A magnetic separation method for isolating and characterizing the biomolecular corona of lipid nanoparticles.

Lipid nanoparticle (LNP) formulations are a proven method for the delivery of nucleic acids for gene therapy as exemplified by the worldwide rollout of LNP-based RNAi therapeutics and mRNA vaccines. However, targeting specific tissues or cells is still a major challenge. After LNP administration, LNPs interact with biological fluids (i.e., blood), components of which adsorb onto the LNP surface forming a layer of biomolecules termed the "biomolecular corona (BMC)" which affects LNP stability, biodistribution, and tissue tropism. The mechanisms by which the BMC influences tissue- and cell-specific targeting remains largely unknown, due to the technical challenges in isolating LNPs and their corona from complex biological media. In this study, we present a new technique that utilizes magnetic LNPs to isolate LNP-corona complexes from unbound proteins present in human serum. First, we developed a magnetic LNP formulation, containing >40 superparamagnetic iron oxide nanoparticles (IONPs)/LNP, the resulting LNPs containing iron oxide nanoparticles (IOLNPs) displayed a similar particle size and morphology as LNPs loaded with nucleic acids. We further demonstrated the isolation of the IOLNPs and their corresponding BMC from unbound proteins using a magnetic separation (MS) system. The BMC profile of LNP from the MS system was compared to size exclusion column chromatography and further analyzed via mass spectrometry, revealing differences in protein abundances. This new approach enabled a mild and versatile isolation of LNPs and its corona, while maintaining its structural integrity. The identification of the BMC associated with an intact LNP provides further insight into LNP interactions with biological fluids.

In Situ Gelling Liquid Crystalline System as Local siRNA Delivery System.

An effective short interfering RNA (siRNA) delivery system protects the siRNA from degradation, facilitates its cellular uptake, and promotes its release into the cytoplasm. Local administration of siRNA presents advantages over systemic administration, such as the possibility to use lower doses and allow local and sustained release. In this context, in situ solidifying organogels based on monoglycerides (MO), polyethylenimine (PEI), propylene glycol (PG) and tris buffer are an attractive strategy for intratumoral delivery of siRNA. In this study, precursor fluid formulation (PFF) composed of MO/PEI/PG/tris buffer at 7.85:0.65:76.5:15 (w/w/w/w) was used to deliver siRNA to tumor cells. The internal structure of the gel obtained from PFF was characterized using small angle X-ray scattering (SAXS). In addition, its ability to complex siRNA, protect it from degradation, and functionally deliver it to tumor cells was investigated. Moreover, in vivo gel formation following intratumoral injection was evaluated. The gel formed in excess water from PFF was found to comprise a mixture of hexagonal and cubic phases. The system was able to complex high amounts of siRNA, protect it from degradation, promote siRNA internalization, and induce gene silencing in vitro in a variety of tumor cell lines. Moreover, a gel formed in situ following intratumoral injection in a murine xenograft model. In conclusion, PFF is a potential delivery system for local and sustained delivery of siRNA to tumor tissue after intratumoral administration.

Lipid Nanoparticles for Nucleic Acid Delivery Beyond the Liver.

Lipid nanoparticles (LNPs) are the most clinically advanced drug delivery system for nucleic acid therapeutics, exemplified by the success of the COVID-19 mRNA vaccines. However, their clinical use is currently limited to hepatic diseases and vaccines due to their tendency to accumulate in the liver upon intravenous administration. To fully leverage their potential, it is essential to understand and address their liver tropism, while also developing strategies to enhance delivery to tissues beyond the liver. Ensuring that these therapeutics reach their target cells while avoiding off-target cells is essential for both their efficacy and safety. There are three potential targeting strategies-passive, active, and endogenous-which can be used individually or in combination to target nonhepatic tissues. In this review, we delve into the recent advancements in LNP engineering for delivering nucleic acid beyond the liver.

