Theme: Nanotechnology in Drug Development
Guest Editors: Nakissa Sadrieh and Banu Zolnick
Challenges in Development of Nanoparticle-Based Therapeutics
Received 1 November 2011; accepted 17 February 2012; published online 10 March 2012
Abstract. In recent years, nanotechnology has been increasingly applied to the area of drug development.
Nanoparticle-based therapeutics can confer the ability to overcome biological barriers, effectively deliver
hydrophobic drugs and biologics, and preferentially target sites of disease. However, despite these
potential advantages, only a relatively small number of nanoparticle-based medicines have been
approved for clinical use, with numerous challenges and hurdles at different stages of development. The
complexity of nanoparticles as multi-component three dimensional constructs requires careful design and
engineering, detailed orthogonal analysis methods, and reproducible scale-up and manufacturing process
to achieve a consistent product with the intended physicochemical characteristics, biological behaviors,
and pharmacological profiles. The safety and efficacy of nanomedicines can be influenced by minor
variations in multiple parameters and need to be carefully examined in preclinical and clinical studies,
particularly in context of the biodistribution, targeting to intended sites, and potential immune toxicities.
Overall, nanomedicines may present additional development and regulatory considerations compared
with conventional medicines, and while there is generally a lack of regulatory standards in the
examination of nanoparticle-based medicines as a unique category of therapeutic agents, efforts are being
made in this direction. This review summarizes challenges likely to be encountered during the
development and approval of nanoparticle-based therapeutics, and discusses potential strategies for
drug developers and regulatory agencies to accelerate the growth of this important field.
KEY WORDS: biodistribution; immune toxicity; nab-paclitaxel; nanoparticle; pharmacokinetics.
Over the last quarter century, the field of nanotechnology
has witnessed tremendous growth and advances. The area of
nanoparticle-based medicine receives particular attention as it
holds the promise to revolutionize medical treatment with more
potent, less toxic, and smart therapeutics that could home into
disease areas like the elusive magic bullet. With substantial
efforts by both academia and the biopharmaceutical industry, a
few nanomedicines have been successfully developed and
approved for clinical use. However, there is no doubt that the
few and far apart. Most nanomedicine research, while exciting
and technically advanced, is in early stages of development and
is only slowly being translated into clinical trials and medical
practice.Thisreview seeks to illustrate the numerous challenges
that are encountered during the development of nanoparticle-
based therapeutics using the commercially available protein-
bound nanoparticle formulation of paclitaxel (Abraxane®,
also adds to the current debate about the future regulatory
requirements for the approval of nanomedicines, and the
challenges to be expected in the development and approval of
generic equivalents of these products.
RATIONALE FOR THE DEVELOPMENT
Nanoparticles often have unique physical and chemical
properties at the cellular, atomic, and molecular levels not
surface to volume ratio. In addition, the ability to create three-
dimensional multi-component structures of nanoparticles also
allows a great degree of flexibility to design drug delivery
systems that may fulfill several desired properties such as the
ability to overcome biological barriers, the ability to deliver
hydrophobic, poorly water-soluble molecules, and the potential
ability to selectively target these nanomedicines to a preferred
site in the body.
Biological Barriers to Drug Delivery
Multiple biological barriers exist for drugs to successfully
reach their intended disease sites. Oral drugs need to have high
stability in the gastrointestinal tract and the ability to penetrate
1Strategic Platforms, Abraxis BioScience, AWholly Owned Subsidiary
California 90025, USA.
2To whom correspondence should be addressed. (e-mail: ndesai@
The AAPS Journal, Vol. 14, No. 2, June 2012 (#2012)
1550-7416/12/0200-0282/0#2012 The Author(s). This article is published with open access at Springerlink.com
intestinal epithelium to achieve high systemic bioavailability (2).
Similarly, skin, nasal, and pulmonary drug delivery requires
drug developmentforsmallmoleculesisfocused on oraldelivery,
with drug chemistry directed towards good oral absorption,
intravenous (IV) administration remains the most direct and
efficient route to deliver drugs that comprise peptides, proteins,
large molecules, and polynucleotides.
Drugs in circulation still need to overcome several
barriers to reach their targets. The blood–brain barrier
(BBB) restricts the diffusion of large or hydrophilic molecules
into the cerebrospinal fluid and is a major obstacle for
treatment of most CNS and brain disorders. Multiple nano-
particle-based strategies, including liposome, nanosphere, and
cationic albumin nanoparticles, are under development to
deliver drugs across the BBB (3).
Overcoming the difficulty of delivering therapeutic agents
tumor vasculature is highly heterogeneous in distribution and
more permeable in some places, however, large areas of tumors
may be poorly perfused (4). Impaired lymphatic drainage in
tumors contributes to increased interstitial fluid pressure (IFP)
(5) adding another barrier to delivery. The elevated IFP has
been described to be one of the main factors contributing to
limited extravasation and transvascular transport of macro-
molecules despite the leaky tumor microvasculature, and
inhibits the transport of molecules in tumor interstitial space
(6).High tumor celldensity and dense tumor stromacan further
hamper the movement of drugs within tumors (4). In particular,
the presence of collagen in tumor extracellular matrix is a major
factor limiting drug penetration in the tumor interstitium, with
pancreatic cancer being a primary example (7). Although
tumors pose these barriers for drug delivery, the pathology
and natural biological mechanisms of tumors, yet unexplored,
leave great opportunities for targeting by nanomedicines.
The high permeability of tumor vasculature and the lack of
proper lymphatic drainage result in the so called enhanced
permeability and retention effect (EPR) in the tumor microen-
vironment (Fig. 1), which has been suggested to improve tumor
drug delivery for nanoparticles and macromolecules that have
the ability to circulate with a long half life (8,9). While
nanomedicines may take advantage of this passive process, the
utilization of an active biological transport process may further
improve drug delivery. For example, Abraxane® (paclitaxel
protein-bound particles for injectable suspension (albumin
bound), also known as nab®-paclitaxel or ABI-007, Abraxis
BioScience/Celgene; Fig. 2) can achieve rapid tumor penetra-
tion and enhanced accumulation by utilizing the albumin
transport pathways, including gp60 albumin receptor and
caveolae-mediated endothelial transcytosis (Fig. 3) across
endothelium of the blood-tumor barrier and potential interac-
tion with albumin binding proteins (e.g., secreted protein, acidic
and rich in cysteine (SPARC)) in the tumor interstitium (10).
Delivery of Hydrophobic Drugs
The efficient and safe delivery of hydrophobic therapeu-
tic compounds remains a serious hurdle for the pharmaceu-
tical industry. The formulation of many hydrophobic drugs
requires toxic solvents and surfactants such as Cremophor
and Tween, which often impair drug distribution and are
associated with severe side effects. For example, Taxol®, the
conventional formulation of the hydrophobic drug paclitaxel
(Bristol-Myers Squibb Co), contains a high concentration of
Cremophor-EL® (BASF, Ludwigshafen, Germany), a solvent
associated with significant toxicities including potentially
lethal hypersensitivity, anaphylactic reactions, and prolonged
peripheral neuropathy (11,12). Cremophor can also sequester
paclitaxel in micelles, which prolongs the systemic exposure
and increases drug toxicity (13). Polysorbate, another com-
monly used solvent for hydrophobic drugs, can also induce
hypersensitivity reactions (14).
