Drug Discovery Today d Volume xxx, Number xx d xxxx 2021 REVIEWS

Stimuli-responsive hydrogels for

POST-SCREEN
intratumoral drug delivery
Ana C. Marques a,⇑, Paulo J. Costa a, Sérgia Velho b,c, Maria H. Amaral a
a
UCIBIO, REQUIMTE, MEDTECH, Laboratory of Pharmaceutical Technology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto,
Porto, Portugal
b
i3S – Instituto de Investigação e Inovação em Saúde, University of Porto, Porto, Portugal
c
IPATIMUP - Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal

The ability of some hydrogels to exhibit a phase transition or change their structure in response to
stimuli has been extensively explored for drug depot formation and controlled drug release. Taking
advantage of the unique features of the tumor microenvironment (TME) or externally applied triggers,
several injectable stimuli-responsive hydrogels have been described as promising candidates for
intratumoral drug delivery. In this review, we provide a brief overview of the TME and highlight the
advantages of intratumoral administration, followed by a summary of the reported strategies to endow
hydrogels with responsiveness to physical (temperature and light), chemical (pH and redox potential),
or biological (enzyme) stimuli.

Keywords: Cancer; Intratumoral administration; Stimuli-responsive hydrogels

Introduction Tumor microenvironment

The standard approach to chemotherapy has been the intra-
venous administration of cytotoxic agents. Notwithstanding,
chemotherapeutic drugs lack specificity for cancer cells, giving
rise to suboptimal dosing and severe adverse effects. Moreover,
this treatment modality is often restricted by dose-limiting toxi-
city and drug resistance.
Despite ongoing efforts to implement targeted therapies,
available systemic treatment options cannot deliver drugs to
tumors while remaining harmless to the rest of the body. Addi-
tionally, current locoregional therapies might help to achieve a
better sparing of healthy tissues but are associated with carcino-
genesis risk, cancer recurrence, or complications of invasive sur-
gical techniques. From this perspective, further research and
investment in developing local and minimally invasive cancer
therapies are imperative.
The tumor microenvironment (TME) is a complex ecosystem
where malignant cells coexist and interact with a heterogeneous
group of non-malignant cells (stromal cells) and noncellular ele-
ments comprising the extracellular matrix (ECM). Although the
latter provides physical support and mechanical integrity to cells,
ECM-degrading enzymes, such as matrix metalloproteinases
(MMPs), are often overexpressed in tumors. This large family of
endopeptidases digests ECM proteins, thereby disrupting physi-
cal barriers that impede cancer cells to leave the primary tumor
site and invade nearby tissues.1
Solid tumors exhaust their blood supply to meet the energetic
demands of proliferating tumor cells and then experience perma-
nent or transient deprivation of oxygen (hypoxia) and nutrients.
Cycles of hypoxia–reoxygenation raise intracellular levels of reac-
tive oxygen species (ROS), and mild oxidative stress stimulates

⇑ Corresponding author. Marques, A.C. (amarques@ff.up.pt)

1359-6446/Ó 2021 Elsevier Ltd. All rights reserved.

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 POST-SCREEN (GREY)
the expression of antioxidant enzymes and reducing species,
such as reduced glutathione (GSH).2,3 The GSH concentration
in some tumor cells and tissues were found to be as high as 1–
10 mM.4
In response to hypoxia and low nutrition, a metabolic switch
from oxidative phosphorylation to aerobic glycolysis takes place.
Cancer cells increase their glucose uptake and convert it to lac-
tate even in the presence of oxygen (the ‘Warburg effect’). Given
that high glycolytic rates lead to excessive production of lactic
acid, tumor cells upregulate the expression of acid-extruding
transporters, maintaining intracellular pH as neutral or slightly
POST-SCREEN
alkaline. As a result, the extracellular pH of solid tumors is more
acidic (typically between 6.5 and 6.9) than that of normal
tissues.5,6
Besides their importance for tumor progression, one can take
advantage of these tumor traits (hypoxia, oxidative stress, and
acidosis) for developing novel therapeutic strategies based on
stimuli-responsive hydrogels.
Intratumoral administration
Upon intravenous administration, sequential obstacles to long
circulation times affect the percentage of free and encapsulated
drugs having access to the tumor. By contrast, intratumoral (IT)
administration bypasses the bloodstream and enables local deliv-
ery of cytotoxic agents to the tumor site, achieving high concen-
trations at target cells and reducing dose requirements. Likewise,
it counteracts drug uptake by nontarget cells, minimizing sys-
temic toxicity.7 Moreover, this administration route is indepen-
dent of the tumor vasculature and, thus, reaches poorly
perfused tumor regions that systemic administration regularly
misses.
Unlike other drug delivery systems, injectable biodegradable
gels do not require surgical procedures and promote retention
of free drugs and drug-loaded particles at the tumor site.
Biodegradable hydrogels can either be degraded after the drug
is completely released by diffusion or release the loaded drug as
they disintegrate into small fragments that are eliminated from
the body. Besides biodegradability, injectable gels are expected
Physical
Temperature
Light
Electric field
Magnetic field
FIGURE 1
Examples of physical, chemical, and biological stimuli responsible for changes in the volume and structure of hydrogels. Physical stimuli include temperature,
light, electric and magnetic fields. In turn, pH, redox potential, and solvent composition are considered chemical stimuli. Finally, enzymes and glucose are two
examples of biological stimuli. Temperature, light, pH, reduction, oxidation, and enzymes have been explored for the development of stimuli-responsive
hydrogels for IT drug administration.
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Drug Discovery Today d Volume xx, Number xx d xxx 2021
to be biocompatible and their biodegradation products to cause
an appropriate host response without cytotoxicity, carcinogene-
sis, and/or mutagenesis. Therefore, biocompatibility is typically
assessed in vitro by cell viability assays [e.g., methylthiazolyl
tetrazolium (MTT) assay] and ex vivo by histological analysis of
tumor sections and major organs after Hematoxylin and Eosin
(H&E) staining. To evaluate biodegradability, hydrogels are often
immersed in phosphate-buffered saline (PBS) solutions at 37 °C
and acidic pH (containing enzymes, redox species, or under light
irradiation if applicable) or subcutaneously injected in mice.
Injectable gels are either preformed gels with shear-thinning
and self-healing properties or in situ-forming gels. The former
can be extruded through a syringe needle after gelation as low
viscosity formulations by application of shear stress. When the
shear is removed, gels self-heal and quickly restore their original
state at the site of injection.8 Conversely, in situ gelling systems
are injected as free-flowing polymer solutions that transform into
gels at the injection site, forming drug depots for sustained drug
release.9 The mechanism of depot formation can be: (i) in situ
crosslinking, usually using a dual-barrel syringe for co-injection
and mixing of gel precursor solutions or an external source of
light to induce gelation (photo-polymerization); or (ii) in situ
phase transition, triggered by certain stimuli, such as changes
in temperature or pH.
