Journal Pre-proof

Delivery of erlotinib for enhanced cancer treatment: An update review on particulate

systems

Duy Hieu Truong, Vu Khanh Hoa Le, Tung Thanh Pham, Anh Hoang Dao, Thi
Phuong Dung Pham, Tuan Hiep Tran

PII: S1773-2247(19)31231-6
DOI: https://doi.org/10.1016/j.jddst.2019.101348
Reference: JDDST 101348

To appear in: Journal of Drug Delivery Science and Technology

Received Date: 19 August 2019

Revised Date: 11 October 2019
Accepted Date: 22 October 2019

Please cite this article as: D.H. Truong, V.K. Hoa Le, T.T. Pham, A.H. Dao, T.P. Dung Pham, T.H. Tran,
Delivery of erlotinib for enhanced cancer treatment: An update review on particulate systems, Journal of
Drug Delivery Science and Technology (2019), doi: https://doi.org/10.1016/j.jddst.2019.101348.

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Graphical Abstract
 Delivery of erlotinib for enhanced cancer treatment: an update review on
particulate systems

Duy Hieu Truonga,1, Vu Khanh Hoa Leb,1, Tung Thanh Phamc,

Anh Hoang Daod, Thi Phuong Dung Phame , Tuan Hiep Tranf,g,*

a
Institute of Research and Development, Duy Tan University, Da Nang, Vietnam
b
NTT Hi-Tech Institute, Nguyen Tat Thanh University, Ho Chi Minh City, Vietnam
c
College of Pharmacy, Yeungnam University, Gyeongsan, Gyeongbuk 38541, South Korea
d
National Institute of Medicinal Materials, Hanoi, Vietnam
e
Faculty of Pharmacy, Dai Nam University, Hanoi, Vietnam
f
Department for Management of Science and Technology Development, Ton Duc Thang

University, Ho Chi Minh City, Vietnam

g
Faculty of Pharmacy, Ton Duc Thang University, Ho Chi Minh City, Vietnam
1
These authors contributed equally to this manuscript.

Authors to whom correspondence should be addressed:

* Corresponding author: Tuan Hiep Tran, Ph.D.

Ton Duc Thang University, Ho Chi Minh City, Vietnam

E-mail: trantuanhiep@tdt.edu.vn

(ORCID: 0000-0002-8732-8230)

First author’s email: truongduyhieu@gmail.com (Duy Hieu Truong)

(ORCID: 0000-0002-1720-1744).

1
 Delivery of erlotinib for enhanced cancer treatment: an update review on
particulate systems

Abstract

Erlotinib is an epidermal growth factor receptor (EGFR) inhibitor that has been approved for
metastatic non-small cell lung cancer and highly aggressive pancreatic cancer. Although
erlotinib contributes progressively to cancer treatment, there is still room to develop a better
platform for erlotinib-based therapy. The progression of the materials science field has offered
many options to incorporate drugs with additional advantages. In this review, we summarize
various strategies, including polymeric, lipidic, inorganic and hybrid-based carriers, to
improve drug pharmacokinetics, reduce interpatient variability and minimize unwanted side
effects. Moreover, the combination of erlotinib with other therapeutic agents or modalities is
further discussed as a potential paradigm for realizing better efficacy and/or lower systemic
toxicity in clinical practice.

Key words: erlotinib; EGFR; nanoparticles; delivery systems; efficacy enhancement;

cancer.

2
1. Introduction

Erlotinib (ERL), a quinazoline derivative with the chemical name N-(3-ethynylphenyl)-6,7-

bis(2-methoxyethoxy)-4-quinazolinamine, is an FDA-approved small-molecule tyrosine
kinase inhibitor for the treatment of metastatic non-small cell lung cancer (NSCLC) and
locally advanced, unresectable or metastatic pancreatic cancer in combination with
gemcitabine [1]. By selectively occupying the adenosine triphosphate (ATP) binding sites of
epidermal growth factor receptor (EGFR), this molecule reversely inhibits EGFR activation,
resulting in the suppression of further downstream signaling processes, mostly through the
phosphatidylinositol 3-kinase (PI3K) and mitogen activated protein kinase (MAPK)
pathways, and consequent inhibition of cellular proliferation, angiogenesis and metastases [2,
3]. To date, the potential of this substance has been expanded to treat other types of cancer,
such as ovarian cancer, head and neck cancer, and glioblastoma [4-8].

As a substance with low aqueous solubility and high permeability (logP of 2.7), ERL is
categorized into class II in the biopharmaceutical classification system [9]. Regarding the oral
route of administration, its poor solubility restricts the dissolution of the drug in the
gastrointestinal fluid that can limit its absorption, thereby causing high fluctuation in its
bioavailability and large interpatient variability [10, 11]. Although the commercial tablet
dosage form (Tarceva®) shows an acceptable bioavailability of approximately 60% in healthy
volunteers upon oral administration of 150 mg [12], similar to most of the compounds used
for chemotherapy, patients receiving this therapy still suffer from drug resistance and severe
toxicities, i.e., skin rash, diarrhea, gastrointestinal perforations and Stevens-Johnson
syndrome, that might hamper its clinical application [13]. These obstacles necessitate the
careful design of suitable delivery systems for not only improved efficacy but also reduced
side effects.

Recently, nanotechnology has been widely used to improve the physicochemical and
pharmacokinetic properties of poorly water-soluble drugs, such as solubility, dissolution,
absorption and bioavailability [14-16]. There have been many different techniques or
strategies that can be exploited for these purposes, e.g., spray drying, freeze drying, wet
milling and high-pressure homogenization, to fabricate various nanoformulations, such as
solid dispersions, drug nanocrystals and solid lipid nanoparticles (SLNs) [17-20]. What if
these technologies can enhance the properties of ERL? What is the outcome of these changes?
What is the potential of those interventions for clinical application? To answer these
questions, in this review, we summarize several approaches to improve the therapeutic
3
efficacy of ERL as a therapeutic agent alone (Table 1) and in combination with other
therapeutic compounds or modalities (Table 2) using various drug delivery systems, mainly
focusing on nanocarriers.

2. Drug delivery systems containing erlotinib alone

2.1. Polymer-based systems

With the advance of chemical engineering, polymers, a large group of compounds with
versatile physical and chemical properties, can be tuned to make nanocarriers for controlling
the temporal or spatial release of therapeutic agents [21]. Among the different types of
polymers, biodegradable polymers, such as poly(D,L-lactic-co-glycolic acid) (PLGA) and
polycaprolactone-derived copolymers, are preferable to use when designing delivery systems
because of their high safety profiles and controlled release capability of cargo [22, 23]. To
evaluate the safety of PLGA for delivering ERL, this drug was loaded into PLGA
nanoparticles (NPs), and then the toxicity of these NPs following oral administration in
Wistar rats was assessed [24]. Compared to free ERL, these ERL-encapsulated NPs showed
significantly lower toxicity and damage to internal organs, as confirmed by hematological and
biological analyses and histopathological examination. In another study, polycaprolactone -
polyethylene glycol - polycaprolactone (PCEC) triblock copolymers were used to prepare
polymeric NPs in which ERL was incorporated into their lipophilic cores [25]. The results
showed that the molar ratios of the polycaprolactone and polyethylene glycol monomers
defined the characteristics of these NPs. In particular, the higher the ratio was, the larger the
NP size and encapsulation efficacy of ERL, and the lower release rate of ERL from the NPs.
Drug-encapsulated NPs showed remarkably higher cytotoxicity than that of the free drug,
especially at higher doses and longer incubation periods, in A549 lung cancer cells.

For a long period of time, cyclodextrins have been used to make inclusion complexes with
various drug compounds to improve their solubility and bioavailability [26-28]. However,
natural cyclodextrins are not highly soluble in aqueous solutions, which limits their practical
applications as delivery vehicles. To overcome this hurdle, Suresh’s group employed one of
the more hydrophilic cyclodextrin derivatives, sulfobutyl ether β-cyclodextrin, to formulate an
inclusion complex with ERL by the freeze drying method [29]. By using the design of
experiments technique, the molar ratio of 1.05:1 between ERL and sulfobutyl ether β-
cyclodextrin was found to be the optimal ratio to easily prepare an inclusion complex.
Compared to free ERL, this formulation demonstrated more than 2-fold higher dissolution at
4
60 minutes and 3.6-fold higher AUC0-∞. Later, this group utilized a β-cyclodextrin-based
nanosponge, which was prepared by cross-linking a β-cyclodextrin monomer and the cross-
linker carbonyldiimidazole, for complexing with ERL [30]. After performing some
preliminary solubility experiments, the formulation with the optimum ERL and nanosponge
ratio of 1:4 was prepared and characterized. In comparison to pure ERL, this optimized
formula approximately doubled the dissolution rate and oral bioavailability and exhibited
more cytotoxic effects on the MIA PaCa-2 and PANC-1 pancreatic cancer cell lines. For lung-
specific delivery of ERL, Momin et al. synthesized glutathione nanosponges through a high-
yielding reaction among β-cyclodextrin, 2-hydroxyethyl disulfide and the cross-linker
pyromellitic dianhydride [31]. ERL-loaded glutathione nanosponges were able to show a
sustained release pattern of ERL (up to 7 days), which was proportionally dependent on the
concentrations of glutathione in the surrounding environment. The drug-loaded glutathione
nanosponges not only were markedly more cytotoxic in A549 NSCLC cells than free ERL but
also demonstrated higher tumor growth inhibition in a xenograft mouse model of lung cancer
with significantly lower biodistribution in the spleen, heart, kidney and liver resulting from
the targeting ability of glutathione. Taken together, these results suggested that complexation
with β-cyclodextrin derivatives could be an effective method to enhance the efficacy of ERL
with reduced side effects.