Nanomedicines which do not require the use of toxic
solvents offer clear advantages over their conventional counter-
parts. The first protein nanotechnology-based chemotherapeu-
tic approved, nab-paclitaxel, is a Cremophor-free, albumin-
bound nanoparticle formulation of paclitaxel with a mean
particle size of approximately 130 nm. By eliminating the
CremophorEL/ethanolvehicleused inTaxoland the associated
toxicities and incorporating nanotechnology, nab-paclitaxel
Fig. 1. Enhanced drug delivery to solid tumors using nanoparticles or macromolecules (8). A Passive
delivery (EPR). After IV injection, nanoparticles accumulate in tumors through leaky and permeable
tumor vasculature and impaired lymphatic system. B EPR + targeted delivery. “Targeted” nanoparticles or
macromolecules bind to cancer cell receptors resulting in potentially improved drug delivery. (Adapted
from Nanomedicine, June 2010, vol. 5, no. 4, pp 597–615 with permission of Future Medicine Ltd.)
283Challenges in Development of Nanoparticle-Based Therapeutics
confers the ability to achieve a 50% higher maximum tolerated
dose (300 vs. 175 mg/m2for solvent-based paclitaxel, every
3 weeks), a shorter infusion time (30 min vs. 3 to 24 h with
solvent-based paclitaxel) without the need for premedication to
prevent solvent-related hypersensitivity reactions. nab-
paclitaxel also has linear pharmacokinetics (PK) due to the
avoidance of micelle entrapment which occurs with solvent-
based paclitaxel. In addition, nab-paclitaxel can use standard
tubing and IV bags (10).
Desire for Targeting
Targeting increases efficacy, decreases side effects, and
reduces systemic drug exposures. The organized structures of
nanoparticles allow for incorporation of various targeting
moieties to enhance drug delivery to the target sites, reduce
off-target organ toxicities, and facilitate cellular uptake of
therapeutic agents (15). The distinctive biology and pathology
of tumors make effective delivery a high priority for antitumor
therapeutics, as well as present an endless list of potential
candidates for targeting (Fig. 1). Most active targeting moieties
of nanoparticles are biologics, including ligands for receptors,
peptides, proteins, and antibodies. Studies have shown that
nanoparticles carrying ligands or monoclonal antibodies tar-
geted to surfacereceptors overexpressedbycancer cells,suchas
the transferrin receptor, the folate receptor and EGFR, can
increase cellular internalization of the agents through endocy-
tosis and improve the efficacy of systemic anticancer therapy
(16,17). In addition, nanoparticles incorporating cell-penetrat-
ing peptides and protein-transduction domains, such as oligo-
arginine and TAT, enable the uptake of agents that otherwise
cannot effectively enter cancer cells (18). Further, some nano-
medicines are designed to target tumor endothelial cells. For
through binding to the integrins α2β3, αvb3 and α5β1, can
direct liposomes and other nanoparticles carrying vascular
targeting and cytotoxic agents to endothelium and tumor cells,
thus achieving significant antitumor activity (19–21).
CHALLENGES IN NANOMEDICINE FORMULATION,
CHARACTERIZATION AND MANUFACTURING
Nanomedicines are likely to be three-dimensional con-
structs of multiple components with preferred spatial arrange-
ments for their functions. As a result, subtle changes in
process or composition can adversely affect the complex
superposition of the components with negative consequences.
A thorough understanding of the components through
detailed physicochemical characterization as well as function-
al tests may be essential in order to support highly reproduc-
ible manufacturing processes for nanomedicines.
Multiple hurdles exist before a nanomedicine can
reach the clinic, starting with detailed characterization and
the successful manufacture of these complex constructs.
Other than the standard criteria for acceptable safety and
efficacy, and desirable pharmaceutical characteristics (e.g.,
stability, ease of administration, etc.) that are applicable
to most drugs, the ideal nanoparticle system or nano-
medicine to be utilized for therapeutic purpose may
embody the following features:
& Detailed understanding of critical components and their
& Identification of key characteristics and their relation to
& Ability to replicate key characteristics under manufacturing
& Easy to produce in a sterile form
& Ability to target or accumulate in the desired site of action
by overcoming the restrictive biological barriers
& Good in-use stability, easy to store and to administer
Nanoparticle Components and Characteristics
The versatility of nanoparticles allows for a potentially
wide variety of choices in the payload, nanoparticle compo-
sition, and targeting moieties. The in vitro and in vivo
properties of nanoparticles depend on a number of key
physicochemical characteristics, including size and size distri-
bution, surface morphology, surface chemistry, surface
charge, surface adhesion, steric stabilization, drug loading
efficiency, drug release kinetics, and hemodynamic properties
of the nanoparticles.
Nanoparticles have been adapted to deliver different
kinds of therapeutic agents, including small molecule drugs,
Fig. 2. Abraxane (nab-paclitaxel) schematic
Fig. 3. nab-paclitaxel (ABI-007) transcytosis across endothelial cells
as an assay demonstrating “biological function” or transport due to
albumin (22). Transcytosis of paclitaxel across human umbilical
vascular endothelial cells was significantly enhanced in nab-paclitaxel
compared with Cremophor-based paclitaxel (Taxol). The enhanced
transcytosis of nab-paclitaxel was inhibited by β-methyl cyclodextrin
(BMC), a known inhibitor of the gp60/caveolar transport
peptides, proteins, oligo- and polynucleotides, and genes. The
nanoparticles used in drug delivery include liposomes,
polymers, proteins, micelles, dendrimers, quantum dots,
nanoshells, nanocrystals, gold nanoparticles, paramagnetic
nanoparticles, and carbon nanotubes (23). Each of these
systems has widely varying architecture and attempting to
generalize key physicochemical characteristics between the
different approaches may be futile. Therefore, it is beneficial
that in each particular system, key characteristics and critical
components that may dictate the performance of the nano-
particle system or particular nanomedicine be defined and
understood by the innovators.
