Criticality of Surface Characteristics of Intravenous Iron-Carbohydrate Nanoparticle Complexes: Implications for Pharmacokinetics and Pharmacodynamics.

Un-complexed polynuclear ferric oxyhydroxide cannot be administered safely or effectively to patients. When polynuclear iron cores are formed with carbohydrates of various structures, stable complexes with surface carbohydrates driven by multiple interacting sites and forces are formed. These complexes deliver iron in a usable form to the body while avoiding the serious adverse effects of un-complexed forms of iron, such as polynuclear ferric oxyhydroxide. The rate and extent of plasma clearance and tissue biodistribution is variable among the commercially available iron-carbohydrate complexes and is driven principally by the surface characteristics of the complexes which dictate macrophage opsonization. The surface chemistry differences between the iron-carbohydrate complexes results in significant differences in in vivo pharmacokinetic and pharmacodynamic profiles as well as adverse event profiles, demonstrating that the entire iron-carbohydrate complex furnishes the pharmacologic action for these complex products. Currently available physicochemical characterization methods have limitations in biorelevant matrices resulting in challenges in defining critical quality attributes for surface characteristics for this class of complex nanomedicines.

Correction: Funk et al. Criticality of Surface Characteristics of Intravenous Iron-Carbohydrate Nanoparticle Complexes: Implications for Pharmacokinetics and Pharmacodynamics. Int. J. Mol. Sci. 2022, 23, 2140.

The authors wish to make the following corrections to this paper [...].

Kupffer Cells and Blood Monocytes Orchestrate the Clearance of Iron-Carbohydrate Nanoparticles from Serum.

Intravenous (IV) iron nanoparticle preparations are widely used to treat iron deficiency. The mechanism of mononuclear phagocyte system-mediated clearance of IV iron nanoparticles is unknown. The early uptake and homeostasis of iron after injection of ferric carboxymaltose (FCM) in mice was studied. An increase in serum iron was observed at 2.5 h followed by a return to baseline by 24 h. An increase in circulating monocytes was observed, particularly Ly6Chi and Ly6Clow. FCM was also associated with a time-dependent decrease in liver Kupffer cells (KCs) and increase in liver monocytes. The increase in liver monocytes suggests an influx of iron-rich blood monocytes, while some KCs underwent apoptosis. Adoptive transfer experiments demonstrated that following liver infiltration, blood monocytes differentiated to KCs. KCs were also critical for IV iron uptake and biodegradation. Indeed, anti-Colony Stimulating Factor 1 Receptor (CSF1R)-mediated depletion of KCs resulted in elevated serum iron levels and impaired iron uptake by the liver. Gene expression profiling indicated that C-C chemokine receptor type 5 (CCR5) might be involved in monocyte recruitment to the liver, confirmed by pharmaceutical inhibition of CCR5. Liver KCs play a pivotal role in the clearance and storage of IV iron and KCs appear to be supported by the expanded blood monocyte population.

Factors influencing safety and efficacy of intravenous iron-carbohydrate nanomedicines: From production to clinical practice.

Iron deficiency is an important subclinical disease affecting over one billion people worldwide. A growing body of clinical records supports the use of intravenous iron-carbohydrate complexes for patients where iron replenishment is necessary and oral iron supplements are either ineffective or cannot be tolerated by the gastrointestinal tract. A critical characteristic of iron-carbohydrate drugs is the complexity of their core-shell structure, which has led to differences in the efficacy and safety of various iron formulations. This review describes parameters influencing the safety and effectiveness of iron-carbohydrate complexes during production, storage, handling, and clinical application. We summarized the physicochemical and biological assessments of commercially available iron carbohydrate nanomedicines to provide an overview of publicly available data. Further, we reviewed studies that described how subtle differences in the manufacturing process of iron-carbohydrate complexes can impact on the physicochemical, biological, and clinical outcomes of original product versus their intended copies or so-called iron "similar" products.

Nano-scale characterization of iron-carbohydrate complexes by cryogenic scanning transmission electron microscopy: Building the bridge to biorelevant characterization.

