In the field of oncology research, a deeper understanding of tumor biology has shed light on the role of environmental conditions surrounding cancer cells. In this regard, targeting the tumor ...microenvironment has recently emerged as a new way to access this disease. In this work, a novel extracellular matrix (ECM)-targeting nanotherapeutic was engineered using a lipid-based nanoparticle chemically linked to an inhibitor of the ECM-related enzyme, lysyl oxidase 1 (LOX), that inhibits the crosslinking of elastin and collagen fibers. We demonstrated that, when the conjugated vesicles were loaded with the chemotherapeutic epirubicin, superior inhibition of triple negative breast cancer (TNBC) cell growth was observed both in vitro and in vivo. Moreover, in vivo results displayed prolonged survival, minimal cytotoxicity, and enhanced biocompatibility compared to free epirubicin and epirubicin-loaded nanoparticles. This all-in-one nano-based ECM-targeting chemotherapeutic may provide a key-enabling technology for the treatment of TNBC.
The advancement of nanotechnology toward more sophisticated bioinspired approaches has highlighted the gap between the advantages of biomimetic and biohybrid platforms and the availability of ...manufacturing processes to scale up their production. Though the advantages of transferring biological features from cells to synthetic nanoparticles for drug delivery purposes have recently been reported, a standardizable, batch‐to‐batch consistent, scalable, and high‐throughput assembly method is required to further develop these platforms. Microfluidics has offered a robust tool for the controlled synthesis of nanoparticles in a versatile and reproducible approach. In this study, the incorporation of membrane proteins within the bilayer of biomimetic nanovesicles (leukosomes) using a microfluidic‐based platform is demonstrated. The physical, pharmaceutical, and biological properties of microfluidic‐formulated leukosomes (called NA‐Leuko) are characterized. NA‐Leuko show extended shelf life and retention of the biological functions of donor cells (i.e., macrophage avoidance and targeting of inflamed vasculature). The NA approach represents a universal, versatile, robust, and scalable tool, which is extensively used for the assembly of lipid nanoparticles and adapted here for the manufacturing of biomimetic nanovesicles.
The microfluidic platform NanoAssemblr is used to synthesize biomimetic nanovesicles. This platform enables the high‐throughput, reproducible, and scalable production of nanoparticles, without affecting their pharmaceutical and biological properties. The versatility of this approach makes it suitable for good‐manufacturing‐practice‐compliant manufacture of biomimetic nanoparticles.
Understanding interactions occurring at the interface between nanoparticles and biological components is an urgent challenge in nanomedicine due to their effect on the biological fate of ...nanoparticles. After the systemic injection of nanoparticles, a protein corona constructed by blood components surrounds the carrier’s surface and modulates its pharmacokinetics and biodistribution. Biomimicry-based approaches in nanotechnology attempt to imitate what happens in nature in order to transfer specific natural functionalities to synthetic nanoparticles. Several biomimetic formulations have been developed, showing superior in vivo features as a result of their cell-like identity. We have recently designed biomimetic liposomes, called leukosomes, which recapitulate the ability of leukocytes to target inflamed endothelium and escape clearance by the immune system. To gain insight into the properties of leukosomes, we decided to investigate their protein corona in vivo. So far, most information about the protein corona has been obtained using in vitro experiments, which have been shown to minimally reproduce in vivo phenomena. Here we directly show a time-dependent quantitative and qualitative analysis of the protein corona adsorbed in vivo on leukosomes and control liposomes. We observed that leukosomes absorb fewer proteins than liposomes, and we identified a group of proteins specifically adsorbed on leukosomes. Moreover, we hypothesize that the presence of macrophage receptors on leukosomes’ surface neutralizes their protein corona-meditated uptake by immune cells. This work unveils the protein corona of a biomimetic carrier and is one of the few studies on the corona performed in vivo.
Despite numerous advances in medical treatment, sepsis remains one of the leading causes of death worldwide. Sepsis is characterized by the involvement of all organs and tissues as a consequence of ...blood poisoning, resulting in organ failure and eventually death. Effective treatment remains an unmet need and novel approaches are urgently needed. The growing evidence of clinical and biological heterogeneity of sepsis suggests precision medicine as a possible key for achieving therapeutic breakthroughs. In this scenario, biomimetic nanomedicine represents a promising avenue for the treatment of inflammatory diseases, including sepsis. We investigated the role of macrophage-derived biomimetic nanoparticles, namely leukosomes, in a lipopolysaccharide-induced murine model of sepsis. We observed that treatment with leukosomes was associated with significantly prolonged survival.
In vitro
studies elucidated the potential mechanism of action of these biomimetic vesicles. The direct treatment of endothelial cells (ECs) with leukosomes did not alter the gene expression profile of EC-associated cell adhesion molecules. In contrast, the interaction of leukosomes with macrophages induced a decrease of pro-inflammatory genes (IL-6, IL-1b, and TNF-α), an increase of anti-inflammatory ones (IL-10 and TGF-β), and indirectly an anti-inflammatory response on ECs. Taken together, these results showed the ability of leukosomes to regulate the inflammatory response in target cells, acting as a bioactive nanotherapeutic.