Particle size and size distribution is one of the most
widely accepted defining characteristic of nanoparticle-based
medicines because size can significantly impact the PK,
biodistribution, and safety. After administration, small nano-
particles with size smaller than 20–30 nm are rapidly cleared
by renal excretion, while particles 200 nm or greater in size
are more efficiently taken up by the mononuclear phagocytic
system (MPS; also known as reticuloendothelial system), with
cells in the liver, spleen, and bone marrow (24). Previous
reports have shown that nanoparticles of 150–300 nm locate
mainly in the liver and spleen (25), and colloids of sizes 200 to
400 nm undergo rapid hepatic clearance (26). It has been well
described that tumor blood vessels are leaky with fenestra-
tions ranging between 0.2 and 1.2 μm, therefore nanoparticles
with size below 200 nm can take advantage of the EPR effect
for enhanced drug accumulation in tumors (27–29). Particles
will always exist in a range of sizes, therefore, size distribution
must also be taken into account when designing a nano-
medicine. Considering a normal size distribution, for the vast
majority of particles to be below 200 nm in size, the mean
nanoparticle may need to be well below 200 nm to confer the
full “benefits” of a nanomedicine. Therefore, the nanoparticle
size and size distribution need to be carefully controlled
during the small-scale preparation and in particular during a
larger-scale manufacturing process.
Nanoparticle surface properties are also critical determi-
nants for nanoparticle behaviors and interaction with proteins
and cells (30). A multitude of surface characteristics (charge,
hydrophobicity, functional groups, etc.) play an important role
innanoparticlestability and theopsonization process (24,31,32).
Upon entering circulation, colloidal nanoparticles are coated
with various blood components (such as albumin, fibrinogen,
IgG, and apolipoproteins) in the opsonization process, which
activates the complement pathway and targets the particles for
clearance by macrophages (26,33). Complement activation by
nanoparticles is also sensitive to surface polymer conformation
(34). As an example, macrophages can directly recognize
nanoparticles via the particle surface. Particles with hydrophilic
surfaces can become more hydrophobic in circulation by the
adsorption of IgG, whereas hydrophobic particles can be
directly taken up by macrophages without opsonization (26).
Poly(ethylene glycol) (PEG) and other polymers can provide a
hydrophilic surface andprotect nanoparticles from opsonization
and immune recognition (35).
The choice of a suitable targeting moiety for the intended
disease could play important roles in enhancing efficacy and
reducing side effects for a nanomedicine. Active and biological
targeting moieties can alter PK, biodistribution, and cellular
uptake. For instance, alteration of the density of surface ligands
can potentially elicit complement activation and other immune
responses, further complicating the composition of multifunc-
tional and multicomponent nanoparticles (36–38).
Identifying the “right” nanoparticle parameters for the
intended indication is crucial. Moving in one direction could
solve a particular problem butmay often lead to another issue. In
this respect the example of Doxil® and Myocet® is quite
illustrative. Conventional liposomes termed as “non-stealth”
have high affinity for the MPS and are rapidly removed from
can significantly decrease their uptake by macrophages. PEGy-
lated liposomal doxorubicin (Doxil®) demonstrates a prolonged
half life, increased tumor drug concentration, better antitumor
efficacy, and fewer side effects than conventional doxorubicin
(39). While the PEG coating of Doxil limits cardiotoxicity, it also
results in higher concentrations in the skin resulting in a side
effect called palmar plantar erythrodysesthesia, more commonly
known as hand–foot syndrome (39). In addition, PEGylated
liposome infusion has shown idiosyncratic non-IgE-mediated
signs of hypersensitivity in humans, which can be reduced by
slowing the rate of infusion or by premedication (40). In contrast,
Myocet® (Enzon Pharmaceuticals) is a non-pegylated liposomal
doxorubicin approved in Europe and Canada for treatment of
metastatic breast cancer in combination with cyclophosphamide
(41). The difference in compositions allows Myocet to have
reduced cardiotoxicity but no hand-foot syndrome (42).
Currently, there are no good in vivo models to predict
the diverse behaviors of the many types of nanoparticles
under investigation, so the development of nanoparticles with
desirable properties has to rely on empirical evidence and
extensive preclinical animal testing.
Analysis and Characterization of Nanoparticle Formulations
Identifying the appropriate analytical tests to fully
characterize nanomedicines, whether physical, chemical or
biological, may be one of the more challenging aspects of
nanomedicine development both from a technical as well as
regulatory perspective. Due to the complex nature of nano-
medicines compared with standard pharmaceuticals, a more
sophisticated level of testing should be required to fully
characterize a nano product. Because each component of a
nanomedicine serves a specific function, it would be essential
to be able to quantify each component and also evaluate the
relationships and interactions between these components in
aspects that include both stoichiometry and their spatial
orientation. Multiple orthogonal characterization techniques
are essential to ensure that the nanomedicines have all the
desired properties for the intended therapeutic purpose.
While the standard analytical tests such as quantification
of active and inactive ingredients, impurities, etc. in pharma-
ceutical products still apply, various additional techniques are
employed specifically for the characterization of nanoparticle
physicochemical properties. These tests involve a broad range
of methods, including visualization of nanoparticles by
microscopy (atomic force microscopy, transmission electron
microscopy (TEM), and scanning electron microscopy);
measurement of particle size and size distribution with light
scattering (static and dynamic), analytical ultracentrifugation,
Challenges in Development of Nanoparticle-Based Therapeutics
capillary electrophoresis, and field flow fractionation (25);
analysis of surface charge or zeta potential; and examination
of surface chemistry by X-ray photoelectron spectroscopy or
Fourier transform infrared spectroscopy (43). The crystalline
state of drugs encapsulated in the nanoparticles can be
assessed by X-ray diffraction and differential scanning
More innovative testing methods are constantly being
developed and applied to the analysis of nanoparticles. The
key parameters, as well as the overall stability of nano-
particles, should be tested with nanoparticles in solid form, in
suspension, and in biological medium, and under accelerated
conditions such as higher temperature to ensure the robust
performance of nanoparticles. However, the above tests may
not be able to functionally differentiate between an “active”
formulation and one that is “inactive” or “less active.”
As an example, consider a nanoparticle carrying a
payload of an active drug and having at its surface moieties
that allow stealth features, such as PEG and moieties that
may allow targeting including peptides, nucleic acids, pro-
teins, or antibody fragments that bind certain receptors.
Quantitative techniques to analyze the individual compo-
nents, while essential, will be missing key information about
the distribution of these moieties on the surface of the
particles or whether indeed these moieties are at the surface
or buried within the structure of the nanoparticle. The spatial
distribution of these moieties is critical for the intended
function of the particular nanoparticle. A different series of
tests may be required to determine these aspects; such tests
are likely to be non-conventional in pharmaceutical use,
including surface analysis, as well as biological function tests
to assure that the manufacturing process produces nano-
particles that are active in cellular uptake, transcytosis
(Fig. 3), or binding to appropriate biological materials. These
“structure-function” tests are essential at least up to the point
where one can validate a highly reproducible manufacturing
process. Bioassays to confirm activity of the product are often
used when testing biologic drugs.
Several proposed nanotherapeutics have complex com-
ponents such as proteins or nucleic acids forming an integral
part of the nanomedicine (44,45), which may be sensitive to
the manufacturing process conditions and in some cases
undergo change in composition as a result of the manufac-
turing. These components may not be the “active” pharma-
ceutical ingredient in the nanomedicine, but their presence
may serve a role in targeting specific cells/biological pathways
or distribution of the active ingredient in the body. These
components cannot be considered as inactive excipients due
to their important role in efficacy or safety of the product, and
should be fully characterized by appropriate analytical tests.