Stimuli-responsive hydrogels
Hydrogels are 3D crosslinked networks of hydrophilic polymers
that absorb and retain large amounts of water while resisting dis-
solution in the aqueous medium. Some polymers respond to
external stimuli by changing their conformation, solubility, or
hydrophilic/lipophilic balance.10 Such changes are reversible
because stimuli-responsive polymers, also named ‘smart’ poly-
mers, return to their initial state once the stimulus ends. Accord-
ing to their nature, stimuli can be classified as physical, chemical,
or biological (Fig. 1).11
In the context of IT drug delivery, stimuli-responsive hydro-
gels can display a volume phase transition, swelling/shrinking,
or assembly/disassembly under exposure to exogenous triggers
Chemical Biological
pH
Enzymes
Redox
Glucose
Solvent composition
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Drug Discovery Today d Volume xx, Number xx d xxxx 2021
(light) or endogenous stimuli in the TME (temperature, pH,
redox, and enzymes). The concept of stimuli responsiveness
extends to hydrogels undergoing degradation in response to
stimuli by reversible or irreversible bond cleavage in the polymer
backbone or crosslinks.
Compared with conventional hydrogels, stimuli-responsive
hydrogels might allow for greater control of the local and dura-
tion of the drug release. Depending on the stimulus, these hydro-
gels offer different advantages for IT drug administration. For
example, thermoresponsive hydrogels exhibiting sol–gel transi-
tion at the tumor site are injected in the sol state with particular
ease and form implants in situ after gelation. The in situ-forming
implants adapt their shape to the surrounding tissues and act as a
reservoir system (i.e., a polymer-coated drug core) that enables
sustained drug release over longer periods compared with pre-
formed hydrogels. In addition to stimuli responsiveness, the
presence of acid-labile bonds, photocleavable moieties, or
enzyme substrates improves or imparts biodegradability to
hydrogels. Moreover, the ‘on/off’ switching of a remotely applied
stimulus enables a pulsatile drug release (versus continuous
release) with negligible release during the ‘off’ state, which can
be useful for certain cancer therapies.
The following sections cover stimuli-responsive hydrogels
developed recently for IT drug administration.
Thermoresponsive hydrogels
Hydrogels capable of responding to temperature changes are
considered thermoresponsive. For instance, the shift from room
temperature (25 °C) to body/tumor temperature (37 °C) can
cause thermoresponsive polymer solutions to exhibit a sol–gel
phase transition. The temperature at which a polymer solution
undergoes a transition from one phase to two phases (an aqueous
and a polymer-enriched phases) is designated the ‘critical solu-
tion temperature’.12
Most thermoresponsive polymers have a lower critical solu-
tion temperature (LCST) in water that is similar to the volume
phase transition temperature of the corresponding hydrogel. At
temperatures below the LCST, these polymers are miscible with
water but become insoluble when raising the temperature
beyond the LCST, separating from the solution and yielding a
gel. In the case of thermoresponsive polymers presenting an
upper critical solution temperature (UCST), heating beyond the
UCST favors the solubilization of polymer chains. Thermore-
sponsive polymers having the phase transition temperature near
body temperature are mainly LCST-type polymers.13
Over the past two decades, a large number of in situ-forming
thermoresponsive hydrogels based on natural (chitosan) or syn-
thetic (polyacrylamides, poloxamers, and polyesters) polymers
have been developed for IT drug administration.
Although atypical in most polysaccharides, the cationic nat-
ure of chitosan (CS) enables valuable interactions with anionic
molecules such as b-glycerophosphate (b-GP) to produce ther-
moresponsive hydrogels. For example, Bragta et al.14 selected IT
administration as an alternative route to intravenous delivery
of carboplatin in melanoma and formulated a CS/b-GP hydrogel
carrying carboplatin-loaded poly(e-caprolactone) (PCL) nanopar-
ticles (NPs). In another study, the efficacy of one IT injection of a
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POST-SCREEN (GREY)
CS/b-GP hydrogel containing paclitaxel (PTX)-loaded poly
(lactide-co-glycolide) (PLGA) microparticles was at least similar
to that of four intraperitoneal injections of a commercial formu-
lation (PaklitaxfilÒ, Fresenius Kabi) in mammary tumor-bearing
mice.15 However, CS/b-GP hydrogels are prone to in vivo degrada-
tion by erosion and the appearance of cracks that impair their
mechanical properties and controlled drug release. Accordingly,
significant efforts have been made to developing CS hydrogels
based on dynamic covalent chemistry with self-repair ability
after damage. To illustrate, Han et al.16 modified a CS/b-GP
hydrogel with dialdehyde-functionalized poly(ethylene glycol)
POST-SCREEN
(PEG) via chemical crosslinking to achieve autonomous self-
healing and sustained release of doxorubicin (DOX), used as a
model drug. Alternatively, a semi-interpenetrating polymer net-
work of N-acetylated glycol CS (or glycol chitin) and poly(acrylic
acid) allows for thermal gelation at 37 °C, as well as superior gel
strength.17
Whereas N-isopropylacrylamide is cytotoxic even at low con-
centrations, the polymerized form is nontoxic and biocompati-
ble with different cell types. Nevertheless, grafting poly(N-
isopropylacrylamide) (PNIPAM) with natural polymers has been
performed to impart biodegradability to the corresponding
hydrogel. Fong et al.18 prepared PNIPAM end-capped with car-
boxylic acid for conjugation with CS and hyaluronic acid (HA),
obtaining a HA-CS-graft-PNIPAM copolymer with an LCST simi-
lar to that of PNIPAM (32 °C).
Among the poloxamers, Poloxamer 407 (P407) or PluronicÒ
F127 (F127) solutions occupy the foreground of thermorespon-
sive polymer-based vehicles for IT drug delivery. For instance,
P407 solutions were mixed with DOX to kill MC-38 colon cancer
cells19 or with a topotecan solution to achieve a long-acting anti-
tumor effect on retinoblastoma.20 Several hybrid systems inte-
grating drug-loaded NPs and F127 hydrogels have also been
reported for this administration route. In an interesting approach
to the treatment of melanoma, Yu et al.21 prepared an F127
hydrogel to intratumorally deliver cisplatin (CDDP)-loaded
poly(a-L-glutamate)-graft-methoxy PEG NPs and microspheres
entrapping losartan potassium, which exerts antifibrotic effects.
The combination of liposomes and a PluronicÒ F127/F68 hydro-
gel was proposed to stabilize the lactone form of 7-ethyl-10-
hydroxycamptothecin and enhance drug retention at H22 hep-
atoma tumors.22 By contrast, mixed micelles comprising F127
and another surfactant, such as SolutolÒ HS1523 or D-a-
tocopherol PEG 1000 succinate,24 have been incorporated into
F127 hydrogels to deliver hydrophobic drugs, namely doc-
etaxel23 and PTX.24 Finally, the addition of N,N,N-trimethyl
CS25 or the use of crosslinkers, such as genipin,26 can improve
the mechanical properties of P407 hydrogels.
More recently, amphiphilic copolymers comprising PEG and
polyesters have been defined as first-line materials for the design
and manufacture of thermoresponsive hydrogels for IT drug
administration. The most used polyesters are polylactic acid
(PLA), PLGA, and PCL.