mRNA delivery systems for cancer immunotherapy: Lipid nanoparticles and beyond.

mRNA-based vaccines are emerging as a promising alternative to standard cancer treatments and the conventional vaccines. Moreover, the FDA-approval of three nucleic acid based therapeutics (Onpattro, BNT162b2 and mRNA-1273) has further increased the interest and trust on this type of therapeutics. In order to achieve a significant therapeutic efficacy, the mRNA needs from a drug delivery system. In the last years, several delivery platforms have been explored, being the lipid nanoparticles (LNPs) the most well characterized and studied. A better understanding on how mRNA-based therapeutics operate (both the mRNA itself and the drug delivery system) will help to further improve their efficacy and safety. In this review, we will provide an overview of what mRNA cancer vaccines are and their mode of action and we will highlight the advantages and challenges of the different delivery platforms that are under investigation.

Biofluid specific protein coronas affect lipid nanoparticle behavior in vitro.

Lipid nanoparticles (LNPs) have successfully entered the clinic for the delivery of mRNA- and siRNA-based therapeutics, most recently as vaccines for COVID-19. Nevertheless, there is a lack of understanding regarding their in vivo behavior, in particular cell targeting. Part of this LNP tropism is based on the adherence of endogenous protein to the particle surface. This protein forms a so-called corona that can change, amongst other things, the circulation time, biodistribution and cellular uptake of these particles. The formation of this protein corona, in turn, is dependent on the nanoparticle properties (e.g., size, charge, surface chemistry and hydrophobicity) as well as the biological environment from which it is derived. With the potential of gene therapy to target virtually any disease, administration sites other than intravenous route are considered, resulting in tissue specific protein coronas. For neurological diseases, intracranial administration of LNPs results in a cerebral spinal fluid derived protein corona, possibly changing the properties of the lipid nanoparticle compared to intravenous administration. Here, the differences between plasma and CSF derived protein coronas on a clinically relevant LNP formulation were studied in vitro. Protein analysis showed that LNPs incubated in human CSF (C-LNPs) developed a protein corona composition that differed from that of LNPs incubated in plasma (P-LNPs). Lipoproteins as a whole, but in particular apolipoprotein E, represented a higher percentage of the total protein corona on C-LNPs than on P-LNPs. This resulted in improved cellular uptake of C-LNPs compared to P-LNPs, regardless of cell origin. Importantly, the higher LNP uptake did not directly translate into more efficient cargo delivery, underlining that further assessment of such mechanisms is necessary. These findings show that biofluid specific protein coronas alter LNP functionality, suggesting that the site of administration could affect LNP efficacy in vivo and needs to be considered during the development of the formulation.

Inhibition of tumor growth by targeted anti-EGFR/IGF-1R nanobullets depends on efficient blocking of cell survival pathways.

The clinical efficacy of epidermal growth factor receptor (EGFR)-targeted inhibitors is limited due to resistance mechanisms of the tumor such as activation of compensatory pathways. Crosstalk between EGFR and insulin-like growth factor 1 (IGF-1R) signaling has been frequently described to be involved in tumor proliferation and resistance. One of the attractive features of nanomedicines is the possibility to codeliver agents that inhibit different molecular targets in one nanocarrier system, thereby strengthening the antitumor effects of the individual agents. Additionally, exposure to healthy tissues and related unwanted side-effects can be reduced. To this end, we have recently developed anti-EGFR nanobody (Nb)-liposomes loaded with the anti-IGF-1R kinase inhibitor AG538, which showed promising antiproliferative effects in vitro. In the present study, we have further evaluated the potential of this dual-active nanomedicine in vitro and for the first time in vivo. As intended, the nanomedicine inhibited EGFR and IGF-1R signaling and subsequent activation of downstream cell proliferation and survival pathways. The degree of inhibition induced by the nanomedicine on a molecular level correlated with cytotoxicity in tumor cell proliferation assays and may even be predictive of the response to nanomedicine treatment in tumor xenograft models. Combination therapy with kinase inhibitor-loaded Nb-liposomes is therefore an appealing strategy for inhibiting the proliferation of tumors that are highly dependent on EGFR and IGF-1R signaling.

