Download Immunotoxicity derived from manipulating leukocytes with lipid

ADR-12332; No of Pages 11
Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
Contents lists available at SciVerse ScienceDirect
Advanced Drug Delivery Reviews
journal homepage: www.elsevier.com/locate/addr
Immunotoxicity derived from manipulating leukocytes with
lipid-based nanoparticles☆
Dan Peer ⁎
Laboratory of Nanomedicine, Department of Cell Research and Immunology, George S. Wise Faculty of Life Science, Tel Aviv University, Tel Aviv 69978, Israel
Center for Nanoscience and Nanotechnology, Tel Aviv University, Tel Aviv 69978, Israel
a r t i c l e
i n f o
Article history:
Received 14 May 2012
Accepted 14 June 2012
Available online xxxx
Keywords:
Lipid-based nanoparticles
Liposomes
Immune response
Leukocytes
RNAi
T cells
Antigen-presenting cells
a b s t r a c t
Lipid-based nanoparticles (LNPs) such as liposomes, micelles, and hybrid systems (e.g. lipid-polymer) are
prominent delivery vehicles that already made an impact on the lives of millions around the globe. A common denominator of all these LNP‐based platforms is to deliver drugs into specific tissues or cells in a pathological setting with minimal adverse effects on bystander cells. All these platforms must be compatible to
the physiological environment and prevent undesirable interactions with the immune system. Avoiding immune stimulation or suppression is an important consideration when developing new strategies in drug and
gene delivery, whereas in adjuvants for vaccine therapies, immune activation is desired. Therefore, profound
understanding of how LNPs elicit immune responses is essential for the optimization of these systems for various biomedical applications. Herein, I describe general concepts of the immune system and the interaction of
subsets of leukocytes with LNPs. Finally, I detail the different immune toxicities reported and propose ways to
manipulate leukocytes’ functions using LNPs.
© 2012 Elsevier B.V. All rights reserved.
Contents
1.
2.
3.
4.
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Immune recognition . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.1.
Pattern recognition receptors (PRRs) . . . . . . . . . . . . . . . . .
2.2.
The adaptive immune arm . . . . . . . . . . . . . . . . . . . . . .
Lipid-based nanoparticles . . . . . . . . . . . . . . . . . . . . . . . . . .
3.1.
Structural and physicochemical properties of LNPs determine the type of
3.1.1.
LNPs size distribution . . . . . . . . . . . . . . . . . . . .
3.1.2.
Formulation additives and contaminations . . . . . . . . . .
3.1.3.
LNPs surface charge . . . . . . . . . . . . . . . . . . . . .
3.1.4.
Aggregation of LNPs. . . . . . . . . . . . . . . . . . . . .
3.1.5.
Route of administration . . . . . . . . . . . . . . . . . . .
3.2.
LNPs may stimulate or suppress the immune system. . . . . . . . . .
Modulating the immune response by LNPs as a potential therapeutic strategy .
4.1.
Immunostimulation by LNPs . . . . . . . . . . . . . . . . . . . . .
4.2.
Modulating the immune response with RNAi-LNPs strategies . . . . . .
4.2.1.
Targeted delivery into pan leukocytes . . . . . . . . . . . .
4.2.2.
Targeting subsets of leukocytes . . . . . . . . . . . . . . .
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immune response
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Abbreviations: mAb, monoclonal antibody; BD, biodistribution; DOTAP, N-[1-(2,3-dioleoyloxy)-propyl]-N,N,N- trimethylammonium methylsulfate; DOTMA, 3b-[N-(N′,N
′-dimethy lainin(ethyl) carbamoyl; dsRNA, double stranded ribonucleic acid; EPR, enhanced permeability and retention; I.V, intravenous; miRNA, micro ribonucleic acid; MPS,
mononuclear phagocytic system; MW, molecular weight; NLR, Nod‐like receptors; NP, nanoparticle; nt, nucleotide; PEG, polyethylene glycol; PEI, polyethyleneimine; PC, phosphatidylcholine; PK, pharmacokinetics; RES, reticuloendothelial system; RISC, RNA-induced silencing complex; RNA, ribonucleic acid; shRNA, short hairpin ribonucleic acid; siRNA,
small interfering RNA; SNALP, stable lipid-nucleic acid particle; T1/2, half time; TH, T helper lymphocytes; TLR, Toll-like receptor.
☆ This review is part of the Advanced Drug Delivery Reviews theme issue on “Nanotoxicity: from bench to bedsite.”
⁎ Tel.: +972 3640 7925; fax: +972 3640 5926.
E-mail address: [email protected].
0169-409X/$ – see front matter © 2012 Elsevier B.V. All rights reserved.
doi:10.1016/j.addr.2012.06.013
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
2
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
5.
Conclusions and future prospective . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Acknowledgement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1. Introduction
Profound understanding of the immune system and the interaction of various materials with subsets of immune cells (leukocytes)
is important for optimizing novel drug delivery systems.
The immune system guards the body from internal and external intruders by holding the host protection mechanism for self-defense
against infections and malignancies. This is a fine-tuning mechanism
that allows leukocytes to respond to cellular and extracellular stimuli,
identify pathogens, and respond to different types of materials with various size, geometry, surface curvature and charge [1–5]. Here, I will describe the general mechanisms by which leukocytes interact with
lipid-based nanoparticles (LNPs) and detail immuno-toxicities caused
by these nanomaterials. Finally, I will suggest ways to manipulate leukocytes’ function using LNPs loaded with various types of drugs.
2. Immune recognition
The immune system can be divided into two divisions: the innate
arm and the adaptive arm. The innate immune arm constitutes the
first host defense line, mainly composed of the phagocytic cells and
circulating macrophages (derived from blood monocytes) [6] and of
the complement system (which is often considered as the link between the innate and the adaptive arms). The “complement cascade”
is a complex multi-component system composed of 26 proteins,
which combine with antibodies or cell surfaces, is constitutive and
nonspecific but must be activated in order to function [7,8]. Macrophages mediate crucial innate immune responses via the clearance
mechanism: caspase-1-dependent processing and secretion of interleukin (IL)-1β and IL-18. The second arm of the immune system,
the adaptive immune arm, is able to response in a highly specific
manner against molecular determinants on pathogens and this process may proceed over weeks [9].