Here, we have demonstrated that biomimetic nanovesicles assembled from macrophages' membrane proteins are similar to macrophage-derived exosomes. The anti-inflammatory activity observed
in vivo
derives from their direct interaction with macrophages.
Inflammatory bowel disease (IBD), which includes Crohn's disease and ulcerative colitis, is a chronic inflammatory condition of the gastrointestinal (GI) tract. Currently, it is treated with ...immunosuppressant or biologics that often induce severe adverse effects. Thus, there is an urgent clinical need for more specific treatments. To provide a valid therapeutic tool for IBD therapy, in this work we developed biomimetic nanovesicles by manipulating leukocyte membranes to exploit mechanisms of T-cell recruitment during inflammation. A subset of T-lymphocytes participates in homing to inflamed tissue in the gastrointestinal tract by overexpressing the α4β7 integrin, which is responsible for binding to its receptor on the endothelial membrane, the mucosal addressin cell adhesion molecule 1. Based on this principle, we engineered biomimetic vesicles, referred to as specialized leukosomes (SLKs), which are leukocyte-like carriers 'doped' with the α4β7 integrin over-induced in purified immune cells. We tested SLKs in an in vivo murine model of IBD induced by treatment with dextran sulfate sodium. Notably, treatment of IBD mice with SLKs allowed us to observe a reduction of inflammation (favorable modulation of both pro- and anti-inflammatory genes, as well as reduction of immune cells infiltration into the colon tissue), and a consequent enhanced intestinal repair (low epithelial damage). In this study, we demonstrate that biological-derived nanoparticles can be used not only as naturally targeted drug delivery systems, but also as nano-therapeutics endowed with intrinsic anti-inflammatory properties.
In the last decades, several approaches were developed to design drug delivery systems to address the multiple biological barriers encountered after administration while safely delivering a payload. ...In this scenario, bio-inspired and bio-mimetic approaches have emerged as promising solutions to evade the mononuclear phagocytic system while simultaneously negotiating the sequential transport across the various biological barriers. Leukocytes freely circulate in the bloodstream and selectively target the inflamed vasculature in response to injury, infection, and cancer. Recently we have shown the use of biomimetic nanovesicles, called leukosomes, which combine both the physical and biological properties of liposomes and leukocytes, respectively, to selectively deliver drugs to the inflamed vasculature. Here we report the use of leukosomes to target and deliver doxorubicin, a model chemotherapeutic, to tumors in syngeneic murine models of breast cancer and melanoma. Exploiting the inflammatory pathway responsible for recruiting immune cells to the site of injury, leukosomes exhibited increased targeting of cancer vasculature and stroma. Furthermore, delivery of doxorubicin with leukosomes enabled significant tumor growth inhibition compared with free doxorubicin in both breast and melanoma tumors. This study demonstrates the promise of using biomimetic nanovesicles for effective cancer management in solid tumors.
Biomimetic nanovesicles deriving from leukocytes membrane proteins, called leukosomes, exhibit increased targeting of cancer vasculature and stroma by exploiting the inflammatory pathway responsible for recruiting immune cells to the cancer lesion.
Activation of the vascular endothelium is characterized by increased expression of vascular adhesion molecules and chemokines. This activation occurs early in the progression of several diseases and ...triggers the recruitment of leukocytes. Inspired by the tropism of leukocytes, we investigated leukocyte-based biomimetic nanoparticles (i.e., leukosomes) as a novel theranostic platform for inflammatory diseases.
: Leukosomes were assembled by combining phospholipids and membrane proteins from leukocytes. For imaging applications, phospholipids modified with rhodamine and gadolinium were used. Leukosomes incubated with antibodies blocking lymphocyte function-associated antigen 1 (LFA-1) and CD45 were administered to explore their roles in targeting inflammation. In addition, relaxometric assessment of NPs was evaluated.
: Liposomes and leukosomes were both spherical in shape with sizes ranging from 140-170 nm. Both NPs successfully integrated 8 and 13 µg of rhodamine and gadolinium, respectively, and demonstrated less than 4% variation in physicochemical features. Leukosomes demonstrated a 16-fold increase in breast tumor accumulation relative to liposomes. Furthermore, quantification of leukosomes in tumor vessels demonstrated a 4.5-fold increase in vessel lumens and a 14-fold increase in vessel walls. Investigating the targeting mechanism of action revealed that blockage of LFA-1 on leukosomes resulted in a 95% decrease in tumor accumulation. Whereas blockage of CD45 yielded a 60% decrease in targeting and significant increases in liver and spleen accumulation. In addition, when administered in mice with atherosclerotic plaques, leukosomes exhibited a 4-fold increase in the targeting of inflammatory vascular lesions. Lastly, relaxometric assessment of NPs demonstrated that the incorporation of membrane proteins into leukosomes did not impact the r
and r
relaxivities of the NPs, demonstrating 6 and 30 mM
s
, respectively.