Fusogenic Coiled-Coil Peptides Enhance Lipid Nanoparticle-Mediated mRNA Delivery upon Intramyocardial Administration.

Heart failure is a serious condition that results from the extensive loss of specialized cardiac muscle cells called cardiomyocytes (CMs), typically caused by myocardial infarction (MI). Messenger RNA (mRNA) therapeutics are emerging as a very promising gene medicine for regenerative cardiac therapy. To date, lipid nanoparticles (LNPs) represent the most clinically advanced mRNA delivery platform. Yet, their delivery efficiency has been limited by their endosomal entrapment after endocytosis. Previously, we demonstrated that a pair of complementary coiled-coil peptides (CPE4/CPK4) triggered efficient fusion between liposomes and cells, bypassing endosomal entrapment and resulting in efficient drug delivery. Here, we modified mRNA-LNPs with the fusogenic coiled-coil peptides and demonstrated efficient mRNA delivery to difficult-to-transfect induced pluripotent stem-cell-derived cardiomyocytes (iPSC-CMs). As proof of in vivo applicability of these fusogenic LNPs, local administration via intramyocardial injection led to significantly enhanced mRNA delivery and concomitant protein expression. This represents the successful application of the fusogenic coiled-coil peptides to improve mRNA-LNPs transfection in the heart and provides the potential for the advanced development of effective regenerative therapies for heart failure.

Physicochemical and biological evaluation of siRNA polyplexes based on PEGylated Poly(amido amine)s.

PURPOSE: Use of RNA interference as novel therapeutic strategy is hampered by inefficient delivery of its mediator, siRNA, to target cells. Cationic polymers have been thoroughly investigated for this purpose but often display unfavorable characteristics for systemic administration, such as interactions with serum and/or toxicity. METHODS: We report the synthesis of a new PEGylated polymer based on biodegradable poly(amido amine)s with disulfide linkages in the backbone. Various amounts of PEGylated polymers were mixed with their unPEGylated counterparts prior to polyplex formation to alter PEG content in the final complex. RESULTS: PEGylation effectively decreased polyplex surface charge, salt- or serum-induced aggregation and interaction with erythrocytes. Increasing amount of PEG in formulation also reduced its stability against heparin displacement, cellular uptake and subsequent silencing efficiency. Yet, for polyplexes with high PEG content, significant gene silencing efficacy was found, which was combined with almost no toxicity. CONCLUSIONS: PEGylated poly(amido amine)s are promising carriers for systemic siRNA delivery in vivo.

Glucocorticoid-loaded core-cross-linked polymeric micelles with tailorable release kinetics for targeted therapy of rheumatoid arthritis.

Polymerizable and hydrolytically cleavable dexamethasone (DEX, red dot in picture) derivatives were covalently entrapped in core-cross-linked polymeric micelles that were prepared from a thermosensitive block copolymer (yellow and gray building block). By varying the oxidation degree of the thioether in the drug linker, the release rate of DEX could be controlled. The DEX-loaded micelles were used for efficient treatment of inflammatory arthritis in two animal models.

Utilizing in vitro drug release assays to predict in vivo drug retention in micelles.

