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A novel nanoparticle (NP) delivery carrier for curcumin based on electrostatic 6-deoxy-6-arginine modified chitosan (DAC) assembled by γ-poly-glutamic acid (γ-PGA) was prepared. The NP structure was evaluated using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Interactions between DAC and γ-PGA were characterized using Fourier transform infrared spectroscopy (FT-IR). The sustained release kinetics of curcumin-loaded NPs was investigated in simulated gastrointestinal fluids. After exposed to heating, pH, and NaCl aqueous solution, the stabilities of both normal and curcumin-loaded NPs were determined. The results showed that NPs achieved a high encapsulation efficiency (79.5%) and loading capacity (11.31%) for curcumin. The curcumin-loaded NPs displayed a sustained release profile under simulated gastrointestinal conditions. Under certain pH (3-9), salt (0-100 mM), and temperature (30 - 60 °C) conditions, the vehicles of curcumin showed better stability. This demonstrates that NPs can be used as stable carriers for curcumin., Competing Interests: 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., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Targeting cisplatin to the sites of action and decreasing its side effects are still major challenges. Here, we introduced a polyglutamic acid-platinum(IV) prodrug nanoconjugates (γ-PGA-CA-Pt(IV)) constructed by polyglutamic acid and modified platinum(IV) prodrug to reserve the anti-tumor efficacy of cisplatin with decreased side effects. We describe the synthesis, physico-chemical characterization, and redox- and pH-sensitive releasing behavior of the nanoconjugate. In vitro studies revealed that, when incubated with glutathione in advance, the γ-PGA-CA-Pt(IV) nanoconjugate induced significant apoptosis in human breast carcinoma MCF-7 cells. From in vivo antitumor efficacy evaluation, the γ-PGA-CA-Pt(IV) nanoconjugate obviously improved the survival rate of tumor-bearing mice with inhibition of the tumor growth compared with cisplatin. Meanwhile, the nanoconjugates showed remarkable improved safety profile than the free cisplatin., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Some diseases could be treated by Tanshinone IIA (TA), which is an isolated component from the Chinese medicinal herb Tanshen (Salvia miltiorrhiza). However, the poor water solubility and low oral bioavailability of TA limited its clinical application. In this paper, TA was encapsulated by water - soluble chitosan/poly - γ - glutamic acid (WCS-γ-PGA) to improve its dissolution and oral bioavailability. The in vitro dissolution and in vivo metabolism of the encapsulated composite in rats were employed to evaluate the efficiency of the improvement. FTIR spectroscopy was applied to confirm the validity of encapsulation for TA by WCS-γ-PGA. The study's results showed that the optimal ratio of TA to drug carrier (WCS + γ-PGA) was 1:5.5 in weight with a reaction time of 1 h at room temperature for the encapsulation. The proper concentrations for WCS and TA in preparing the encapsulated composite using γ-PGA 0.125 mg ml -1 were 6 mg ml -1 and 1 mg ml -1 , respectively; The encapsulation efficiency and drug loading efficiency of WCS-γ-PGA-TA composite were (93.99 ± 2.20)% and (10.73 ± 0.75)%, respectively. The cumulative release of TA from the WCS-γ-PGA-TA encapsulated composite reached to 81% within 60 min, which was 5.56 times of that of the original TA in vitro dissolution. The peak concentration C max of TA from the encapsulated composite in rat blood as measured by an ultracentrifugation test of an intra - gastric administration was 4.43 times that of the original TA concentration, and the area under the drug-time curve AUC (0-t) and AUC (0-∞) (p<0.01) of the WCS-γ-PGA-TA encapsulated composite were 4.56 and 4.20 times that of the original TA, respectively. It indicated that the encapsulation of TA with WCS-γ-PGA improved its solubility and bioavailability significantly.