Shi et al.27 used a poly(D,L-lactide)-PEG-poly(D,L-lactide)
(PDLLA-PEG-PDLLA) triblock copolymer to prepare a thermore-
sponsive hydrogel coloaded with gemcitabine and CDDP for syn-
ergistic combination therapy in pancreatic cancer. The IT
injection of a norcantharidin-loaded PDLLA-PEG-PDLLA hydro-
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 POST-SCREEN (GREY) Drug Discovery Today d Volume xx, Number xx d xxx 2021

gel prolonged drug retention and improved its curative effect on
hepatocellular carcinoma.28 By incorporating free bevacizumab
and DOX-encapsulated PDLLA-PEG-PDLLA micelles into a
PDLLA-PEG-PDLLA hydrogel, Darge et al.29 induced normaliza-
tion of the tumor vasculature and potentiated the chemothera-
peutic effects of DOX. Zhou et al.30 developed a hydrogel
composite comprising erlotinib-loaded hollow mesoporous silica
NPs and a PDLLA-PEG-PDLLA hydrogel for localized treatment of
non-small cell lung cancer (NSCLC).
Shen et al.31 constructed a PEG-polyester-drug conjugate via
covalent linking of the Pt(IV) prodrug to the hydrophobic ends
POST-SCREEN
release of tamoxifen.32 Regarding commercial formulations,
OncoGelTM (MacroMed Inc.) is an injectable and thermorespon-
sive hydrogel formulated with a PLGA-PEG-PLGA triblock
copolymer (RegelTM) to deliver PTX to the tumor site in solid
malignancies. Despite evidence of preliminary efficacy in a Phase
IIa study, IT administration of OncoGelTM failed to reveal any
impact on overall tumor response to neoadjuvant radiotherapy
and intravenous chemotherapy (CDDP and 5-fluorouracil) in a
Phase IIb study (NCT00573131) in patients with esophageal
cancer.
Amphiphilic triblock copolymers have been yielded lately by

of two methoxy PEG (mPEG)-PLGA diblock copolymers. The bi
(mPEG-PLGA)-Pt(IV) conjugate self-assembled into core shell-
type micelles, the hydrophobic core of which functioned as
reservoir for PTX. At concentrated solutions, this thermorespon-
sive polymer–Pt(IV) conjugate underwent sol–gel transition
upon heating, accomplishing an IT and synchronous co-
delivery of Pt(IV) and PTX for treatment of ovarian cancer. In
addition, one IT injection of drug-loaded hydrogels based on
PLGA-PEG-PLGA triblock copolymers enabled the sustained
copolymerizing e-caprolactone33 and lactide34 in the presence
of PEG. Whereas a PEG-PCL-PEG hydrogel containing CDDP
and PTX-loaded mPEG-PCL micelles was proposed for combina-
tion chemotherapy in cervical cancer,33 a (PCL-co-lactide)-PEG-
(PCL-co-lactide) hydrogel incorporating curcumin-loaded
mPEG-PLA nanopolymersomes was fabricated to treat glioma.34
Table 1 summarizes the described thermoresponsive hydro-
gels regarding their phase transition temperature and their
in vitro and in vivo cancer models.

TABLE 1
a
Thermoresponsive hydrogels recently developed for IT drug delivery.
Composition Drug Phase Cancer cell Cancer (in vivo) Refs
transition (in vitro)
temperatureb
CS/b-GP, PCL NPs Carboplatin 37 ± 0.8 °C B16F1 Melanoma 14
CS/b-GP, PLGA microparticles PTX – – Mammary adenocarcinoma 15
CS/DF-PEG/b-GP DOX 31.51 ± 0.31 °C – Hepatocellular carcinoma 16
Glycol chitin/PAA, PLGA microspheres PTX 17 °Cc HepG2/C3A – 17
(hepatocellular
carcinoma)
HA-CS-g-PNIPAM, folate-conjugated GO DOX 30.07 °C MCF-7 Breast cancer 18
P407 DOX – MC-38 (colon – 19
cancer)
Topotecan 37.23 ± 0.473 °C – Retinoblastoma 20
F127, PLG-g-mPEG nanoparticles and microspheres CDDP, losartan 30 °C – Melanoma 21
F127/F68, liposomes SN-38 – – Hepatocellular carcinoma 22
F127/ P188, F127 and SolutolÒ HS15 micelles DTX 37 °C – Colon cancer 23
F127/HA, F127, and TPGS micelles PTX, DOX 35 °C – – 24
F127/TMC DTX 37 °C U87MG Glioblastoma 25
PDLLA-PEG-PDLLA Gemcitabine, CDDP 35 °C Bxpc-3 Pancreatic cancer 27
Norcantharidin – – Hepatocellular carcinoma 28
PDLLA-PEG-PDLLA, PDLLA-PEG-PDLLA micelles DOX, bevacizumab 37 °C HeLa Cervical carcinoma 29
PDLLA-PEG-PDLLA, hollow mesoporous silica NPs Erlotinib 35.7°Cd36.4 °C A549 NSCLC 30
Bi(mPEG-PLGA)-Pt (IV) Pt(IV) prodrug, PTX 35 °C SKOV-3 Ovarian cancer 31
PLGA-PEG-PLGA Tamoxifen 37.14 ± 0.66 °C – Breast cancer 32
PEG-PCL-PEG, mPEG-PCL micelles CDDP, PTX 30 °C – Cervical cancer 33
PCLA-PEG-PCLA, mPEG-PLA nanopolymersomes Curcumin 32 °C – Glioma 34
a
Abbreviations: b-GP, b-glycerophosphate; Bi(mPEG-PLGA) Pt(IV), bi(poly(ethylene glycol) methyl ether-poly(lactic-co-glycolic acid))-Pt(IV) conjugate; CDDP, cisplatin; CS, chitosan; DF-PEG,
dialdehyde-functionalized poly(ethylene glycol); DOX, doxorubicin; DTX, docetaxel; F68, PluronicÒ F68; F127, PluronicÒ F127; GO, graphene oxide; HA, hyaluronic acid; HA-CS-g-PNIPAM, hyaluronic
acid-chitosan-graft-poly(N-isopropylacrylamide); mPEG-PCL, poly(ethylene glycol) methyl ether-poly(e-caprolactone); mPEG-PLA, poly(ethylene glycol) methyl ether-polylactic acid; NSCLC, non-
small cell lung cancer; P188, poloxamer 188; P407, poloxamer 407; PAA, poly(acrylic acid); PCL, poly(e-caprolactone); PCLA-PEG-PCLA, poly(e-caprolactone-co-lactide)-poly (ethylene glycol)-poly(e-
caprolactone-co-lactide): PDLLA PEG PDLLA, poly(D,L-lactide)-poly(ethylene glycol)-poly(D,L-lactide); PEG-PCL-PEG, poly(ethylene glycol)-poly(e-caprolactone)-poly(ethylene glycol); PLG-g-mPEG,
poly(a-L-glutamate)-graft-poly(ethylene glycol) methyl ether; PLGA, poly(lactic-co-glycolic acid); PLGA-PEG-PLGA, poly(lactic-co-glycolic acid)-poly(ethylene glycol)-poly(lactic-co-glycolic acid); PTX,
paclitaxel; SN-38, 7-ethyl-10-hydroxycamptothecin; TMC, N,N,N-trimethyl chitosan; TPGS, D-a-tocopherol poly(ethylene glycol) 1000 succinate.
b
The volume phase transition temperatures (sol–gel transition or gelation temperatures) were determined by the test tube inversion method, rheology, differential scanning calorimetry, or using
a thermometer.
c
This formulation comprises agent A (glycol chitin/PAA solution containing PTX-loaded PLGA microspheres, crosslinker, and initiator) and agent B (promoter). The gel forms at 37 °C upon co-
injection of agents A and B, which are liquid and stored separately in the refrigerator.
d
Incorporation of erlotinib at a higher concentration (6 mg/ml) led to a slight increase in the phase transition temperature of the hydrogel compared with that of a low concentration of erlotinib
(2 mg/ml).