Iron deficiency and iron deficiency anemia pose significant health challenges worldwide. Iron carbohydrate nanoparticles administered intravenously are a mainstay of treatment to deliver elemental iron safely and effectively. However, despite decades of clinical use, a complete understanding of their physical structure and the significance for their behavior, particularly at the nano-bio interface, is still lacking, underscoring the need to employ more sophisticated characterization methods. Our study used cryogenic Scanning Transmission Electron Microscopy (cryo-STEM) to examine iron carbohydrate nanoparticle morphology. This method builds upon previous research, where direct visualization of the iron cores in these complexes was achieved using cryogenic Transmission Electron Microscopy (cryo-TEM). Our study confirms that the average size of the iron cores within these nanoparticles is approximately 2 nm across all iron-based products studied. Furthermore, our investigation revealed the existence of discernible cluster-like morphologies, not only for ferumoxytol, as previously reported, but also within all the examined iron-carbohydrate products. The application of cryo-STEM for the analyses of product morphologies provides high-contrast and high-resolution images of the nanoparticles, and facilitates the characterization at liquid nitrogen temperature, thereby preserving the structural integrity of these complex samples. The findings from this study offer valuable insights into the physical structure of iron-carbohydrate nanoparticles, a crucial step towards unraveling the intricate relationship between the structure and function of this widely used drug class in treating iron deficiency. Additionally, we developed and utilized the self-supervised machine learning workflow for the image analysis of iron-carbohydrate complexes, which might be further expanded into a useful characterization tool for comparability studies.

Iron-carbohydrate complexes treating iron anaemia: Understanding the nano-structure and interactions with proteins through orthogonal characterisation.

Intravenous (IV) iron-carbohydrate complexes are widely used nanoparticles (NPs) to treat iron deficiency anaemia, often associated with medical conditions such as chronic kidney disease, heart failure and various inflammatory conditions. Even though a plethora of physicochemical characterisation data and clinical studies are available for these products, evidence-based correlation between physicochemical properties of iron-carbohydrate complexes and clinical outcome has not fully been elucidated yet. Studies on other metal oxide NPs suggest that early interactions between NPs and blood upon IV injection are key to understanding how differences in physicochemical characteristics of iron-carbohydrate complexes cause variance in clinical outcomes. We therefore investigated the core-ligand structure of two clinically relevant iron-carbohydrate complexes, iron sucrose (IS) and ferric carboxymaltose (FCM), and their interactions with two structurally different human plasma proteins, human serum albumin (HSA) and fibrinogen, using a combination of cryo-scanning transmission electron microscopy (cryo-STEM), x-ray diffraction (XRD), small-angle x-ray scattering (SAXS) and small-angle neutron scattering (SANS). Using this orthogonal approach, we defined the nano-structure, individual building blocks and surface morphology for IS and FCM. Importantly, we revealed significant differences in the surface morphology of the iron-carbohydrate complexes. FCM shows a localised carbohydrate shell around its core, in contrast to IS, which is characterised by a diffuse and dynamic layer of carbohydrate ligand surrounding its core. We hypothesised that such differences in carbohydrate morphology determine the interaction between iron-carbohydrate complexes and proteins and therefore investigated the NPs in the presence of HSA and fibrinogen. Intriguingly, IS showed significant interaction with HSA and fibrinogen, forming NP-protein clusters, while FCM only showed significant interaction with fibrinogen. We postulate that these differences could influence bio-response of the two formulations and their clinical outcome. In conclusion, our study provides orthogonal characterisation of two clinically relevant iron-carbohydrate complexes and first hints at their interaction behaviour with proteins in the human bloodstream, setting a prerequisite towards complete understanding of the correlation between physicochemical properties and clinical outcome.

Putting square pegs in round holes: Why traditional pharmacokinetic principles cannot universally be applied to iron-carbohydrate complexes.