Pulmonary delivery is increasingly seen as an attractive, non-invasive route for the delivery of forthcoming protein therapeutics. In this context, here we describe protein complexes with a new 'complexing excipient' - vitamin B 12 -targeted poly(ethylene glycol)-block-poly(glutamic acid) copolymers. These form complexes in sub-200nm size with a model protein, suitable for cellular targeting and intracellular delivery. Initially we confirmed expression of vitamin B 12 -internalization receptor (CD320) by Calu-3 cells of the in vitro lung epithelial model used, and demonstrated enhanced B 12 receptor-mediated cellular internalization of B 12 -targeted complexes, relative to non-targeted counterparts or protein alone. To develop an inhalation formulation, the protein complexes were spray dried adopting a standard protocol into powders with aerodynamic diameter within the suitable range for lower airway deposition. The cellular internalization of targeted complexes from dry powders applied directly to Calu-3 model was found to be 2-3 fold higher compared to non-targeted complexes. The copolymer complexes show no complement activation, and in vivo lung tolerance studies demonstrated that repeated administration of formulated dry powders over a 3 week period in healthy BALB/c mice induced no significant toxicity or indications of lung inflammation, as assessed by cell population count and quantification of IL-1β, IL-6, and TNF-α pro-inflammatory markers. Importantly, the in vivo data appear to suggest that B 12 -targeted polymer complexes administered as dry powder enhance lung retention of their protein payload, relative to protein alone and non-targeted counterparts. Taken together, our data illustrate the potential developability of novel B 12 -targeted poly(ethylene glycol)-poly(glutamic acid) copolymers as excipients suitable to be formulated into a dry powder product for the inhalation delivery of proteins, with no significant lung toxicity, and with enhanced protein retention at their in vivo target tissue., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Constructing intracellular degradable drug delivery vehicles is critical to fully exert the function of loaded drugs. Considering the poly (amino acid) is sensitively degradable to acid and enzyme which indwell in the mature lysosome, we here presented the poly(amino acid) capsules constructed by the synthetic poly(amino acid), (poly-glutamic acid, PGA and poly-ornithine, POR). The fabrication of Dox loaded poly (amino acid) capsules was demonstrated, and was thoroughly characterized by various techniques, including Zetasizer, SEM, TEM, fluorescent microscopy, and confocal laser scan microscopy. By controlling fabrication process, we tuned the carriers with different physical properties (charges and stiffness). Then, we thoroughly investigated the effects of these properties on the intracellular uptake and anti-cancer abilities of various carriers@Dox. In addition, the degradability of poly(amino acid) capsules was studied to reveal the release profiles of the carriers with or without templates from the side aspect. We found the positively charged and stiffer carriers mainly contributed to the cellular uptake process and amount, while both the uptake amount and degradability of the endocytosed carriers@Dox played a critical role on the cytotoxicity. We believe the findings here could pave the way for designing poly(amino acid) capsules or other degradable polymers based on poly(amino acid) as the drug delivery vehicles., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Recently, novel approaches for the delivery of therapeutic antibodies have attracted much attention, especially sustained release formulations. However, sustained release formulations capable of carrying a high antibody load remain a challenge for practical use. In this study, a novel injectable hydrogel composed of maleimide-modified γ-polyglutamic acid (γ-PGA-MA) and thiol end-functionalized 4-arm poly(ethylene glycol) (4-arm PEG-SH) was developed for the subcutaneous delivery of trastuzumab. γ-PGA-MA and 4-arm PEG-SH formed a hydrogel through thiol-maleimide reactions, which had shear-thinning properties and reversible rheological behaviors. Moreover, a high content of trastuzumab (>100 mg/mL) could be loaded into this hydrogel, and trastuzumab demonstrated a sustained release over several weeks through electrostatic attraction. In addition, trastuzumab released from the hydrogel had adequate stability in terms of its structural integrity, binding bioactivity, and antiproliferative effect on BT-474 cells. Pharmacokinetic studies demonstrated that trastuzumab-loaded hydrogel (Her-hydrogel-10, composed of 1.5% γ-PGA-MA, 1.5% 4-arm PEG-SH, and 10 mg/mL trastuzumab) and trastuzumab/Zn-loaded hydrogel (Her/Zn-hydrogel-10, composed of 1.5% γ-PGA-MA, 1.5% 4-arm PEG-SH, 5 mM ZnCl 2 , and 10 mg/mL trastuzumab) could lower the maximum plasma concentration (C max ) than the trastuzumab solution. Furthermore, Her/Zn-hydrogel-10 was better able to release trastuzumab in a controlled manner, which was ascribed to electrostatic attraction and formation of trastuzumab/Zn nanocomplexes. In a BT-474 xenograft tumor model, Her-hydrogel-10 had a similar tumor growth-inhibitory effect as that of the trastuzumab solution. By contrast, Her/Zn-hydrogel-10 exhibited a superior tumor growth-inhibitory capability due to the functionality of Zn. This study demonstrated that this hydrogel has potential as a carrier for the local and systemic delivery of proteins and antibodies. STATEMENT OF SIGNIFICANCE: Recently, novel sustained-release formulations of therapeutic antibodies have attracted much attention. However, these formulations should be able to carry a high antibody load owing to the required high dose, and these formulations remain a challenge for practical use. In this study, a novel injectable chemically cross-linked hydrogel was developed for the subcutaneous delivery of trastuzumab. This novel hydrogel possessed ideal characteristics of loading high content of trastuzumab (>100 mg/mL), sustained release of trastuzumab over several weeks, and maintaining adequate stability of trastuzumab. In vivo studies demonstrated that a trastuzumab-loaded hydrogel possessed the ability of controlled release of trastuzumab and maintained antitumor efficacy same as that of trastuzumab. These results implied that a γ-PGA-MA and 4-arm PEG-SH-based hydrogel has great potential in serving as a carrier for the local or systemic delivery of therapeutic proteins or antibodies., (Copyright © 2019 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Colorectal cancer (CRC) is a major malignancy characterized by a high metastasis rate. Systematic chemotherapy is important for patients with advanced CRC. However, many limitations (e.g., side effects to normal organs, shorter circulation time, and unsatisfactory tumor inhibition results) of traditional chemotherapy restrict its further application. Thus, it is necessary to find a method to overcome these challenges and improve the efficacy of CRC treatment. In this study, 20(S)-ginsenoside (Rg3) co-loaded poly(ethylene glycol)-block-poly(L-glutamic acid-co-L-phenylalanine) (mPEG-b-P(Glu-co-Phe)) nanoparticles (Rg3-NPs) were prepared. mPEG-b-P(Glu-co-Phe)-based drug delivery systems are pH sensitive that can target cancer cells and circulate for longer in blood. Rg3 could be released rapidly from the nanoparticles within tumor cells. A subcutaneous colon cancer mouse model was developed to evaluate the anticancer efficiency of the Rg3-NPs. The in vivo study indicated that the Rg3-NPs could significantly inhibit tumor proliferation by decreasing the expressions of proliferating cell nuclear antigen, resulting in tumor apoptosis through the increased expressions of caspase-3. Our study demonstrated the marked potential of the Rg3-NPs to treat CRC.