Albumin has been proven as a safe vehicle that is approved in many clinical products for drug
delivery and bioimaging purposes, the most notable of which is Abraxane® (paclitaxel
albumin-bound particles for injectable suspension) [32]. By using the same concept, Noorani
and colleagues synthesized ERL-loaded albumin NPs by desolvation followed by a thermal
cross-linking method [33]. The use of an aqueous solution of albumin at pH 5 and an
acetone/dimethyl sulfoxide mixture with a 5:1 (v/v) ratio as the organic solvent resulted in a
high drug loading capacity (approximately 27%) of ERL in the albumin NPs with
hydrophobic interactions and hydrogen bonds between the two components. In both the
ASPC-1 and PANC-1 pancreatic cancer cell lines, the ERL-loaded albumin NPs demonstrated
higher cytotoxicity than that of free drug. In another work, Shen and Li coated ERL albumin-
bound NPs with a hyaluronic acid layer to specifically target CD44-overexpressing tumor
cells [34]. Due to the presence of the coating layer, the NPs showed excellent stability after 3
months and slower drug release than their uncoated counterparts. Additionally, compared to
the uncoated NPs and free ERL, the NPs whose surface was modified by hyaluronic acid
exhibited significantly enhanced uptake into A549 lung cancer cells, thereby further

5
suppressing their proliferation. Importantly, the coated NPs demonstrated remarkable tumor
growth inhibition in tumor-bearing mice without relapse within 30 days posttreatment.

In another effort to enhance the oral bioavailability of ERL and reduce the variation in
bioavailability between the fasted and fed states, a novel nanoparticulation method using fat
and supercritical fluid (NUFSTM) technology was applied [35]. In this method, ERL was
melted with a mixture of myristyl alcohol and polyvinylpyrrolidone at 130°C before being
rapidly cooled to 15°C for the preparation of nanoparticulated ERL. Then, solidified myristyl
alcohol was removed by supercritical CO2 to obtain the formulation as a dried powder. This
formulation showed higher dissolution rates under both fasted and fed conditions and a 5.5-
fold higher AUC in the fasted state than did Tarceva. Additionally, the ratio of fasted AUC to
fed AUC for the formulation was 1:1.8, which was significantly lower than that of Tarceva
(1:5.8). In particular, compared to Tarceva, the formulation markedly improved antitumor
efficacy in the A549 xenograft tumor model, which likely resulted from the enhanced
dissolution and potent inhibition of EGFR signaling.

2.2. Lipid-based systems

Lipid-based delivery carriers have demonstrated great promise in enhancing the delivery of
poorly water-soluble drugs, with numerous successful commercial products, such as Neoral®
(self-emulsifying formulations of cyclosporine), Onivyde® (liposomal irinotecan) and
AmBisome® (liposomal amphotericin B). Given the similarity in solubility between these
drugs and ERL, the abovementioned products suggest several potential options of lipid
carriers that can be used to develop ERL-containing formulations with improved profiles.

Our work sought to utilize a self-emulsifying formulation to deliver ERL [36]. By using the
ternary phase diagram, we chose the optimized formulation, consisting of 5% w/w of oil
(linoleoyl polyoxyl-6 glycerides or Labrafil® M2125CS), 65% w/w of surfactant
(caprylocaproyl polyoxyl-8 glycerides or Labrasol®) and 30% w/w of cosolvent (diethylene
glycol monoethyl ether or Transcutol® HP), to dissolve ERL and then solidified the resultant
solution by spray-drying with dextran 40 or Aerosil® 200 as the inert carrier. The
incorporation of ERL into these formulations in the amorphous form or as a molecular
dispersion remarkably improved its dissolution and bioavailability compared to those of the
free drug.

6
Zhou and coworkers incorporated ERL into the hydrophobic layer of a PEGylated liposomal
formulation by the thin-film hydration method [37]. In contrast to non-PEGylated ERL-
loaded liposomes, the PEGylated ERL-loaded liposomes exhibited slightly higher cellular
cytotoxicity and a 2-fold increase in bioavailability without causing any apparent damage to
tissues of vital internal organs owing to the enhanced permeation and retention effect.

The most well-known indication of ERL is to treat NSCLC; hence, direct delivery of ERL to
lung cancer tissues and cells is highly desirable. For that purpose, a microparticulate
formulation of inhalable dry powder (IDP) containing ERL-encapsulated SLNs was prepared
[38]. The optimal SLNs, composed of Compritol® 888 ATO, Tween® 80 and Poloxamer 407
at a weight ratio of 24:1:20, were spherical particles with sizes of less than 100 nm. The ERL-
encapsulated SLNs showed enhanced cytotoxic effects of ERL on A549 human NSCLC cells.
For the delivery of ERL to deep lung tissues, the ERL-loaded SLNs were spray-dried into
mannitol, forming microparticles (1-5 µm) with suitable flowability and aerodynamic
properties, as confirmed by Hausner ratio, Carr’s index and the results of the pharmacopoeial
Next Generation Impactor® (NGI) test. Another SLN-loaded IDP for the pulmonary delivery
of ERL was developed by Naseri et al. using different components [39]. The oily phase,
which was composed of glycerol monostearate, Transcutol®, poloxamer 188 and ERL, was
mixed in distilled water to form SLNs. Similar to the abovementioned study, these drug-
loaded SLNs showed higher cytotoxicity than that of the free drug in the A549 cell line. The
fine particle fraction (FPF) and mass median aerodynamic diameter (MMAD) values of three
mannitol-based powders were approximately 10% and 4.5 - 6.7 µm, respectively, similar to
those of commercial products, demonstrating the suitability of these inhalable formulations
for lung cancer treatment.

In addition to using polymers to improve ERL’s efficacy [29, 30], Suresh’s group also
cooperated with Jain’s group to employ phosphatidyl choline for developing a molecular
complex with ERL [40]. A drug-to-phospholipid ratio of 1 to 2 promoted stable complexation
where the drug was engulfed by the phospholipid, thereby transforming the drug into a less
crystalline phase and improving its hydrophilicity, which led to 5.4-fold and 1.7-fold
increases in the aqueous solubility and oral bioavailability of ERL, respectively. Along with
exhibiting higher cytotoxicity and apoptotic index in two human pancreas adenocarcinoma
cell lines (i.e., MIA PaCa-2 and PANC-1) owing to efficient internalization, the ERL-
phospholipid complex also demonstrated better safety profiles in blood and normal tissues

7
than those of free ERL, which might result from the protective and free radical scavenging
effects of the phospholipid.

2.3. Inorganic systems

Magnetic NPs have versatile applications in cancer therapy, such as diagnosis, drug delivery
and theranostics [41-43]. Xu et al. prepared carbon-coated iron magnetic NPs to combine the
effect of hyperthermal therapy induced by radio frequency with the effect of ERL attached
onto the carbon shells [44]. Because of the strong affinity of the drug for the magnetic NPs, as
shown by the high loading capacity, only 30% of the drug was released at pH 7.4, with a
slight increase to approximately 41% at pH 5.5 resulting from its protonation. Under a radio
frequency of 350 kHz, the release of ERL from the NPs was also amplified, especially in an
acidic pH environment. In PANC-1 pancreatic cells treated with the magnetic NPs, although
radio frequency did not significantly increase apoptosis, it halved cell proliferation to 25.4%.

Theranostics, which integrate diagnostics and therapeutics into a single platform, have
become increasingly important in clinical oncology and personalized medicine [45].
Theranostic agents can simultaneously provide the results of diagnostic tests and monitor drug
effects in real time for early disease detection and optimization of personal treatment.
Considering the dual role of ERL as a therapeutic and EGFR-targeting agent, Chen’s
laboratory designed a theranostic platform based on ERL-conjugated ultrasmall
superparamagnetic iron oxide nanoparticles (FeDC-E NPs) [46]. With ERL on the surface, the
FeDC-E NPs preferentially accumulated inside EGFR-overexpressing cells, intracellularly
released ERL and productively eradicated these cells. These conjugated NPs showed
remarkable suppression of the EGFR-extracellular signal-regulated kinase-nuclear factor
kappa B (EGFR-ERK-NF-κB) signaling pathways, which subsequently resulted in inhibition
of the migration and invasion of CL1-5-F4 lung adenocarcinoma cells. Notably, in a
xenograft-bearing mouse model, the FeDC-E NPs markedly decreased tumor growth and
reduced the normalized T2-weighted MRI signal intensities within the tumor. Recently, this
group combined the MRI technique with a nuclear factor kappa-B (NF-κB) reporter gene
system to further evaluate the FeDC-E NP treatment response for the potential clinical
application of these NPs against NSCLC and to understand their roles in the apoptotic
mechanisms (Fig. 1A) [47]. They found that the cellular uptake and intratumoral distribution
of this probe could be noninvasively monitored by using ∆R2* values from MRI.
Furthermore, because of the NF-κB reporter gene system, the apoptotic mechanism of the
8
FeDC-E NPs was shown to effectively suppress antiapoptotic effects through NF-κB
signaling and significantly induce intrinsic and extrinsic apoptotic pathways in vitro and in
vivo. These results demonstrated the potential of this theranostic probe for clinical translation
in the treatment of lung cancer.

To enhance the targeting to EGFR-overexpressing pancreatic cancer cells and combine

multimodal imaging techniques for early cancer diagnosis and treatment, ERL was conjugated
to iron oxide-gold nanoclusters to form a Fe3O4@AuNCs@ERL nanocomposite (Fig. 1B)
[48]. The core-shell nanocomposite improved the uptake and cytotoxicity of ERL in PANC-1
pancreatic cells owing to the appearance of ERL while still preserving the intense
fluorescence property of the gold nanoclusters, which can be employed to monitor these cells
before surgery by confocal laser scanning microscopy imaging.

2.4. Hybrid systems

Mandal et al. developed core-shell-type lipid–polymer hybrid NPs in which an ERL-

incorporated polymeric polycaprolactone core was covered by a phospholipid monolayer [49].
Optimization of the formulation and process parameters resulted in an optimized formula with
a size of 170 nm, making it suitable for intravenous injection. The CellTitre-Glo® luminescent
cell viability assay showed that in comparison with ERL solution, the ERL-loaded hybrid NPs
remarkably reduced the viability of A549 cells with a 25-fold lower IC50 value (100 nM),
which was consistent with the result of a colony formation assay.