Intravenous iron-carbohydrate complexes are nanomedicines that are commonly used to treat iron deficiency and iron deficiency anemia of various etiologies. Many challenges remain regarding these complex drugs in the context of fully understanding their pharmacokinetic parameters. Firstly, the measurement of the intact iron nanoparticles versus endogenous iron concentration fundamentally limits the availability of data for computational modeling. Secondly, the models need to include several parameters to describe the iron metabolism which is not completely defined and those identified (e.g. ferritin) exhibit considerable interpatient variability. Additionally, modeling is further complicated by the lack of traditional receptor/enzyme interactions. The known parameters of bioavailability, distribution, metabolism, and excretion for iron-carbohydrate nanomedicines will be reviewed and future challenges that currently prevent the direct application of physiologically-based pharmacokinetic or other computational modeling techniques will be discussed.

Uncovering the dynamics of cellular responses induced by iron-carbohydrate complexes in human macrophages using quantitative proteomics and phosphoproteomics.

Iron-carbohydrate complexes are widely used to treat iron deficiencies. Macrophages play a crucial role in the uptake and fate of these nanomedicines, however, how complexed iron carbohydrates are taken up and metabolized by macrophages is still not fully understood. Using a (phospho-)proteomics approach, we assessed differences in protein expression and phosphorylation in M2 macrophages triggered by iron sucrose (IS). Our results show that IS alters the expression of multiple receptors, indicative of a complex entry mechanism. Besides, IS induced an increase in intracellular ferritin, the loss of M2 polarization, protective mechanisms against ferroptosis, and an autophagic response. These data indicate that macrophages can use IS as a source of iron for its storage and later release, however, the excess of iron can cause oxidative stress, which can be successfully regulated by the cells. When comparing IS with ferric carboxymaltose (FCM) and iron isomaltoside-1000 (IIM), complexes with a higher carbohydrate ligand stability, we observed that FCM and IIM are metabolized at a slower rate, and trigger M2 polarization loss to a lower extent. These results indicate that the surface characteristics of the iron-carbohydrate complexes may influence the cell responses. Our data show that the application of (phospho-)proteomics can lead to a better understanding of metabolic processes, including the uptake, biodegradation and bioavailability of nanomedicines.

Critical nanomaterial attributes of iron-carbohydrate nanoparticles: Leveraging orthogonal methods to resolve the 3-dimensional structure.

Intravenous iron-carbohydrate nanomedicines are widely used to treat iron deficiency and iron deficiency anemia across a wide breadth of patient populations. These colloidal solutions of nanoparticles are complex drugs which inherently makes physicochemical characterization more challenging than small molecule drugs. There have been advancements in physicochemical characterization techniques such as dynamic light scattering and zeta potential measurement, that have provided a better understanding of the physical structure of these drug products in vitro. However, establishment and validation of complementary and orthogonal approaches are necessary to better understand the 3-dimensional physical structure of the iron-carbohydrate complexes, particularly with regard to their physical state in the context of the nanoparticle interaction with biological components such as whole blood (i.e. the nano-bio interface).

Nanoparticle iron medicinal products - Requirements for approval of intended copies of non-biological complex drugs (NBCD) and the importance of clinical comparative studies.

Currently, most countries apply the standard generic approach for the approval of intended copies of originator nanoparticle iron medicinal products, requiring only demonstration of bioequivalence to a reference medicinal product by bioavailability studies. However, growing evidence suggests that this regulatory approach is not appropriate. Clinical and non-clinical studies have shown that intended copy preparations of nanoparticle iron medicinal products can differ substantially from the originator product in their efficacy and potentially in their safety profile. An adapted regulatory pathway (separate from the standard generic approach) with defined data requirements is needed for approval of intended copies of iron medicinal products. Here, we discuss the difficulties involved in assessing therapeutic equivalence of nanoparticle iron medicinal products and suggest key concepts of a regulatory approach. Standardized non-clinical comparative studies are necessary but, as demonstrated in the reported clinical data, they may not be sufficient to demonstrate a comparable efficacy and safety profile. Validated, prospective, comparative clinical studies might be needed, in addition to non-clinical studies, in order to enable appropriate assessment of therapeutic equivalence. Furthermore, including brand names in addition to the International Non-proprietary Names (INNs) in safety reports could enable effective safety monitoring of intended copies and originator products.