The immune response against a potential threat (internal or external) can be divided into stimulation or suppression of the immune system. Immune stimulating via activation of the innate or adaptive
immune arms can be caused by nanoparticles (NPs) made from different types of materials, having various sizes, shapes and surface charges
in a variety of molecular recognition mechanisms [1,3,8,10,11]. It is possible to utilize the type of response (either stimulation or suppression)
for therapeutic purposes [1,12]. When provoking a stimulation response the impact of the therapeutic efficacy might be effected, such
as in the case of cancer treatment and vaccine efficacy. In contrast, robust immune stimulation might cause an undesirable response, for example, when lipid-based nanoparticles (LNPs) interact with specific cell
surface receptors on subsets of leukocytes [1,10,13,14] and can induce
cytokine storm, interferon response and/or lymphocyte activation causing severe adverse effects that may diminish the therapeutic effect. Correspondingly, a suppression response can be desired and as such may
enhance the therapeutic benefits of treatments for allergies and autoimmune diseases as well as prevent rejection of transplanted organs
[1,3–5] or may be inadvertent and lower the body's defense against infection and cancerous cells.
0
0
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tissue. This interaction may lead to signal cascades upon activation of
pattern recognition receptors (PRRs) [15–17]. PRRs are proteins
expressed by cells of the innate immune arm to identify
pathogen-associated molecular patterns (PAMPs), which are associated with microbial pathogens or cellular stress. Other types of receptors are the damage-associated molecular patterns (DAMPs), which
are associated with cell components released during cell damage
(Fig. 1). On the bases of their function, PRRs may be divided into signaling PRRs or endocytic PRRs.
Signaling PRRs include the large families of membrane-bound
Toll-like receptors (TLRs) [18–20] that can recognize nucleic acids
(single and double stranded), parts of bacteria's well such as
lipopolysaccharides (LPS), several types of charged phospholipids
and more [13,21]. In addition, cytoplasmic NOD-like receptors
(NLRs) for nucleotide oligomerization domain receptors are cytoplasmic proteins that may have a variety of functions in regulation of inflammation and apoptotic responses [15,22]. Endocytic PRRs promote
the attachment, engulfment and destruction of microorganisms by
phagocytes, without relaying an intracellular signal. These types of
PRRs recognize carbohydrates and include mannose receptors of
macrophages, glucan receptors present on all phagocytes and scavenger receptors that recognize charged ligands and are found on all
phagocytes and mediate removal of apoptotic cells [23,24]. If the
NPs indeed interact similar to some pathogens (such as in the case
of several cationic lipids [13,21]) then it is highly possible that a similar cascade that may activate PAMPs and DAMPs by a pathogen can
occur by interaction between cationic NPs in the PRRs (Fig. 1). This
highly possible activation is described below.
Upon interaction of the NPs with endothelial cells or cells in a particular tissue, PAMPs and DAMPs bind to Toll-like receptors (TLRs)
expressed by innate immune cells, such as dendritic cells (DCs)
[17]. This binding promotes DC maturation and migration to lymph
nodes. In the lymph nodes, the mature DCs release cytokines
(proinflammatory), such as interleukin-12 (IL-12), which induces T
helper 1 (TH1) cell differentiation, and IL-6, which works together
with transforming growth factor-β (TGF-β) to promote the differentiation of TH17 cells. Then, T cells that undergone differentiation
leave the lymph nodes and migrate through the bloodstream to tissues, where they are further activated by encounter with self‐antigens presented by antigen-presenting cells. Then, in the already
inflamed tissues, TH17 cells are further activated by a panel of cytokines such as IL-1 and IL-23, but also by IL-17 and IL-21 produced
early in the immune response by subsets of T cells (γδ T cells) and
other innate lymphoid cells (ILCs). Granulocyte–macrophage
colony-stimulating factor (GM-CSF) that is produced by both TH1
and TH17 cells activates CD11b + myeloid cells, DCs and macrophages.
Cytokines produced by TH1 and TH17 cells activate macrophages, DCs
and monocytes, promoting the release of proinflammatory mediators,
such as IL-1β, tumor necrosis factor (TNF), matrix metalloproteinases
(MMPs) and reactive oxygen species (ROS), which all mediate tissue
damage. IL-17 induces the production of chemokines, especially
CXC-chemokine ligand 2 (CXCL2; also known MIP2) and IL-8 (also
known as CXCL8), which recruit neutrophils to the site of inflammation (Fig. 1).
2.1. Pattern recognition receptors (PRRs)
2.2. The adaptive immune arm
Immune stimulation often initiates when NPs interact with cells of
the innate immune arm such as dendritic cells and macrophages in a
similar manner to a pathogen infection or being sensed as a damaged
The second defense line, the adaptive arm, involves
antigen-mediated T- and B-lymphocytes carrying antigen-specific
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
surface receptors (e.g. TH cells and cytotoxic T lymphocytes; CTLs). TH
cells are composed of several subsets of T cells such as TH1 [25], TH2
[26], TH17 [25,27], Tregs [27], TH9 [28] and the newly identified TH22
cells [29]. These subsets of T cells are defined according to their
3
production of lineage-indicating cytokines and functions. For example, TH1 cells mediate a cellular immune response, which is mainly
a proinflammatory response by producing cytokines such as interleukin (IL)-2, interferon (IFN) γ and tumor necrosis factor (TNF)-α. TH2
Fig. 1. Effector cells signaling cascades upon interaction of cationic LNPs with leukocytes. (A) Due to LNP interaction with a specific tissue, which might be similar to a pathogen
interaction, pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPS) bind to Toll-like receptors (TLRs) expressed by innate immune
cells, such as dendritic cells (DCs). This promotes DC maturation and migration to the lymph nodes. (B) In the lymph nodes, mature DCs release proinflammatory cytokines, including interleukin-12 (IL-12), which induces T helper 1 (TH1) cell differentiation, and IL-6, which works together with transforming growth factor-β (TGF-β) to promote the differentiation of TH17 cells. Differentiated autoreactive T cells exit the lymph nodes and migrate through the blood to tissues, where they are further activated by encounter with
self-antigens presented by antigen-presenting cells. (C) In the inflamed tissues, TH17 cells are further activated by IL-1 and IL-23, but also by IL-17 and IL-21 produced early in
the immune response by γδ T cells and other innate lymphoid cells (ILCs). Granulocyte–macrophage colony-stimulating factor (GM-CSF) that is produced by both TH1 and TH17
cells activates CD11b+ myeloid cells. DCs and macrophages and appears to be essential for the development of autoimmunity. Cytokines produced by TH1 and TH17 cells activate
macrophages, DCs and other cells, promoting the release of proinflammatory mediators, such as IL-1β, tumor necrosis factor (TNF), matrix metalloproteinases (MMPs) and reactive
oxygen species (ROS), which mediate tissue damage. IL-17 induces the production of chemokines, especially CXC-chemokine ligand 2 (CXCL2; also known MIP2) and IL-8 (also
known as CXCL8), which recruit neutrophils to the site of inflammation.