Polysaccharides represent a versatile class of building blocks that are used in macromolecular design. By choosing the appropriate saccharide block, various physico-chemical and biological properties can be introduced both at the level of the polymer chains and the resulting self-assembled nanostructures. Here, we synthetized amphiphilic diblock copolymers combining a hydrophobic and helical poly(γ-benzyl-L-glutamate) PBLG and two polysaccharides, namely hyaluronic acid (HA) and laminarin (LAM). The copolymers could self-assemble to form particles in water by nanoprecipitation. In addition, hybrid particles containing both HA and LAM in different ratios were obtained by co-nanoprecipitation of the two copolymers. By controlling the self-assembly process, five particle samples with different morphologies and compositions were developed. The interaction between the particles and biologically relevant proteins for HA and LAM, namely CD44 and Dectin-1 respectively, was evaluated by surface plasmon resonance (SPR). We demonstrated that the particle-protein interaction could be modulated by the particle structure and composition. It is therefore suggested that this method based on nanoprecipitation is a practical and versatile way to obtain particles with controllable interactions with proteins, hence with the appropriate biological properties for biomedical applications such as drug delivery.

Non-muscle invasive bladder cancer is one of the most frequent forms of cancer. Commonly, the superficial tumor is removed by a minimal invasive surgery, mostly followed by the intravesical instillation of cytostatic agents. Although the initial treatment is usually successful, the recurrence rate is extraordinary high, which might be grounded in limiting factors during instillation, such as the high resistance of the bladder wall and the constant dilution with the permanently produced urine reducing the probability of interaction between the drug and the target cell. To overcome these limitations, we introduce a lectin-mediated drug delivery system consisting of wheat germ agglutinin as targeter, poly-l-glutamic acid as backbone and the active pharmaceutical ingredient (API) doxorubicin that should prolong the residence time in the bladder cavity. The optimized WGA-PGA-Doxo conjugate revealed an API load of 81 molecules per mole backbone and convinced in urothelial cell monolayer studies with an up to 56-fold higher cell-binding potential than pure doxorubicin. Additionally, the highly substituted drug delivery system not only internalized into urothelial cells, but also inhibited cell viability up to 99%. The combination of the high drug load with the promising cell-associative properties, which also revealed a higher affinity to malignant than to healthy urothelial cells, and the proven cytotoxic potential might lead to an improved efficacy of intravesical chemotherapy., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Podophyllotoxin (PPT) is a chemotherapeutic agent which has shown significant activity against P-glycoprotein (P-gp) mediated multi drug resistant cancer cells. However, because of the poor aqueous solubility and high toxicity, PPT cannot be used in clinical cancer therapy. In order to enhance the efficiency and reduce side effect of PPT, a polypeptide based PPT conjugate PLG-g-mPEG-PPT was developed and used for the treatment of multi drug resistant breast cancer. The PLG-g-mPEG-PPT was prepared by conjugating PPT to poly(l-glutamic acid)-g-methoxy poly(ethylene glycol) (PLG-g-mPEG) via ester bonds. The PPT conjugates self-assembled into nanoparticles with average sizes about 100 nm in aqueous solution. Western blotting assay showed that the PLG-g-mPEG-PPT could effectively inhibit the expression of P-gp in the multiple drug resistant MCF-7/ADR cells. In vitro cytotoxicity assay indicated that the resistance index (RI) values of PLG-g-mPEG-PPT on different drug-resistant cancer cell lines exhibited 57-270 folds reduction than of traditional microtubule inhibitor chemotherapeutic drug PTX or DTX. Hemolysis assay demonstrated that the conjugation greatly decreased the hemolytic activity of free PPT. Maximum tolerated dose (MTD) of PLG-g-mPEG-PPT increased greatly (13.3 folds) as compared to that of free PPT. In vivo study showed that the PLG-g-mPEG-PPT conjugate remarkably enhanced the antitumor efficacy against MCF-7/ADR xenograft tumors with a tumor suppression rate (TSR) of 82.5%, displayed significantly improved anticancer efficacy as compared to free PPT (TSR = 37.1%) with minimal toxicity when both of the two formulations were used in MTD., Statement of Significance: The development of multiple drug resistance (MDR) of cancer cells is the main cause of chemotherapy failure. The over-expression of P-glycoprotein (P-gp) has been recognized to be the most important cause of MDR in cancer. Podophyllotoxin (PPT) is a chemotherapeutic agent which has shown strong activity against P-gp mediated multidrug resistant cancer cells by simultaneously inhibiting the over-expression of P-gp and the growth of cancer cells. However, PPT can not be used in clinical cancer treatment due to its poor aqueous solubility and high toxicity. Herein, we developed a polypeptide based PPT conjugate PLG-g-mPEG-PPT by conjugating PPT to poly(l-glutamic acid)-g-methoxy poly(ethylene glycol). The PLG-g-mPEG-PPT shows significantly decreased hemolytic activity, greatly improved maximum tolerated dose and remarkably enhanced antitumor efficacy against MCF-7/ADR xenograft tumors as compared to free PPT., (Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Poly (γ-glutamic acid) (PGA) is a natural biomaterial with numerous good properties such as non-toxicity, non-immunogenicity, and water holding. So, various forms of PGA-based materials have been developed for bio-applications, including sheet, cross-linked hydrogel, and nanoparticle except injectable hydrogel type. Considering inherent advantages of injectable