In addition to finished product tests, it is useful to
institute appropriate in-process tests during early manufac-
turing development that can provide essential information
during the development of a robust manufacturing process.
These are tests that can be conducted with relatively quick
turnaround and are designed to give an early read into how
varying process conditions can affect the nanomedicine
composition at intermediate stages of the process.
In summary, due to the complex nature of nanomedi-
cines, it is reasonable to expect that the level of analytical
characterization, testing and release required to adequately
understand and define the physicochemical or biological
nature of these products is more sophisticated and burden-
some than for standard pharmaceutical products. The better
one can understand these products in early stages of
development, the more likely it is that a successful reproducible
manufacturing process will be achieved.
Scale-up and Manufacturing
The successful scale-up and manufacturing of a nano-
medicine present unique challenges in pharmaceutical develop-
ment. Conventional pharmaceutical manufacturing does not
typically create three dimensional multicomponent systems in
the nanometer scale and as such this requirement presents a
series of obstacles for the scale-up of nanomedicines.
Since most nanoparticles are complex multicomponent
products with specific arrangement of components, a full
understanding of the components and their interactions is
essential to defining the key characteristics of the product.
Identifying these characteristics early in development in turn,
greatly helps define larger scale manufacturing in order to
establish critical process steps and analytical criteria that
ensure reproducibility of the product.
The methods of nanoparticle preparation can be broadly
categorized into “top–down” and “bottom–up” approaches.
Top–down approaches seek to create smaller entities from
larger ones, such as grinding of particles using the milling
technique. In contrast, bottom–up approaches arrange
smaller components into more complex assemblies, which
often involve polymerization of monomers or molecular self-
assembly to cause single-molecule components to automati-
cally arrange themselves into a useful conformation (43).
The nanoparticle formulation process often involves the
use of organic solvents, high-speed homogenization, sonication,
milling, emulsification, crosslinking, evaporation of organic
solvents, centrifugation, filtration, and lyophilization. During
early development, at the lab- or small scale, it is useful to
consider what approach may be useful if the product were to be
scaled up. Identification of important process conditions is
critical to achieve key attributes andfunctions.Theseconditions
may involve the ratio of polymers, drugs, targeting moieties, the
type of organic solvent, and emulsifier/stabilizer/crosslinker, the
oil-to-water phase ratio, mixing, temperature, pressure, and the
pH (43). Depending on conditions, the process may lead to
altered chemical structure of the active and the other compo-
nents and substantial amount of impurities. For macromole-
cules, particularly biologics, it may result in changes in chemical
structure and conformation, denaturation, crosslinking, coagu-
lation, and degradation. Importantly, nanoparticles are not
simple additions and mixtures of individual components.
Rather, they are integral and highly structured compositions.
The structural integrity andphysicochemicalproperties ofintact
nanoparticles must be preserved throughout the formulation
process to the finished product.
be robust to ensure high reproducibility, and be streamlined to
allow for the ease of scale-up production. The manufacture of
nanoparticles often requires multiple process steps involving
multi-component systems. Although small-scale processes may
achieve reproducibility with well characterized components,
once beyond the early prototype, the reproducibility and
consistency of the constructs remain a constant challenge for
the scale-up and manufacturing process. The manufacturing
plan needs to define acceptable limits for key nanoparticle
attributes and identify process conditions that are critical to
achieve these key attributes and functions. These critical
conditions must be identified at the small scale through extensive
experimentation to gain a full understanding of how process
conditions can impact the product both from a physicochemical
and biological perspective. This necessitates that the physico-
chemical and biological tests be sensitive enough to identify
discrepancies in the product that could affect performance (see
“Analysis and Characterization of Nanoparticle Formulations”).
In multistep processes, in-process testing for critical parameters
with a rapid and reliable analytical method is often very
informative about how well the process is controlled. Building
up a database of information with suitably targeted in-process
tests may be vital to ensure success at the manufacturing scale.
Liposomes provide a good example to illustrate issues of
stability, scale-up and importance of critical process parame-
ters. Differing preparation methods can create liposomes with
multilamellar vesicles, small unilamellar vesicles, or large
unilamellar vesicles. Each of these systems may be unstable
due to high-surface free energy and the tendency to
aggregate. The use of amphipathic PEG on the surface can
inhibit liposome aggregation by reducing interfacial free
energy with water and acting as a steric barrier, therefore
improving stability in addition to extending in vivo circulation
half-life of liposomes (46–48). The physical stability of
liposome drug products is determined by the integrity, size
distribution, and composition of liposome vesicles. The FDA
has highlighted the importance and challenges in maintaining
a close control over the manufacturing process in a draft
guidance for liposome drug products stating that “liposome
drug products are sensitive to changes in the manufacturing
conditions, including changes in scale. This should be
considered during the development process, and critical
manufacturing parameters (e.g., scale, shear force, and
temperature) should be identified and evaluated” (49).
Nanomedicines that are to be utilized by a route of
administration that requires a sterile product will face
particular challenges dependent on their particle size and
composition. Nanomaterials are known to be at increased risk
for being damaged by sterilization techniques such as gamma
irradiation or autoclaving, especially when biological materi-
als are involved (50–52). If the structure of the particles is
flexible or malleable, such as in case of some liposomal
preparations, then sterilization through conventional sterile
filters may not be problematic especially if the starting
particle size is well below 220 nm (0.22 μm). In the case of
rigid structures such as polymeric, silica-based, metal and
other nanoparticles, sterilization by filtration may be the only
option, however, wide particle distributions and particle sizes
closer to 220 nm can result in tremendous difficulty in
filtration due to the nominal pore size of standard filtration
membranes. If the mean particle size is not well below
220 nm, then substantial amounts of the active ingredient
could be lost on filtration. While aseptic manufacturing is
always an option, this can be quite complicated, in particular
for a multiple step process that involves handling and transfer
of materials in a “sterile” environment.
