyes showed increased lipid peroxidation, reduction of glutathione peroxidase-4 expression and mitochondrial deformity in the photoreceptor as cristae loss. These findings strongly suggest the involvement of ferroptotic process in the photoreceptor of OT. In addition, deferiprone, an FDA-approved iron chelator, reduced the iron uptake but also ameliorated toxoplasma-induced retinochoroiditis by reducing retinal inflammation. In conclusion, the iron levels in the VH could serve as diagnostic markers and iron chelators as potential treatments for OT.

Exploring Antibiotic-Potentiating Effects of Tobramycin-Deferiprone Conjugates in Pseudomonas aeruginosa.

Metal ions, including Fe3+, affect the target site binding of some antibiotics and control the porin- and siderophore-mediated uptake of antibiotics. Amphiphilic tobramycins are an emerging class of antibiotic potentiators capable of synergizing with multiple classes of antibiotics against Gram-negative bacteria, including Pseudomonas aeruginosa. To study how the antibiotic-potentiating effect of amphiphilic tobramycins is affected by the presence of intermolecular iron chelators, we conjugated the FDA-approved iron chelator deferiprone (DEF) to tobramycin (TOB). Three TOB-DEF conjugates differing in the length of the carbon tether were prepared and tested for antibacterial activity and synergistic relationships with a panel of antibiotics against clinical isolates of P. aeruginosa. While all TOB-DEF conjugates were inactive against P. aeruginosa, the TOB-DEF conjugates strongly synergized with outer-membrane-impermeable antibiotics, such as novobiocin and rifampicin. Among the three TOB-DEF conjugates, 1c containing a C12 tether showed a remarkable and selective potentiating effect to improve the susceptibility of multidrug-resistant P. aeruginosa isolates to tetracyclines when compared with other antibiotics. However, the antibacterial activity and antibiotic-potentiating effect of the optimized conjugate was not enhanced under iron-depleted conditions, indicating that the function of the antibiotic potentiator is not affected by the Fe3+ concentration.

Drugs That Mimic Hypoxia Selectively Target EBV-Positive Gastric Cancer Cells.

Latent infection of Epstein-Barr virus (EBV) is associated with lymphoid and epithelial cell cancers, including 10% of gastric carcinomas. We previously reported that hypoxia inducible factor-1α (HIF-1α) induces EBV's latent-to-lytic switch and identified several HIF-1α-stabilizing drugs that induce this viral reactivation. Here, we tested three classes of these drugs for preferential killing of the EBV-positive gastric cancer AGS-Akata cell line compared to its matched EBV-negative AGS control. We observed preferential killing with iron chelators [Deferoxamine (DFO); Deferasirox (DFX)] and a prolyl hydroxylase inhibitor (BAY 85-3934 (Molidustat)), but not with a neddylation inhibitor [MLN4924 (Pevonedistat)]. DFO and DFX also induced preferential killing of the EBV-positive gastric cancer AGS-BDneo and SNU-719 cell lines. Preferential killing was enhanced when low-dose DFX (10 μM) was combined with the antiviral prodrug ganciclovir. DFO and DFX induced lytic EBV reactivation in approximately 10% of SNU-719 and 20-30% of AGS-Akata and AGS-BDneo cells. However, neither DFO nor DFX significantly induced synthesis of lytic EBV proteins in xenografts grown in NSG mice from AGS-Akata cells above the level observed in control-treated mice. Therefore, these FDA-approved iron chelators are less effective than gemcitabine at promoting EBV reactivation in vivo despite their high specificity and efficiency in vitro.

Mechanism-Based Pharmacokinetic Modeling of Absorption and Disposition of a Deferoxamine-Based Nanochelator in Rats.