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Drug Discovery Today d Volume xx, Number xx d xxxx 2021
Photo-responsive hydrogels
These hydrogels are typically based on polymers containing a
photoreactive moiety that responds to light by way of a photo-
chemical reaction, such as cleavage and isomerization.35 As an
example, Zhao et al.36 prepared an injectable self-assembled
DOX-loaded hydrogel based on a four-arm star polymer, PEG-
poly(c-o-nitrobenzyl-L-glutamate). The presence of o-
nitrobenzyl groups in the polymer backbone rendered hydrogels
photodegradable by irreversible photocleavage of o-nitrobenzyl
ester linkages after irradiation with ultraviolet (UV) light. By con-
trast, azobenzene moieties undergo reversible trans-to-cis isomer-
ization when irradiated with UV light, returning to trans isomers
upon heating or higher wavelength irradiation. Given the sub-
stantial limitations to the use of UV light (poor penetration
and risk of damaging tissues), approaches involving upconver-
sion of low-energy photons (e.g., near-infrared; NIR) into high-
energy photons (e.g., UV) are favorable.37 Chen et al.38 devel-
oped an injectable ‘click’-crosslinked gelatin hydrogel loaded
with DOX and poly(acrylic acid)-coated upconversion NPs. The
amount of DOX released from the hydrogel was controllable
with an ‘on/off’ switchable operation of NIR irradiation that trig-
gered photo-isomerization of the azobenzene crosslinker.
Nonetheless, NIR-I light (650–950 nm) penetrates <1 cm into
the tissues, which is suitable for superficial thin tumors only.
Given that the tissue penetration depth of NIR-II light (1000–
1700 nm) is 3–5 cm, the treatment of large superficial tumors
might be feasible, but its clinical application to deep tumors is
still restricted.39 In general, increasing the power density (W/
cm2) and exposure time to irradiation leads to greater
penetration.
In terms of IT drug administration, some photoresponsive
hydrogels are supramolecular hydrogels incorporating pho-
tothermal agents. These hydrogels are 3D networks of molecular
building blocks self-assembled via noncovalent interactions,
which exhibit a gel–sol transition under light irradiation in
response to the heat generated by photothermal agents. In par-
ticular, Ruan et al.40 demonstrated that poly(N-phenylglycine)
building blocks could act as NIR-II-absorbing mediators while
tethering PEG chains to form a hydrogel via host–guest interac-
tion with a-cyclodextrin. Meanwhile, PEGylated poly(N-
phenylglycine) was used as photothermal polymer backbone to
construct a photoresponsive hydrogel for IT delivery of DOX.41
Another strategy to achieve reversible photo-induced phase
transition uses NIR-absorbing nanostructures embedded within
the matrix of hydrogels based on thermoresponsive polymers.
Specifically, a light-responsive polysaccharide-based hydrogel
was obtained by dispersing small CuS NPs in a gellan aqueous
solution at elevated temperature, followed by mixing with
DOX and cooling to room temperature. After IT injection of
the hydrogel in mice, CuS NPs transformed irradiated NIR light
(808 nm, 1.0 W/cm2, 3 min) into heating beyond the UCST of
gellan, which elicited a slow gel–sol transition and increased
the release of DOX at the tumor site.42 Instead, the release rate
of DOX can be precisely modulated by light activation of black
phosphorus nanosheets integrating an agarose hydrogel that
undergoes reversible hydrolysis and softening under irradiation
with an 808-nm laser.43 Departing from the use of inorganic
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POST-SCREEN (GREY)
photothermal agents, Hou et al.44 used a derivative of an organic
carbon source, humic acid, to endow an injectable agarose
hydrogel with photothermal conversion property. In a different
work, Wang et al.45formulated an injectable self-healing hydro-
gel via dynamic covalent enamine bond between polyetherimide
and a four-arm star copolymer, poly[2-(dimethylamino)ethyl
methacrylate-co-2-hydroxyethyl methacrylate] modified with
tert-butyl acetoacetate. The blank hydrogel was loaded with
DOX and polydopamine NPs before IT injection in 4T1 breast
tumor-bearing mice. Under NIR irradiation (808 nm, 1.0 W/
cm2, 10 min), the temperature rose well above the LCST
POST-SCREEN
(40 °C) of the copolymer because of the photothermal effect
of polydopamine NPs, leading to shrinking and rapid release of
DOX.
pH-responsive hydrogels
Referring to acidosis of tumor tissues, two strategies have been
reported for the design of pH-responsive hydrogels for IT drug
administration: (i) the selection of pH-responsive polymers with
numerous weak basic (e.g., amine) groups attached to the
hydrophobic backbone, thereby being considered polybases (ca-
tionic); or (ii) the incorporation of acid-labile linkages within the
polymer network or between the polymer and drug, the cleavage
of which enables the release of entrapped or anchored drugs.46
When the external pH is less than the pKa of a cationic poly-
mer (e.g., acidic pH), amine pendant groups are positively
charged (NH3+) by protonation. The electrostatic repulsion
between charges leads to polymer chain expansion and
swelling.47 In turn, the enlarged mesh of swollen hydrogels facil-
itates drug diffusion through the network. In a different
approach, Raza et al.48 synthesized a pH-responsive FEFEFRFK
octapeptide that self-assembled at neutral pH into an injectable
hydrogel for IT administration of PTX. At pH 5.5, protonation
of arginine and electrostatic repulsion between the amino acid
side chains induced a b-sheet-coil transition of peptide fibers, cul-
minating in hydrogel disassembly and sustained release of PTX.
The second strategy involves pH-responsive bonds, which are
cleaved in the extracellular fluid or, after endocytosis, in tumor
cell endosomes or lysosomes. Dynamic covalent bonds, namely
hydrazone bonds, can be used by virtue of their hydrolytic stabil-
ity at normal physiological pH (pH 7.4) and rapid cleavage in
mildly acidic pH. Before gel preparation, Sharma et al.49 conju-
gated part of eight-arm PEG-hydrazine to DOX via its ketone
moiety and covalently entrapped PEG-DOX conjugates into the
polymer matrix. The resulting hydrogel provides an excellent
platform for sustained long-term delivery of DOX, with 81.33%
of the conjugate being released after 40 days at pH 6.4. Given
that variable pH ranges (still acidic but other than 6.5–6.9) are
described for tumor tissues, one can expect this subtle change
in the pH value not to impair the stimuli responsiveness of the
hydrogel. There are also slight differences between the reported
values of other stimuli, such as redox species or enzyme
concentrations.