system, injectable hydrogel based on PGA can broaden the bio-application range of PGA. In this study, a PGA-based injectable hydrogel system was prepared by physically mixing it with a small amount of chitosan with two different water solubility and molecular weight. The prepared hydrogel system showed a thermo-sensitivity through sol-gel transition behavior in the body temperature change. The mechanical properties and in vitro stability could be modulated by varying the ratio of two types of chitosan. In vitro protein (human bFGF) delivery using this injectable formulation showed a sustained release for ∼2 weeks while preserving its bioactivity. In addition, the in situ formation of this PGA-based hydrogel formulation upon subcutaneous injection was demonstrated in vivo. The hydrogel formed in vivo also showed a long term (∼2 weeks) stability without noticeable inflammatory response. Therefore, the present formulation can be applied as a promising injectable hydrogel system for tissue engineering or drug delivery., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: The objective of this study was to survey the therapeutic function of curcumin-encapsulated poly(gamma-benzyl l-glutamate)-poly(ethylene glycol)-poly(gammabenzyl l-glutamate) (PBLG-PEG-PBLG) (P) on diabetic cardiomyopathy (DCM) via cross regulation effect of calcium-sensing receptor (CaSR) and endogenous cystathionine-γ-lyase (CSE)/hydrogen sulfide (H 2 S)., Methods: Diabetic rats were preconditioned with 20 mg/kg curcumin or curcumin/P complex continuously for 8 weeks. The blood and myocardiums were collected, the level of serum H 2 S was observed, and the [Ca 2+ ] i content was measured in myocardial cells, and hematoxylin-eosin, CaSR, CSE, and calmodulin (CaM) expression were detected., Results: Both curcumin and curcumin/P pretreatment alleviated pathological morphological damage of myocardium, increased H 2 S and [Ca 2+ ] i levels, and upregulated the expression of CaSR, CSE, and CaM as compared to DCM group, while curcumin/P remarkably augmented this effect., Conclusion: PBLG-PEG-PBLG could improve water-solubility and bioactivity of curcumin and curcumin/PBLG-PEG-PBLG significantly alleviated diabetic cardiomyopathy., Competing Interests: Disclosure The authors report no conflicts of interest in this study.

Polymeric nanoparticles (NPs) represent an effective platform for drug delivery systems, albeit with various limitations including low drug loading capacity, cytotoxicity and specificity. NPs composed of the negatively charged Polypeptide, poly gamma glutamic acid (γ-PGA) and a designed amphiphilic and cationic β-sheet Peptide (denoted PoP-NPs) loaded with the drug lonidamine (LND), denoted LND-PoP-NPs were previously used in our lab to successfully target the mitochondria when coated with the peptide (LND-mPoP-NPs). In this study, we improved the drug capacity of the LND-mPoP-NPs in addition to lowering non-specific toxicity associated with the drug deficient mPoP-NPs. LND concentrations in LND-mPoP-NPs were increased (h-LND-mPoP-NPs) and the peptide coating concentration was decreased. The new h-LND-mPoP-NPs formulation shows the ability to carry the drug to the proximity of the mitochondria despite the NP's negative zeta potential., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Injectable hydrogels have great potential in minimally invasive delivery. In this work, novel injectable hydrogels were prepared via self-crosslinking of aldehyde hyaluronic acid (HA-CHO) and hydrazide-modified poly (γ-glutamic acid) (γ-PGA-ADH) for proteins delivery. The HA/γ-PGA hydrogels could be formed in situ as fast as 9s with high swelling ratios. Rheological properties illustrated a wide processing range and good mechanical properties, which were reflected by broad linear viscoelastic region and higher threshold shear stress (σ c ) and storage modulus (G'). Meanwhile, the gelation time, swelling ratio, rheological properties, as well as the protein release behavior could be modulated conveniently. Bovine serum albumin (BSA) was designed as a model drug to study the release behavior. We found that the release mechanisms were either diffusion or Case-II relaxation depending on the different hydrogel components. The HA/γ-PGA hydrogels also showed good biocompatibility. Therefore, the HA/γ-PGA hydrogels have great potential as promising injectable biomaterials for controlled protein delivery., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Computational techniques are useful to predict interaction models and molecular properties for the design of drug delivery systems, such as dendrimers. This work evaluated the impact of surface modifications of mannosamine-conjugated multifunctional poly(glutamic acid) (PG)-dendrimers as nanocarriers of the tumour associated antigens (TAA) MART-1, gp100:44 and gp100:209. Molecular dynamics simulations and docking studies were performed. Nitrobenzoxadiazole (NBD)-PG-G4-dendrimer displayed 64 carboxylic groups, however, the Frontier Molecular Orbital Theory study evidenced that only 32 of those were available to form covalent bonds. When the number of mannosamines conjugated to dendrimer was increased from 16 to 32, the dendrimer interacted with the receptor with higher affinity. However, 16 mannosamines-NBD-PG-G4-dendrimer was chosen to conjugate TAA for added functionality as no carboxylic end groups were available for further conjugation in the 32 mannosamines-dendrimer. Docking results showed that the majority of TAA-conjugated NBD-PG-G4-dendrimer demonstrated a favourable interaction with mannosamine binding site on mannose receptor, thus constituting a promising tool for TAA targeted delivery. Our in silico approach effectively narrows down the selection of the best candidates for the synthesis of functionalised PG-dendrimers with desired functionalities. These results will significantly reduce the time and efforts required to experimentally synthesise modified dendrimers for optimal antigen delivery.