Deferoxamine (DFO) is an effective FDA-approved iron chelator. However, its use is considerably limited by off-target toxicities and an extremely cumbersome dose regimen with daily infusions. The recent development of a deferoxamine-based nanochelator (DFO-NP) with selective renal excretion has shown promise in ameliorating animal models of iron overload with a substantially improved safety profile. To further the preclinical development of this promising nanochelator and to inform on the feasibility of clinical development, it is necessary to fully characterize the dose and administration-route-dependent pharmacokinetics and to develop predictive pharmacokinetic (PK) models describing absorption and disposition. Herein, we have evaluated the absorption, distribution, and elimination of DFO-NPs after intravenous and subcutaneous (SC) injection at therapeutically relevant doses in Sprague Dawley rats. We also characterized compartment-based model structures and identified model-based parameters to quantitatively describe the PK of DFO-NPs. Our modeling efforts confirmed that disposition could be described using a three-compartment mamillary model with elimination and saturable reabsorption both occurring from the third compartment. We also determined that absorption was nonlinear and best described by parallel saturable and first-order processes. Finally, we characterized a novel pathway for saturable SC absorption of an ultrasmall organic nanoparticle directly into the systemic circulation, which offers a novel strategy for improving drug exposure for nanotherapeutics.

Iron Neurotoxicity and Protection by Deferoxamine in Intracerebral Hemorrhage.

Intracerebral hemorrhage (ICH) is a subtype of stroke that is characterized by high morbidity and mortality, for which clinical outcome remains poor. An extensive literature indicates that the release of ferrous iron from ruptured erythrocytes in the hematoma is a key pathogenic factor in ICH-induced brain injury. Deferoxamine is an FDA-approved iron chelator that has the capacity to penetrate the blood-brain barrier after systemic administration and binds to iron. Previous animal studies have shown that deferoxamine attenuates ICH-induced brain edema, neuronal death, and neurological deficits. This review summarizes recent progress of the mechanisms by which deferoxamine may alleviate ICH and discusses further studies on its clinical utility.

Tuning the Solid- and Solution-State Fluorescence of the Iron-Chelator Deferasirox.

Deferasirox, an FDA-approved iron chelator, has gained increasing attention for use in anticancer and antimicrobial applications. Recent efforts by our group led to the identification of this core as an easy-to-visualize aggregation-induced emission platform, or AIEgen, that provides a therapeutic effect equivalent to deferasirox (J. Am. Chem. Soc. 2021, 143, 3, 1278-1283). However, the emission wavelength of the first-generation system overlapped with that of Syto9, a green emissive dye used to indicate live cells. Here, we report a library of deferasirox derivatives with various fluorescence emission profiles designed to overcome this limitation. We propose referring to systems that show promise as both therapeutic and optical imaging agents as "illuminoceuticals". The color differences between the derivatives were observable to the unaided eye (solid- and solution-state) and were in accord with the Commission Internationale de L'Eclairage (CIE) chromaticity diagram 1913. Each fluorescent derivative successfully imaged the respective spherical and rod shapes of methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa. They also displayed iron-dependent antibiotic activity. Three derivatives, ExNMe2 (3), ExTrisT (11), and ExDCM (13), display emission features that are sufficiently distinct so as to permit the multiplex (triplex) imaging of both MRSA and P. aeruginosa via stimulated emission depletion microscopy. The present deferasirox derivatives allowed for the construction of a multi-fluorophore sensor array. This array enabled the successful discrimination between Gram-positive/Gram-negative and drug-sensitive/drug-resistant bacteria. Antibiotic sensitivity and drug-resistant mutants from clinically isolated strains could also be identified and differentiated.

Deferoxamine accelerates endothelial progenitor cell senescence and compromises angiogenesis.