In the present work, we aim at developing an in vitro release assay to predict circulation times of hydrophobic drugs loaded into polymeric micelles (PM), upon intravenous (i.v.) administration. PM based on poly (ethylene glycol)-b-poly (N-2-benzoyloxypropyl methacrylamide) (mPEG-b-p(HPMA-Bz)) block copolymer were loaded with a panel of hydrophobic anti-cancer drugs and characterized for size, loading efficiency and release profile in different release media. Circulation times in mice of two selected drugs loaded in PM were evaluated and compared to the in vitro release profile. Release of drugs from PM was evaluated over 7 days in PBS containing Triton X-100 and in PBS containing albumin at physiological concentration (40 g/L). The results were utilized to identify crucial molecular features of the studied hydrophobic drugs leading to better micellar retention. For the best and the worst retained drugs in the in vitro assays (ABT-737 and BCI, respectively), the circulation of free and entrapped drugs into PM was examined after i.v. administration in mice. We found in vivo drug retention at 24 h post-injection similar to the retention found in the in vitro assays. This demonstrates that in vitro release assay in buffers supplemented with albumin, and to a lesser degree Triton X-100, can be employed to predict the in vivo circulation kinetics of drugs loaded in PM. Utilizing media containing acceptor molecules for hydrophobic compounds, provide a first screen to understand the stability of drug-loaded PM in the circulation and, therefore, can contribute to the reduction of animals used for circulation kinetics studies.

Anti-PEG antibodies compromise the integrity of PEGylated lipid-based nanoparticles via complement.

PEGylation of lipid-based nanoparticles and other nanocarriers is widely used to increase their stability and plasma half-life. However, either pre-existing or de novo formed anti-PEG antibodies can induce hypersensitivity reactions and accelerated blood clearance through binding to the nanoparticle surfaces, leading to activation of the complement system. In this study, we investigated the consequences and mechanisms of complement activation by anti-PEG antibodies interacting with different types of PEGylated lipid-based nanoparticles. By using both liposomes loaded with different (model) drugs and LNPs loaded with mRNA, we demonstrate that complement activation triggered by anti-PEG antibodies can compromise the bilayer/surface integrity, leading to premature drug release or exposure of their mRNA contents to serum proteins. Anti-PEG antibodies also can induce deposition of complement fragments onto the surface of PEGylated lipid-based nanoparticles and induce the release of fluid phase complement activation products. The role of the different complement pathways activated by lipid-based nanoparticles was studied using deficient sera and/or inhibitory antibodies. We identified a major role for the classical complement pathway in the early activation events leading to the activation of C3. Our data also confirm the essential role of amplification of C3 activation by alternative pathway components in the lysis of liposomes. Finally, the levels of pre-existing anti-PEG IgM antibodies in plasma of healthy donors correlated with the degree of complement activation (fixation and lysis) induced upon exposure to PEGylated liposomes and mRNA-LNPs. Taken together, anti-PEG antibodies trigger complement activation by PEGylated lipid-based nanoparticles, which can potentially compromise their integrity, leading to premature drug release or cargo exposure to serum proteins.

Functional siRNA Delivery by Extracellular Vesicle-Liposome Hybrid Nanoparticles.

The therapeutic use of RNA interference is limited by the inability of siRNA molecules to reach their site of action, the cytosol of target cells. Lipid nanoparticles, including liposomes, are commonly employed as siRNA carrier systems to overcome this hurdle, although their widespread use remains limited due to a lack of delivery efficiency. More recently, nature's own carriers of RNA, extracellular vesicles (EVs), are increasingly being considered as alternative siRNA delivery vehicles due to their intrinsic properties. However, they are difficult to load with exogenous cargo. Here, EV-liposome hybrid nanoparticles (hybrids) are prepared and evaluated as an alternative delivery system combining properties of both liposomes and EVs. It is shown that hybrids are spherical particles encapsulating siRNA, contain EV-surface makers, and functionally deliver siRNA to different cell types. The functional behavior of hybrids, in terms of cellular uptake, toxicity, and gene-silencing efficacy, is altered as compared to liposomes and varies among recipient cell types. Moreover, hybrids produced with cardiac progenitor cell (CPC) derived-EVs retain functional properties attributed to CPC-EVs such as activation of endothelial signaling and migration. To conclude, hybrids combine benefits of both synthetic and biological drug delivery systems and might serve as future therapeutic carriers of siRNA.