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
4
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
cells secret anti-inflammatory cytokines such as IL-4, IL-5, IL-10 and
IL-13 and also support the production of circulating antibodies. Another set of newly discovered subgroup is the TH17 group that is responsible for regulation of the inflammatory response and these
subsets of T cells secret mainly IL-17, IL-6 and transforming growth
factor (TGF)‐β [30]. T regulatory cells (Tregs) are CD4 +CD25 +FOXP3 +
cells and are involved in shutting down immune responses after they
have successfully tackled invading organisms, and also in regulating
immune responses that may potentially attack one's own tissues
[27]. The TH1–TH2–TH17 cell paradigm now includes a fourth subset
of IL-9 producing effector T cells, TH9 cells [28,31], raising questions
about the plasticity of T helper cell subsets. TH9 cells are generated
under the presence of IL-4 and TGF-β1, but the costimulatory signals
that induce TH9 cell differentiation and the transcriptional regulation
of these cells are still not known.
Newly identified TH22 cells [29] are a subset of human TH cells that
infiltrates the epidermis in individuals with inflammatory skin disorders and are characterized by the secretion of IL-22 and TNF-α, but
not IFN-γ, IL-4, or IL-17. These cells might have a role in skin diseases
and thus might be important to study when applying topically
nanoparticles in the context of skin disorders.
At least two types of signals are required for the activation of T
lymphocytes to proliferate and differentiate into functional effectors
cells: A specific signal of T cells recognition by T cell receptors peptide
fragments bound to the major histocompatibility complex (MHC)
class I and class II molecules and an interaction signal of receptors
with costimulatory molecules on antigen-presenting cell (APC) surface (e.g., B7 family and inducible costimulators ligands (ICOS)). The
extra signals activate the priming and differentiation of cytotoxic T
lymphocytes (CTLs) and Th cells. Th cells deliver antigen‐specific B
cells resulting in antibody production [6,27,32].
3. Lipid-based nanoparticles
Liposomes, micelles, solid lipid-base nanoparticles (SLNPs) and
polymersomes are nano-scale lipid-based particles. Liposomes, the
most veterans delivery vehicles, were described more than 40 years
ago by Bangham et al. [33–35] and were the first to be approved by
the FDA to carry several types of chemotherapeutics (see Table 1)
[36,37]. Liposomes are spherical, self-closed structures formed by
one or several concentric lipid bilayers with aqueous phases inside
and between the lipid bilayers. The properties of liposomes are highly
attributable to their physicochemical properties such as size, surface
charge, composition, rigidity of bilayer and preparation methods
[38]. These integrated liposome features enable the encapsulation,
embedding or association with a wide range of molecules (i.e., small
molecules drugs, antigens, proteins and nucleotides) as well as enhance the delivery of therapeutic payloads into specific tissues and
cells. Liposomes also improve in vitro and in vivo stability and reduce
adverse effects [39]. Several challenges exist with these carriers, including rapid clearance, serum instability (dependent on the specific
formulation), and nonspecific uptake by the mononuclear phagocytic
system (MPS), whose function is to remove foreign materials from
the circulation. To overcome these limitations, poly (ethylene glycol)
(PEG) was introduced for coating lipid-based carriers to provide a hydrophilic layer resulting in increased circulation time [40,41]. To further enhance the selectivity to cell surface markers, targeting
moieties were attached to the liposomal surface via a PEG spacer
arm, which reduced steric hindrances [42]. Instances of selective
targeting using lipid-based carriers include anti-CD19 and
anti-CD20 modified long-circulating liposomes that deliver doxorubicin to B cell lymphoma [43] and anti-Her2 liposomes that show
effective targeting to breast cancer cells overexpressing erbB2
receptor [44]. Other carriers, including folate-targeted liposomes,
have been also proposed as vehicles for boron neutron capture therapy [45]. Transferrin (Tf)-mediated liposome delivery is an emerging
strategy to target tumors overexpressing the Tf receptors; Tf-coated
liposomes entrapping doxorubicin have been developed to target C6
glioma cells [46] and solid tumors [47]. Additional approaches include
the combination of fusogenic peptides, long-circulating agents such
as PEG, targeting moieties, and pH sensitive lipids that will cause liposome disruption in the endosome [48,49]. The length of shelf life is
one of the challenges facing drug-containing liposomes. Existing liposomal formulations can be stored as suspensions (with a limited shelf
life) or as lyophilized powder. In the latter, the lyophilized formulation includes up to 30% v/v sugars as cryo-protectants to maintain
the carriers’ nano-scale dimensions upon rehydration. High sugar
content in the nanocarrier system may limit the number of patients
who can receive these nanocarriers, particularly those patients who
suffer from diabetes. Recent attempts to address these challenges involve the covalent immobilization of high molecular weight
hyaluronan (HA) on unilamellar liposomes. This serves to
cryo-protect nano-scale liposomes in the process of lyophilization
and rehydration [50]. In addition, the HA coatings improve circulation
time and enhance targeting to HA receptor-expressing tumors (CD44
and RHAMM) [51–56]. In addition, HA was also used as a scaffold and
a linker for monoclonal antibody binding to nanoliposomes’ surface
directing these particles into subsets of leukocytes in vivo while delivering RNAi payloads [14,57,58].