Senescence reduces the circulating number and angiogenic activity of endothelial progenitor cells (EPCs), and is associated with aging-related vascular diseases. However, it is very time-consuming to obtain aged cells (~1 month of repeated replication) or animals (~2 years) for senescence studies. Here, we established an accelerated senescence model by treating EPCs with deferoxamine (DFO), an FDA-approved iron chelator. Four days of low-dose (3 μM) DFO induced senescent phenotypes in EPCs, including a senescent pattern of protein expression, impaired mitochondrial bioenergetics, altered mitochondrial protein levels and compromised angiogenic activity. DFO-treated early EPCs from young and old donors (< 35 vs. > 70 years old) displayed similar senescent phenotypes, including elevated senescence-associated β-galactosidase activity and reduced relative telomere lengths, colony-forming units and adenosine triphosphate levels. To validate this accelerated senescence model in vivo, we intraperitoneally injected Sprague-Dawley rats with DFO for 4 weeks. Early EPCs from DFO-treated rats displayed profoundly senescent phenotypes compared to those from control rats. Additionally, in hind-limb ischemic mice, DFO pretreatment compromised EPC angiogenesis by reducing both blood perfusion and capillary density. DFO thus accelerates EPC senescence and appears to hasten model development for cellular senescence studies.

Deferasirox (ExJade): An FDA-Approved AIEgen Platform with Unique Photophysical Properties.

Deferasirox, ExJade, is an FDA-approved iron chelator used for the treatment of iron overload. In this work, we report several fluorescent deferasirox derivatives that display unique photophysical properties, i.e., aggregation-induced emission (AIE), excited state intramolecular proton transfer, charge transfer, and through-bond and through-space conjugation characteristics in aqueous media. Functionalization of the phenol units on the deferasirox scaffold afforded the fluorescent responsive pro-chelator ExPhos, which enabled the detection of the disease-based biomarker alkaline phosphatase (ALP). The diagnostic potential of these deferasirox derivatives was supported by bacterial biofilm studies.

Abstract 170: Deferoxamine Treatment Decreases Levels Of Reactive Oxygen Species And Cellular Apoptosis In Irradiated Skin

Purpose: Radiation therapy is a cornerstone of oncologic treatment. Unfortunately, radiation can lead to pain and ulceration in the acute phase, as well as stiffness and fibrosis in the chronic phase. Deferoxamine (DFO) is an FDA approved iron chelator that has been shown to increase skin vascularization by stabilizing hypoxia-inducible factor 1-alpha levels and improve radiation-induced fibrosis. However, it is unclear whether there are other mechanisms underlying the therapeutic effects that DFO exhibits on the skin. Previous work has shown that DFO delivered transdermally improves healing in diabetic ulcer wounds by decreasing oxidative stress and cellular apoptosis. This study evaluates the effects of transdermal DFO on reactive oxygen species and cell death in the context of irradiation (IR) to skin. Methods: CD-1 nude immunodeficient mice (n=8) were randomized to one of two conditions: 1. Control patch (IR only) (n=4) and 2. DFO patch (n=4). The study period consisted of a 14-day pre-IR period, followed by a 12-day course of fractionated IR which delivered a total of 30 Gray to all mice. The control group received patches without DFO and the DFO group received patches containing DFO throughout the experiment with patches changed daily. Mouse scalp skin was harvested one day after the completion of IR. Tissue samples were either fixed in 4% PFA, snap frozen in OCT, or directly stored at -80°C. Fixed samples were embedded in paraffin, sectioned, and stained for iron with Prussian blue. Snap frozen samples were cryosectioned at 10um, mounted, stained with dihydroethidium (DHE), and imaged with confocal microscopy. DHE detects reactive oxygen species. Stored samples were mechanically homogenized and enzyme-linked immunosorbent assays (ELISAs) were performed to measure levels of human Bax and Cleaved Caspase-3, markers of apoptosis. Results: Treatment with DFO was associated with decreased levels of iron in the skin, confirming DFO’s role as an iron chelator. DHE fluorescence was more intense in the control patch group than in the DFO patch group (***p<0.0001), indicating that ROS generation decreased with DFO treatment. Similarly, apoptosis was highest in the control group and lowest in the DFO group as shown by higher human Bax protein (*p=0.028) and Cleaved Caspase-3 (**p=0.0038) levels. Conclusions: DFO delivered transdermally leads to a dose-dependent decrease in reactive oxygen species and apoptotic markers. These results provide further insight into the mechanisms underlying its restorative effects on irradiated skin. Future work will investigate the relationship of reactive oxygen species and fibroblast subpopulations in the skin.