Multi-parametric assessment of the anti-angiogenic effects of liposomal glucocorticoids.

Inflammation plays a prominent role in tumor growth. Anti-inflammatory drugs have therefore been proposed as anti-cancer therapeutics. In this study, we determined the anti-angiogenic activity of a single dose of liposomal prednisolone phosphate (PLP-L), by monitoring tumor vascular function and viability over a period of one week. C57BL/6 mice were inoculated subcutaneously with B16F10 melanoma cells. Six animals were PLP-L-treated and six served as control. Tumor tissue and vascular function were probed using MRI before and at three timepoints after treatment. DCE-MRI was used to determine K(trans), v(e), time-to-peak, initial slope and the fraction of non-enhancing pixels, complemented with immunohistochemistry. The apparent diffusion coefficient (ADC), T(2) and tumor size were assessed with MRI as well. PLP-L treatment resulted in smaller tumors and caused a significant drop in K(trans) 48 h post-treatment, which was maintained until one week after drug administration. However, this effect was not sufficient to significantly distinguish treated from non-treated animals. The therapy did not affect tumor tissue viability but did prevent the ADC decrease observed in the control group. No evidence for PLP-L-induced tumor vessel normalization was found on histology. Treatment with PLP-L altered tumor vascular function. This effect did not fully explain the tumor growth inhibition, suggesting a broader spectrum of PLP-L activities.

Betulinic acid delivered in liposomes reduces growth of human lung and colon cancers in mice without causing systemic toxicity.

Betulinic acid (BetA) is a plant-derived pentacyclic triterpenoid with potent anticancer capacity that targets the mitochondrial pathway of apoptosis. BetA has a broad efficacy in vitro against prevalent cancer types, including lung, colorectal, prostate, cervix and breast cancer, melanomas, neuroblastomas, and leukemias. The cytotoxic effects of the compound against healthy cells are minimal, rendering BetA a promising potential anticancer drug. However, because of the weak hydrosolubility of BetA, it has been difficult to study its efficacy in vivo and a pharmaceutical formulation is not yet available. We report the development of a liposome formulation of BetA and show its successful application in mice. Large liposomes, assembled without cholesterol to reduce their rigidity, efficiently incorporated BetA. Nude mice xenografted with human colon and lung cancer tumors were treated intravenously with the BetA-containing liposomes. Tumor growth was reduced to more than 50% compared with the control treatment, leading to an enhanced survival of the mice. Oral administration of the liposomal formulation of BetA also slowed tumor growth. Any signs of systemic toxicity caused by BetA treatment were absent. Thus, liposomes are an efficient formulation vehicle for BetA, enabling its preclinical development as a nontoxic compound for the treatment of cancers.

Liposomes targeting tumour stromal cells.

Liposomes have found clinical application in cancer therapy in the delivery of cytostatic agents. As a result of the targeted delivery of these toxic molecules to the tumour cells coupled to avoidance of toxicity-sensitive tissues, the therapeutic window is widened. Over the past years the focus of cancer therapy has shifted towards the stromal cells that are present in the tumour. It appears that clinically relevant tumours have acquired the ability to modulate the microenvironment in such a way that a chronic pro-inflammatory and pro-angiogenic state is achieved that contributes to invasion and metastasis and continued proliferation. Over the past years, liposomal formulations have been designed that target key stromal cell types that contribute to tumour growth. At the same time, many promising cell types have not been targeted yet and most of the studies employ drugs that aim at depleting stromal cells rather than modulating their activity towards an anti-tumour phenotype. In this review these target cell types will be addressed. Complementing these targeted formulations with the appropriate drugs to optimally suppress tumour-promoting signals while preserving anti-tumour action will be the challenge for the future.

Polymeric delivery systems for nucleic acid therapeutics: Approaching the clinic.