One of the other challenges is related to the reproduc-
ibility of in situ preparation of nanomedicines. Some
nanomedicines have utilized the concept of “self-assembly”
and “in situ” preparation where two or more components as
intermediates ready to use at the “bedside” are brought
together under appropriate conditions to create structures or
complexes as the final finished product for human use (53–
55). This approach, by manufacturing individual components,
circumvents the otherwise complex manufacturing steps of
creating a finished nanomedicine as a stable pharmaceutical
product and may significantly reduce the cost of manufactur-
ing and forego the complex development work it would
involve. Despite these advantages, the in situ strategy raises
certain questions. If the three-dimensional complex structure
of nanomedicines is critical to their function, then can one
rely on the vagaries of the individuals in the hospitals or
doctor’s offices to create a “reproducible” finished product
for administration in these different settings? Are these
individuals performing a critical part of the manufacturing
step essentially at the bedside of the patient? Would the in
situ final products be subject to the same stringent standards
that may be applied to a shelf stable nanomedicine product to
ensure reproducibility and control of the manufacturing
process? Or, should there be qualifying release tests for in
situ products as is required for a shelf stable product when
released for distribution? Some of these issues need careful
consideration as policies and guidelines evolve for nano-
An additional issue for the manufacture of nanoparticles
is environmental safety. The handling of dry materials in the
nanometer size scale demands special caution as airborne
nanoparticles distribute as aerosols. Lung deposition of such
nanoparticles can lead to pulmonary toxicities (56,57). During
dosing solution preparation, aerosolization of solutions needs
to be avoided to prevent unintended exposure. Some nano-
particles are capable of penetrating the skin barrier, making
dermal exposure a potential risk so adequate protection of
personnel is essential (56). In this respect, nanoparticles that
are created entirely within a liquid environment may have
significantly lower environmental impact, presumably no
different from standard manufacturing of liquid pharmaceu-
Challenges in the Development of nab-paclitaxel
The development of nab-paclitaxel illustrates well the
challenges in formulation, manufacturing, and testing of a
nanoparticle with appropriate physicochemical properties. As
the first approved protein-based nanomedicine, nab-paclitax-
el underwent extensive preliminary testing in small scale
formulation. Awide range of conditions were investigated for
the manufacturing process, as well as protein from different
sources with variations in quality and purity. Differing
conditions of preparation often resulted in suboptimal
preparations, a challenge that could be overcome only with
large amounts of trial and error. These hurdles for successful
scale-up production of nab-paclitaxel were further illustrated
by unsuccessful attempts in the marketplace to copy the nab-
paclitaxel formulation. This aspect is discussed in the
“Regulatory Challenges to Nanomedicine Development”
section of this review. A large number (several hundred) of
287Challenges in Development of Nanoparticle-Based Therapeutics
optimization runs/batches were conducted to define compo-
nents and compositions of the nanoparticle and to develop a
robust process that performs with consistency and reproduc-
ibility for scale-up manufacture.
As a result, the nab-paclitaxel nanoparticles displayed many
features desirable for an injectable nanomedicine. The size
with a mean particle size of 130 nm as determined by dynamic
laser light scattering (58). TEM and cryo-TEM images revealed
(Fig.4).The albumin surfaceofnab-paclitaxelnanoparticleshasa
albumin, prevents agglomeration and provides the nanoparticles
with good stability in suspension. X-ray powder diffraction
revealed that paclitaxel within the nanoparticles is non-crystalline
(amorphous), making the drug readily bioavailable without the
time lag needed to dissolve crystalline paclitaxel as is well known
for nanocrystals (59). nab-paclitaxel comprises nanoparticles of
drug coated with a layer of albumin crosslinked to a specific level,
with paclitaxel non-covalently bound to albumin via hydrophobic
interaction, which allows for high bioavailability and rapid tissue
distribution. This is in contrast with most of other albumin-based
nanoparticles reported in literature (60–62), which involve the
addition of glutaraldehyde or other crosslinking agents during
nanoparticle formation and require enzymatic degradation of
albumin for drug release in vivo.
In summary, the careful selection of key components,
identification of key characteristics and understanding of
critical manufacturing steps are determinants of whether the
nanoparticles will have the desired pharmacodynamics, PK,
and safety profiles to achieve the intended therapeutic effects.
Multiple orthogonal analysis methods are required for
appropriate in-process quality controls and tests for finished
products. Deviations from key nanoparticle parameters and
processes could have serious negative impacts on safety and
efficacy of a nanomedicine.
PHARMACOLOGY AND SAFETY CHALLENGES
Because the pharmacological and safety profiles of
nanomedicines are influenced by the cumulative contribution
of physicochemical characteristics, subtle changes in compo-
sition arising from small deviations in the manufacturing
process could result in substantial changes in pharmacology
and toxicity of nanomedicines.
Pharmacology Issues Related to Nanomedicines
It is essential for a successful nanomedicine to achieve
the desired pharmacological profile and PK profile suitable
for the intended indication. However, several challenges are
associated with trying to apply the standard criteria of small
molecule PK to the PK of nanomedicines.
Usually, only a small fraction of the administered drug
reaches its intended location and because of this, the standard
approach of determining PK in the blood or plasma as the
sole measure of in vivo behavior of nanoparticles may be
inherently flawed. While small molecule drugs may diffuse
more readily through “biological barriers” and hence a blood
level may be somewhat in equilibrium and related to
achievable target tissue levels, applying this logic for larger
macromolecular complexes and nanomedicines cannot be
assumed to be correct. It is well recognized that small or
subtle compositional differences can affect the biodistribution
of nanoparticles or nanomedicines (1,63,64). With the wide
variety of potential nanomedicines on the horizon, it would
be highly unlikely that standard pharmacological approaches
would be appropriate to characterize their behavior. Accord-
ingly, pharmacology at the site of action may be more
appropriate to define if a particular nanomedicine approach
achieves reliable target tissue concentrations.
The PK and biodistribution of the active drug within the
nanomedicine may be affected by several factors. Subtle
compositional aspects, size, shapes, and physicochemical prop-
erties of nanoparticles can result in altered PK and greater PK
variations compared with conventional small molecule
approaches. Nanomedicines may allow for novel routes of
delivery including oral, pulmonary, and dermal administration,
which requires high bioavailability through the biological
barriers. The PK of both the nanomedicine as a whole as
compared with just the “free” drug may be highly relevant.
The clearance of nanoparticles in vivo is a complicated
process controlled by numerous characteristics such as
particle size, surface properties and possibly other composi-
tional characteristics that are not fully understood at this time.
Smaller nanoparticles are cleared by the kidney, whereas
larger particles are cleared by the Kupffer cells and macro-
phages of MPS located predominantly in the liver and spleen
(24). The nanoparticle clearance is facilitated by the opsoni-
zation of blood components and complement proteins on the
particle surface (31). The inhibition of opsonization and
evasion of detection by macrophages with approaches such
as pegylation prolong the circulation of nanoparticles in the
case of liposomal doxorubicin (Doxil) (65).
The size and composition can affect the release kinetics of
the active drug from the nanomedicine carrier. Whereas
monodisperse nanoparticles or particles with narrow distribu-
tion are desirable for consistency, it has also been proposed that
mixed sizes to intentionally introduce different rates of drug
release for sustained delivery over time. Additionally, both
passive targeting (e.g., clearance by the MPS in cases where the
Fig. 4. Cryo TEM image of nab-paclitaxel showing spherical
liver is the target or where longer circulation time for EPR is
desired) and active targeting approaches may significantly
enhance the distribution and accumulation of nanomedicine
drugs at the intended target sites.