As a strategy for active targeting of ERL to lung cancer cells with EGFR gene mutation—a
phenomenon that can cause drug resistance and limit long-term efficacy—Li et al. anchored
EGFR-binding aptamer-conjugated chitosan onto liposomes, forming multifunctional
liposomal complexes [50]. The aptamer-modified complexes showed enhanced recognition
and internalization by EGFR-mutated H1975 and PC-9 cells compared to EGFR-negative
Helf cells and an increased drug release percentage in the acidic tumor microenvironment
compared to that of their unmodified counterparts. These results likely contributed to the
observably increased antiproliferation and induction of cell cycle arrest and apoptosis by the
aptamer-modified complexes in these lung cancer cells, and the effects on drug-resistant
H1975 cells were much more pronounced than those on drug-sensitive PC-9 cells, supporting
the usage of the selected aptamer to overcome erlotinib resistance caused by EGFR mutation.

9
Recently, tumor microenvironment-responsive NPs have attracted great attention in cancer
therapy due to their capability to specifically target cancer sites by exploiting the different
signals and conditions in the tumor microenvironment compared to healthy tissues, such as
acidic pH and abnormal levels of enzymes, glutathione, and reactive oxygen species [51-56].
Kim et al. used lipid nanocapsules with a PEGylated polypeptide shell, which can prolong
systemic circulation, and a pH-responsive poly(L-aspartic acid) core, which can accelerate the
release of encapsulated cargo in acidic environments, for ERL delivery [57]. As expected, the
lipid nanocapsules exhibited efficient cellular uptake into cancer cells and higher drug release
at pH 5.0 than at pH 7.4. In addition, in comparison with the untreated control and free ERL,
the ERL-encapsulated nanocapsules were dose-dependently cytotoxic to HCC-827 and NCI-
H358 lung cancer cells and significantly retarded tumor growth in mice bearing an HCC-827
xenograft tumor.

Regarding the application of thermoresponsive NPs, Fathi et al. exploited a widely used
thermosensitive polymer, poly(N-isopropylacrylamide) (PNIPAAm), and chemosensitivity-
induced oleic acid for developing a polymer-gold hybrid nanomaterial in which biodegradable
chitosan served as the reducing agent of auric salt (HAuCl4) and steric stabilizer for the
resulting NPs [58]. In addition to demonstrating excellent stability in the pH range from 3 to 8
for several months, the polymer-gold hybrid nanomaterials exhibited temperature-dependent
release of ERL, in which drug release was clearly faster above the lower critical solution
temperature. The efficient internalization of the hybrid NPs into A549 cells (approximately
86% over 3 hours of incubation) led to greater cytotoxicity in this cell line. To further enhance
the active targeting ability of thermosensitive micelles based on PNIPAAm, this group
modified the surface of these NPs with folic acid [59]. As expected, the folate-modified
micelles were significantly more cytotoxic to the folate receptor-positive OVCAR-3 ovarian
carcinoma cells than to the folate receptor-negative A549 lung adenocarcinoma cells, which
was consistent with enhanced internalization into the OVCAR-3 cells. Additionally, drug-
loaded surface-modified NPs demonstrated higher cytotoxicity than the corresponding free
drug, suggesting that receptor-mediated endocytosis of the targeted NPs might help the P-
glycoprotein substrate ERL bypass the efflux mechanism through this transporter.

To enhance ERL delivery to cancer sites using multiple stimuli, Tan et al. prepared pH-
sensitive and redox-responsive NPs (PAA-ERL-NPs) based on poly(acrylic acid)-cystamine-
oleic acid (PAA-ss-OA), which was formed through the conjugation among pH-sensitive
poly(acrylic acid), oleic acid and cystamine [60]. The outer layer composed of poly(acrylic
10
acid) decelerated drug release from the PAA-ERL-NPs, which likely resulted from the fluid-
phase endocytosis/pinocytosis of these NPs into endosomal compartments before releasing
ERL and inducing more cytotoxicity than that of free ERL and ERL-loaded nonlayered NPs
in A549 and NCI-H460 NSCLC cells. Additionally, the PAA-ERL-NPs exhibited the highest
antitumor effect in a lung cancer xenograft mouse model, supporting the efficacy of the dual-
targeting approach in lung cancer treatment. In another study, a folic acid-poly(ethyleneimine)
conjugate was attached to the surface of magnetic mesoporous silica NPs to prepare a pH-
sensitive nanocarrier [61]. After four days, the percentage of ERL released from the coated
nanocarrier at pH 5.5 (approximately 63%) was nearly double that at pH 7.4, confirming the
pH-responsive properties of the nanocarrier. Moreover, despite exhibiting excellent
biocompatibility with red blood cells, these coated NPs demonstrated a significant cytotoxic
effect on HeLa cervical carcinoma cells owing to the overexpression of the folate receptor on
this cell line.

To repurpose the use of the potent but toxic kinase inhibitor ERL from therapy to safer
prevention of liver cancer, versatile myofibroblast-targeting NPs were fabricated (Fig. 2) [62].
The proof of concept was to conjugate ERL-loaded NPs with a short peptide that can
specifically bind to platelet-derived growth factor receptor β (PDGFRB) uniquely expressed
on the surface of hepatic myofibroblasts. Due to the polyethylene glycol layer coating, the
NPs showed a sustained release pattern of ERL with stable drug concentrations in rat plasma
over the period of 12 days (Fig. 2D, E), which is much longer than the half-life of ERL
(approximately 36 hours) [1]. Compared to non-PGFRB-expressing Hep G2 hepatoma cells,
two types of PGFRB-expressing myofibroblasts, TWNT-4 and LX-2 cells, demonstrated
higher uptake of the targeted NPs and dose-dependent phospho-EGFR inhibition. In
particular, by using a cirrhosis-driven liver cancer model in rats, the authors observed a
significant decrease in tumor size by 7.3-fold and reduced hepatocyte damage in the group
treated with the myofibroblast-targeting NPs compared with the group systemically treated
with the free drug at an equivalent dose.

3. Combination with other therapeutic agents or modalities

Combination therapies that combine two or more therapeutic agents or modalities, such as
photodynamic or photothermal therapies, possess several advantages over monotherapies in
cancer treatment. They are able to enhance efficacy and reduce tumor resistance by

11
simultaneously targeting various anticancer mechanisms, decrease costs and effort toward the
development of new therapeutic compounds, minimize side effects, etc. [63, 64].

Gupta et al. coloaded paclitaxel and ERL into the solid lipid core of nanocapsules having a
PEGylated block copolymer shell for dual-drug delivery to NSCLC cells [65]. The authors
found that these two drugs could be released in a pH-dependent and controlled manner.
Additionally, by using this delivery system, both drugs are efficiently taken up by NCI-H23
cells, resulting in remarkably higher cytotoxicity than that of the free drugs individually or in
combination.

To prevent or delay acquired resistance to ERL, a liquisolid formulation containing ERL and
valproic acid was prepared by using polyethylene glycol 400 as the solvent [66]. With the
assistance of calcium silicate as a microporous adsorbent, the optimized formula, in which the
molar ratio between ERL and valproic acid was 1:2, demonstrated a significant enhancement
in the dissolution and bioavailability of ERL owing to the ionic interaction between the
components. Importantly, this formulation exhibited substantially higher cytotoxicity and
apoptosis percentage in ERL-resistant HCC827 lung adenocarcinoma cells than those of ERL
alone by downregulating the expression of survivin.

To reverse hypoxia-induced drug resistance in lung cancer, Gao’s laboratory codelivered ERL
and perfluorooctylbromide—a high-capacity oxygen carrier—in aptamer-conjugated
chitosan-anchored liposomal complexes [67]. Owing to the presence of the anti-EGFR
aptamer, these multifunctional NPs were specifically recognized by EGFR-positive A549,
H1975 and PC-9 NSCLC cells, and compared to EGFR-negative Helf cells, these EGFR-
positive cells exhibited enhanced uptake of the NPs. While ERL-loaded complexes showed a
lower total percentage of ERL release under hypoxic conditions than under normoxic
conditions, these coloaded complexes still demonstrated similar release rates under both
conditions, which might result from the promotion of internal pressure by oxygen. In addition,
the coloaded NPs revealed strong effects on the inhibition of proliferation and induction of
apoptosis of NSCLC cells and a significant inhibitory effect on the expression of EGFR, p-
EGFR and hypoxia-inducible factor-1α (HIF-1α), especially under hypoxic conditions. In vivo
experiments showed profound tumor growth suppression and relatively lower distribution in
the spleen and heart in the groups of mice treated with the coloaded NPs compared to the
other groups. Later, this laboratory attempted to overcome ERL resistance in EGFR mutation-
positive NSCLC by synergizing the effects of chloroquine on normalizing the disturbed tumor
vascular structure and downregulating the anti-apoptotic protein survivin [68]. In this study,
12
the authors coloaded survivin-recombinant short hairpin RNA-expressing plasmid (survivin-
shRNA) with ERL into anti-EGFR aptamer-modified polyamidoamine dendrimers.
Chloroquine was shown to markedly amplify the intracellular uptake and gene transfection
efficiency of these coloaded nanocomplexes, thereby enhancing their cytotoxicity in EGFR-
mutated NSCLC cells. In addition, cotreatment with chloroquine and the nanocomplexes
exhibited dramatic tumor suppression, which might partly stem from the decreased tumor
vessel leakage, improved tumor vessel maturation of and elevated intratumoral drug
concentration. Very recently, these authors developed a nanotheranostic platform based on
chitosan to encapsulate ERL and a heptamethine cyanine derivative (Cy7) as a near-infrared
photosensitizer for photodynamic therapy (Fig. 3) [69]. These NPs demonstrated high release
of these two cargos in an acidic medium with lysozyme and high uptake into PC-9 cells. With
the combination of near-infrared irradiation and the coloaded NPs, profound generation of
reactive oxygen species, marked upregulation of tumor suppressor p53 and downregulation of
survivin expression were observed in three subtypes of NSCLC cells, especially in PC-9 cells.
Importantly, this synergistic combination significantly inhibited tumor growth in a mouse
model implanted with PC-9 cells, which might result from the specific distribution of the NPs
in the lung.