Interactions of Alginate-Deferoxamine Conjugates With Blood Components and Their Antioxidation in the Hemoglobin Oxidation Model.

While deferoxamine (DFO) has long been used as an FDA-approved iron chelator, its proangiogenesis ability attracts increasing number of research interests. To address its drawbacks such as short plasma half-life and toxicity, polymeric conjugated strategy has been proposed and shown superiority. Owing to intravenous injection and application in blood-related conditions, however, the blood interactions and antioxidation of the DFO-conjugates and the mechanisms underlying these outcomes remain to be elucidated. In this regard, incubating with three different molecular-weight (MW) alginate-DFO conjugates (ADs) red blood cells (RBCs), coagulation system, complement and platelet were investigated. To prove the antioxidant activity of ADs, we used hemoglobin oxidation model in vitro. ADs did not cause RBCs hemolysis while reversible aggregation and normal deformability ability were observed. However, the coagulation time, particularly APTT and TT, were significantly prolonged in a dose-dependent manner, and fibrinogen was dramatically decreased, suggesting ADs could dominantly inhibit the intrinsic pathways in the process of coagulation. The dose-dependent anticoagulation might be related with the functional groups along the alginate chains. The complements, C3a and C5a, were activated by ADs in a dose-dependent manner through alternative pathway. For platelet, ADs slightly suppressed the activation and aggregation at low concentration. Based on above results, the cross-talking among coagulation, complement and platelet induced by ADs was proposed. The antioxidation of ADs through iron chelation was proved and the antioxidant activity was shown in a MW-dependent manner.

Abstract QS12: Deferoxamine Diminishes Breast Cancer Proliferation and Enhances Irradiated Breast Reconstruction: An Antitumorigenic Mechanism Defined

PURPOSE: Radiation plays an essential role in the oncologic management of triple-negative breast cancer, but patients who undergo radiotherapy experience significantly more wound complications during the reconstructive process. Deferoxamine is an FDA-approved iron chelator with immense potential to up-regulate angiogenesis and improve reconstructive outcomes. More specifically, the ability of deferoxamine to mitigate the deleterious effects of radiation on skin and soft tissue has been shown to enhance expander-based irradiated breast reconstruction, increase fat graft retention, and facilitate irradiated transverse rectus abdominis myocutaneous flap survival. The purpose of this study is to determine the impact of deferoxamine on breast cancer cell proliferation in-vitro, in order to delineate oncologic safety concerns regarding the utilization of deferoxamine as a regenerative therapeutic. METHODS: Triple-negative breast cancer cells, which do not respond to hormone or HER2 blocking therapy, were utilized in this study as patients with triple-negative breast cancer commonly receive chemoradiation and are potential candidates to receive deferoxamine as a regenerative therapeutic during the reconstructive process. The effect of radiation and deferoxamine on two triple-negative breast cancer cell lines (MDA-MB-231, MDA-MB-468) and one female fibroblast control cell line was determined via MTS (percent cell viability) analysis. Radiation (0, 5, 10 Gy) and deferoxamine (0, 25, 50, 75, 100 µM) were delivered individually and jointly, and all experiments were completed in triplicate. Intracellular iron concentration was determined via QuantiChromTMassay, NF-κB localization was determined via dual-luciferase reporter assay, and apoptosis/necrosis was determined via annexin V assay in order to delineate mechanism. ANOVA statistical analysis was performed using SPSS (p<0.05). RESULTS: Both triple-negative breast cancer cell lines exemplified a significant decrease in percent viability following exposure to 10 Gy of radiation (p<0.05) or 25 µM deferoxamine (p<0.01). The administration of radiation (10 Gy) in combination with deferoxamine (100 µM) to triple-negative breast cancer cells resulted in significant reduction of percent cell viability compared to the administration of radiation alone (p<0.05). Fibroblasts exemplified no significant response to individually administered radiation (10 Gy) or deferoxamine (100 µM), while joint administration of radiation (10Gy) and deferoxamine (25µM) resulted in a significant increase in percent cell viability compared to the administration of radiation alone (p<0.05). In both triple-negative breast cancer cell lines, individual administration of deferoxamine (100µM) significantly decreased intracellular iron concentration (p<0.05). Joint administration of radiation and deferoxamine suppressed NF-κB activation indicating a cessation of cell proliferation and amplified cellular apoptosis (p<0.01) with no notable increase in cellular necrosis. CONCLUSIONS: The administration of radiation and/or deferoxamine imparts a significant decrease in triple-negative breast cancer cell proliferation, without significantly impeding growth of female fibroblast control cells in-vitro. Mechanistically, deferoxamine administration decreases intracellular iron concentration leading to decreased cell proliferation and increased cell death via apoptosis. Taken together, these findings suggest deferoxamine may be safely utilized to facilitate improved reconstructive outcomes among triple-negative breast cancer survivors and warrant future investigations to quantify in-vivoeffects.