Redox-responsive hydrogels
The basic principle of designing reduction-responsive hydrogels
is the incorporation of disulfide linkages directly into polymers
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 POST-SCREEN (GREY)
or via disulfide-containing crosslinkers, namely cystamine and
its derivatives.50 Disulfide bonds can be cleaved in the presence
of a reducing agent, such as GSH, which, acting as an electron
donor, is oxidized to glutathione disulfide. Zou et al.51 also
described self-assembled hydrogels based on a reducible dipheny-
lalanine dipeptide derivative for IT delivery of 5-aminolevulinic
acid hydrochloride prodrug. Circular dichroism spectra of 10-
mM GSH-treated hydrogels suggested a change in their sec-
ondary structure because of the reduction of disulfide bonds in
the dipeptide. Alternatively, diselenide bonds are expected to
impart more sensitivity than disulfide linkages because of their
POST-SCREEN
lower bond energy. Generally, diselenide bonds can be cleaved
through the action of both reductants and oxidants (e.g., hydro-
gen peroxide, H2O2), being reduced to selenol or oxidized to sele-
ninic acid. Considering the potential of diselenide-containing
hydrogels for a dual redox response, Gong et al.52 produced an
injectable PEG hydrogel via coupling reaction between the N-
hydroxysuccinimide ester ends of four-arm PEG and amine
groups of the selenocystamine crosslinker. Under the stimuli of
low concentrated GSH (0.1 mg/ml) or H2O2 (0.01%) solutions,
the breakage of diselenide bonds in selenocystamine led to dis-
ruption of the crosslinked network and fast release of the encap-
sulated drug.
Oxidation-responsive hydrogels depend on ROS to enable a
concurrent behavior of hydrogel degradation and drug release.
These oxidative species can be generated endogenously (e.g., dur-
ing oxidative phosphorylation in the mitochondria) or exoge-
nously (e.g., using a photosensitizer in photodynamic therapy).
In addition to oxygen free radicals, namely superoxide (O2 )
and hydroxyl radical (OH), ROS include nonradical species, such
H2O2, usually ranging from 100 mM to 1 mM in tumor cells.53 In
vitro studies are typically performed with H2O2 in PBS solutions
to create an oxidative environment.
TABLE 2
Dual and multiple stimuli-responsive injectable hydrogels for IT drug delivery.
Stimuli Composition
Temperature; pH PDMAEMA, DOX, 131
I-
BSA
CS-g-(mPEG-b-PCL),
DOX, Curcumin
P(NIPAM-co-IA), CS/b-
GP, DOX
pH; enzyme; light; Gela NP, PDA NP, DOX
temperature
pH; ROS; GSH; light; HSF, Cy7, DOX
temperature
a
Abbreviations: CS-g-(mPEG-b-PCL), chitosan-graft-(poly(ethylene glycol) methyl ether-block-poly(e caprolactone)); CS/b-GP, chitosan/b-glycerophosphate; Cy7, cyanine-7; DOX, doxorubicin; GdL,
glucono delta-lactone; Gela, gelatin; GSH, glutathione; HSF, hydrophilic silk fibroin; 131I-BSA, iodine-131-labeled bovine serum albumin; IA, itaconic acid; LCST, lower critical solution temperature;
MMP, matrix metalloproteinase; NIR, near-infrared; NP, nanoparticles; PDA, polydopamine; PDMAEMA, poly(N,N-dimethyl aminoethyl methacrylate); PNIPAM, poly(N-isopropylacrylamide); P(NIPAM-
co-IA), poly(N-isopropylacrylamide-co-itaconic acid); ROS, reactive oxygen species.
6 www.drugdiscoverytoday.com
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One possible strategy to produce oxidation-responsive hydro-
gels involves thioketal linkages that are readily cleaved by ROS
into thiol and ketone groups. Xu et al.54 fabricated a L-
phenylalanine-based low-molecular-weight gelator containing
thioketal and a control gelator without ROS-cleavable bonds.
The corresponding hydrogels coloaded with DOX and a photo-
sensitizer were intratumorally injected in 4T1 breast
tumor = bearing mice to assess their in vivo antitumor efficacy.
Compared with the groups treated with drug-loaded ROS-
responsive gels, those receiving IT injections of control gels
(without thioketal) showed little inhibition of tumor growth,
owing to the absence of response to a ROS-rich
microenvironment.
Enzyme-responsive hydrogels
Enzyme responsiveness is usually introduced into hydrogels by
means of enzyme-mediated crosslinkages or enzyme-cleavable
moieties.
The enzymatic degradation of hydrogels for IT drug adminis-
tration requires the incorporation of an enzyme substrate or sub-
strate mimic, which can be: (i) the polymer itself in the case of
biodegradable polymers, such as polysaccharides and polyesters,
through the breakage of glycosidic and ester bonds, respectively;
or (ii) a peptide linker specifically recognized and cleaved by
MMPs overexpressed in many tumor tissues. To date, 23 MMPs
have been identified in humans and their concentration in
tumor tissues varies according to the type of cancer. For example,
MMP-9 and MMP-2 are reported to be the most abundant in
breast cancer, in a concentration of 32.27 and 17.08 ng/mg,
respectively.55 Li et al.56 prepared DOX-encapsulated PDLLA-
PEG-PDLLA micelles and selected a peptide linker containing
an MMP-2-labile sequence (GPQGIWGQ) to develop an in situ
crosslinked MMP-responsive HA hydrogel. According to in vitro
a
Highlights Refs
PDMAEMA solutions transformed into nanogels at 37 °C 60
At pH 5.8, electrostatic repulsion within polymer network led to swelling and fast in vitro
release of DOX
Sol–gel phase transition occurred at pH 6.9 and physiological temperature 61
CS-g-(mPEG-b-PCL) hydrogels released both hydrophilic and hydrophobic drugs in sustained
manner for up to 2 weeks
Incorporation of hydrophilic moiety (IA) elevated LCST of PNIPAM toward body temperature 62
Swollen state of cationic hydrogel quickened release of DOX
GdL-induced acidification caused charge reversal of Gela NP to form colloidal Gela/PDA 63
hydrogel
Degradation of Gela NP by proteases anticipated responsiveness to MMP-2/9 and enzymatic
promotion of DOX release
PDA NP converted NIR light into heat, eliciting gel–sol transition and increased release of DOX
HSF molecules self-assembled into hydrogel via physical forces 64
Acidity and ROS damaged b-sheet structure, whereas GSH cleaved intra- and intermolecular
disulfide bonds in HSF molecules
Hyperthermia generated by Cy7 upon NIR irradiation enhanced molecular thermal motion,
disrupting physical interactions within hydrogel
HSF hydrogels responded to multiple stimuli that together increased release rate of DOX
Drug Discovery Today d Volume xx, Number xx d xxxx 2021
release studies in the presence of MMP-2 (1.5 lg/ml), diffusion
and degradation by peptide cleavage were implicated in the
release of DOX. Meanwhile, a HS-MMP-SH peptide crosslinker
(GCREG-PQGIWGQ-ERCG) was introduced in another HA
hydrogel, further loaded with nano DOX and nano indocyanine
green (photosensitizer) for chemophototherapy of head and
neck squamous cell carcinoma.57
A second prerequisite for enzymatic degradation is the acces-
sibility of enzymes to substrate within the hydrogel because it
can considerably interfere with the kinetics of enzyme-
catalyzed reactions.58 This can be illustrated by a pentablock
copolymer (denoted as L-Pep-NC) synthesized from a L-
peptide-PEG macroinitiator to link core-crosslinked micelles
and yield an injectable enzyme-responsive hydrogel. The peptide
midblock was cleaved after 160 min of exposure to a type IV col-
lagenase (0.5 units/ml), used as a model enzyme for MMP-2 and
MMP-9. By contrast, it took 7 days for the hydrogel crosslinked
with L-Pep-NC to be completely degraded at an enzyme concen-
tration of 30 units/ml, probably because of the limited access of
collagenase to peptide block in hydrogel network.59
Dual and multiple stimuli-responsive hydrogels
Dual or multiple stimuli-responsive hydrogels respond in a speci-
fic and distinct manner to two or more stimuli available in the
TME and/or assisted by an external source. Compared with their
single stimulus-responsive counterparts, these hydrogels offer
greater control over their responsive behavior and an opportu-
nity to improve overall performance and applicability to IT drug
delivery.