In this study, nano-doped calcium phosphate cement delivery systems (poly (γ-glutamic acid)/β-tricalcium phosphate/calcium phosphate ceramics and nano (γ-glutamic acid)/β-tricalcium phosphate/calcium phosphate ceramic) were fabricated, and low doses (10 µg/g) of two growth factors, insulin-like growth factor-1 and bone morphogenetic protein-2, were encapsulated then sequentially released. We characterized the delivery systems using Fourier transform infrared spectroscopy and X-ray diffraction and measured washout resistance and compressive strength, and thus optimized the most appropriate proportioning of delivery systems for the two growth factors. One of the growth factors was absorbed by the nano-poly (γ-glutamic acid)/β-tricalcium phosphate, which was then mixed into the calcium phosphate ceramic solid phase to create a new solid phase calcium phosphate ceramic. Nano-poly (γ-glutamic acid)/β-tricalcium phosphate/calcium phosphate ceramic carriers were then prepared by blending the new calcium phosphate ceramic solid phase powder with a solution of the remaining growth factor. The effects of different release patterns (studying sequential behavior) of insulin-like growth factor-1 and bone morphogenetic protein-2 on osteogenic proliferation and differentiation of the MC3t3-E1 mouse osteoblast cell were investigated. This combinational delivery system provided a controlled release of the two growth factors, in which nano-doping significantly affected their release kinetics. The incorporation of dual growth factors could potentially stimulate bone healing and promoting bone ingrowth processes at a low dose.

Diabetic cerebral infarction is with poorer prognosis and high rates of mortality. Ginsenoside Rg1 (Rg1) has a wide variety of therapeutic values for central nervous system (CNS) diseases for the neuron protective effects. However, the blood-brain barrier (BBB) restricts Rg1 in reaching the CNS. In this study, we investigated the therapeutic effects of Rg1 nanoparticle (PHRO, fabricated with γ-PGA, L-PAE (H), Rg1, and OX26 antibody), targeting transferrin receptor, on the diabetes rats complicated with diabetic cerebral infarction in vitro and in vivo. Dynamic light scattering analysis shows the average particle size of PHRO was 79±18 nm and the polydispersity index =0.18. The transmission electron microscope images showed that all NPs were spherical in shape with diameters of 89±23 nm. PHRO released Rg1 with sustained release manner and could promote the migration of cerebrovascular endothelial cells and tube formation and even penetrated the BBB in vitro. PHRO could penetrate the BBB with high concentration in brain tissue to reduce the cerebral infarction volume and promote neuronal recovery in vivo. PHRO was promising to be a clinical treatment of diabetes mellitus with cerebral infarction., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

The conventional chemotherapeutics could not be traced in vivo and provide timely feedback on the clinical effectiveness of drugs. In this study, poly(L-γ-glutamyl-glutamine)-paclitaxel (PGG-PTX), as a model polymer, was chemically conjugated with Gd-DTPA (Gd-diethylenetriaminepentaacetic acid), a T 1 -contrast agent of MRI, to prepare a Gd-DTPA-conjugated PGG-PTX (PGG-PTX-DTPA-Gd) delivery system used for tumor theranostics. PGG-PTX-DTPA-Gd can be self-assembled to NPs in water with a z-average hydrodynamic diameter about 35.9 nm. The 3 T MRI results confirmed that the relaxivity of PGG-PTX-DTPA-Gd NPs (r 1  = 18.98 mM -1 S -1 ) was increased nearly 4.9 times compared with that of free Gd-DTPA (r 1  = 3.87 mM -1 S -1 ). The in vivo fluorescence imaging results showed that PGG-PTX-DTPA-Gd NPs could be accumulated in the tumor tissue of NCI-H460 lung cancer animal model by EPR effect, which was similar to PGG-PTX NPs. The MRI results showed that compared with free Gd-DTPA, PGG-PTX-DTPA-Gd NPs showed significantly enhanced and prolonged signal intensity in tumor tissue, which should be attributed to the increased relaxivity and tumor accumulation. PGG-PTX-DTPA-Gd NPs also showed effective antitumor effect in vivo. These results indicated that PGG-PTX-DTPA-Gd NPs are an effective delivery system for tumor theranostics, and should have a potential value in personalized treatment of tumor.