There is no uniformly effective approach to design a
nanomedicine to achieve a desired PK profile. A common
approach has been to try to get long circulation times with
nanomedicines to take advantage of EPR effect or targeting
(9). However, this approach may not always be appropriate
for the desired indication, and in some cases could reduce
therapeutic efficacy and unnecessarily increase systemic
exposure. Alternate approaches may also have benefit as in
the case of nab-paclitaxel. Rather than utilizing passive
transport via the EPR effect for drug delivery to tumor,
nab-paclitaxel relies on active albumin transport pathways
including the gp60/caveolae-mediated albumin transcytosis
across tumor blood vessel endothelium and potential associ-
ation with tumoral SPARC to achieve enhanced drug
accumulation in tumors (10). Desai et al. have shown that
compared with Cremophor-based paclitaxel (Taxol), nab-
paclitaxel formulation increased the endothelial binding of
paclitaxel by 9.9-fold (P<0.0001) and the transport of
paclitaxel across microvessel endothelial cell monolayers by
4.2-fold (P<0.0001) (16). Compared to non-linear PK of
conventional Cremophor-based paclitaxel (Taxol®), nab-pacli-
taxel exhibits a linear PK profile with faster clearance and
increased volume of distribution, which is contrary to the typical
approach for nanoparticle-based medicines (66). Caveolar
transport utilizing nab formulations has been demonstrated in
vivo in a live rat model of transpulmonary absorption. The
gp60 and caveolar transport (67–69). Intratracheal administra-
tion of nab-paclitaxel showed a rapid uptake into the blood
stream with a plasma concentration profile from pulmonary
delivery of these nanoparticles essentially matching the plasma
concentration for the intravenously delivered drug (70). These
results suggested that the caveolar transport process was a
As discussed above, standard PK may not be sufficient
for evaluating nanoparticle-based medicines, as plasma PK is
not necessarily representative of PK in tumors and disease
sites, and therefore cannot predict clinical activity. The PK at
site of action is more relevant and can be better correlated
with therapeutic outcome. This is especially true for targeted
nanomedicines. For example, despite faster clearance of nab-
paclitaxel from circulation, intravenously administered nab-
paclitaxel achieved 33% higher intratumoral paclitaxel con-
centration than equal dose of Taxol in tumor xenografts (16).
In a retrospective analysis of non-small cell lung cancer
patients treated with the combination of nab-paclitaxel and
carboplatin, no correlation was observed between drug
exposure based on plasma PK parameters and clinical efficacy
as measured by overall response rate and progression-free
survival (Abraxis BioScience, unpublished data). This could
be due to the lack of correlation between tumor drug
exposure and plasma PK, and potential differences in drug
resistance among patients. Further, the interpatient PK
variability of a liposomal CKD-602 was observed to be
several-fold higher compared with a conventional formulation
variability in PK due to variable clearance mechanisms.
In summary, the development of nanoparticle-based
medicines faces numerous pharmacological challenges. The
PK profile of nanoparticle-based medicines can be influenced
by variations in many different parameters and therefore,
nanomedicines may require different PK approaches for
different indications. Furthermore, instead of standard testing
of plasma PK, physicochemical properties and drug concen-
trations or accumulation at the disease site may be more
relevant to evaluate the reproducibility and activity of nano-
medicines. In the future, these methods may also be useful for
evaluating bioequivalency of nanotechnology-based products.
Safety Challenges in Nanomedicine Development
In recent years there has been increasing attention to
toxicities unique to nanoparticle-based medicines (71,72).
Regulatory bodies have engaged in several public discussions
on this topic and in some cases have published their findings
(73). The consensus has been that each product may have its
own case by case issues requiring particular investigations. In
general, the standard battery of formal toxicology analyses in
the preclinical setting that are conducted for any new drug
should be sufficient to catch any tissue specific adverse
outcome with a nanomedicine. This may be a good guiding
principle, however, it should be recognized that additional
testing in the preclinical setting may be required that is
specific for the behavior of the particular product. As an
example, in case of the materials that are persistent, not
readily excreted, eliminated or metabolized, or reside in
particular tissues for extended periods, it is reasonable to
expect a regulatory agency to require that the consequences
of the longer persistence be fully evaluated. In contrast,
nanomaterials that can be proven to be rapidly eliminated
from the body may not require protracted testing.
Important and unique to nanomedicines is the safety of
the nanoparticulate system as a whole. International stan-
dard-setting bodies have recognized this implication and
agreed that “as a minimum set of measurements—size, zeta
potential (surface charge), and solubility” of nanoparticles
should be used as predictors of nanoparticle toxicity (74). For
example, when inhaled, nanomaterials less than 100 nm can
induce pulmonary inflammation and oxidative stress (56) and
disrupt distal organ functions through mechanisms including
hydrophobic interactions, redox cycling, and free radical
formation. Unstable nanoparticles may form large aggregates
in micrometer size scale, which can be entrapped in the
capillary bed of the lungs and pose a serious danger to
patients. Notwithstanding these suggestions, it should be
recognized that standard toxicology studies required before
moving a product into the clinic will more than likely pick up
any manifestation of such toxicities due to the extensive
Immunological Challenges of Nanomedicines
However, one set of toxicities that cannot readily carry over
from preclinical testing to humans is immunotoxicity. The
immune response can be elicited by different sources. Biologics
such as proteins, peptides, antibody fragments, and nucleic acids
in nanoparticles can serve as antigens. Interaction of drug and
carrier can result in conformation changes that increase
289 Challenges in Development of Nanoparticle-Based Therapeutics
immunogenicity. For example, immunological issues can arise
from paclitaxel interacting with albumin (75). In the case of nab-
paclitaxel, an immunological type response was observed in pigs
with nab-paclitaxel drug but not the albumin control, supporting
the observations reported by Trynda-Lemiesz et al. (72). The
(BSA) resulted in the generation of particle-specific antibodies
and was used for immunization (76,77). Polyamidoamine den-
drimers conjugated to BSA also showed increased antigenic
potentials and induced dendrimer-specific antibody (78). The
complex manufacturing process of nanoparticle-based medicines
presents many opportunities for endotoxin contamination, which
is also a source for immune response.
Nanoparticles can be antigenic themselves, with the
immunogenicity of nanoparticles being affected by their size,
surface characteristics, charge, hydrophobicity, and solubility.
Depending on these properties, some nanoparticles can be
opsonized by plasma proteins and recognized as foreign bodies,
resulting in the activation of complement pathway. The
complement activation can lead to rapid phagocytosis and
clearance by macrophages of the MPS system in the liver and
spleen, such as in the cases of superparamagnetic iron oxide
nanoparticles Ferumoxtran-10 (Combidex) and ferumoxytol
(Feraheme) (79). More importantly, complement activation
may also provoke undesirable consequences including life-
threatening allergic, anaphylacticandhypersensitivity reactions,
as well as activation of humoral and cellular immune responses
against the nanoparticles (72,80,81). For example, Abrams et al.
reported that liposomal siRNA delivery vehicle LNP201
induced cytokine release typical of unregulated innate immune
“cytokine storm” (82). While PEG or other types of polymers
can shield nanoparticles from immune recognition, reports have
shown that administration of PEG-coated liposomes results in
the formation of PEG-specific antibodies, which accelerate
clearance of the PEG-liposomes and alters their PK and safety
profiles (83–85). This is well recognized for liposomal doxoru-
bicin, which requires premedication as a part of the administra-
tion regimen to ameliorate the cytokine effects and also shows a
change in PK overtime due to increased clearance (86).