Some examples of dual and multiple stimuli-responsive inject-
able hydrogels are presented in Table 2.
Concluding remarks
A better understanding of the TME, together with the advance-
ments in materials chemistry, provided a solid background for
the design of stimuli-responsive hydrogels for IT drug delivery.
Their proof-of-concept has been demonstrated by growth inhibi-
tion of several tumors in different mouse models. However, sev-
eral obstacles to clinical translation remain because: (i) some
materials lack enough biocompatibility and biodegradability or
need further research into these properties; (ii) the synthetic
routes of some polymers are too complex for industrial scale-up
production; and (iii) the responsiveness of hydrogels to the stim-
References
1 P. Pittayapruek, J. Meephansan, O. Prapapan, M. Komine, M. Ohtsuki, Role of
matrix metalloproteinases in photoaging and photocarcinogenesis, Int J Mol Sci
17 (2016) 868.
2 W.L. Chen, C.C. Wang, Y.J. Lin, C.P. Wu, C.H. Hsieh, Cycling hypoxia induces
chemoresistance through the activation of reactive oxygen species-mediated B-
cell lymphoma extra-long pathway in glioblastoma multiforme, J Transl Med 13
(2015) 389.
3 H. Wang, H. Jiang, M. Van de Gucht, M. De Ridder, Hypoxic radioresistance: can
ROS be the key to overcome It?, Cancers 11 (2019) 112
4 G. Boysen, The glutathione conundrum: Stoichiometric disconnect between its
formation and oxidative stress, Chem Res Toxicol 30 (2017) 1113–1116.
5 A.P. Andersen, J.M. Moreira, S.F. Pedersen, Interactions of ion transporters and
channels with cancer cell metabolism and the tumour microenvironment, Philos
Trans R Soc Lond B Biol Sci 369 (2014) 20130098.
Please cite this article in press as: A.C. Marques et al., Drug Discovery Today (2021), https://doi.org/10.1016/j.drudis.2021.04.012
POST-SCREEN (GREY)
uli in human tumors might be similar, but not the same as those
observed in the laboratory.
To understand the latter, one should consider that the in vitro
conditions sometimes fail to mimic the real environment of
tumors. In particular, if enzymes (e.g., MMPs) or biomolecules
(e.g., GSH or ROS) are used in higher concentrations than those
reported for tumor tissues, a decrease in the sensitivity of the
hydrogel is likely to be observed in vivo. Also, the contribution
of concomitant stimuli to the hydrogel response is often
neglected. Even when the in vitro studies are performed under
pathophysiological conditions, endogenous cues are somewhat
POST-SCREEN
different between individuals or at different stages of cancer,
which interferes with the reproducibility of the response
in vivo. Accordingly, additional efforts should be made to set
the range of values (e.g., pH, concentration, etc.) that trigger a
desirable response for each stimuli-responsive hydrogel and
develop novel hydrogels that can change dynamically with sub-
tle variations in the environment. Nevertheless, one can predict
the next generation of stimuli-responsive hydrogels to be multi-
ple stimuli-responsive hydrogels that allow for independent tun-
ing of the individual response to each stimulus. In turn, the
combined effect of multiple stimuli would result in more precise
and programmable drug delivery and, ultimately, a better thera-
peutic effect.
Finally, most tumor models have been established by subcuta-
neous inoculation of human cancer cells in mice, which do not
reflect the TME accurately. Instead, orthotopic mouse models
would be more clinically relevant because tumor xenografts are
placed into the tissue/organ of origin. Therefore, future preclini-
cal studies should be conducted in orthotopic tumor models and
one can expect encouraging results to inspire further evaluation
of these formulations in clinical trials.
Declaration of Competing Interest
The authors declare that they have no known competing
financial interests or personal relationships that could have
appeared to influence the work reported in this paper.
Acknowledgments
This work was supported by the Applied Molecular Bio-
sciences Unit-UCIBIO, which is financed by national funds from
FCT/MCTES (UID/Multi/04378/2019).
6 K.M. Bailey, J.W. Wojtkowiak, H.H. Cornnell, M.C. Ribeiro, Y. Balagurunathan, A.
I. Hashim, et al., Mechanisms of buffer therapy resistance, Neoplasia 16 (2014)
354–364.
7 A. Fakhari, Subramony J. Anand, Engineered in-situ depot-forming hydrogels for
intratumoral drug delivery, J Control Release 220 (2015) 465–475.
8 M.H. Chen, L.L. Wang, J.J. Chung, Y.H. Kim, P. Atluri, J.A. Burdick, Methods to
assess shear–thinning hydrogels for application as injectable biomaterials, ACS
Biomater Sci Eng 3 (2017) 3146–3160.
9 T. Thambi, Y. Li, D.S. Lee, Injectable hydrogels for sustained release of therapeutic
agents, J Control Release 267 (2017) 57–66.
10 W.B. Liechty, D.R. Kryscio, B.V. Slaughter, N.A. Peppas, Polymers for drug
delivery systems, Annu Rev Chem Biomol Eng 1 (2010) 149–173.
11 V.A. Ganesh, A. Baji, S. Ramakrishna, Smart functional polymers – a new route
towards creating a sustainable environment, RSC Adv 4 (2014) 53352–53364.
www.drugdiscoverytoday.com 7
 POST-SCREEN (GREY)
12 D.D. Chakraborty, L.K. Nath, P. Chakraborty, Recent progress in smart polymers:
behavior, mechanistic understanding and application, Polym Plast Technol Eng
57 (2018) 945–957.
13 L. Klouda, Thermoresponsive hydrogels in biomedical applications: a seven-year
update, Eur J Pharm Biopharm 97 (2015) 338–349.
14 P. Bragta, R.K. Sidhu, K. Jyoti, A. Baldi, U.K. Jain, R. Chandra, J. Mada, et al.,
Intratumoral administration of carboplatin bearing poly (e-caprolactone)
nanoparticles amalgamated with in situ gel tendered augmented drug delivery,
cytotoxicity, and apoptosis in melanoma tumor, Colloids Surf B 166 (2018) 339–
348.
15 J.I. Pesoa, M.J. Rico, V.R. Rozados, O.G. Scharovsky, J.A. Luna, L.N. Mengatto,
Paclitaxel delivery system based on poly(lactide-co-glycolide) microparticles and
chitosan thermo–sensitive gel for mammary adenocarcinoma treatment, J Pharm
Pharmacol 70 (2018) 1494–1502.