3.1. Structural and physicochemical properties of LNPs determine the
type of immune response
Physicochemical properties of LNPs such as size, geometry, lipid
composition, charge, surface characteristics, bilayer packing and defects, as well as LNPs quantity, dose and way of administration, can
all act as immunological adjuvant and trigger a robust immune response [11] (see also Table 2). When developing an LNP formulation
for clinical application, the prevention of unwanted adverse immune
effects can be ruled out by taking into consideration several crucial
parameters.
3.1.1. LNPs size distribution
Size distribution has been suggested as the leading parameter that
determines the potential to induce cytokine expression [5,59–61]. For
example, particles that were prepared at a narrow size range (e.g., 20,
40, 49, 67, 93, 101, and 123 nm in diameter) were shown to determine whether antigens loaded into NPs enhance cellular type I interferon (IFN) γ response or antibody type II (IL-4) cytokine response.
After a single immunization in mice with 40 and 49 nm beads
containing ovalbumin (OVA) antigen, IFNγ response was significantly
lower than other particles sizes tested. In contrast, IL-4 response to
OVA was higher after immunization with OVA conjugated to larger
beads (93, 101, and 123 nm) [60].
Table 1
FDA approved liposomal drugs on the market (many more are under clinical
evaluation).
Compound
Commercial
name ™
Indications
Daunorubicin
Doxorubicin
DaunoXome Kaposi's sarcoma
Mycet
Combinational therapy of recurrent breast
cancer
Doxorubicin
Doxil/
Refractory Kaposi's sarcoma; recurrent breast
(PEG-liposomes) Caelyx
cancer; ovarian cancer, multiple myeloma
Amphotericin B
AmBisome
Fungal infections
Cytarabine
DepoCyt
Lymphomatous meningitis
Vincristine
Onco TCS
Relapsed aggressive non-Hodgkin's lymphoma
(NHL)
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
3.1.2. Formulation additives and contaminations
Several studies demonstrated that induction of cytokines can be
triggered not only by the liposomes but also by surfactants or bacterial endotoxins present in the liposome formulation [75,76]. Schöler
et al. showed that cytotoxicity was strongly influenced by the surfactant used being marketed with cetylpyridinium chloride (CPC) coated
solid lipid nanoparticles (SLN) at a concentration of 0.001% and
above. All other SLN formulations, containing Poloxamine 908
(P908), Poloxamer 407 (P407), Poloxamer 188 (P188), Solutol HS15
(HS15), Tween 80 (T80), Lipoid S75 (S75), sodium cholate (SC), or sodium dodecylsulfate (SDS), used at the same concentrations, modestly reduced the cell viability [75]. Therefore, it is important to quantify
the presence of byproducts and endotoxins contaminants before administration of any formulation into a vertebrate.
3.1.3. LNPs surface charge
Among non-viral delivery systems for oligonucleotides (e.g., plasmid
DNA, DNA and RNA anti-sense, siRNAs and other oligonucleotides), cationic lipids have been widely used as part of the formulation [49,63,77].
Cationic lipids are lipid-bilayer‐forming vesicles with a positive surface
charge. Common cationic lipids that were extensively studied included
1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and N-(1-(2,3Dioleoyloxy)propyl)-N,N,N-trimethyl-ammoniummethyl
sulphate
(DOTMA), 3b-[N-(N′,N′-dimethy lainin(ethyl) carbamoyl cholesterol
(DC-Chol), and dimethyldioctadecylammonium bromide (DDAB) but
the variety of cationic lipids and lipid-like novel materials is enormous
[78,79].
Cationic liposomes showed a superior adjuvant effect compared to
anionic or neutral liposomes as demonstrated in vivo in animal
models [64,66]. Since cationic liposomes can activate the complement
system and cause rapid clearance by macrophages of the mononuclear phagocytic system (MPS) [63], these cationic particles can be used
to enhance and modulate the immune response in a desirable direction and represent an efficient tool when designing tailor-made adjuvant for a specific disease target. The use of cationic liposomes in vivo
elicited dose-dependent toxicity, as demonstrated by the multivalence cationic liposome, LipofectAMINE2000 a comparison with the
monovalence cationic lipids, such as DOTAP [62,65]. While it was
found that DOTAP-based cationic liposomes also caused severe damage to the mitochondria [67], the specific mechanism by which this
cytotoxicity occur is still unknown and might involve different cellular pathways depending on the particular cell type. In addition, mice
treated with positively charged LNP‐containing DOTAP showed increased liver enzyme release and mild body weight loss compared
to mice treated with neutral or negatively charged LNPs [1,13]. Intravenous administration of cationic LNPs induced type I IFN response
and elevated mRNA levels of interferon responsive genes 15–25 fold
higher than neutral and negatively charged NPs in mouse
splenocytes. Treatment with cationic LNPs provoked a dramatic
Table 2
Different types of immune responses generated when LNPs interact with subsets of
leukocytes.
Type of an immune response
Liposome
feature
Parameter
examined
Induction of various types of
cytokines
Proinflammatory response
inducing TH1 cytokine
expression
Complement activation
Liposome size
Diameter > 100 nm [59–61]
Liposome
surface's
charged
Liposome
aggregation
Induced a robust high IgG1, Route of
IgG2a and IFN-γ response
administration
Lymphocyte activation
Liposomes’
surface charge
Cationic lipids:
DOTAP
DOTMA
PEG
References
[13,62–67]
[11,68–70]
Intralymphatic
[71,72]
injection
Negatively charged [73,74]
PS-containing
liposomes
5
proinflammatory response by inducing TH1 cytokine expression
(IL-2, IFNγ and TNFα) 10- to 75-fold higher than treatment with control particles (neutral or negatively charged particles). Using TLR4
knockout mice, it was shown that the induction of cytokines and
IFN response to cationic liposomes decreased dramatically [13],
strengthening the hypothesis suggested by Ruysschaert several
years ago that cationic liposomes can agonize TLR4 [80].
PEGylated liposomes, which are mildly negatively charged under
physiological conditions [81], have been extensively used as drug delivery carriers. Their advantage in improve circulation time of the
entrapped therapeutic payload is well known [82,83]. However, it
was observed that when PEGylated liposomes were injected into
mice, rats and rhesus monkeys repeatedly, they lost this long circulation feature and was accumulated in the liver, a phenomenon which
was termed accelerated blood clearance (ABC) [84,85]. Ishida et al.
suggested a possible mechanism of action for the ABC phenomenon.