Nanoparticles have also been reported to increase
antigenicity of weak antigens and thus serve as adjuvants.
This proves to be useful in the development of nanoparticle-
based vaccines but is detrimental to nanomedicines for other
indications. The adsorption of high molecular weight materi-
als to colloidal particles is used clinically to increase antibody
titer (72). A preclinical study has demonstrated that lecithin
nanoparticles serve as an adjuvant to increase an immune
response against protein antigens more than six times
stronger than the aluminum hydroxide adjuvant (87), and
Kreuter et al. have reported the use of polymethylmethacrylate
nanoparticles as adjuvants (88).
Nanoparticles have also been associated with other
hematologic safety concerns, including hemolysis and throm-
bogenicity. The hemolysis can be immune-mediated through
nanoparticle-specific antibody or non-immunogenic through
nanoparticle–erythrocyte interaction (81). Studies have
shown that positive surface charge from cationic surface
groups such as unprotected primary amines increases the
erythrocyte damage and hemolytic potency of nanoparticles
(89,90). Nanoparticles with a combination of hydrophobic
and hydrophilic areas on the surfaces can act as surfactants to
disrupt erythrocyte membrane (91). Hemolysis induced by
nanoparticles can lead to severe side effects such as anemia.
The released hemoglobin and cell debris can in turn attach to
the nanoparticles, causing rapid clearance and potential
immune response (92,93).
Thrombogenicity of nanoparticles is the outcome of
interactions between nanoparticles and blood coagulation
components and can induce blood clotting and partial or
complete occlusion of blood vessels. This concern is further
exacerbated by nanoparticles engineered to have longer circu-
lation time. In the most severe form, nanoparticles can cause
potentially fatal disseminated intravascular coagulation (DIC).
Greish et al. reported that amine-terminated dendrimers
triggered DIC in mice whereas carboxyl- and hydroxyl-termi-
nated dendrimers of similar sizes were tolerated (94).
These toxicity concerns of nanoparticles present significant
challenges in ensuring the safety of a nanoparticle-based
medicine. As shown above, the complexity of nanoparticle
systems can lead to a broad spectrum of toxicities, which are
directly related to specific aspects of nanoparticle properties. A
favorable safety profile would require careful adjustments of
components and parameters during the development of nano-
particles. Subtle differences in composition/conformation aris-
ing from variations in manufacturing could alter the toxicities of
nanoparticles, which means that a manufacturing process that is
not well controlled for highly selective parameters is at risk for
variations in immunogenic potential from batch to batch. In
addition this has implications for potential future generic
versions of nanomedicines, which may have the same compo-
nents as the innovator, but could result in a different immuno-
logical profile as a result of subtle variations in composition
resulting from a different manufacturing process or from
inadequate control of the manufacturing process.
The testing of nanoparticle toxicity constitutes another
challenge. Currently, there is no standard list of required
tests. A list of in vitro assays can test the interaction of
nanoparticles with the immune system, which include assays
for hemolysis, platelet aggregation, plasma coagulation,
complement activation, plasma protein binding, phagocytosis,
CFU-GM, leukocyte proliferation, nitric oxide production by
macrophages, and chemotaxis (81). Rodents generally are not
predictive of immunological responses, while rabbits are
hypersensitive to antigens. Overall, the strength of immune
responses in different species follows the order of mouse<rat
<dog<primate<human<rabbit. Therefore, preclinical toxici-
ty study results, particularly those related to immunotoxicity,
can not accurately predict the safety of nanomedicines in
human. In clinical studies evaluating nanomedicine toxicity,
special attention needs to be paid to side effects related to
immune response, particularly for nanomedicines that contain
a biologic component such as proteins, peptides, antibodies,
and antibody fragments. It is likely that for nanomedicine
products, immunological studies may have to be carried out in
human clinical trials.
REGULATORY CHALLENGES TO NANOMEDICINE
Due to the complexity and large potential diversity of
nanoparticle-based products, it may seem apparent that the
regulatory pathway for nanomedicines may face several
hurdles. Currently, the FDA, EMA, and other regulatory
agencies examine each new nanoparticle-based drug on a
product-by-product basis. There is generally a lack of stand-
ards in the examination of nanomedicines as a unique
category of therapeutic agents. Recent movements towards
establishing some definitions and guidelines (73,95,96) are
first steps in determining if additional regulation will be
applied to nanomedicines.
The complex nature of nanoparticle-based medicines
with their multiple components, where more than one
component can affect pharmacological behavior of the active,
contrasts against standard drugs where there is usually a
single active agent and the other components mostly serve as
inactive formulation aids (excipients). It is reasonable to
expect that nanomedicine raises complicated regulatory
strategies, and processes are likely to be significantly more
complex. A few fundamental and logical questions may help
simplify the discussion around potential regulatory complex-
ity of nanomedicine products while maintaining the basic
principles guiding the regulatory agencies to protect public
safety while providing access to new and innovative
1. Physico-chemical characteristics: what are the key charac-
teristics of the product that are essential for its activity and
safety, and are those critical characteristics of the product
reproduced within acceptable pharmaceutical tolerances in
2. Biodistribution: are there any particular properties of
the product that one would expect unusual biodistribu-
tion or more importantly cause persistence of the
product in particular tissues over extended periods of
time, intentionally or otherwise? If so, what are their
3. Clinical: what human clinical data should be collected
to evaluate potential immunological responses to the
nanomedicine, whether acute or on a longer term basis
upon repeated administration?