: Our study demonstrates the ability of leukosomes to target activated vasculature and exhibit superior accumulation in tumors and vascular lesions. The versatility of the phospholipid backbone within leukosomes permits the incorporation of various contrast agents. Furthermore, leukosomes can potentially be loaded with therapeutics possessing diverse physical properties and thus warrant further investigation toward the development of powerful theranostic agents.
Targeted nanoparticle delivery is a promising strategy for increasing efficacy and limiting side effects of therapeutics. When designing a targeted liposomal formulation, the in vivo biodistribution ...of the particles must be characterized to determine the value of the targeting approach. Peroxisome proliferator-activated receptor (PPAR) agonists effectively treat metabolic syndrome by decreasing dyslipidemia and insulin resistance but side effects have limited their use, making them a class of compounds that could benefit from targeted liposomal delivery. The adipose targeting sequence peptide (ATS) could fit this role, as it has been shown to bind to adipose tissue endothelium and induce weight loss when delivered conjugated to a pro-apoptotic peptide. To date, however, a full assessment of ATS in vivo biodistribution has not been reported, leaving important unanswered questions regarding the exact mechanisms whereby ATS targeting enhances therapeutic efficacy. We designed this study to evaluate the biodistribution of ATS-conjugated liposomes loaded with the PPARα/γ dual agonist tesaglitazar in leptin-deficient ob/ob mice. The ATS-liposome biodistribution in adipose tissue and other organs was examined at the cellular and tissue level using microscopy, flow cytometry, and fluorescent molecular tomography. Changes in metabolic parameters and gene expression were measured by target and off-target tissue responses to the treatment. Unexpectedly, ATS targeting did not increase liposomal uptake in adipose relative to other tissues, but did increase uptake in the kidneys. Targeting also did not significantly alter metabolic parameters. Analysis of the liposome cellular distribution in the stromal vascular fraction with flow cytometry revealed high uptake by multiple cell types. Our findings highlight the need for thorough study of in vivo biodistribution when evaluating a targeted therapy.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Macrophages are important regulators of obesity-associated inflammation and PPARα and -γ agonism in macrophages has anti-inflammatory effects. In this study, we tested the efficacy with which ...liposomal delivery could target the PPARα/γ dual agonist tesaglitazar to macrophages while reducing drug action in common sites of drug toxicity: the liver and kidney, and whether tesaglitazar had anti-inflammatory effects in an
model of obesity-associated dysmetabolism.
: Male leptin-deficient (
) mice were administered tesaglitazar or vehicle for one week in a standard oral formulation or encapsulated in liposomes. Following the end of treatment, circulating metabolic parameters were measured and pro-inflammatory adipose tissue macrophage populations were quantified by flow cytometry. Cellular uptake of liposomes in tissues was assessed using immunofluorescence and a broad panel of cell subset markers by flow cytometry. Finally, PPARα/γ gene target expression levels in the liver, kidney, and sorted macrophages were quantified to determine levels of drug targeting to and drug action in these tissues and cells.
: Administration of a standard oral formulation of tesaglitazar effectively treated symptoms of obesity-associated dysmetabolism and reduced the number of pro-inflammatory adipose tissue macrophages. Macrophages are the major cell type that took up liposomes with many other immune and stromal cell types taking up liposomes to a lesser extent. Liposome delivery of tesaglitazar did not have effects on inflammatory macrophages nor did it improve metabolic parameters to the extent of a standard oral formulation. Liposomal delivery did, however, attenuate effects on liver weight and liver and kidney expression of PPARα and -γ gene targets compared to oral delivery.
: These findings reveal for the first time that tesaglitazar has anti-inflammatory effects on adipose tissue macrophage populations
. These data also suggest that while nanoparticle delivery reduced off-target effects, yet the lack of tesaglitazar actions in non-targeted cells such (as hepatocytes and adipocytes) and the uptake of drug-loaded liposomes in many other cell types, albeit to a lesser extent, may have impacted overall therapeutic efficacy. This fulsome analysis of cellular uptake of tesaglitazar-loaded liposomes provides important lessons for future studies of liposome drug delivery.
Despite an improved understanding of its pathophysiology and a wide range of new treatments, cardiovascular disease (CVD) remains a serious public health issue and the number one cause of mortality ...in the United States. Conditions that promote chronic systemic inflammation, such as obesity, cancer, and autoimmune and infectious diseases, are now known to play an important role in promoting CVD by inducing the expression of endothelial adhesion molecules and chemokines; these in turn promote leukocyte adherence and infiltration, which initiates and spurs the progression of CVD. In response to this new understanding, researchers are evaluating the potential cardiovascular benefits of new-generation therapies based on endogenous molecules with anti-inflammatory properties. Similarly, targeted approaches that leverage the phenotypic differences between non-inflamed and inflamed endothelia have the potential to selectively deliver therapeutics and decrease the morbidity and mortality of CVD patients. In this review, we discuss the role of inflammation in CVD and explore the therapeutic potential of targeting inflamed vasculature through conventional and biomimetic approaches.
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