Gene therapy using nucleic acids has many clinical applications for the treatment of diseases with a genetic origin as well as for the development of innovative vaccine formulations. Since nucleic acids in their free form are rapidly degraded by nucleases present in extracellular matrices, have poor pharmacokinetics and hardly pass cellular membranes, carrier systems are required. Suitable carriers that protect the nucleic acid payload against enzymatic attack, prolong circulation time after systemic administration and assist in cellular binding and internalization are needed to develop nucleic acid based drug products. Viral vectors have been investigated and are also clinically used as delivery vehicles. However, some major drawbacks are associated with their use. Therefore there has been substantial attention on the use of non-viral carrier systems based on cationic lipids and polymers. This review focuses on the properties of polymer-based nucleic acid formulations, also referred as polyplexes. Different polymeric systems are summarized, and the cellular barriers polyplexes encounter and ways to tackle these are discussed. Finally attention is given to the clinical status of non-viral nucleic acid formulations.

Multimodal clinical imaging to longitudinally assess a nanomedical anti-inflammatory treatment in experimental atherosclerosis.

Atherosclerosis is an inflammatory disease causing great morbidity and mortality in the Western world. To increase the anti-inflammatory action and decrease adverse effects of glucocorticoids (PLP), a nanomedicinal liposomal formulation of this drug (L-PLP) was developed and intravenously applied at a dose of 15 mg/kg PLP to a rabbit model of atherosclerosis. Since atherosclerosis is a systemic disease, emerging imaging modalities for assessing atherosclerotic plaque are being developed. (18)F-Fluoro-deoxy-glucose positron emission tomography and dynamic contrast enhanced magnetic resonance imaging, methods commonly used in oncology, were applied to longitudinally assess therapeutic efficacy. Significant anti-inflammatory effects were observed as early as 2 days that lasted up to at least 7 days after administration of a single dose of L-PLP. No significant changes were found for the free PLP treated animals. These findings were corroborated by immunohistochemical analysis of macrophage density in the vessel wall. In conclusion, this study evaluates a powerful two-pronged strategy for efficient treatment of atherosclerosis that includes nanomedical therapy of atherosclerotic plaques and the application of noninvasive and clinically approved imaging techniques to monitor delivery and therapeutic responses. Importantly, we demonstrate unprecedented rapid anti-inflammatory effects in atherosclerotic lesions after the nanomedical therapy.

Local release of siRNA using polyplex-loaded thermosensitive hydrogels.

One of the challenges for the clinical translation of RNA interference (RNAi)-based therapies concerns the deposition of therapeutically effective doses of the nucleic acids, like siRNA, at a local tissue level without severe off-target effects. To address this issue, hydrogels can be used as matrices for the local and sustained release of the siRNA cargo. In this study, the formation of polyplexes based on siRNA and poly(2-dimethylaminoethyl methacrylate) (PDMAEMA)-based polymers was investigated, followed by their loading in a thermosensitive hydrogel to promote local siRNA release. A multifunctional NPD triblock copolymer consisting of a thermosensitive poly(N-isopropylacrylamide) (PNIPAM, N), a hydrophilic poly(ethylene glycol) (PEG, P), and a cationic PDMAEMA (D) block was used to study the binding properties with siRNA taking the non-thermosensitive PD polymer as control. For both polymers, small polyplexes with sizes ranging from 10-20 nm were formed in aqueous solution (HBS buffer, 20 mM HEPES, 150 mM NaCl, pH 7.4) when prepared at a N/P charge ratio of 5 or higher. Formulating the siRNA into NPD or PD polyplexes before loading into the thermosensitive PNIPAM-PEG-PNIPAM hydrogel resulted in a more controlled and sustained release compared to free siRNA release from the hydrogel. The polyplexes were released for 128 hours in HBS, when changing the release medium twice a day, while free siRNA was completely released within 50 hours with already 40% being released after changing the release medium just once. The release of the polyplexes was dependent on the dissolution rate of the hydrogel matrix. Moreover, intact polyplexes were released from the hydrogels with a similar size as before loading, suggesting that the hydrogel material did not compromise the polyplex stability. Finally, it was shown that the released polyplexes were still biologically active and transfected FaDu cells, which was observed by siRNA-induced luciferase silencing in vitro. This study shows the development of an injectable thermosensitive hydrogel to promote local and sustained release of siRNA, which can potentially be used to deliver siRNA for various applications, such as the treatment of tumors.