To overcome the multidrug resistance to DOX and other chemotherapeutics in cancer cells
and to reduce drug side effects, giant liposomes that are coloaded with three drugs (DOX,
ERL and 17-AAG), DNA nanostructures, nanomagnetite and gold nanorods were assembled
with hydrophilic thermally oxidized porous silicon NPs on a microfluidic chip to generate a
nano-in-micro platform [70]. The use of porous silicon NPs allowed high drug loading and
sustained release of the hydrophobic drug ERL as well as control over the drug ratio in the
liposomes, whereas the nanomagnetite and gold nanorods rendered the platform magnetically
and photothermally responsive, which might facilitate targeted drug delivery at tumor sites. In
particular, the combination of DNA nanostructures and the three drugs achieved synergistic
effects on killing DOX-resistant MCF-7 breast cancer cells with a significantly lower
percentage of cell viability than that achieved with other combinations.

Considering the possibility that resistance to EGFR inhibition can be induced by the
activation of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3
(STAT3) signaling [71, 72] or Notch signaling [73, 74], the combination of ERL and JAK2
inhibitors or Notch inhibitors might be a rational approach to overcome this resistance. In one
study, Chen et al. loaded both ERL and a JAK2 inhibitor—fedratinib—into poly(L-lactide)-b-
13
polyethylene glycol (PEG-b-PLA) NPs [75]. Compared to single drug-loaded NPs, the dual
drug-loaded NPs exerted significantly higher cytotoxic effects on two ERL-resistant lung
adenocarcinoma cell lines, H1975 and H1650, as well as stronger tumor growth suppression
ability in H1650 tumor-bearing mice. In another study, Wan and colleagues coencapsulated
ERL and a gamma secretase inhibitor—DAPT—into poly(L-lactide) (PLA)-based NPs [76].
To enhance their tumor targeting and penetrating abilities, the prepared NPs were
functionalized with a conjugate of a tumor-homing peptide and a cell-penetrating peptide.
Due to the presence of an acid-sensitive hydrazone linkage between these two peptides, the
modified NPs showed the highest cellular uptake at an acidic pH of 6.0 and the highest
cytotoxicity in MDA-MB-231 triple-negative breast cancer cells. Notably, among all mice
groups tested, tumor-bearing mice injected with these modified NPs exhibited the lowest
tumor size without weight loss resulting from the highest tumor accumulation and lowest
distribution in other organs.
Long-term pretreatment with the EGFR inhibitor ERL was found to significantly enhance or
sensitize the efficacy of the DNA-damaging agent doxorubicin (DOX) in NSCLC cells and
triple-negative breast cancer cells [77] and in MCF-7 and MDA-MB-468 human breast cancer
cells [29]. Similar phenomena were also observed with other combinations of drug(s) and/or
gene therapy in other cancer models [78-80], suggesting that the efficacy of combination
therapies does not depend simply on their compositions but rather critically on the sequence
of their component’s delivery. Bearing that in mind, to deliver both ERL and DOX to the
same tumor cells, Morton et al. encapsulated them in the exterior hydrophobic layer and
interior hydrophilic compartment of folate-coated liposomes, respectively [81]. The folate
coating enhanced the uptake of the liposomes and DOX into folate receptor-overexpressing
A549 NSCLC and BT-20 triple-negative breast cancer cell lines. The core-shell design
facilitated the release of ERL first and DOX later from the dual drug-loaded liposomes, which
demonstrated a significant increase in apoptosis and prolonged inhibition of EGFR in these
two cell lines compared to those induced by DOX-loaded liposomes. Notably, although the
dual drug combination reduced the tumor burden in A549 and BT-20 xenograft-bearing mice,
the two drugs must be packed in the same vehicle to achieve the desired efficacy. In another
study, He et al. codelivered a combination of ERL and DOX by using a pH-sensitive charge
conversion nanocarrier (designated as M-HHG2C18-L) [82]. In this system, DOX was loaded
into the core of amino-functionalized mesoporous silica NPs (MSN-NH2), which were then
shielded by an ERL-incorporated lipid bilayer to form M-HHG2C18-L(E+D) (Fig. 4). Under

14
the low pH in the tumor extracellular and intracellular environments, the surface potential of
the dual drug-loaded NPs shifted from negative to positive due to the protonation of the
zwitterionic oligopeptide lipid in the external layer, thereby enhancing the internalization of
the NPs into cancer cells and improving the release of both drugs. Notably, the sequential
release of the two drugs remarkably synergized their antiproliferative and apoptotic effects on
A549 cells and dramatically delayed tumor growth in tumor-bearing mice with an excellent
safety profile. Recently, by using a different one-compartment carrier design, Zhou et al.
coloaded ERL and the hydrophobic form of DOX into poly(L-lactide)-b-polyethylene glycol
(PLA-b-PEG) NPs after ion-pairing of positively charged DOX with a negatively charged
lipid, 1,2-dioleoyl-sn-glycero-3-phosphate [83]. This complexation significantly retarded the
release rate of DOX while exhibiting burst release of ERL (approximately 80% in 4 hours)
that increased the efficacy of the coloaded NPs against MDA-MB-468 breast cancer cells.
Furthermore, an in vivo biodistribution study demonstrated that the polymeric NPs
considerably accumulated inside the tumor with marginal distribution in the major organs,
confirming their efficacy and safety. To control the time lag of drug administration even more
precisely for optimal efficacy, two separate gold nanocages with pH-sensitive poly(acrylic
acid) and thermosensitive poly(N-isopropylacrylamide-co-acrylamide) shells were developed
to carry ERL and DOX, respectively [84]. Upon accumulation in the tumor microenvironment
via the enhanced permeability and retention effect, the poly(acrylic acid)-covered gold
nanocages loosened their shell because of the acidic pH and released ERL. Then, after 6
hours, under exogenous NIR irradiation, the gold nanocages generated heat to make the
poly(N-isopropylacrylamide-co-acrylamide) shell collapse and release DOX. The
combination of chemotherapy and phototherapy demonstrated a synergistic tumor-killing
effect with more potent efficacy in highly EGFR-expressing A431 cells than in weakly
EGFR-expressing MCF-7 cells in both in vitro and in vivo models attributed to the caspase-8
signaling pathway.

4. Current challenges

Despite some initial clinical success, acquired (secondary) resistance to erlotinib usually
develops in most NSCLC cases after 10-13 months [85]. The resistance can stem mainly from
the acquisition of a secondary EGFR mutation and amplification of the hepatocyte growth
factor receptor (HGFR/c-MET) oncogene (Fig. 5) as well as various conditions in the tumor
microenvironment. First, the gatekeeper T790 mutation is the most common type of mutation,
15
occurring in approximately 50% of all resistant NSCLC cases, whereas amplification of c-
MET accounts for approximately 20% of mutations [3, 86, 87]. Several second-generation
EGFR tyrosine kinase inhibitors (e.g., afatinib and dacomitinib) and third-generation
inhibitors (e.g., osimertinib) that bind irreversibly to EGFR and selectively to T790M EGFR,
respectively, and c-MET inhibitors (e.g., crizotinib) have been developed to overcome this
type of resistance. Second, hypoxia, which refers to the low oxygen levels in tumor tissues,
and disturbed tumor tissue vasculature are two major factors in the tumor microenvironment
that substantially limit drug penetration and antitumor activity in tumor tissues [46, 88, 89].
These factors can be bypassed by delivering oxygen directly into tumor sites and by using
vascular normalization agents to restore the functions of tumor vessels, respectively [67, 68].
In addition, the upregulation and activation of some proteins, such as survivin, STAT3 and
phosphoglycerate dehydrogenase (PHGDH), in the tumor microenvironment were found to be
positively associated with drug resistance and poor prognosis in lung cancer [90-92]. This can
be addressed by silencing these genes using specific short interfering RNAs or gene
suppressors [68, 90, 92, 93].

5. Authors’ outlook

In this review, we have discussed several approaches for delivering erlotinib by using
numerous carrier systems to enhance its efficacy. First, polymer-based, lipid-based, inorganic
and hybrid systems were used to improve erlotinib’s characteristics, such as its aqueous
solubility, dissolution, bioavailability and toxicity. Second, NPs delivering combinations of
erlotinib and other therapeutic agents or modalities were exploited to prevent the development
of tumor resistance and improve the effects of erlotinib. The advantages and disadvantages of
delivery systems of erlotinib are summarized in Table 3.

These promising results encourage the translation of these nanoparticulate systems into the
clinic. Nevertheless, there are some challenges that delay their appearance on the market.
First, the large-scale manufacturing of NPs according to GMP guidelines is a laborious and
expensive process because of the high costs of materials, low production yield, incomplete
purification of the final products, etc. Second, in the preclinical phase, there is a need for
developing standardized protocols to evaluate the toxicity of NPs in vitro, in vivo and in
animal models and to understand their interaction with cells/tissues/organs, etc. Third, due to
the complex nature of NPs, there is a lack of appropriate government regulations for
nanoparticulate systems that would help guide the manufacturing practices, quality control
and design of optimal clinical trials to evaluate their safety/toxicity and therapeutic efficacy in
16
humans [94]. Moreover, patents and intellectual properties can be serious problems that halt
the commercialization of nanomaterials because their nomenclature is much more
complicated than that of conventional products on the market. Therefore, by addressing all the
abovementioned issues, pharmaceutical companies might successfully develop an oral
formulation of erlotinib with improved pharmacokinetics and toxicity profiles for enhanced
clinical efficacy and patient compliance.

Conflict of interest

The authors declare no conflict of interest.

Acknowledgements

This research received no specific grant from any funding agency in the public, commercial,
or not-for-profit sectors.

References

[1] U.S. Food and Drug Administration, Ful prescribing information of Erlotinib (Tarceva).
http://www.accessdata.fda.gov/drugsatfda_docs/label/2016/021743s025lbl.pdf. Accessed on
September 25, 2018.

[2] J.D. Moyer, E.G. Barbacci, K.K. Iwata, L. Arnold, B. Boman, A. Cunningham, C. DiOrio,
J. Doty, M.J. Morin, M.P. Moyer, Induction of apoptosis and cell cycle arrest by CP-358,774,
an inhibitor of epidermal growth factor receptor tyrosine kinase, Cancer Res. 57(21) (1997)
4838-4848.

[3] V.D. Cataldo, D.L. Gibbons, R. Pérez-Soler, A. Quintás-Cardama, Treatment of non–

small-cell lung cancer with erlotinib or gefitinib, New England J. Med. 364(10) (2011) 947-
955.