Context: Poly-l-glutamic acid (PGA) is an anionic polymer with a large number of carboxyl groups that can interact electrostatically with cationic drugs such as doxorubicin (DOX)., Objective: For stable encapsulation of DOX into liposomes, we prepared triethylamine (TEA)-PGA-liposomes using PGA as an internal trapping agent., Methods: We prepared TEA-PGA-liposomes by remote loading of DOX with a TEA gradient into preformed liposomes prepared with 1, 2, or 4 mg/mL PGA (molecular weights 4800, 9800, and 20 500), and evaluated their biodistribution and antitumor effects on Lewis lung carcinoma (LLC) tumor-bearing mice., Results: TEA-PGA-liposomes using the higher the molecular weight or concentration of PGA showed a slower release of DOX from the liposomes. TEA-PGA-liposomes prepared with a high concentration of PGA could enhance DOX accumulation in tumors and prolonged DOX circulation in the serum, indicating that DOX may be retained stably in the liposomal interior by interaction with PGA. Furthermore, injection of TEA-PGA-liposomes prepared with 4 mg/mL of PGA 9800 or 2 mg/mL PGA 20500 strongly inhibited tumor growth in LLC tumor-bearing mice., Conclusions: PGA may be a potential trapping agent for liposomal DOX for tumor drug delivery.

Boron neutron capture therapy is a promising tumor treatment method, though its wide application has been limited due to the poor tumor selectivity and intracellular delivery of 10 B-compounds. Here, block copolymer-boron cluster conjugate based on the clinically used sodium borocaptate (BSH) and poly(ethylene glycol)-b-poly(glutamic acid) copolymer have been developed for effectively penetrating tumor tissues and homogeneously delivering the boron clusters into cancer cells towards safe and efficient boron neutron capture therapy. The PEGylated block copolymer-boron cluster (BSH) conjugate has demonstrated significant higher cellular uptake and tumor accumulation when compared to the non-PEGylated formulations and BSH. Moreover, the enhanced delivery to tumors of the conjugates, as well as their superior intratumoral penetration, which facilitated reaching the intracellular space of most cells in tumors, allowed the effective ablation of tumors after neutron irradiation., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Injectable and stimuli-responsive hydrogels have attracted attention in molecular imaging and drug delivery because encapsulated diagnostic or therapeutic components in the hydrogel can be used to image or change the microenvironment of the injection site by controlling various stimuli such as enzymes, temperature, pH, and photonic energy. In this study, we developed a novel injectable and photoresponsive composite hydrogel composed of anticancer drugs, imaging contrast agents, bio-derived collagen, and multifaceted anionic polypeptide, poly (γ-glutamic acid) (γ-PGA). By the introduction of γ-PGA, the intrinsic temperature-dependent phase transition behavior of collagen was modified to a low viscous sol state at room temperature and nonflowing gel state around body temperature. The modified temperature-dependent phase transition behavior of collagen/γ-PGA hydrogels was also evaluated after loading of near-infrared (NIR) fluorophore, indocyanine green (ICG), which could transform absorbed NIR photonic energy into thermal energy. By taking advantage of the abundant carboxylate groups in γ-PGA, cationic-charged doxorubicin (Dox) and hydrophobic MnFe 2 O 4 magnetic nanoparticles were also incorporated successfully into the collagen/γ-PGA hydrogels. By illumination of NIR light on the collagen/γ-PGA/Dox/ICG/MnFe 2 O 4 hydrogels, the release kinetics of Dox and magnetic relaxation of MnFe 2 O 4 nanoparticles could be modulated. The experimental results suggest that the novel injectable and NIR-responsive collagen/γ-PGA hydrogels developed in this study can be used as a theranostic platform after loading of various molecular imaging probes and therapeutic components., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Neural cell adhesion molecule (NCAM) expression is known to be associated with an aggressive biological behavior, increased metastatic capacity and expression of stem-cell markers in several tumor types. NCAM was also found to be expressed on tumor endothelial cells while forming new capillary-like tubes, but not on normal endothelial cells. An NCAM-targeted polymer-drug conjugate can be used both to target tumors expressing high levels of NCAM as well as the angiogenic vessels and cancer stem cells populations characterized by NCAM expression within tumors. Here, we describe the design, synthesis, physico-chemical characterization and the biological evaluation of an NCAM-targeted conjugate of polyglutamic acid with paclitaxel that was developed and evaluated on neuroblastoma, a high NCAM-expressing tumor. This conjugate inhibited tumor growth to a higher extent compared to the control conjugates and was less toxic than free paclitaxel. The dose of the conjugate could be increased at least twice than the maximum tolerated dose of paclitaxel to achieve better activity without aggravating toxicity. This work presents evidence that NCAM targeting can highly increase the efficacy of nanomedicines in the appropriate tumor models., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Stimuli-responsive anticancer formulations can promote drug release and activation within the target tumour, facilitate cellular uptake, as well as improve the therapeutic efficacy of drugs and reduce off-target effects. In the present work, indocyanine green (ICG)-containing polyglutamate (PGA) nanoparticles were developed and characterized. Digestion of nanoparticles with cathepsin B, a matrix metalloproteinase overexpressed in the microenvironment of advanced tumours, decreased particle size and increased ICG cellular uptake. Incorporation of ICG in PGA nanoparticles provided the NIR-absorbing agent with time-dependent altered optical properties in the presence of cathepsin B. Having minimal dark toxicity, the formulation exhibited significant cytotoxicity upon NIR exposure. Combined use of the formulation with saporin, a ribosome-inactivating protein, resulted in synergistically enhanced cytotoxicity attributed to the photo-induced release of saporin from endo/lysosomes. The results suggest that this therapeutic approach can offer significant therapeutic benefit in the treatment of superficial malignancies, such as head and neck tumours.