Probing Subcellular Iron Availability with Genetically Encoded Nitric Oxide Biosensors.

Cellular iron supply is required for various biochemical processes. Measuring bioavailable iron in cells aids in obtaining a better understanding of its biochemical activities but is technically challenging. Existing techniques have several constraints that make precise localization difficult, and the lack of a functional readout makes it unclear whether the tested labile iron is available for metalloproteins. Here, we use geNOps; a ferrous iron-dependent genetically encoded fluorescent nitric oxide (NO) biosensor, to measure available iron in cellular locales. We exploited the nitrosylation-dependent fluorescence quenching of geNOps as a direct readout for cellular iron absorption, distribution, and availability. Our findings show that, in addition to ferrous iron salts, the complex of iron (III) with N,N'-bis (2-hydroxybenzyl)ethylenediamine-N,N'-diacetic acid (HBED) can activate the iron (II)-dependent NO probe within intact cells. Cell treatment for only 20 min with iron sucrose was also sufficient to activate the biosensor in the cytosol and mitochondria significantly; however, ferric carboxymaltose failed to functionalize the probe, even after 2 h of cell treatment. Our findings show that the geNOps approach detects available iron (II) in cultured cells and can be applied to assay functional iron (II) at the (sub)cellular level.

Need for Expansion of Pharmacy Education Globally for the Growing Field of Nanomedicine.

The emerging landscape of nanomedicine includes a wide variety of active pharmaceutical ingredients and drug formulations. Their design provides nanomedicines with unique features leading to improved pharmacokinetics and pharmacodynamics. They are manufactured using conventional or biotechnological manufacturing processes. Their physical characteristics are vastly different from traditional small-molecule drugs. Pharmacists are important members of the multi-disciplinary team of scientists involved in their development and clinical application. Consequently, their training should lead to an understanding of the complexities associated with the production and evaluation of nanomedicines. Therefore, student pharmacists, post-doctoral researchers, and trainees should be given more exposure to this rapidly evolving class of therapeutics. This commentary will provide an overview of nanomedicine education within the selection of pharmacy programs globally, discuss the current regulatory challenges, and describe different approaches to incorporate nanomedicine science in pharmacy programs around the world.

The Growing Field of Nanomedicine and Its Relevance to Pharmacy Curricula.

The field of nanomedicine is a rapidly growing scientific domain. Nanomedicine encompasses a diverse number of active pharmaceutical ingredients. Submissions of Investigational New Drugs and New Drug Applications have risen dramatically over the last decade. There are over 50 nanomedicines approved for use by the US Food and Drug Administration (FDA). Because of the fundamental role pharmacists will play in therapeutic and administrative decisions regarding nanomedicines, it is imperative for future pharmacists to gain exposure early in their training to this rapidly evolving class of drugs. This commentary describes nanomedicines, discusses current regulatory challenges, and provides recommendations for judicious incorporation of nanomedicine topics into the Doctor of Pharmacy curriculum based on emerging pharmaceutical and clinical science applications.

Tackling the challenges of nanomedicines: are we ready?

PURPOSE: This review provides an overview of the proceedings of the symposium "Tackling the Challenges of Nanomedicines: Are We Ready?" organized by the International Pharmaceutical Federation (FIP) Hospital Pharmacy Section and Non-Biological Complex Drugs (NBCDs) Working Group at the 2019 FIP World Congress of Pharmacy and Pharmaceutical Sciences. Debate centered on reasons underlying the current complex regulatory landscape for nanomedicines and their follow-on products (referred to as nanosimilars) and the pivotal role of hospital pharmacists in selecting, handling, and guiding usage of nanomedicines and nanosimilars. SUMMARY: The evaluation and use of nanomedicines are recognized among scientific, pharmaceutical, and regulatory bodies as complex. Interchangeability and substitutability of nanomedicines and nanosimilars are confounded by a lack of pharmaceutical and pharmacological equivalence, reflecting the inherent complex nature of these drug products and manufacturing processes. Consequences include implications for clinical safety and efficacy and, ultimately, comparability. Local regulatory approvals of some nanomedicines have occurred, but there is no standard to ensure streamlined evaluation and use of consistent measures of therapeutic equivalence of reference products and their nanosimilars. Hospital pharmacists are expected to be experts in the selection, handling, and substitution of nanomedicines and familiarize themselves with the limitations of current methods of assessing pharmaceutical and clinical equivalence of nanosimilars in order to ensure informed formulary decision-making and eventual patient benefit. CONCLUSION: Supportive guidance for pharmacists focusing on the substitutability and/or interchangeability of nanomedicines and their nanosimilars is needed. Current FIP guidance for pharmacists on therapeutic interchange and substitution should be extended to include nanomedicines and nanosimilars.