[4] H. Hirte, A. Oza, K. Swenerton, S. Ellard, R. Grimshaw, B. Fisher, M. Tsao, L. Seymour,

A phase II study of erlotinib (OSI-774) given in combination with carboplatin in patients with
recurrent epithelial ovarian cancer (NCIC CTG IND. 149), Gynecol. Oncol. 118(3) (2010)
308-312.

[5] E. Despierre, I. Vergote, R. Anderson, C. Coens, D. Katsaros, F.R. Hirsch, B. Boeckx, M.

Varella-Garcia, A. Ferrero, I. Ray-Coquard, Epidermal growth factor receptor (EGFR)
17
pathway biomarkers in the randomized phase III trial of erlotinib versus observation in
ovarian cancer patients with no evidence of disease progression after first-line platinum-based
chemotherapy, Targeted Oncol. 10(4) (2015) 583-596.

[6] A.S.M. Anisuzzaman, A. Haque, D. Wang, M.A. Rahman, C. Zhang, Z. Chen, Z.G. Chen,
D.M. Shin, A.R. Amin, In vitro and in vivo synergistic antitumor activity of the combination
of BKM120 and erlotinib in head and neck cancer: Mechanism of apoptosis and resistance,
Mol. Cancer Therapeut. 16(4) (2017) 729-738.

[7] J. Raizer, P. Giglio, J. Hu, M. Groves, R. Merrell, C. Conrad, S. Phuphanich, V.K.

Puduvalli, M. Loghin, N. Paleologos, A phase II study of bevacizumab and erlotinib after
radiation and temozolomide in MGMT unmethylated GBM patients, J. Neuro-oncol. 126(1)
(2016) 185-192.

[8] J.J. Raizer, L.E. Abrey, A.B. Lassman, S.M. Chang, K.R. Lamborn, J.G. Kuhn, W.A.
Yung, M.R. Gilbert, K.A. Aldape, P.Y. Wen, A phase II trial of erlotinib in patients with
recurrent malignant gliomas and nonprogressive glioblastoma multiforme postradiation
therapy, Neuro-oncol. 12(1) (2009) 95-103.

[9] N. Budha, A. Frymoyer, G. Smelick, J. Jin, M. Yago, M. Dresser, S. Holden, L. Benet, J.

Ware, Drug absorption interactions between oral targeted anticancer agents and PPIs: is
pH‐dependent solubility the Achilles heel of targeted therapy?, Clin. Pharmacol. Ther. 92(2)
(2012) 203-213.

[10] F. Thomas, P. Rochaix, M. White-Koning, I. Hennebelle, J. Sarini, A. Benlyazid, L.

Malard, J.-L. Lefebvre, E. Chatelut, J.P. Delord, Population pharmacokinetics of erlotinib and
its pharmacokinetic/pharmacodynamic relationships in head and neck squamous cell
carcinoma, Eur. J. Cancer 45(13) (2009) 2316-2323.

[11] A. Gruber, M. Czejka, P. Buchner, M. Kitzmueller, N.K. Baroian, C. Dittrich, A.S.

Hrgovcic, Monitoring of erlotinib in pancreatic cancer patients during long-time
administration and comparison to a physiologically based pharmacokinetic model, Cancer
Chemother. Pharmacol. 81(4) (2018) 763-771.

[12] J.R. Johnson, M. Cohen, R. Sridhara, Y.-F. Chen, G.M. Williams, J. Duan, J. Gobburu,
B. Booth, K. Benson, J. Leighton, Approval summary for erlotinib for treatment of patients
with locally advanced or metastatic non–small cell lung cancer after failure of at least one
prior chemotherapy regimen, Clin. Cancer Res. 11(18) (2005) 6414-6421.

18
[13] M. D’Arcangelo, F. Cappuzzo, Erlotinib in the first-line treatment of non-small-cell lung
cancer, Exp. Rev. Anticancer Ther. 13(5) (2013) 523-533.

[14] S. Kumar, N. Dilbaghi, R. Saharan, G. Bhanjana, Nanotechnology as emerging tool for

enhancing solubility of poorly water-soluble drugs, BioNanoSci. 2(4) (2012) 227-250.

[15] P. Chi Lip Kwok, H.-K. Chan, Nanotechnology versus other techniques in improving
drug dissolution, Cur. Pharm. Design 20(3) (2014) 474-482.

[16] M. Sharma, R. Sharma, D.K. Jain, Nanotechnology based approaches for enhancing oral
bioavailability of poorly water soluble antihypertensive drugs, Scientifica 2016 (2016).

[17] J. Hu, K.P. Johnston, R.O. Williams III, Nanoparticle engineering processes for
enhancing the dissolution rates of poorly water soluble drugs, Drug Dev. Ind. Pharm. 30(3)
(2004) 233-245.

[18] H. Chen, C. Khemtong, X. Yang, X. Chang, J. Gao, Nanonization strategies for poorly
water-soluble drugs, Drug Dis. Today 16(7-8) (2011) 354-360.

[19] C.M. Keck, R.H. Müller, Drug nanocrystals of poorly soluble drugs produced by high
pressure homogenisation, Eur. J. Pharm. Biopharm. 62(1) (2006) 3-16.

[20] H. Bunjes, Lipid nanoparticles for the delivery of poorly water‐soluble drugs, J. Pharm.
Pharm. 62(11) (2010) 1637-1645.

[21] O. Pillai, R. Panchagnula, Polymers in drug delivery, Cur. Opin. Chem. Biol. 5(4) (2001)
447-451.

[22] H.K. Makadia, S.J. Siegel, Poly lactic-co-glycolic acid (PLGA) as biodegradable
controlled drug delivery carrier, Polymers 3(3) (2011) 1377-1397.

[23] D. Mondal, M. Griffith, S.S. Venkatraman, Polycaprolactone-based biomaterials for

tissue engineering and drug delivery: Current scenario and challenges, Int. J. Polym. Mat.
Polym. Biomat. 65(5) (2016) 255-265.

[24] G. Marslin, C.J. Sheeba, V. Kalaichelvan, R. Manavalan, P. Neelakanta Reddy, G.

Franklin, Poly (D, L-lactic-co-glycolic acid) nanoencapsulation reduces Erlotinib-induced
subacute toxicity in rat, J. Biomed. Nanotechnol. 5(5) (2009) 464-471.

[25] L. Barghi, D. Asgari, J. Barar, A. Nakhlband, H. Valizadeh, Synthesis, characterization

and in vitro anti-tumoral evaluation of Erlotinib-PCEC nanoparticles, Asian Pac. J. Cancer
Prev. 15(23) (2014) 10281-7.
19
[26] W. Saenger, Cyclodextrin inclusion compounds in research and industry, Angewandte
Chem. Int. Ed. English 19(5) (1980) 344-362.

[27] J. Szejtli, Cyclodextrin inclusion complexes, Cyclodextrin technology, Springer1988, pp.

79-185.

[28] M.E. Davis, M.E. Brewster, Cyclodextrin-based pharmaceutics: past, present and future,
Nat. Rev. Drug Dis. 3(12) (2004) 1023-1035.

[29] N. Devasari, C.P. Dora, C. Singh, S.R. Paidi, V. Kumar, M.E. Sobhia, S. Suresh,
Inclusion complex of erlotinib with sulfobutyl ether-β-cyclodextrin: Preparation,
characterization, in silico, in vitro and in vivo evaluation, Carbohyd. Polym. 134 (2015) 547-
556.

[30] C.P. Dora, F. Trotta, V. Kushwah, N. Devasari, C. Singh, S. Suresh, S. Jain, Potential of
erlotinib cyclodextrin nanosponge complex to enhance solubility, dissolution rate, in vitro
cytotoxicity and oral bioavailability, Carbohyd. Polym. 137 (2016) 339-349.

[31] M.M. Momin, Z. Zaheer, R. Zainuddin, J.N. Sangshetti, Extended release delivery of
erlotinib glutathione nanosponge for targeting lung cancer, Artif. Cells Nanomed. Biotechnol.
46(5) (2018) 1064-1075.

[32] F.-F. An, X.-H. Zhang, Strategies for preparing albumin-based nanoparticles for
multifunctional bioimaging and drug delivery, Theranostics 7(15) (2017) 3667-3689.

[33] M. Noorani, N. Azarpira, K. Karimian, H. Heli, Erlotinib-loaded albumin nanoparticles:

A novel injectable form of erlotinib and its in vivo efficacy against pancreatic
adenocarcinoma ASPC-1 and PANC-1 cell lines, Int. J. Pharm. 531(1) (2017) 299-305.

[34] Y. Shen, W. Li, HA/HSA co-modified erlotinib–albumin nanoparticles for lung cancer
treatment, Drug Design Dev. Ther. 12 (2018) 2285-2292.

[35] K.M. Yang, I.C. Shin, J.W. Park, K.-S. Kim, D.K. Kim, K. Park, K. Kim,
Nanoparticulation improves bioavailability of Erlotinib, Drug Dev. Ind. Pharm. 43(9) (2017)
1557-1565.

[36] D.H. Truong, T.H. Tran, T. Ramasamy, J.Y. Choi, H.H. Lee, C. Moon, H.-G. Choi, C.S.
Yong, J.O. Kim, Development of solid self-emulsifying formulation for improving the oral
bioavailability of erlotinib, AAPS PharmSciTech 17(2) (2016) 466-473.

20
[37] X. Zhou, H. Tao, K.-H. Shi, Development of a nanoliposomal formulation of erlotinib
for lung cancer and in vitro/in vivo antitumoral evaluation, Drug Design Dev. Ther. 12 (2018)
1-8.

[38] Z. Bakhtiary, J. Barar, A. Aghanejad, A.A. Saei, E. Nemati, J. Ezzati Nazhad Dolatabadi,
Y. Omidi, Microparticles containing erlotinib-loaded solid lipid nanoparticles for treatment of
non-small cell lung cancer, Drug Dev. Ind. Pharm. 43(8) (2017) 1244-1253.