In the past decade, poly-γ-glutamic acid (γ-PGA)-based micro/nanoparticles have garnered remarkable attention as antimicrobial agents and for drug delivery, owing to their controlled and sustained-release properties, low toxicity, as well as biocompatibility with tissue and cells. γ-PGA is a naturally occurring biopolymer produced by several gram-positive bacteria that, due to its biodegradable, non-toxic and non-immunogenic properties, has been used successfully in the medical, food and wastewater industries. Moreover, its carboxylic group on the side chains can offer an attachment point to conjugate antimicrobial and various therapeutic agents, or to chemically modify the solubility of the biopolymer. The unique characteristics of γ-PGA have a promising future for medical and pharmaceutical applications. In the present review, the structure, properties and micro/nanoparticle preparation methods of γ-PGA and its derivatives are covered. Also, we have highlighted the impact of micro/nanoencapsulation or immobilisation of antimicrobial agents and various disease-related drugs on biodegradable γ-PGA micro/nanoparticles.

Background and Purpose: Poly-l-glutamic acid (PG) has been used widely as a carrier to deliver anticancer chemotherapeutics. This study evaluates PG as a selective renal drug carrier., Experimental Approach: 3 H-deoxycytidine-labeled PGs (17 or 41 kDa) and 3 H-deoxycytidine were administered intravenously to normal rats and streptozotocin-induced diabetic rats. The biodistribution of these compounds was determined over 24 h. Accumulation of PG in normal kidneys was also tracked using 5-(aminoacetamido) fluorescein (fluoresceinyl glycine amide)-labeled PG (PG-AF). To evaluate the potential of PGs in ferrying renal protective anti-oxidative stress compounds, the model drug 4-(2-aminoethyl)benzenesulfonyl fluoride hydrochloride (AEBSF) was conjugated to 41 kDa PG to form PG-AEBSF. PG-AEBSF was then characterized and evaluated for intracellular anti-oxidative stress efficacy (relative to free AEBSF)., Results: In the normal rat kidneys, 17 kDa radiolabeled PG (PG-Tr) presents a 7-fold higher, while 41 kDa PG-Tr shows a 15-fold higher renal accumulation than the free radiolabel after 24 h post injection. The accumulation of PG-AF was primarily found in the renal tubular tissues at 2 and 6 h after an intravenous administration. In the diabetic (oxidative stress-induced) kidneys, 41 kDa PG-Tr showed the greatest renal accumulation of 8-fold higher than the free compound 24 h post dose. Meanwhile, the synthesized PG-AEBSF was found to inhibit intracellular nicotinamide adenine dinucleotide phosphate oxidase (a reactive oxygen species generator) at an efficiency that is comparable to that of free AEBSF. This indicates the preservation of the anti-oxidative stress properties of AEBSF in the conjugated state., Conclusion/implications: The favorable accumulation property of 41 kDa PG in normal and oxidative stress-induced kidneys, along with its capabilities in conserving the pharmacological properties of the conjugated renal protective drugs, supports its role as a potential renal targeting drug carrier., Competing Interests: The authors report no conflicts of interest in this work.

In this research, we synthesized bioderived poly(amino acid) hydrogel particles that showed pH-dependent membrane-disrupting properties and controlled cytosolic delivery of antitumor drugs. Poly(γ-glutamic acid) (γ-PGA) that has been produced extensively using bacteria, especially those of Bacillus subtilis species, was modified with cholesterol (γ-PGA/Chol), and the γ-PGA/Chol conjugates were used to form polymeric nanoparticles the size of 21.0±1.1 nm in aqueous solution. When the polymeric nanoparticles were mixed with doxorubicin (Dox), raspberry-like hydrogel particles (RBHPs) were formed by the electrostatic interaction between the cationically charged Dox and the anionically charged nanoparticles. The average size and surface charge of the RBHPs in aqueous solution were 444.9±122.5 nm and -56.44 mV, respectively. The loaded amount of Dox was approximately 63.9 μg/mg of RBHPs. The RBHPs showed controlled drug release behavior in both in vitro and ex vivo cell-based experiments. Through fluorescence microscopy and fluorescence-activated cell sorting, the cellular uptake of RBHPs into human cervical cancer cells (HeLa) was analyzed. The cytotoxic effect, evaluated by the methyl tetrazolium salt assay, was dependent on both the concentration of RBHPs and the treatment time. The pH-dependent membrane-disrupting properties of the RBHPs and the subsequent cytosolic delivery of Dox were evaluated using a standard hemolysis assay. Upon an increase in hydrophobicity at the lysosomal acidic pH, RBHPs could easily interact, penetrate cell membranes, and destabilize them. Taken together, the data suggested that RBHPs could be used as drug delivery carriers after loading with other therapeutic drugs, such as proteins or small interfering RNA for cancer therapy., Competing Interests: The authors report no conflicts of interest in this work.