POST-SCREEN
16 X. Han, X. Meng, Z. Wu, Z. Wu, X. Qi, Dynamic imine bond cross-linked self-
healing thermosensitive hydrogels for sustained anticancer therapy via
intratumoral injection, Mater Sci Eng C Mater Biol Appl 93 (2018) 1064–1072.
17 H.R. Lin, Y.T. Chen, Y.C. Wu, Y.J. Lin, Glycol chitin/PAA hydrogel composite
incorporated bio-functionalized PLGA microspheres intended for sustained
release of anticancer drug through intratumoral injection, J Biomater Sci Polym
Ed 29 (2018) 1839–1858.
18 Y.T. Fong, C.H. Chen, J.P. Chen, Intratumoral delivery of doxorubicin on folate–
conjugated graphene oxide by in-situ forming thermo-sensitive hydrogel for
breast cancer therapy, Nanomaterials 7 (2017) 388.
19 C.K. Chung, J. García-Couce, Y. Campos, D. Kralisch, K. Bierau, A. Chan, et al.,
Doxorubicin loaded poloxamer thermosensitive hydrogels: chemical,
pharmacological and biological evaluation, Molecules 25 (2020) 2219.
20 Y. Huo, Q. Wang, Y. Liu, J. Wang, Q. Li, Z. Li, et al., A temperature-sensitive
phase-change hydrogel of topotecan achieves a long-term sustained antitumor
effect on retinoblastoma cells, OncoTargets Ther 12 (2019) 6069–6082.
21 M. Yu, C. Zhang, Z. Tang, X. Tang, H. Xu, Intratumoral injection of gels
containing losartan microspheres and (PLG-g-mPEG)-cisplatin nanoparticles
improves drug penetration, retention and anti-tumor activity, Cancer Lett 442
(2019) 396–408.
22 R. Bai, X. Deng, Q. Wu, X. Cao, T. Ye, S. Wang, Liposome-loaded thermo-
sensitive hydrogel for stabilization of SN-38 via intratumoral injection:
Optimization, characterization, and antitumor activity, Pharm Dev Technol 23
(2018) 106–115.
23 M. Xu, Y. Mou, M. Hu, W. Dong, X. Su, R. Wu, et al., Evaluation of micelles
incorporated into thermosensitive hydrogels for intratumoral delivery and
controlled release of docetaxel: a dual approach for in situ treatment of tumors,
Asian J Pharm Sci 13 (2018) 373–382.
24 M. Rezazadeh, V. Akbari, E. Amuaghae, J. Emami, Preparation and
characterization of an injectable thermosensitive hydrogel for simultaneous
delivery of paclitaxel and doxorubicin, Res Pharm Sci 13 (2018) 181–191.
25 M.H. Turabee, T.H. Jeong, P. Ramalingam, J.H. Kang, Y.T. Ko, N, N, N-trimethyl
chitosan embedded in situ Pluronic F127 hydrogel for the treatment of brain
tumor, Carbohydr Polym 203 (2019) 302–309.
26 Kelly H, Duffy G, Rossi S, Hastings C. A thermo-responsive hydrogel for
intratumoral administration as a treatment in solid tumor cancers. Royal
College of Surgeons in Ireland. EP3706718
27 K. Shi, B. Xue, Y. Jia, L. Yuan, R. Han, F. Yang, et al., Sustained co-delivery of
gemcitabine and cis-platinum via biodegradable thermo-sensitive hydrogel for
synergistic combination therapy of pancreatic cancer, Nano Res 12 (2019) 1389–
1399.
28 B. Xue, M. Lei, K. Shi, M. Wang, Y. Hao, Y. Xiao, et al., Intratumoral injection of
norcantharidin-loaded poly(D, L–lactide)-b-poly(ethylene glycol)-b-poly(D, L-
lactide) thermosensitive hydrogel for the treatment of primary hepatocellular
carcinoma, J Biomed Nanotechnol 15 (2019) 2025–2044.
29 H.F. Darge, A.T. Andrgie, E.Y. Hanurry, Y.S. Birhan, T.W. Mekonnen, H.Y. Chou,
et al., Localized controlled release of bevacizumab and doxorubicin by thermo-
sensitive hydrogel for normalization of tumor vasculature and to enhance the
efficacy of chemotherapy, Int J Pharm 572 (2019) 118799.
30 X. Zhou, X. He, K. Shi, L. Yuan, Y. Yang, Q. Liu, et al., Injectable thermosensitive
hydrogel containing erlotinib–loaded hollow mesoporous silica nanoparticles as
a localized drug delivery system for NSCLC therapy, Adv Sci 7 (2020) 2001442.
31 W. Shen, X. Chen, J. Luan, D. Wang, L. Yu, J. Ding, Sustained codelivery of
cisplatin and paclitaxel via an injectable prodrug hydrogel for ovarian cancer
treatment, ACS Appl Mater Interfaces 9 (2017) 40031–40046.
32 D. Meng, H. Lei, X. Zheng, Y. Han, R. Sun, D. Zhao, et al., A temperature-sensitive
phase-change hydrogel of tamoxifen achieves the long-acting antitumor
activation on breast cancer cells, OncoTargets Ther 12 (2019) 3919–3931.
8 www.drugdiscoverytoday.com
Please cite this article in press as: A.C. Marques et al., Drug Discovery Today (2021), https://doi.org/10.1016/j.drudis.2021.04.012
Drug Discovery Today d Volume xx, Number xx d xxx 2021
33 S. Xu, X. Du, G. Feng, Y. Zhang, J. Li, B. Lin, et al., Efficient inhibition of cervical
cancer by dual drugs loaded in biodegradable thermosensitive hydrogel
composites, Oncotarget 9 (2018) 282–292.
34 M. Babaei, J. Davoodi, R. Dehghan, M. Zahiri, K. Abnous, S.M. Taghdisi, et al.,
Thermosensitive composite hydrogel incorporated with curcumin-loaded
nanopolymersomes for prolonged and localized treatment of glioma, J Drug
Deliv Sci Technol 59 (2020) 101885.
35 I. Tomatsu, K. Peng, A. Kros, Photoresponsive hydrogels for biomedical
applications, Adv Drug Deliv Rev 63 (2011) 1257–1266.
36 D. Zhao, Q. Tang, Q. Zhou, K. Peng, H. Yang, X. Zhang, A photo-degradable
injectable self-healing hydrogel based on star poly(ethylene glycol)-b-polypeptide
as a potential pharmaceuticals delivery carrier, Soft Matter 14 (2018) 7420–7428.
37 A. Barhoumi, Q. Liu, D.S. Kohane, Ultraviolet light-mediated drug delivery:
principles, applications, and challenges, J Control Release 219 (2015) 31–42.
38 Y. Chen, Y. Hao, Y. Huang, W. Wu, X. Liu, Y. Li, et al., An injectable, near-
infrared light-responsive click cross-linked azobenzene hydrogel for breast cancer
chemotherapy, J Biomed Nanotechnol 15 (2019) 1923–1936.
39 X. Wang, Z. Xuan, X. Zhu, H. Sun, J. Li, Z. Xie, Near-infrared photoresponsive
drug delivery nanosystems for cancer photo-chemotherapy, J
Nanobiotechnology 18 (2020) 108.