[39] N. Naseri, P. Zakeri-Milani, H. Hamishehkar, Y. Pilehvar-Soltanahmadi, H. Valizadeh,

Development, in vitro characterization, antitumor and aerosol performance evaluation of
respirable prepared by self-nanoemulsification method, Drug Res. 67 (2017) 343-348.

[40] C.P. Dora, V. Kushwah, S.S. Katiyar, P. Kumar, V. Pillay, S. Suresh, S. Jain, Improved
oral bioavailability and therapeutic efficacy of erlotinib through molecular complexation with
phospholipid, Int. J. Pharm. 534(1-2) (2017) 1-13.

[41] P. Martinkova, M. Brtnicky, J. Kynicky, M. Pohanka, Iron Oxide Nanoparticles:

Innovative Tool in Cancer Diagnosis and Therapy, Adv. Healthcare Mat. 7(5) (2018)
1700932.

[42] S. Fathi Karkan, M. Mohammadhosseini, Y. Panahi, M. Milani, N. Zarghami, A.

Akbarzadeh, E. Abasi, A. Hosseini, S. Davaran, Magnetic nanoparticles in cancer diagnosis
and treatment: a review, Artif. cell. Nanomed. Biotechnol. 45(1) (2017) 1-5.

[43] M. Arruebo, R. Fernández-Pacheco, M.R. Ibarra, J. Santamaría, Magnetic nanoparticles

for drug delivery, Nano today 2(3) (2007) 22-32.

[44] Y. Xu, A. Karmakar, W.E. Heberlein, T. Mustafa, A.R. Biris, A.S. Biris, Multifunctional
magnetic nanoparticles for synergistic enhancement of cancer treatment by combinatorial
radio frequency thermolysis and drug delivery, Adv. Healthcare Mat. 1(4) (2012) 493-501.

[45] S. Luo, X. Yang, C. Shi, Newly Emerging Theranostic Agents for Simultaneous
Cancertargeted Imaging and Therapy, Cur. Med. Chem. 23(5) (2016) 483-497.

[46] A.A.A. Ali, F.-T. Hsu, C.-L. Hsieh, C.-Y. Shiau, C.-H. Chiang, Z.-H. Wei, C.-Y. Chen,
H.-S. Huang, Erlotinib-conjugated iron oxide nanoparticles as a smart cancer-targeted
theranostic probe for MRI, Sci. Rep. 6 (2016) 36650.

[47] F.-T. Hsu, H.-S. Liu, A.A.A. Ali, P.-H. Tsai, Y.-C. Kao, C.-F. Lu, H.-S. Huang, C.-Y.
Chen, Assessing the selective therapeutic efficacy of superparamagnetic erlotinib
nanoparticles in lung cancer by using quantitative magnetic resonance imaging and a nuclear
21
factor kappa-B reporter gene system, Nanomedicine: Nanotechnol. Biol. Med. 14(3) (2018)
1019-1031.

[48] J. Nebu, J.A. Devi, R. Aparna, K. Abha, G. Sony, Erlotinib conjugated gold nanocluster
enveloped magnetic iron oxide nanoparticles–A targeted probe for imaging pancreatic cancer
cells, Sensor. Actuators B: Chem. 257 (2018) 1035-1043.

[49] B. Mandal, N.K. Mittal, P. Balabathula, L.A. Thoma, G.C. Wood, Development and in
vitro evaluation of core–shell type lipid–polymer hybrid nanoparticles for the delivery of
erlotinib in non-small cell lung cancer, Eur. J. Pharm. Sci. 81 (2016) 162-171.

[50] F. Li, H. Mei, X. Xie, H. Zhang, J. Liu, T. Lv, H. Nie, Y. Gao, L. Jia, Aptamer-
conjugated chitosan-anchored liposomal complexes for targeted delivery of erlotinib to
EGFR-mutated lung cancer cells, AAPS J. 19(3) (2017) 814-826.

[51] R. Mo, Z. Gu, Tumor microenvironment and intracellular signal-activated nanomaterials

for anticancer drug delivery, Mat. Today 19(5) (2016) 274-283.

[52] S. Uthaman, K.M. Huh, I.-K. Park, Tumor microenvironment-responsive nanoparticles

for cancer theragnostic applications, Biomat. Res. 22 (2018) 22.

[53] H.B. Ruttala, T. Ramasamy, T. Madeshwaran, T.T. Hiep, U. Kandasamy, K.T. Oh, H.-G.
Choi, C.S. Yong, J.O. Kim, Emerging potential of stimulus-responsive nanosized anticancer
drug delivery systems for systemic applications, Arch. Pharm. Res. 41(2) (2018) 111-129.

[54] H.B. Ruttala, N. Chitrapriya, K. Kaliraj, T. Ramasamy, W.H. Shin, J.-H. Jeong, J.R.
Kim, S.K. Ku, H.-G. Choi, C.S. Yong, Facile construction of bioreducible crosslinked
polypeptide micelles for enhanced cancer combination therapy, Acta Biomat. 63 (2017) 135-
149.

[55] T. Ramasamy, H.B. Ruttala, N. Chitrapriya, B.K. Poudal, J.Y. Choi, S.T. Kim, Y.S.
Youn, S.K. Ku, H.-G. Choi, C.S. Yong, Engineering of cell microenvironment-responsive
polypeptide nanovehicle co-encapsulating a synergistic combination of small molecules for
effective chemotherapy in solid tumors, Acta Biomat. 48 (2017) 131-143.

[56] T. Ramasamy, H.B. Ruttala, B. Gupta, B.K. Poudel, H.-G. Choi, C.S. Yong, J.O. Kim,
Smart chemistry-based nanosized drug delivery systems for systemic applications: a
comprehensive review, J. Control. Release 258 (2017) 226-253.

[57] J. Kim, T. Ramasamy, J.Y. Choi, S.T. Kim, Y.S. Youn, H.-G. Choi, C.S. Yong, J.O.
Kim, PEGylated polypeptide lipid nanocapsules to enhance the anticancer efficacy of
22
erlotinib in non-small cell lung cancer, Colloid. Surfaces B: Biointerfaces 150 (2017) 393-
401.

[58] M. Fathi, P.S. Zangabad, J. Barar, A. Aghanejad, H. Erfan-Niya, Y. Omidi, Thermo-

sensitive chitosan copolymer-gold hybrid nanoparticles as a nanocarrier for delivery of
erlotinib, Int. J. Biol. Macromol. 106 (2018) 266-276.

[59] M. Fathi, P.S. Zangabad, A. Aghanejad, J. Barar, H. Erfan-Niya, Y. Omidi, Folate-

conjugated thermosensitive O-maleoyl modified chitosan micellar nanoparticles for targeted
delivery of erlotinib, Carbohyd. Polymers 172 (2017) 130-141.

[60] S. Tan, G. Wang, Redox-responsive and pH-sensitive nanoparticles enhanced stability

and anticancer ability of erlotinib to treat lung cancer in vivo, Drug Design, Devel. Ther. 11
(2017) 3519-3529.

[61] N. Avedian, F. Zaaeri, M.P. Daryasari, H.A. Javar, M. Khoobi, pH-sensitive

biocompatible mesoporous magnetic nanoparticles labeled with folic acid as an efficient
carrier for controlled anticancer drug delivery, J. Drug Deliv. Sci. Technol. 44 (2018) 323-
332.

[62] M. Deshmukh, S. Nakagawa, T. Higashi, A. Vincek, A. Venkatesh, M.R. De Galarreta,

A.P. Koh, N. Goossens, H. Hirschfield, C.B. Bian, Cell type-specific pharmacological kinase
inhibition for cancer chemoprevention, Nanomed. Nanotechnol. Biol. Med. 14(2) (2018) 317-
325.

[63] J.A. DiMasi, R.W. Hansen, H.G. Grabowski, The price of innovation: new estimates of
drug development costs, J. Health Eco. 22(2) (2003) 151-185.

[64] R.B. Mokhtari, T.S. Homayouni, N. Baluch, E. Morgatskaya, S. Kumar, B. Das, H.

Yeger, Combination therapy in combating cancer, Oncotarget 8(23) (2017) 38022-38043.

[65] B. Gupta, B.K. Poudel, S. Regmi, S. Pathak, H.B. Ruttala, M. Gautam, G.J. An, J.-H.
Jeong, H.-G. Choi, C.S. Yong, Paclitaxel and Erlotinib-co-loaded Solid Lipid Core
Nanocapsules: Assessment of Physicochemical Characteristics and Cytotoxicity in Non-small
Cell Lung Cancer, Pharm. Res. 35(5) (2018) 96.

[66] K. Patel, R. Doddapaneni, M. Patki, V. Sekar, A. Bagde, M. Singh, Erlotinib-Valproic

Acid Liquisolid Formulation: Evaluating Oral Bioavailability and Cytotoxicity in Erlotinib-
Resistant Non-small Cell Lung Cancer Cells, AAPS PharmSciTech 20 (2019) 135.

23
[67] F. Li, H. Mei, Y. Gao, X. Xie, H. Nie, T. Li, H. Zhang, L. Jia, Co-delivery of oxygen and
erlotinib by aptamer-modified liposomal complexes to reverse hypoxia-induced drug
resistance in lung cancer, Biomat. 145 (2017) 56-71.

[68] T. Lv, Z. Li, L. Xu, Y. Zhang, H. Chen, Y. Gao, Chloroquine in combination with
aptamer-modified nanocomplexes for tumor vessel normalization and efficient
erlotinib/Survivin shRNA co-delivery to overcome drug resistance in EGFR-mutated non-
small cell lung cancer, Acta Biomat. 76 (2018) 257-274.

[69] Y. Gao, H. Zhang, Y. Zhang, T. Lv, L. Zhang, Z. Li, X. Xie, F. Li, H. Chen, L. Jia,
Erlotinib-guided self-assembled trifunctional click nanotheranostics for distinguishing
druggable mutations and synergistic therapy of NSCLC, Mol. Pharm. 15(11) (2018) 5146-
5161.

[70] F. Kong, X. Zhang, H. Zhang, X. Qu, D. Chen, M. Servos, E. Mäkilä, J. Salonen, H.A.
Santos, M. Hai, Inhibition of Multidrug Resistance of Cancer Cells by Co‐Delivery of DNA
Nanostructures and Drugs Using Porous Silicon Nanoparticles@ Giant Liposomes, Adv.
Funct. Mat. 25(22) (2015) 3330-3340.