Liposomes with asymmetric bilayers produced from inverse emulsions for nucleic acid delivery.

Asymmetrical lipid nanoparticles are interesting nanocarriers for charged molecules, like nucleic acids. They promise control over inner and outer charge. High charge density on the inside is favourable for efficient condensation and charge neutralisation of highly charged biopharmaceuticals, while a neutral or slightly negative outer layer promotes biocompatibility. The main goal of this work was the development and characterisation of asymmetric liposomes, prepared using water-in-oil (w/o) nanoemulsions of phospholipids (PLs) and squalene in a centrifugal field. This method enables the control over the lipid composition of each monolayer. Liposomes were prepared by passing PL w/o nanoemulsions through an oil-water interface previously saturated with PLs. We used N-(7-Nitrobenz-2-Oxa-1,3-Diazol-4-yl)-1,2-Dihexadecanoyl-sn-Glycero-3-Phosphoethanolamine (NBD-PE) or N-(7-Nitrobenz-2-Oxa-1,3-Diazol-4-yl)-1,2-Dihexadecanoyl-sn-Glycero-3- phosphocholine (NBD-PC) as a fluorescent marker for either the inner or outer lipid layer and plasmid DNA (pDNA) as nucleic acid payload. The final liposomes had sizes below 200 nm and polydispersity indexes of 0.3 and had a bilayer asymmetry of 70%, thus shielding the charge of positive PLs in the inner bilayer leaflet. Final formulations were examined using negative staining transmission electron microscopy (TEM). Plasmid encapsulation efficiency of the method was 10-15%. Our results indicate that the w/o nanoemulsion-centrifugation method allows the successful production of liposomes with tailored features for encapsulation of nucleic acid therapeutics.

Liposomal dexamethasone inhibits tumor growth in an advanced human-mouse hybrid model of multiple myeloma.

Glucocorticoids are the cornerstone in the clinic for treatment of hematological malignancies, including multiple myeloma. Nevertheless, poor pharmacokinetic properties of glucocorticoids require high and frequent dosing with the off-target adverse effects defining the maximum dose. Recently, nanomedicine formulations of glucocorticoids have been developed that improve the pharmacokinetic profile, limit adverse effects and improve solid tumor accumulation. Multiple myeloma is a hematological malignancy characterized by uncontrolled growth of plasma cells. These tumors initiate increased angiogenesis and microvessel density in the bone marrow, which might be exploited using nanomedicines, such as liposomes. Nano-sized particles can accumulate as a result of the increased vascular leakiness at the bone marrow tumor lesions. Pre-clinical screening of novel anti-myeloma therapeutics in vivo requires a suitable animal model that represents key features of the disease. In this study, we show that fluorescently labeled long circulating liposomes were found in plasma up to 24 h after injection in an advanced human-mouse hybrid model of multiple myeloma. Besides the organs involved in clearance, liposomes were also found to accumulate in tumor bearing human-bone scaffolds. The therapeutic efficacy of liposomal dexamethasone phosphate was evaluated in this model showing strong tumor growth inhibition while free drug being ineffective at an equivalent dose (4 mg/kg) regimen. The liposomal formulation slightly reduced total body weight of myeloma-bearing mice during the course of treatment, which appeared reversible when treatment was stopped. Liposomal dexamethasone could be further developed as monotherapy or could fit in with existing therapy regimens to improve therapeutic outcomes for multiple myeloma.