The empty PEGylated liposomes elicited PEG-specific IgM response,
which are produced in the spleen in response to the first injection, selectively bind to the PEG on a second injected liposome dose and subsequently activate the complement system. An opsonization of
liposomes by C3 fragment occurred, which lead to an enhance uptake
of the liposomes by specialized macrophages in the liver known as
the Kupffer cells [86].
The surface charge of the LNPs affects also the tissue specificity of
the particles uptake. Macrophages seem to preferentially take up negatively charged LNPs [73,81,87]. Different malignant cell lines have
different uptake patterns with respect to positive, neutral, or negative
charges, and in vivo uptake patterns can be further different [87]. Anionic liposomes interact with a limited fraction of dendritic cells in
vitro, whereas cationic liposomes interact with a high percentage of
the dendritic cells, probably by means of electrostatic binding to the
negatively charged surface heparane sulfate proteoglycans resulting
in intracellular localization [88].
1,2-Dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC), a neutral
phospholipid‐forming liposome, was used to deliver siRNA payloads
in vivo into tumor cells [77,89–92] 10- and 30-fold more effectively
than cationic, DOTAP-based liposomes and naked siRNA, respectively
[93]. DOPC-based nano-scaled liposomes (~ 100 nm in diameter) did
not induce any detectable toxicity and were found to be safe in
orthotopic mouse models, making them highly attractive for further
therapeutic development [94].
Phagocytic cells fervently take up anionic liposomes in an unclear
mechanism. When entrapping nucleic acids such as siRNAs or DNA
the loading efficiency into these liposomes is quite low, probably
due to the negative charge of these molecules [1,87,95].
Clearly, a complete understanding of the best liposomal design for
delivery of therapeutic substances is still evolving. It is possible that
with nucleic acid delivery the use of a neutral lipid, such as DOPC,
which will enter the cells via a mechanism that will not involve
endosomes, will allow a balance between efficient uptake of the therapeutic payload into a liposome at preparation, uptake of the liposome into a particular cell type and the release of the payload from
the liposome inside the cell cytoplasm, as was demonstrated for multiple targets using siRNAs entrapped in DOPC‐based particles
[91,92,96].
3.1.4. Aggregation of LNPs
Aggregation might cause complement activation even in negligible quantities. It was shown that Doxil® (pegylated liposomes
entrapping doxorubicin) or Danuxome® (daunorubicin entrapped
in nanoliposomes) activate the complement, despite the fact that
both drugs are located within the liposome particles, apparently
shielded from plasma, and aggregates are present within these formulations [68]. It is thus recommended that all new liposomalbased formulation candidates undergo adequate investigations into
their interactions with the immune system prior to their use in
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
6
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
clinical trials [97,98]. Recent excellent reviews and commentaries on
LNPs that activate the complement can be found elsewhere and are
beyond the scope of this report [8,11,70,99].
3.1.5. Route of administration
Routes of administrations such as subcutaneous (S.C.), intradermal (I.D.), intramuscular (I.M.) and intralymphatic (I.L.) and their influence on the immune response were studied by Mohanan et al. [71].
The researchers used the ovalbumin (OVA) in three different particulate antigen-delivery systems: dimethyldioctadecylammonium bromide (DDA) liposomes, N-trimethyl chitosan (TMC) NPs, and poly
lactic-co-glycolide (PLGA) microspheres for evaluating the influence
of the administration routes on the immune response. Whether the
vaccine was administrated S.C., I.D., I.M., or I.L., a minor effect was observed, inducing an antibody response of IgG1 subclass, associated
with TH2-type (anti-inflammatory) response. In contrast, the administration route strongly affected both kinetic and magnitude of the
IgG2a subclass that is associated with TH1-type (proinflammatory)
response: A single I.L. administration of all the tested delivery systems induced a robust IgG2a response. The I.D. and I.M. routes generated intermediate IgG2a titers. A thorough investigation into the
rough of administration of different LNPs (made from various lipids)
and characterized based on their size, geometry and surface charge
will be a substantial contribution to this field and will add important
knowledge correlating the administration route to structural properties of the LNPs.
3.2. LNPs may stimulate or suppress the immune system
Despite advantages of increased stability, facilitated delivery and
protection of its cargo, following systemic administration, LNPs may
trigger the innate immune system, especially the complement cascade and macrophage clearance mechanisms [11] (Fig. 2). As other
circulating particles, liposomes, for example, are often first taken up
by phagocytic cells (such as blood monocytes and macrophages of
the liver, spleen and bone marrow) [100]. There might be undesirable
interactions between the liposomes and the immune system, such as
immunostimulation or immunosuppression. Liposomes have been
well documented as agonists of TLRs and can be also internalized
into macrophages by scavenger receptors, but there is lack of immunological knowledge about the interaction of liposomes and other
LNPs with NLRs and specifically with the inflammasome. Liposomes
can also activate complement or induce an ‘educational’ event with
the adaptive immune system. Interaction of LNPs with TH and
B-lymphocytes is less characterized and has a tremendous potential
to explore new avenues in the adaptive immune system.
The inadvertent recognition of liposomes as foreign entities by leukocytes may result in a multilevel immune response against the liposomes and eventually lead to toxicity in the host and/or lack of
therapeutic benefit [1,100]. A harmful activation of the complement
cascade may occur in some types of particles. This event may lead to hypersensitivity reactions and anaphylaxis [4]. Szebeni et al. showed that
an intravenous injection of LNPs could cause acute hypersensitivity reactions (HSRs) in up to 45% of patients, with hemodynamic, respiratory
and cutaneous manifestations. The phenomenon can be explained with
activation of the complement system on the surface of lipid particles,
leading to anaphylatoxin (C5a and C3a) liberation and subsequent release reactions of mast cells, basophiles and possibly other inflammatory cells in the blood [99].
LNPs decorated with polyethylene glycol (PEG) and entrapping
doxorubicin (Doxil™) also activate the complement system. The
reported frequency of HSRs to Doxil™ is up to 25% of all the treated
patients. Unlike IgE-mediated (type I) allergy, these reactions occur
mostly at the first exposure to the formulation without prior sensitization [68]. An additional example is a harmful activation of the complement system at tumor sites, which may stimulate tumor-
associated immune cells and promote their conversion into a
tumor-supportive phenotype, thereby stimulating cancer progression
[4,70,101].