[71] D. Harada, N. Takigawa, N. Ochi, T. Ninomiya, M. Yasugi, T. Kubo, H. Takeda, E.

Ichihara, K. Ohashi, S. Takata, JAK 2‐related pathway induces acquired erlotinib resistance in
lung cancer cells harboring an epidermal growth factor receptor‐activating mutation, Cancer
Sci. 103(10) (2012) 1795-1802.

[72] B.D. Looyenga, D. Hutchings, I. Cherni, C. Kingsley, G.J. Weiss, J.P. MacKeigan,
STAT3 is activated by JAK2 independent of key oncogenic driver mutations in non-small cell
lung carcinoma, PloS one 7(2) (2012) e30820.

[73] R.R. Arasada, J.M. Amann, M.A. Rahman, S.S. Huppert, D.P. Carbone, EGFR blockade
enriches for lung cancer stem–like cells through Notch3-dependent signaling, Cancer Res.
74(19) (2014) 5572-5584.

[74] A.T. Baker, A. Zlobin, C. Osipo, Notch-EGFR/HER2 bidirectional crosstalk in breast

cancer, Front. Oncol. 4 (2014) 360.

[75] D. Chen, F. Zhang, J. Wang, H. He, S. Duan, R. Zhu, C. Chen, Y. Chen, L. Yin,
Biodegradable Nanoparticles Mediated Co-Delivery of Erlotinib (ELTN) and Fedratinib
(FDTN) toward the Treatment of ELTN-Resistant Non-Small Cell Lung Cancer (NSCLC) via
Suppression of the JAK2/STAT3 Signaling Pathway, Front. Pharm. 9 (2018) 1214.
24
[76] X. Wan, C. Liu, Y. Lin, J. Fu, G. Lu, Z. Lu, pH sensitive peptide functionalized
nanoparticles for co-delivery of erlotinib and DAPT to restrict the progress of triple negative
breast cancer, Drug Delivery 26(1) (2019) 470-480.

[77] M.J. Lee, S.Y. Albert, A.K. Gardino, A.M. Heijink, P.K. Sorger, G. MacBeath, M.B.
Yaffe, Sequential application of anticancer drugs enhances cell death by rewiring apoptotic
signaling networks, Cell 149(4) (2012) 780-794.

[78] L. Zhang, Z. Lu, Q. Zhao, J. Huang, H. Shen, Z. Zhang, Enhanced chemotherapy

efficacy by sequential delivery of siRNA and anticancer drugs using PEI‐grafted graphene
oxide, Small 7(4) (2011) 460-464.

[79] X. Qian, Y. Ren, Z. Shi, L. Long, P. Pu, J. Sheng, X. Yuan, C. Kang, Sequence-
dependent synergistic inhibition of human glioma cell lines by combined temozolomide and
miR-21 inhibitor gene therapy, Mol. Pharm. 9(9) (2012) 2636-2645.

[80] D.B. Pacardo, F.S. Ligler, Z. Gu, Programmable nanomedicine: synergistic and
sequential drug delivery systems, Nanoscale 7(8) (2015) 3381-3391.

[81] S.W. Morton, M.J. Lee, Z.J. Deng, E.C. Dreaden, E. Siouve, K.E. Shopsowitz, N.J. Shah,
M.B. Yaffe, P.T. Hammond, A nanoparticle-based combination chemotherapy delivery
system for enhanced tumor killing by dynamic rewiring of signaling pathways, Sci. Signal.
7(325) (2014) ra44.

[82] Y. He, Z. Su, L. Xue, H. Xu, C. Zhang, Co-delivery of erlotinib and doxorubicin by pH-
sensitive charge conversion nanocarrier for synergistic therapy, J. Control. Release 229
(2016) 80-92.

[83] Z. Zhou, C. Kennell, M. Jafari, J.-Y. Lee, S.J. Ruiz-Torres, S.E. Waltz, J.-H. Lee,
Sequential delivery of erlotinib and doxorubicin for enhanced triple negative Breast cancer
treatment using polymeric nanoparticle, Int. J. Pharm. 530(1-2) (2017) 300-307.

[84] Y. Feng, Y. Cheng, Y. Chang, H. Jian, R. Zheng, X. Wu, K. Xu, L. Wang, X. Ma, X. Li,
Time-staggered delivery of erlotinib and doxorubicin by gold nanocages with two smart
polymers for reprogrammable release and synergistic with photothermal therapy, Biomat. 217
(2019) 119327.

[85] M. Zhang, B. Sai, P. Cao, Z. Li, L. Zhang, C. Shuai, X. Wang, J. Wang, G. Li, J. Xiang,
Iron Oxide Nanoparticles Synergize with Erlotinib to Suppress Refractory Non-Small Cell

25
Lung Cancer Cell Proliferation Through the Inhibition of ErbB/PI3K/AKT and PTEN
Activation, J. Biomed. Nanotechnol. 13(4) (2017) 458-468.

[86] A. Ahsan, Mechanisms of resistance to EGFR tyrosine kinase inhibitors and therapeutic
approaches: an update, Lung Cancer Personal. Med., Springer2016, pp. 137-153.

[87] R.M. Jotte, D.R. Spigel, Advances in molecular‐based personalized non‐small‐cell lung
cancer therapy: targeting epidermal growth factor receptor and mechanisms of resistance,
Cancer Med. 4(11) (2015) 1621-1632.

[88] E.C. Finger, A.J. Giaccia, Hypoxia, inflammation, and the tumor microenvironment in
metastatic disease, Cancer Metast. Rev. 29(2) (2010) 285-293.

[89] O. Trédan, C.M. Galmarini, K. Patel, I.F. Tannock, Drug resistance and the solid tumor
microenvironment, J. Nat. Cancer Insti. 99(19) (2007) 1441-1454.

[90] K. Okamoto, I. Okamoto, E. Hatashita, K. Kuwata, H. Yamaguchi, A. Kita, K.

Yamanaka, M. Ono, K. Nakagawa, Overcoming erlotinib resistance in EGFR mutation–
positive non–small cell lung cancer cells by targeting survivin, Mol. Cancer Ther. 11(1)
(2012) 204-213.

[91] D. Harada, N. Takigawa, K. Kiura, The role of STAT3 in non-small cell lung cancer,
Cancers 6(2) (2014) 708-722.

[92] J.-K. Dong, H.-M. Lei, Q. Liang, Y.-B. Tang, Y. Zhou, Y. Wang, S. Zhang, W.-B. Li, Y.
Tong, G. Zhuang, Overcoming erlotinib resistance in EGFR mutation-positive lung
adenocarcinomas through repression of phosphoglycerate dehydrogenase, Theranostics 8(7)
(2018) 1808-1823.

[93] R. Li, Z. Hu, S.-Y. Sun, Z.G. Chen, T.K. Owonikoko, G.L. Sica, S.S. Ramalingam, W.J.
Curran, F.R. Khuri, X. Deng, Niclosamide overcomes acquired resistance to erlotinib through
suppression of STAT3 in non–small cell lung cancer, Mol. Cancer Ther. 12(10) (2013) 2200-
2212.

[94] S. Tinkle, S.E. McNeil, S. Mühlebach, R. Bawa, G. Borchard, Y.C. Barenholz, L.

Tamarkin, N. Desai, Nanomedicines: addressing the scientific and regulatory gap, An. New
York Acad. Sci. 1313(1) (2014) 35-56.

26
Table 1. Nanoparticles for delivery of erlotinib as a single therapeutic agent.

Type Materials Experiments Results Ref.

1. Polymer-based poly(D,L-lactic-co-glycolic acid) in vitro Decrease of toxicity and damage to internal organs [24]
particles (PLGA)
polycaprolactone - polyethylene in vitro Increase of cytotoxicity in A549 NSCLC cells [25]
glycol - poly caprolactone (PCEC)
sulfobutyl ether β-cyclodextrin in vitro, in vivo Increase of dissolution and bioavailability by 2-fold and 3.6-fold, [29]
respectively
cyclodextrin-based nanosponge in vitro, in vivo Increase of dissolution and bioavailability by 2-fold, and cytotoxic effects [30]
in MIA PaCa-2 and PANC-1 pancreatic cell lines
glutathione-nanosponge in vitro, in vivo Increase of cytotoxicity in A549 NSCLC cells and inhibition of tumor [31]
growth, and decrease distribution in vital organs
albumin-bound nanoparticles in vitro Increase of cytotoxicity in ASPC-1 and PANC-1 pancreatic cancer cells [33]
albumin-bound nanoparticles in vitro, in vivo Decrease of proliferation of A549 NSCLC cells and inhibition of tumor [34]
growth
polyvinylpyrrolidone in vitro, in vivo Increase of dissolution, bioavailability and antitumor effect in A549 [35]
xenograft model, and decrease of fasted/fed variability

2. Lipid-based self-emulsifying drug delivery in vitro, in vivo Increase of dissolution and bioavailability [36]
particles system
liposomes in vitro, in vivo Increase of cytotoxicity in A549 NSCLC cells and bioavailability [37]
solid lipid nanoparticles in vitro Increase of cytotoxicity in A549 NSCLC cells [38]
solid lipid nanoparticles in vitro Increase of cytotoxicity in A549 NSCLC cells [39]
molecular complex in vitro, in vivo Increase of cytotoxicity in MIA PaCa-2 and PANC-1 pancreas [40]
adenocarcinoma cells, solubility and bioavailability

3. Inorganic carbon-coated, iron magnetic in vitro Decrease of proliferation of PANC-1 pancreatic cells, especially under [44]
particles nanoparticles radio frequency
ultra-small superparamagnetic iron in vitro, in vivo Suppression of migration and invasion of CL1-5-F4 lung adenocarcinoma [46]
oxide nanoparticles cells, and inhibition of tumor growth
 ultra-small superparamagnetic iron in vitro, in vivo Dose-dependent cytotoxicity in CL1-5-F4 NSCLC, and inhibition of [47]
oxide nanoparticles tumor growth and NF-κB activity
iron oxide-gold nanoclusters in vitro Increase of uptake and cytotoxicity in PANC-1 pancreatic cells [48]