Hypoxia Signaling in the Skeleton: Implications for Bone Health.

We reviewed recent literature on oxygen sensing in osteogenic cells and its contribution to development of a skeletal phenotype, the coupling of osteogenesis with angiogenesis and integration of hypoxia into canonical Wnt signaling, and opportunities to manipulate oxygen sensing to promote skeletal repair. Oxygen sensing in osteocytes can confer a high bone mass phenotype in murine models; common and unique targets of HIF-1α and HIF-2α and lineage-specific deletion of oxygen sensing machinery suggest differentia utilization and requirement of HIF-α proteins in the differentiation from mesenchymal stem cell to osteoblast to osteocyte; oxygen-dependent but HIF-α-independent signaling may contribute to observed skeletal phenotypes. Manipulating oxygen sensing machinery in osteogenic cells influences skeletal phenotype through angiogenesis-dependent and angiogenesis-independent pathways and involves HIF-1α, HIF-2α, or both proteins. Clinically, an FDA-approved iron chelator promotes angiogenesis and osteogenesis, thereby enhancing the rate of fracture repair.

Abstract 2911: The propitious dual roles of deferoxamine in head and neck cancer management

Abstract Introduction: For head and neck cancer survivors, radiotherapy (XRT) improves survivorship at a cost of impaired quality of life. XRT-induced pathologic fractures (fxs) and associated nonunions are devastating iatrogenic sequelae associated with microvascular damage in bone, yet there are no pharmacologic strategies to prevent or treat these maladies. Deferoxamine (DFO), an FDA-approved iron chelator, has been extensively investigated for its ability to promote tissue vascularization through upregulation of the HIF-1α pathway. Our laboratory has previously demonstrated improved fx healing and prevention of nonunions in radiated bone utilizing localized DFO administration. However, the role of DFO in head and neck squamous cell carcinoma (HNSCC) tumorigenesis is currently unknown. While iron is essential for normal cell proliferation, excessive iron is associated with multiple oncogenic pathways. Thus, localized iron chelation may have beneficial antitumorigenic properties. Elucidating the role of iron metabolism and correlations between iron chelation and angiogenic signaling may offer opportunities for diagnostic and treatment purposes, while simultaneously improving tissue engineering efforts for HNSCC survivors. Methods: We surveyed the UALCAN survivorship data base for iron-related genes in HNSCC. In vitro, established MDA1986 (HPV+) and UMSCC108 (HPV-) cell lines were grown in culture. The effects of DFO were probed on 3-D tumorspheres. HIF-1α was examined by IHC, and Western blot (WB) was used to elucidate iron mediators. In vivo, MDA-1986 buccal xenografts were developed in Nu/Nu mice. Control, DFO, and XRT groups were generated, and DFO and XRT were administered as previously described. IVIS imaging and tumor volumes were used to monitor growth. UALCAN data guided the selection of iron-regulatory proteins identified by WB in excised tumors. Results: TFRC and FTH-1 were highly upregulated, suggesting that dysregulation of iron metabolism contributes to HNSCC development. In vitro, we observed a dose-dependent decrease in 3-D tumorsphere formation and no alterations in HIF-1α expression. WB demonstrated TFRC downregulation in response to 100µM DFO in both HPV+ and - cells. In vivo, a significant decrease in tumor growth and radiance was observed with DFO treatment (comparable to XRT) when compared to untreated controls. Excised tumor WB demonstrated downregulation of TFRC, identifying it as a pivotal indicator of response to iron-chelation therapy. Conclusion: In vitro and in vivo studies reveal that DFO exhibits antitumorigenic effects, and suggest that HIF-1α is not affected by iron chelation in HNSCC cells. Moreover, TFRC was identified as a biomarker for indicating treatment response. Our findings signify that localized DFO may have antitumorigenic effects, in addition to the previously established regenerative therapeutic effects for the treatment of XRT-induced bone pathologies. Citation Format: Alexis Donneys, Chitra Subramanian, Jeremy Lynn, Kevin Urlaub, Kevin Kovatch, Halil S. Uygur, Mark S. Cohen, Steven R. Buchman. The propitious dual roles of deferoxamine in head and neck cancer management [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2911.