40 C. Ruan, C. Liu, H. Hu, X.L. Guo, B.P. Jiang, H. Liang, et al., NIR-II light-
modulated thermosensitive hydrogel for light–triggered cisplatin release and
repeatable chemo-photothermal therapy, Chem Sci 10 (2019) 4699–4706.
41 C. Liu, X. Guo, C. Ruan, H. Hu, B.P. Jiang, H. Liang, et al., An injectable
thermosensitive photothermal-network hydrogel for near-infrared-triggered drug
delivery and synergistic photothermal–chemotherapy, Acta Biomater 96 (2019)
281–294.
42 Y. Zheng, Y. Liang, D. Zhang, Z. Zhou, J. Li, X. Sun, et al., Fabrication of injectable
CuS nanocomposite hydrogels based on UCST-type polysaccharides for NIR-
triggered chemo-photothermal therapy, Chem Commun 54 (2018) 13805–
13808.
43 M. Qiu, D. Wang, W. Liang, L. Liu, Y. Zhang, X. Chen, et al., Novel concept of the
smart NIR-light-controlled drug release of black phosphorus nanostructure for
cancer therapy, Proc Natl Acad Sci U S A 115 (2018) 501–506.
44 M. Hou, R. Yang, L. Zhang, L. Zhang, G. Liu, Z. Xu, et al., Injectable and natural
humic acid/agarose hybrid hydrogel for localized light-driven photothermal
ablation and chemotherapy of cancer, ACS Biomater Sci Eng 4 (2018) 4266–4277.
45 C. Wang, N. Zhao, W. Yuan, NIR/thermoresponsive injectable self-healing
hydrogels containing polydopamine nanoparticles for efficient synergistic
cancer thermochemotherapy, ACS Appl Mater Interfaces 12 (2020) 9118–9131.
46 M. Norouzi, B. Nazari, D.W. Miller, Injectable hydrogel-based drug delivery
systems for local cancer therapy, Drug Discov Today 21 (2016) 1835–1849.
47 M. Rizwan, R. Yahya, A. Hassan, M. Yar, A.D. Azzahari, V. Selvanathan, et al., pH
Sensitive hydrogels in drug delivery: brief history, properties, swelling, and
release mechanism, material selection and applications, Polymers 9 (2017) 137.
48 F. Raza, Y. Zhu, L. Chen, X. You, J. Zhang, A. Khan, et al., Paclitaxel-loaded pH
responsive hydrogel based on self-assembled peptides for tumor targeting,
Biomater Sci 7 (2019) 2023–2036.
49 P.K. Sharma, Y. Singh, Glyoxylic hydrazone linkage-based PEG hydrogels for
covalent entrapment and controlled delivery of doxorubicin, Biomacromolecules
20 (2019) 2174–2184.
50 J. Zhang, X. Jiang, X. Wen, Q. Xu, H. Zeng, Y. Zhao, et al., Bio-responsive smart
polymers and biomedical applications, J Phys Mater 2 (2019) 032004.
51 Q. Zou, R. Chang, R. Xing, C. Yuan, X. Yan, Injectable self-assembled bola-
dipeptide hydrogels for sustained photodynamic prodrug delivery and enhanced
tumor therapy, J Control Release 319 (2020) 344–351.
52 C. Gong, M. Shan, B. Li, G. Wu, Injectable dual redox responsive diselenide-
containing poly(ethylene glycol) hydrogel, J Biomed Mater Res A 105 (2017)
2451–2460.
53 D. Wang, R. Shi, J. Zhou, S. Shi, H. Wu, P. Xu, et al., Photo-enhanced singlet
oxygen generation of Prussian blue-based nanocatalyst for augmented
photodynamic therapy, iScience 9 (2018) 14–26.
54 L. Xu, M. Zhao, Y. Yang, Y. Liang, C. Sun, W. Gao, et al., A reactive oxygen species
(ROS)-responsive low molecular weight gel co-loaded with doxorubicin and Zn(ii)
phthalocyanine tetrasulfonic acid for combined chemo-photodynamic therapy, J
Mater Chem B 5 (2017) 9157–9164.
55 K.J. Isaacson, M. Martin Jensen, N.B. Subrahmanyam, H. Ghandehari, Matrix–
metalloproteinases as targets for controlled delivery in cancer: An analysis of
upregulation and expression, J Control Release 259 (2017) 62–75.
56 W. Li, C. Tao, J. Wang, Y. Le, J. Zhang, MMP-responsive in situ forming hydrogel
loaded with doxorubicin-encapsulated biodegradable micelles for local
chemotherapy of oral squamous cell carcinoma, RSC Adv 9 (2019) 31264–31273.
Drug Discovery Today d Volume xx, Number xx d xxxx 2021
57 H.H. Wang, Z.G. Fu, W. Li, Y.X. Li, L.S. Zhao, L. Wen, et al., The synthesis and
application of nano doxorubicin–indocyanine green matrix metalloproteinase-
responsive hydrogel in chemophototherapy for head and neck squamous cell
carcinoma, Int J Nanomed 14 (2019) 623–638.
58 R. Chandrawati, Enzyme-responsive polymer hydrogels for therapeutic delivery,
Exp Biol Med 241 (2016) 972–979.
59 M. Najafi, H. Asadi, J. van den Dikkenberg, M.J. van Steenbergen, M.H.A.M. Fens,
W.E. Hennink, et al., Conversion of an injectable MMP–degradable hydrogel into
core-cross-linked micelles, Biomacromolecules 21 (2020) 1739–1751.
60 D. Maiti, Y. Chao, Z. Dong, X. Yi, J. He, Z. Liu, et al., Development of a
thermosensitive protein conjugated nanogel for enhanced radio-chemotherapy
of cancer, Nanoscale 10 (2018) 13976–13985.
61 N. Jommanee, C. Chanthad, K. Manokruang, Preparation of injectable hydrogels
from temperature and pH responsive grafted chitosan with tuned gelation
Please cite this article in press as: A.C. Marques et al., Drug Discovery Today (2021), https://doi.org/10.1016/j.drudis.2021.04.012
POST-SCREEN (GREY)
temperature suitable for tumor acidic environment, Carbohydr Polym 198 (2018)
486–494.
62 M. Fathi, M. Alami-Milani, M.H. Geranmayeh, J. Barar, H. Erfan-Niya, Y. Omidi,
Dual thermo-and pH-sensitive injectable hydrogels of chitosan/(poly(N-
isopropylacrylamide-co-itaconic acid)) for doxorubicin delivery in breast cancer,
Int J Biol Macromol 128 (2019) 957–964.
63 G. He, S. Chen, Y. Xu, Z. Miao, Y. Ma, H. Qian, et al., Charge reversal induced
colloidal hydrogel acts as a multi–stimuli responsive drug delivery platform for
synergistic cancer therapy, Mater Horizons 6 (2019) 711–716.
64 S. Gou, D. Xie, Y. Ma, Y. Huang, F. Dai, C. Wang, et al., Injectable, thixotropic,
and multiresponsive silk fibroin hydrogel for localized and synergistic tumor
therapy, ACS Biomater Sci Eng 6 (2020) 1052–1063.
POST-SCREEN
www.drugdiscoverytoday.com 9