4. Modulating the immune response by LNPs as a potential therapeutic strategy
Nano-scale liposomes that can modulate an immune response can
be divided into two categories: liposomes encapsulating an antigen
type I, which are designed to elicit an immune response, and a type
II polymer coated liposomes, which are designed to prevent an immune recognition [4]. One example of immunosuppressive liposomes
is the liposomal alendronate (LA). LA is a bisphosphonate drug used
for osteoporosis and other types of bone diseases. It has been
shown that partial systemic inactivation and transient depletion of
monocytes and macrophages is caused by liposomal bisphosphonates
(BP), which reduce neointimal hyperplasia and restenosis in animal
models [102]. The anti-inflammatory effect resulting from inhibition
of tissue macrophages by LA has been documented in experimental
arthritis [103], delayed graft rejection [104] and tumor angiogenesis
models [105]. Haber et al. demonstrated that a high dose of LA
(10 mg/kg) administrated either intraperitoneal (I.P.) or intravenous
(I.V.) exhibited a significant depletion of the circulating monocytes in
the pathological conditions of restenosis and endometriosis. The
mode of liposome administration (I.P. or I.V.) determines whether
systemic or local inhibition of monocytes or macrophages will occur
[100].
An additional, elegant example for liposomes, which induce an
anti-inflammatory effect, is demonstrated by the phosphatidylserine
(PS)-presenting liposomes strategy [81]. Following myocardial infarction (MI) resident and recruitment of macrophages remove necrotic
and apoptotic cells, secreted cytokines, and modulated angiogenesis
at the infarct site [106]. In a rat model of acute MI, targeting of
PS-presenting liposomes to infarct macrophages after i.v. injection
Fig. 2. The interaction of different LNPs with subsets of leukocytes can suppress or activate the immune response. The first line of defense by the innate immune arm includes different pattern recognition receptors such as membrane‐bound Toll-like
receptors (TLRs), cytoplasmic NOD-like receptors (NLRs) and scavenger receptors on
innate immune cells such as monocytes, macrophages and dendritic cells. The second
line of defense includes the adaptive immune arm with several important T helper subsets such as TH1, TH2, TH17, Tregs, TH9 and TH22 cells. Each subset of leukocytes can interact differently with different types of nanoparticles made from different materials,
sizes, geometry, and surface charges. Adapted with permission from ref. [5].
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
demonstrated improved infract repair by mimicking the antiinflammatory effects of apoptotic cells. Following PS-liposome
uptake by macrophages in vitro and in vivo, the cells secreted high
levels of anti-inflammatory cytokines (e.g., TGF-β and IL-10) and
upregulated the expression of mannose-CD206, concomitant with
downregulation of proinflammatory markers such as TNF-α and the
costimulatory molecule CD86. The treatment promoted angiogenesis,
preservation of small scars and prevented ventricular dilatation and
remodeling. The strategy of resolving inflammation in this case was
to mimic anti-inflammatory effects of apoptotic cell clearance with
macrophages by the use of natural, well-defined materials [107].
One important note is that strongly negatively charged particles
may also increase cytotoxicity and promote cell death; thus, prior to
translating this strategy into clinical practice several important investigations need to be done in order to understand the level of the
anti-inflammatory response and the depletion of important innate
immune players such as monocytes and macrophages.
4.1. Immunostimulation by LNPs
In order to induce a robust immune response, it is necessary to develop vaccine strategies involving the generation of cytotoxic (CD8 +)
T lymphocytes (CTL) to possess a mechanism by which administrated
exogenous antigens can be presented not only on MHC class II, but
also using MHC class I molecules on antigen‐presenting cells
[32,108,109]. A drug delivery system based on the oligomannosecoated liposomes (OML) has been developed and tested in an attempt
to increase the effect of generating CTLs against the delivered antigens. The OML‐based vaccine could reject transplanted tumor cells,
prevent progression of encephalitis and vertical transmission and reduce offspring mortality of Neospora caninum as shown in the studies
by Ikehara et al. [110–112]. OML-based vaccine produces strong
adjuvanicity for CTLs, as liposomes coated by oligomannose are exclusively taken up by F4/80 + intraperitoneal mononuclear cells and
gathered at extranodal lymphoid tissues [113]. The OML-based vaccine mechanism of action is based on the immune-surveillance system for detecting pathogens invading the abnormal cavity in either
a mannose dependent or mannose independent manner. Taken together, a line of clearance process for OML may associate with strong
adjuvanticity that induced CTLs [114]. Another strategy built on induction of CTL and antibody response was detailed by the groups of
Barenholz and Kedar [115]. This strategy is based on a novel
polycationic sphingolipid (CCS)/cholesterol liposomal formulation
that markedly enhanced the hepatitis A virus (HAV)-specific antibody
response at the intestinal interface, particularly when delivered
intrarectally or intranasally, to Balb/c mice at low HAV doses. Although the specific mechanism is not discussed, no dramatic adverse
effects were observed and this strategy is now under clinical
investigation.
An additional formulation that is now under clinical evaluation is
the CAF01 [116]. This is a cationic liposomal formulation that incorporates synthetic mycobacterial cord factor and primes protective TH1,
TH17, and antibody responses in animal models of bacterial, viral,
and parasitic infections. Using CAF01 as a backbone, a recent study
demonstrated that incorporating the TLR3 ligand polyinosinic/
polycytidylic acid, poly(I:C), primes CD8 + T cells specific to the
SIINFEKL epitope of the model antigen ovalbumin. The CAF01/
poly(I:C) can induce CD8 + T cells that efficiently lyse target cells
and significantly reduce tumor growth in two different mouse
tumor models [117].
Adjuvant utilizing liposomes is not always effective, as demonstrated by Amin et al. The impact of chitosan-coated liposomes
(CCL) as a nasal vaccine vehicle for eliciting viral specific humoral
mucosal and cellular immune response was studied. CCL interacted
with nasal associated lymphoid tissue failed to increase the residence
7
time of the particles in the nasal cavity, nor induced mucosal and systemic responses as efficiently as non-coated liposomes [118].