Iron deposition is associated with differential macrophage infiltration and therapeutic response to iron chelation in prostate cancer

Immune cells such as macrophages are drivers and biomarkers of most cancers. Scoring macrophage infiltration in tumor tissue provides a prognostic assessment that is correlated with disease outcome and therapeutic response, but generally requires invasive biopsy. Routine detection of hemosiderin iron aggregates in macrophages in other settings histologically and in vivo by MRI suggests that similar assessments in cancer can bridge a gap in our ability to assess tumor macrophage infiltration. Quantitative histological and in vivo MRI assessments of non-heme cellular iron revealed that preclinical prostate tumor models could be differentiated according to hemosiderin iron accumulation—both in tumors and systemically. Monitoring cellular iron levels during “off-label” administration of the FDA-approved iron chelator deferiprone evidenced significant reductions in tumor size without extensive perturbation to these iron deposits. Spatial profiling of the iron-laden infiltrates further demonstrated that higher numbers of infiltrating macrophage iron deposits was associated with lower anti-tumor chelation therapy response. Imaging macrophages according to their innate iron status provides a new phenotypic window into the immune tumor landscape and reveals a prognostic biomarker associated with macrophage infiltration and therapeutic outcome.

Escherichia coli Free Radical-Based Killing Mechanism Driven by a Unique Combination of Iron Restriction and Certain Antibiotics.

Bacterial resistance to antibiotics is precipitating a medical crisis, and new antibacterial strategies are being sought. Hypothesizing that a growth-restricting strategy could be used to enhance the efficacy of antibiotics, we determined the effect of FDA-approved iron chelators and various antibiotic combinations on invasive and multidrug-resistant extraintestinal pathogenic Escherichia coli (ExPEC), the Gram-negative bacterium most frequently isolated from the bloodstreams of hospitalized patients. We report that certain antibiotics used at sublethal concentrations display enhanced growth inhibition and/or killing when combined with the iron chelator deferiprone (DFP). Inductively coupled plasma optical emission spectrometry reveals abnormally high levels of cell-associated iron under these conditions, a response that correlates with an iron starvation response and supraphysiologic levels of reactive oxygen species (ROS). The high ROS level is reversed upon the addition of antioxidants, which restores bacterial growth, suggesting that the cells are inhibited or killed by excessive free radicals. A model is proposed in which peptidoglycan-targeting antibiotics facilitate the entry of lethal levels of iron-complexed DFP into the bacterial cytoplasm, a process that drives the generation of ROS. This new finding suggests that, in addition to restriction of access to iron as a general growth-restricting strategy, targeting of cellular pathways or networks that selectively disrupt normal iron homeostasis can have potent bactericidal outcomes. The prospect that common bacteria will become resistant to all antibiotics is challenging the medical community. In addition to the development of next-generation antibiotics, new bacterial targets that display cytotoxic properties when altered need to be identified. Data presented here demonstrate that combining subinhibitory levels of both iron chelators and certain antibiotics kills pathogenic Escherichia coli. The mechanism of this effect is the production of supraphysiologic levels of reactive oxygen species, likely powered by the excessive import of iron. These findings were consistent for both clinically relevant and no longer clinically used antibiotics and may extend to Staphylococcus aureus as well.