4. Definition: does the product meet the criteria of an
acceptable, scientifically sound description or definition
(these are still evolving) of what can be considered a
nanomedicine (certain size constraints as well as unique
It should be noted that we have recently entered the era
of generic nanomedicines. Both generic drug manufacturers
and drug regulators will be faced with major challenges in
defining what studies will be required to demonstrate that the
generic nanomedicine is bioequivalent to the innovator and
that the products have the same physicochemical properties
and are safe and effective. For example, there have been
several unsuccessful attempts in the marketplace to copy the
nab-paclitaxel formulation. These attempted formulations
which the manufacturers claimed were copies of approved
nab-paclitaxel, when tested, failed to reproduce size distribu-
tion, stability, potency, or physicochemical characteristics of
nab-paclitaxel, which could potentially lead to undesirable
and unsafe effects. In one case, the claimed copy had high
endotoxin and residual solvent levels greatly exceeding
recognized safety limits. In a separate case, the attempted
formulation showed substantial inter-batch variations in
particle size, suggesting poor reproducibility in the manufac-
turing process. There was also a wide size distribution with a
large portion of particles over 200 nm, resulting in significant
drug loss after filtration through a 220-nm sterile filter. The
reconstituted nanoparticles also displayed poor stability
under accelerated conditions of 40°C and formed large
precipitates and aggregates of several micrometers in size
within 24 h (Fig. 5a, b), unlike nab-paclitaxel which was stable
under these conditions. Such tests suggest that fundamental
differences in the behavior of these formulations result from
differences in composition and manufacturing. These exam-
ples also illustrate how generic drug manufacturers and
health authorities are going to face unique challenges in the
development, regulation and approval of nanomedicines that
claim to be equivalent to the innovator products. These issues
will likely be no less challenging than the difficulties
surrounding the development and regulation of biosimilar
FDA has recently begun to consider relevant approval
standards for generic copies of nanomedicines. Several
Fig. 5. Undesirable precipitation and instability seen with an
attempted albumin-paclitaxel formulation modeled as a copy for
nab-paclitaxel. Reconstituted suspensions in saline were monitored
for 24 h a 40°C. No precipitation was seen with nab-paclitaxel. a
Image for the copied product at magnification of ×400 showing large
visible particles; b higher magnification image of copied product at
×1,000 clearly shows needle-shaped crystals of paclitaxel
291 Challenges in Development of Nanoparticle-Based Therapeutics
liposomal type products such as those containing the drugs
amphotericin and doxorubicin have recently gone off patent.
Understandably, the complexity of such products should
necessitate a different standard of “equivalence” testing than
what is required for standard drugs. In the absence of critical
information relating to composition, three dimensional con-
figuration of components, and critical parameters that are
essential for function of nanomedicine products, there may be
the risk that “generic” versions are approved using the
conventional chemistry, manufacturing and controls and
bioequivalence standards for generic drug approvals which
may result in substandard products in the marketplace.
Equivalence in formulation and/or in standard blood phar-
macokinetics (bioequivalence) may not adequately represent
the function of the nanomedicine at the site of action, as is
assumed for most standard formulations. Therefore, it is
imperative that an exhaustive physicochemical understanding
of complex nanomedicine products and identification of
critical parameters that affect their functions be conducted
early in development to lay the ground rules for potential
nanomedicine generics in the future. Indeed, the FDA has
recently issued a guidance for liposomal doxorubicin (Doxil)
consistent with this approach (63).
In general, nanomedicines are complicated multicompo-
nent and multifunctional drug-delivery systems. There is an
urgent need for the regulatory agencies to develop a
comprehensive list of tests and a streamlined approval
process that covers the whole range of particle characteriza-
tion, pharmacology, and toxicology issues. The overall
behavior, PK, and safety profile of nanoparticles is the
combined results of interplay of all nanoparticle components,
parameters, and spatial composition. There is inadequate
understanding of the connection between nanoparticle phys-
icochemical properties and its clinical PK and safety, and
conventional animal models may be insufficient to correctly
extrapolate and predict nanoparticle biodistribution and
toxicity in humans. This is especially relevant when compar-
ing a novel nanoparticle-based drug with conventional
formulations, and when evaluating a generic version of an
approved nanomedicine versus the innovator product.
Bioequivalence of a generic and innovator nanomedicine
cannot be assumed by similar results observed in general PK
and toxicity studies, or by a simple comparison of the
composition of the drug products. Rather, disease models
should be applied when possible to reflect the pharmacology
of nanomedicines at the intended target sites. Although
clinical trials can assess the short-term toxicity of nano-
medicines, long-term effects of their accumulation and
chronic exposure may require continued monitoring over
extended period of time.
Finally, many of the nanoparticle-based medicines in
development consider a targeting approach to a particular
receptor or protein that may be expressed, e.g., by tumors. It
would be advantageous to consider a “personalized” ap-
proach to treatment and identify these subgroups of patients
that are more likely to respond. Such an approach could
considerably reduce the size of the required clinical trial and
could be considered favorable by the regulatory agencies
since a larger fraction of the patients treated under this
approach would benefit from treatment. Hence, a personal-
ized approach to nanoparticle-based medicine may not only
reduce costs of running large clinical studies but could
provide a speedier path to approval.
SUMMARY AND CONCLUSIONS
In summary, a new nanoparticle-based medicine
needs to successfully overcome several hurdles before it
is approved for marketing. These include the development
of the nanostructure with appropriate components and
properties, engineering of a reproducible manufacturing
process, selection of orthogonal analysis methods for
adequate characterization, a favorable pharmacology and
toxicity profile, and demonstration of safety and efficacy
in clinical trials. While conceptually these are similar
hurdles that may be faced by any new drug, the particular
complexity and multicomponent nature of nanomedicines
introduce large number of additional variables that may
substantially increase the level of difficulty in controlling
processes and predictability of behavior in a biological
system. Additional regulatory and development consider-
ations arise when generic nanomedicines are presented for
health authority approval with claims of equivalence to
the innovator drug.
The further development of nanomedicines will likely
include a personalized medicine approach as an integral part
of the clinical development strategy to identify subgroups of
that particularly benefit from therapy. In addition to the many
benefits of this approach and particularly true in oncology, is
the reduction in size of clinical trials potentially leading to
approval. Additionally, in the face of the natural biological
barriers to the delivery of drugs, tissues with aberrant
pathology, such as tumors, are often very efficient in
harnessing active biological mechanisms for high nutrient
supply and rapid growth. Elucidation of these active transport
mechanisms and the ability to harness these with nano-
medicines could provide a step forward in the treatment of
cancer and other diseases.
Finally, health authorities that are charged with respon-
sibility for the approval of safe and effective medicines will
need to respond to the challenges posed by the emergence of
products based on new technologies. Appropriate processes
to develop definitions, quality standards, and requirements
fordevelopment studies including clinicaltrials,mustbeinplace
to proactively address rapid advances in drug development.
The expert assistance of Shihe Hou in compiling this
manuscript is deeply appreciated. Figure 1 of this manuscript
has been adapted from an illustration originally published in
Nanomedicine, June 2010, Vol. 5, No. 4, Pages 597–615. We
thank Future Medicine Ltd. for kindly granting permission to
reproduce the material, and thank Dr. Russell J. Mumper and
Dr. Xiaowei Dong for generously allowing the use of
illustration from their publication. We thank Willard Foss,
Viktor Peykov, Tapas De and Jon Cordia at Celgene for
providing data on attempted copies of nab-paclitaxel. We
thank Richard Girards, Gad Soffer, Mitch Clark, Renu Vaish
and Anita Schmid for their valuable comments on the
Open Access This article is distributed under the terms of the
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distribution, and reproduction in any medium, provided the
original author(s) and the source are credited.
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