A pragmatic regulatory approach for complex generics through the U.S. FDA 505(j) or 505(b)(2) approval pathways.

The diverse nature of complex drug products poses challenges for the development of regulatory guidelines for generic versions. While complexity is not new in medicines, the technical capacity to measure and analyze data has increased. This requires a determination of which measurements and studies are relevant to demonstrate therapeutic equivalence. This paper describes the views of the NBCD Working Group and provides pragmatic solutions for approving complex generics by making best use of existing U.S. Food and Drug Administration's abbreviated approval pathways 505(j) and 505(b)(2). We argue that decisions on the appropriateness of submitting a 505(j) or 505(b)(2) application can build on the FDA's complex drug product classification as well as the FDA's much applauded guidance document for determining whether to submit an ANDA or a 505(b)(2) application. We hope that this paper contributes to the discussions to increase the clarity of regulatory approaches for complex generics, as well as the predictability for complex generic drug developers, to facilitate access to much-needed complex generics and to promote the sustainability of the healthcare system.

How Do Hospital Pharmacists Approach Substitution of Nanomedicines? Insights from a Qualitative Pilot Study and a Quantitative Market Research Analysis in Five European Countries.

We conducted research to assess hospital pharmacists' familiarity with/interpretation of data requirements for the different regulatory approval frameworks and the impact of this on their approach to substitution in the formulary. The online questionnaire included a small molecule (acetylsalicylic acid-follow-ons approved via the generic pathway), two biologic drugs (insulin glargine and etanercept-follow-ons approved via the biosimilar pathway), a non-biologic complex drug (NBCD; glatiramer acetate-follow-ons approved via the hybrid pathway) and a nanomedicine, ferric carboxymaltose (no follow-ons approved as yet). The study was conducted in two phases: an initial qualitative pilot study with 30 participants, followed by a quantitative stage involving 201 pharmacists from five European countries. Most expected negligible safety/efficacy differences between reference and follow-on products. Head-to-head clinical data showing therapeutic equivalence as a prerequisite for reference product/follow-on substitution was perceived to be needed most for biologics (47%), followed by NBCDs (44%)/nanomedicines (39%) and small molecules (23%). Overall, 28% did not know the data requirements for follow-on approval via the hybrid pathway; 16% were familiar with this pathway, compared with 50% and 55% for the generic and biosimilar pathways, respectively. Overall, 19% of respondents thought the European Medicines Agency (EMA) was responsible for defining the substitutability of follow-ons. Education is required to increase hospital pharmacist's knowledge of regulatory approval frameworks and their relevance to substitution practices.

Nanomedicines: The magic bullets reaching their target?

Nanomedicines, since the approval of the first one in the 1950s, have been accompanied by expectations of higher efficiency and efficacy, compared to less complex drugs. The fulfilment of those expectations has been slower than anticipated, due to the high complexity of nanomedicine drugs combined with a lack of scientific understanding of nanomedicine interactions with biological systems. The unique properties of their size and their surface composition create difficulties in their physicochemical characterization, and as a consequence, difficulty in assessing the similarity of follow-on products (nanosimilars) to originator nanomedicines. During the 2018 European Federation for Pharmaceutical Sciences (EUFEPS) annual meeting "Crossing the barrier for future medicines" in Athens, there were several sessions on nanomedicines organised by the EUFEPS Nanomedicine Network. This review focuses on the session "Nanomedicines and nanosimilars: how to assess similar?", discussing the nature of nanomedicines, the regulatory aspects of the topic and the impact of practical use and handling of such medicinal products. Emphasis is put on the consequences their nanosize-related properties have on the establishment of their critical quality attributes and how this affects the demonstration of bioequivalence of nanosimilars to their originator products. The lack of an appropriate and harmonized regulatory evaluation procedure and the absence of corresponding education are also discussed, especially the uncertainty surrounding the practical use of nanosimilars, including the higher healthcare cost due to less than satisfactory number of safe and efficacious nanosimilars in the market.