4. Hybrid polymeric polycaprolactone; in vitro Decrease of cellular viability in A549 NSCLC cells [49]
particles phospholipid
aptamer-conjugated chitosan; in vitro Increase of antiproliferation, induction of cell cycle arrest and apoptosis in [50]
liposome EGFR-mutated H1975 and PC-9 cells
poly(L-aspartic acid); in vitro, in vivo Increase of uptake and cytotoxicity in HCC-827 and NCI-H358 lung [57]
poly(ethylene glycol)-b-poly(L- cancer cells, and inhibition of tumor growth
aspartic acid) (PEG-b-PAsp)
polymer-gold hybrid nanomaterial in vitro Increase of uptake and cytotoxicity in A549 NSCLC cells [58]
thermosensitive micelles in vitro Increase of cytotoxicity in OVCAR-3 ovarian carcinoma cells [59]
poly(acrylic acid)-cystamine-oleic in vitro, in vivo Increase of cytotoxicity in A549 and NCI-H460 NSCLC cells, and [60]
acid (PAA-ss-OA) antitumor efficacy in a lung cancer xenograft model
magnetic mesoporous silica Increase of drug release at acidic pH by 2-fold and cytotoxicity in HeLa [61]
nanoparticles; folic acid- cervical carcinoma cells
poly(ethyleneimine) conjugate
myofibroblast-targeting in vitro, in vivo Sustained drug release over 12 days, and decrease of tumor burden by 7.3- [62]
nanoparticles fold and hepatocyte damage
Table 2. Nanoparticles for delivery of erlotinib and other therapeutic agent/modalities.
Type
1. Hybrid particles
2. Lipid-based particles
Material
solid lipid core nanocapsules;
polymer shell
aptamer-conjugated chitosan;
liposomes
aptamer-conjugated chitosan;
liposomes
liposomes; hydrophilic
thermally oxidized porous
silicon nanoparticles
amino-functionalized
mesoporous silica
nanoparticles; 1,5-
dioctadecyl-L-glutamyl 2-
histidyl-hexahydrobenzoic
acid (HHG2C18)
gold nanocages; poly(acrylic
acid); poly(N-
isopropylacrylamide-co-
acrylamide)
liquisolid formulation
liposomes
Other active Experiments Results Ref.
agents/modalities
paclitaxel in vitro Increase of cytotoxicity in NCI-H23 cells [65]
perfluorooctylbromide in vitro, in vivo Increase of proliferation inhibition and [67]
induction of apoptosis of NSCLC cells, and
significant decrease of tumor growth
survivin-recombinant short in vitro, in vivo Increase of gene transfection efficiency and [68]
hairpin RNA-expressing toxicity in EGFR mutated NSCLC cells, and
plasmid; chloroquine inhibition of tumor growth
doxorubicin; in vitro Synergistic effects on killing DOX-resistant [70]
MCF-7 breast cancer cells
17-AAG; DNA
nanostructures; nano-
magnetite; gold nanorods
doxorubicin in vitro, in vivo Increase of proliferation and apoptosis in A549 [82]
NSCLC cells, and inhibition of tumor growth
doxorubicin in vitro, in vivo Synergistic tumor-killing effect in vitro and in [84]
vivo with stronger efficacy in highly EGFR-
expressing A431 cells than in weakly EGFR-
expressing MCF-7 cells
valproic acid in vitro, in vivo Increase of cytotoxicity in ERL-resistant [66]
HCC827 lung adenocarcinoma cells and
bioavailability
doxorubicin in vitro, in vivo Increase of apoptosis, decrease of EGFR [81]
expression in A549 NSCLC and BT-20 triple-
negative breast cancer cells, and inhibition of
 tumor growth

3. Polymer-based particles chitosan a heptamethine cyanine in vitro, in vivo Increase of reactive oxygen species levels and [69]
derivative tumor suppressor p53, and decrease of survivin
expression in EGFR mutated NSCLC cells and
tumor growth
poly(L-lactide)-b- fedratinib in vitro, in vivo Increase of cytotoxicity in two ERL-resistant [75]
polyethylene glycol lung adenocarcinoma cell lines and tumor
growth suppression in H1650 tumor-bearing
mice
poly(L-lactide) DAPT in vitro, in vivo Increase of cytotoxicity in MDA-MB-231 [76]
triple-negative breast cancer cells, and
inhibition of tumor growth
poly(L-lactide)-b- doxorubicin in vitro, in vivo Increase of cytotoxicity in MDA-MB-468 breast [83]
polyethylene glycol cancer cells and accumulation in tumor sites
Table 3. Advantages and disadvantages associated with different types of ERL-loaded delivery systems.

Delivery systems Advantages Disadvantages

Polymeric NPs Enhance bioavailability Poor drug loading capacity

Enhance toxicity to tumor cells and reduce toxicity to internal organs Use of organic solvent in
formulation

Lipidic NPs

Solid self-emulsifying Enhance bioavailability Poor drug loading capacity

formulation
Improve storage stability

Liposomes Enhance bioavailability Poor storage stability

Ease of functionalization to enhance targeting and stability
Deliver both hydrophilic and hydrophobic agents

Solid lipid nanoparticles (SLNs) Use of biocompatible ingredients Poor storage stability
Ease of scale-up to industrial scale Poor drug loading capacity

Inorganic NPs Versatile applications (e.g. drug delivery, theranostics) Possible metabolism-associated
issues
High cellular uptake
Biocompatibility

Hybrid NPs Deliver multiple therapeutic agents Poor storage stability

Ease of functionalization to enhance targeting and control drug release
Fig 1. (A) Dual functional FeDC-E NPs may serve both therapeutic effect and diagnostic utility in the same time
in NSCLC. Reprinted with permission from [47]. (B) Schematic representation of the formation of
Fe3O4@AuNCs@ERL nanocomposite. Reprinted with permission from [48].
Fig. 2. Myofibroblast-targeting nanoparticles. (A) Schematic presentation of erlotinib-loaded nanoparticles that
target PDGFRB-expressing myofibroblasts (PPB-NP-erlotinib). Erlotinib is released after cellular internalization
and inhibits intracellular kinase domain of EGFR protein. In the 3D structural modeling, PPB fits the
extracellular domain of PDGFRB with close proximity to the site of PDGF ligand binding. (B) Morphology of
the nanoparticles in rat plasma over time by TEM imaging. Scale bar indicates 200 nm. (C) Size of nanoparticles
in rat plasma over time measured on TEM images (median of 59 nanoparticles at each time point). Boxes
represent 75th and 25th percentile, horizontal line is median, and whiskers mark lowest and highest values.
Outliers outside 1.5× of inter-quartile range are shown as open circles. (D) Concentration of erlotinib released
from the nanoparticles over time in vitro in rat plasma. (E) Cumulative amount (%) of erlotinib released from the
nanoparticles over time in vitro in rat plasma. PDGFRB: platelet-derived growth factor receptor-β, PPB:
PDGFRB-binding peptide, NP: nanoparticle, GMBS: N–maleimidobutyryl-oxysuccinimide ester, PEG:
polyethylene glycol, EGFR: epidermal growth factor receptor. Reprinted with permission from [62].
Fig. 3. Schematic illustration of the preparation and the action of chitosan-based self-assembled
nanoparticles CE7Ns. (a) Self-assembly and structural composition of CE7Ns. The CE7Ns were composed of
biodegradable polymer Cs, molecular targeted drug erlotinib, and NIRF dye Cy7 with PDT effects. (b)
Schematic illustration of the self-assembled nanoparticles as a robust nanoplatform for molecular targeted drug-
mediated imaging of druggable mutations and combinational therapy of drug-resistant lung tumor. Reprinted
with permission from [69]. Copyright (2018) American Chemical Society.
Fig. 4. Schematic illustration of preparation of erlotinib/DOX combination co-delivery nanocarriers and
synergistic therapy of erlotinib and DOX. The surface of mesoporous silica nanoparticles (A) were modified
with 3-aminopropyltriethoxysilane (APTES) to form MSN-NH2 (B). MSN-NH2 were then loaded with DOX to
obtain DOX-loaded MSN-NH2 (C), followed by supported with erlotinib-loaded lipid bilayer (D) to yield M-
HHG2C18-L(E+D) (E). After injection of M-HHG2C18-L(E+D), the nanoparticles accumulated at the tumor site
through enhanced permeability and retention effect in tumor blood vessels. M-HHG2C18-L(E+D) were positively
charged at extracellular environment (F) leading to easy internalization by tumor cells. After cell uptake, as
erlotinib was loaded in the exterior lipid bilayer and the controlled release ability of MSN-NH2, erlotinib
released faster than DOX. Additionally, the lipid bilayer was more positive and induced a strong repulsion with
MSN-NH2 in intracellular environment (G), enhancing sequential staggered release of erlotinib and DOX, which
was pretreated and staggered to inhibit kinase domain of EGFR in cell membrane (H) and was targeted to cell
nucleus (I) respectively, thus maximized the synergistic therapy. Effect of extracellular and intracellular pH (pHe
and pHi), respectively on the HHG2C18 (J). Reprinted with permission from [82].
Fig. 5. Mechanisms of acquired resistance to gefitinib/erlotinib in EGFR‐mutated NSCLC. EGFR, epidermal
growth factor receptor; ErbB3, v‐erb‐b2 avian erythroblastic leukemia viral oncogene homolog 3; NSCLC,
non‐small‐cell lung cancer; RTK, receptor tyrosine kinase; MET, met proto‐oncogene; AXL, AXL receptor
tyrosine kinase; mAb, monoclonal antibody; TKI, tyrosine kinase inhibitor; NF‐κB, nuclear factor
kappa‐light‐chain‐enhancer of activated B cells; AKT, v‐akt murine thymoma viral oncogene homolog 1; STAT,
signal transducers and activation of transcription; ERK, extracellular signal‐regulated kinase; BIM, BCL2‐like
11 (apoptosis facilitator). Reprinted with permission from [87] under a Creative Commons Attribution 4.0
International License (CC BY 4.0).
Conflict of interest
The authors declare no conflict of interest.