The complex interplay of iron, biofilm formation, and mucoidy affecting antimicrobial resistance ofPseudomonas aeruginosa

Pseudomonas aeruginosa is a Gram-negative opportunistic bacterial pathogen that is refractory to a variety of current antimicrobial therapeutic regimens. Complicating treatment for such infections is the ability of P. aeruginosa to form biofilms, as well as several innate and acquired resistance mechanisms. Previous studies suggest iron plays a role in resistance to antimicrobial therapy, including the efficacy of an FDA-approved iron chelator, deferasirox (DSX), or Gallium, an iron analog, in potentiating antibiotic-dependent killing of P. aeruginosa biofilms. Here, we show that iron-replete conditions enhance resistance of P. aeruginosa nonbiofilm growth against tobramycin and tigecycline. Interestingly, the mechanism of iron-enhanced resistance to each of these antibiotics is distinct. Whereas pyoverdine-mediated iron uptake is important for optimal resistance to tigecycline, it does not enhance tobramycin resistance. In contrast, heme supplementation results in increased tobramycin resistance, while having no significant effect on tigecycline resistance. Thus, nonsiderophore bound iron plays an important role in resistance to tobramycin, while pyoverdine increases the ability of P. aeruginosa to resist tigecycline treatment. Lastly, we show that iron increases the minimal concentration of tobramycin, but not tigecycline, required to eradicate P. aeruginosa biofilms. Moreover, iron depletion blocks the previous observed induction of biofilm formation by subinhibitory concentrations of tobramycin, suggesting iron and tobramycin signal through overlapping regulatory pathways to affect biofilm formation. These data further support the role of iron in P. aeruginosa antibiotic resistance, providing yet another compelling case for targeting iron acquisition for future antimicrobial drug development.

Promotion of airway anastomotic microvascular regeneration and alleviation of airway ischemia by deferoxamine nanoparticles

Airway tissue ischemia and hypoxia in human lung transplantation is a consequence of the sacrifice of the bronchial circulation during the surgical procedure and is a major risk factor for the development of airway anastomotic complications. Augmented expression of hypoxia-inducible factor (HIF)-1α promotes microvascular repair and alleviates allograft ischemia and hypoxia. Deferoxamine mesylate (DFO) is an FDA-approved iron chelator which has been shown to upregulate cellular HIF-1α. Here, we developed a nanoparticle formulation of DFO that can be topically applied to airway transplants at the time of surgery. In a mouse orthotopic tracheal transplant (OTT) model, the DFO nanoparticle was highly effective in enhancing airway microvascular perfusion following transplantation through the production of the angiogenic factors, placental growth factor (PLGF) and stromal cell-derived factor (SDF)-1. The endothelial cells in DFO treated airways displayed higher levels of p-eNOS and Ki67, less apoptosis, and decreased production of perivascular reactive oxygen species (ROS) compared to vehicle-treated airways. In summary, a DFO formulation topically-applied at the time of surgery successfully augmented airway anastomotic microvascular regeneration and the repair of alloimmune-injured microvasculature. This approach may be an effective topical transplant-conditioning therapy for preventing airway complications following clinical lung transplantation.