4.2. Modulating the immune response with RNAi-LNPs strategies
Gene silencing by RNA interference (RNAi) was discovered by Fire
and Mello in 1998 [119]. This discovery has rapidly emerged as a
promising new strategy for drug target validation, and it is currently
evaluated in clinical trials as a potential therapy for diseases caused
by undruggable therapeutic targets [120].
The use of RNAi for therapeutics holds a great promise to change
the therapeutic modality in many diseases; however, it is a daunting
task to deliver small interfering RNA (siRNA) molecules into specific
cell types in a safe manner. A naked siRNA has a limited ability to
cross the cell membrane due to its 13 KDa molecular weight and
the polyanionic nature (~ 40 negative phosphate residues) [121].
Unmodified, naked siRNA is unstable in blood and serum, as it is rapidly degraded by endo- and exonucleases, with a very short half-life
in vivo. In addition, unless chemically modified, siRNA can induce immune responses and may lead to off-target effects. In vertebrates, an
immune response can be induced by dsRNA, which is part of a defense mechanism against viral infection. dsRNA can be sensed in the
cytoplasmic and endosomal compartments by RNA-dependent kinases (PKRs) and induce [122] an interferon (IFN) response that
will end up in production of proinflammatory cytokines, by activating
NF-kappa-B dependent pathway. Other known mechanisms, which
may cause NF-kappa-B activation and IFN production, are helicase
retinoid-acid-inducible gene I (RIG-I) and melanoma differentiationassociated gene 5 (MDA-5). Studies have found that both chemically
synthesized siRNAs and siRNAs generated from in vitro transcription
with various lengths can lead to the activation of PKR. The dsRNA
length increased the IFN induction levels. For example, Rossi et al.
demonstrated that a 5′ triphosphate on T7 polymerase-transcribed
RNA molecules is a potent inducer of IFN response [123].
Non-self RNA can also be recognized on the cell surface by TLRs. Recognition of RNA by TLRs activates cellular signaling pathways that lead to
the activation of NF-κB and production of proinflammatory cytokines.
Kleinman et al. showed that siRNA against VEGF, a pro-angiogenic factor
that is involved in age-related macular degeneration in the eye, can antagonize TLR3 on the cell surface of fibroblasts. The observed effect of decreased blood vessel growth was demonstrated regardless of the
sequence, which meant that sequence-independent RNAi triggered
TLR3 and subsequent local inflammation that lead to a therapeutic benefit
was not part of a specific silencing of VEGF target [124].
In order to avoid or reduce unfavorable immune response, siRNAs
need to be optimized in a way that evades triggering TLRs responses
as well as intracellular responses such as with the inflammasome. For
example, avoid GU-rich sequences, optimizing the length and/or reducing the dosing of siRNA and perform an experimental screening
of multiple siRNAs against the same target in order to identify the optimal candidate with minimal immune response. There are several
exceptional reviews on the chemical modifications of RNAi payloads
[125,126], which are beyond the scope of this report.
It is widely known that efficient intracellular RNAi delivery to specific target cells following systemic administration is the challenge for
widespread use of RNAi in the clinic [121]. Among many studied delivery systems, only few seem to hold more promise. One of the reasons is the toxicity and immunogenicity caused by the interactions
of the particles with leukocytes.
As with other therapeutics delivered via systemic strategy, this approach can be divided into two major categories: passive and active
(targeted) delivery. Passive delivery utilizes the inherited propensity
of liposomes and other LNPs to accumulate in cancerous or highly inflamed tissues due to the enhanced permeability and retention (EPR)
effect. In this effect an increased permeability of blood vessels in solid
tumors and highly inflamed tissues with dysfunctional lymphatic
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
8
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
drainage is occurring, which retain the accumulated liposomes/LNPs
in close proximity to the tumor or the inflamed tissue [127]. Different
studies have shown that the efficacy of extravasation into tumors is
higher with smaller particles (b200 nm) [128–131].
Liposomes and NPs also tend to accumulate in organs of the
mononuclear phagocytic system (MPS). The MPS, part of the immune
system, consists of phagocytic cells located in reticular connective tissue, primarily monocytes, dendritic cells and macrophages. These
cells accumulate in the spleen, liver and lymph nodes and take up foreign particles such as bacteria, viruses, and parasites with different
sizes and surface charges [132,133].
Active siRNA delivery is based on specific antibodies and ligands
that direct the nanocarriers to specific target cells and tissues in
order to achieve maximal therapeutic effect, decrease the amount of
RNAi payload required and avoid nonspecific silencing and toxicity
in bystander cells [128]. Tremendous progress has been made in systemic siRNA delivery and those have been reviewed extensively elsewhere by us and by others [1,95,134–136]. Below we will only detail
example of modulating the immune response by passive and by active delivery strategies targeted to leukocytes (see Table 3).
4.2.1. Targeted delivery into pan leukocytes
The systemic delivery of RNAi for specific gene silencing to target
leukocytes is an unconquered and extremely challenge task that
holds a great potential for treatment of leukocyte-implicated diseases
such as inflammation, blood cancers (lymphoma, leukemia, myeloma), and leukocyte-tropic viral infections (HIV, ebola, dengue).
Since leukocytes are resistant to conventional transfection reagents and are dispersed throughout the body, it becomes a major
challenge to achieve successful delivery of RNAi payloads selectively
in leukocytes.
Several strategies are under different research stages. For example, PEGylated liposomes are being used for delivery and targeting
of siRNA in vitro with ligands covalently attached to the liposomal
surface that bind to receptors expressed on leukocytes [139]. Unfortunately, these PEGylated particles induced a significant increase in
serum cytokine levels in an unclear mechanism, thus reducing the
therapeutic potential concealed with these particles. Nevertheless,
cytokine induction might be considered as a therapeutic advantage,
when an immune activation could strengthen the therapeutic effect.
Table 3
Immune modulation by selected delivery platforms.