Differences between intravenous iron products: focus on treatment of iron deficiency in chronic heart failure patients.

Iron deficiency is the leading cause of anaemia and is highly prevalent in patients with chronic heart failure (CHF). Iron deficiency, with or without anaemia, can be corrected with intravenous (i.v.) iron therapy. In heart failure patients, iron status screening, diagnosis, and treatment of iron deficiency with ferric carboxymaltose are recommended by the 2016 European Society of Cardiology guidelines, based on results of two randomized controlled trials in CHF patients with iron deficiency. All i.v. iron complexes consist of a polynuclear Fe(III)-oxyhydroxide/oxide core that is stabilized with a compound-specific carbohydrate, which strongly influences their physico-chemical properties (e.g. molecular weight distribution, complex stability, and labile iron content). Thus, the carbohydrate determines the metabolic fate of the complex, affecting its pharmacokinetic/pharmacodynamic profile and interactions with the innate immune system. Accordingly, i.v. iron products belong to the new class of non-biological complex drugs for which regulatory authorities recognized the need for more detailed characterization by orthogonal methods, particularly when assessing generic/follow-on products. Evaluation of published clinical and non-clinical studies with different i.v. iron products in this review suggests that study results obtained with one i.v. iron product should not be assumed to be equivalent to other i.v. iron products that lack comparable study data in CHF. Without head-to-head clinical studies proving the therapeutic equivalence of other i.v. iron products with ferric carboxymaltose, in the highly vulnerable population of heart failure patients, extrapolation of results and substitution with a different i.v. iron product is not recommended.

Medication practice in hospitals: are nanosimilars evaluated and substituted correctly?

INTRODUCTION: This study investigates the drug selection and dispensing behaviour of hospital pharmacists of intravenous iron products including iron sucrose and iron sucrose similar, with special emphasis on substitution and interchangeability in France and Spain. Iron-carbohydrate complex drugs represent different available intravenous iron drugs and are part of the non-biological complex drug (NBCD) class, an expanding drug class with up to 30 brands available in intravenous pharmacotherapy and over 50 in clinical development. Follow-on versions of iron sucrose have appeared in some markets such as France and Spain, which were authorised by the generic approval pathway. However, differences in clinical efficacy and safety of iron sucrose similars compared with the reference originator drug Venofer have been observed, putting a question mark on their equivalence as assessed for authorisation and consequently their substitutability and interchangeability. METHOD: 70 French and 70 Spanish hospital pharmacists were surveyed via an online questionnaire on their formulary selection and dispensing behaviour of intravenous iron medicines. RESULTS: There is little awareness about the characteristics of this class of drugs and the reported differences in safety and efficacy between iron sucrose and iron sucrose similars. In approximately 85% of cases the intravenous iron is chosen according to the hospital formulary. In 30% (France) and 34% (Spain) of cases an iron sucrose similar was dispensed because the formulary requires dispensing an alternative lower cost drug when available. In 26% (France) and 52% (Spain) of cases the physician is not informed on such a medication change using a similar product. CONCLUSIONS: Evaluation of NBCD similars for substitution and interchange by hospital pharmacists is rarely based on scientific and clinical criteria but rather on cost aspects only, which does not ensure safe, efficacious and cost-effective use of such drugs.

Correction to: Reflections on FDA Draft Guidance for Products Containing Nanomaterials: Is the Abbreviated New Drug Application (ANDA) a Suitable Pathway for Nanomedicines?

In the published article the given name and the family name for each author is listed in the incorrect order and therefore cited incorrectly. The correct order (given name followed by family name) of names is listed above.