Type of immune
modulation
Target cells
LNP
Dimensions of
the LNPs (nm in technology
diameter)
References
Anti-inflammatory
(depletion of the
circulating
monocytes)
Anti-inflammatory
Macrophages,
monocytes,
dendritic cells
~180
Liposomal
aldronate P
[100]
Infarct
macrophages
1200
[81]
~1053
PS‐
presenting
liposomes⁎
OML⁎
[111,114]
2300
CCL⁎
[118]
~100
PEGylated
[86]
liposomes⁎
[137]
P-selectin
coated
liposomes⁎⁎
I-tsNP⁎⁎
[138]
CTL adjuvanicity
Cytotoxic T
lymphocytes
Failure in inducing Lymphoid
tissue
mucosal and
systemic
adjuvant
responses
Cytokine
Leukocytes
induction
Inflammation
T cells
Gut inflammation
⁎
⁎⁎
Gut
lymphocytes
Passive delivery.
Active targeted delivery.
100–150
~100
Huang et al. demonstrated another delivery platform to silence
genes in circulating cells with the P-selectin coated liposomes.
P-selectin can be absorbed onto the surface of a blood-compatible
microrenathane tube. P-selectin-coated surface could successfully
captured P-selectin receptor-positive stem cells from physiological
shear flow in vitro and from the bloodstream in vivo, although previous studies have shown that P-selectin-based cell capture is also associated with inflammation of the liver or lung in vivo [137].
4.2.2. Targeting subsets of leukocytes
An elegant approach for the delivery of RNAi molecules into dendritic cells that is based on liposomes decorated with monoclonal antibody against DEC-205, a dendritic cell specific protein and
CD40-siRNA was reported. After i.v. administration to mice, a functional silencing of CD40 was demonstrated [140]. This approach can
be used to target CD40 siRNA specifically to dendritic cells in vivo
and consequently induce antigen-specific immune suppression and
may be feasible for siRNA-based clinical therapy.
Leukocyte integrins mediate the adhesive interaction between endothelial cells and leukocytes, one of the crucial steps in the migration of
cells to the inflamed site. Antibodies such as efalizumab (Raptiva) and
natalizumab (Tysabri) were developed in order to block these adhesive
interactions and are examples of FDA approved, humanized,
integrin-blocking antibodies for treatment of autoimmune diseases. The
potential use of integrins as receptor targets for RNAi delivery to lymphocytes is based on the fact that two of the family members, β2 and β7
integrins, are expressed exclusively on leukocytes and thus enable the selective targeting. Moreover, these integrins are constitutively internalized
and recycled. Thus, integrin recycling supports the internalization of
bound antibodies, a prerequisite for siRNA-mediated activation of the
RNAi pathway. The most unique feature of integrins is their ability to
bind ligands via a conformational change from a low affinity conformation in resting cells to a high affinity conformation upon activation. A
proof-of-concept for utilizing leukocyte integrins as receptor targets for
RNAi delivery was demonstrated first by the use of scFv against the
high affinity form of the integrin lymphocyte function-associated antigen
I (LFA-I), which was fused to protamine, a highly positively charged protein that nucleates DNA in the sperm. This fusion protein selectively delivered siRNAs into activated lymphocytes, both in vitro and in vivo, but not
to resting lymphocytes [138]. Based on this strategy, a sophisticated
method was devised termed the integrin‐targeted and stabilized
nanoparticles (I-tsNPs) [14,57,58,141]. Using this approach, it was demonstrated that a single i.v. injection of I-tsNP-encapsulated siRNA
(2.5 mg/kg) targeted against the DNA repair-associated gene Ku70 was
determined to be sufficient to reduce target gene expression by approximately 80% in β7 integrin-positive cells of gut lymphocytes [14]. The same
platform was used for validation of the role of the cell cycle regulator, cyclin D1, as a potential anti-inflammatory target. Cyclin D1 was found to be
upregulated in T lymphocytes at sites of inflammation such as in inflammatory bowel diseases. An i.v. injection of β7 I-tsNP-entrapping cyclin
D1-siRNAs in colitis-induced mice reduced cyclin D1 mRNA and protein
levels and caused a remarkable reduction in intestinal inflammation
[14] without activating lymphocytes, inducing an interferon response
and provoking a proinflammatory cytokine secretion with minimal
off-target effects and toxicities. This strategy is now widely exploited in
several different types of blood cancer, gut inflammation and HIV
infection.
5. Conclusions and future prospective
Understanding the complicated nature of the immune system
with its unique arms can open the door for utilizing LNPs to manipulate the immune response. In a recent study [16], it was demonstrated
that hydrophobicity can dictate the immune response. A small library
of gold NPs was synthesized and characterized. These gold NPs had
increased hydrophobic carbon chains on their surface and had almost
Please cite this article as: D. Peer, Immunotoxicity derived from manipulating leukocytes with lipid-based nanoparticles, Adv. Drug Deliv.
Rev. (2012), doi:10.1016/j.addr.2012.06.013
D. Peer / Advanced Drug Delivery Reviews xxx (2012) xxx–xxx
a linear behavior (both in vitro and in vivo) when interacted with
subsets of leukocytes. LNPs are one of the most veteran delivery vehicles used already in clinical practice for many years and several new
classes of LNPs including hybrid systems (lipid-polymers) are under
clinical evaluation with therapeutic and imaging payloads. Adequate
and comprehensive understanding of the specific interaction between LNPs with T cells, B cells, DCs, macrophages and monocytes
at the cellular and molecular level, and appropriate assays to probe
immune suppression or stimulation such as cytokine induction, interferon response, lymphocyte activation, coagulation cascades and
complement activation, will aid in designing safer vehicles for therapeutic and imaging strategies that will benefit not only the patients,
but also the entire pharmaceutical and biotech industry. Those
might experience shorter development path for new chemical entities that are entrapped within LNPs.
Acknowledgement
Dan Peer wishes to thank Ms. Varda Wexler for her help with the
graphics and illustrations and the Peer laboratory members for helpful discussions. This work was supported in part by grants from the
Marie Curie IRG-FP7 of the European Union, Lewis Family Trust, Israel
Science Foundation (Award #181/10), The MAGNET program of the
Israeli OCS, the Kenneth Rainin Foundation, the Israeli Centers of Research Excellence (I-CORE), Gene Regulation in Complex Human Disease, Center No 41/11 and by the FTA: Nanomedicine for Personalized
Theranostics awarded to D.P.
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