Mesoporous silica nanoparticles (MSNs) are highly attractive as supports in the design of controlled delivery systems that can act as containers for the encapsulation of therapeutic agents, overcoming common issues such as poor water solubility and poor stability of some drugs and also enhancing their bioavailability. In this context, we describe herein the development of polyglutamic acid (PGA)-capped MSNs that can selectively deliver rhodamine B and doxorubicin. PGA-capped MSNs remain closed in an aqueous environment, yet they are able to deliver the cargo in the presence of pronase because of the hydrolysis of the peptide bonds in PGA. The prepared solids released less than 20% of the cargo in 1 day in water, whereas they were able to reach 90% of the maximum release of the entrapped guest in ca. 5 h in the presence of pronase. Studies of the PGA-capped nanoparticles with SK-BR-3 breast cancer cells were also undertaken. Rhodamine-loaded nanoparticles were not toxic, whereas doxorubicin-loaded nanoparticles were able to efficiently kill more than 90% of the cancer cells at a concentration of 100 μg/mL.

Unlabelled: The linear, Y-shaped, and linear-dendritic block copolymers of methoxy poly(ethylene glycol)-block-polyamidoamine-block-poly(l-glutamic acid) (MPEG-b-PAMAM-b-PGA) with one, two, four, and eight PGA arms but similar MPEG/PGA weight ratios (W/W) (named as P1PA, P2PA, P4PA and P8PA, respectively) were synthesized and comparatively investigated for doxorubicin hydrochloride (DOX) delivery. All the obtained block copolymers were highly biocompatible and could efficiently load DOX into nanoparticles (NPs) through electrostatic interaction. The NPs formed by linear (P1PA) or Y-shaped (P2PA) block copolymers and DOX were spherically shaped with smaller sizes, while the NPs formed from linear-dendritic block copolymers (P4PA and P8PA) were irregular in shape and larger in size. The P1PA/DOX and P2PA/DOX NPs exhibited better DOX protection and slower DOX release profile. However, cell cytotoxicity assays indicated that all the DOX-loaded NPs exhibited similar cytotoxicities with free DOX, indicating effective DOX release after cellular uptake. The NPs from linear and Y-shaped block copolymers greatly extended the blood circulation time, and displayed more accumulation in tumor site and less accumulation in the liver and kidney compared with the linear-dendritic counterparts. In addition, the P1PA/DOX and P2PA/DOX NPs also exhibited higher anti-tumor efficacy and less toxicity than the other DOX formulations. All these results indicated that the linear and Y-shaped MPEG-b-PAMAM-b-PGA block copolymers displayed better DOX delivery ability in anti-tumor treatment than the linear-dendritic copolymers., Statement of Significance: Polymeric NPs derived from block copolymers have emerged as effective vehicles for drug delivery. However, the majority of the researches in this field have involved simple linear block copolymers and there are very few comparative studies on the self-assembly, in vitro, and in vivo drug delivery by the block copolymers with similar composition but different architectures. In this study, a series of linear, Y-shaped, and linear-dendritic polypeptide-based block copolymers were prepared and thoroughly investigated for DOX delivery. These block polymers loaded DOX into NPs with different sizes and morphologies, and exhibited different anti-tumor capabilities both in vitro and in vivo. The results indicated that the architecture of the block copolymers played an important role in their drug delivery behaviors., (Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Polyelectrolyte layer-by-layer assembled lipid nanoparticles (NPs) were prepared to improve their stability against lipolysis in gastrointestinal tract, and efficiency of oral absorption of doxorubicin (DOX). The lipid NPs were prepared by hot melt-probe sonication method. The polyelectrolyte layer-by-layer assembled lipid NPs (DOX-NPs/CS/γ-PGA) was prepared by layer-by-layer self-assembling polyelectrolytes cationic chitosan (CS) and anionic poly (γ-glutamic acid) (γ-PGA) on the surface of lipid NPs based on electrostatic interaction. The particle size, polydispersity index (PDI) and zeta potential of lipid NPs and DOX-NPs/CS/γ-PGA were determined by dynamic light scattering (DLS). The in vitro drug release was determined in p H 1.2 HCl solution and p H 6.8 phosphate buffer solution (PBS). The stability of lipid NPs against lipolysis was evaluated in simulated gastrointestinal medium containing lipase. The cellular uptake of lipid NPs and DOX-NPs/CS/γ-PGA was evaluated in Caco-2 cell model. The pharmacokinetic of DOX after oral absorption was studied in SD rats. Results showed that the average particle size and zeta potential of DOX-NPs/CS/γ-PGA were 180.6 ± 5.4 nm and-38.53 ± 0.29 m V, respectively. The DOX-NPs/CS/γ-PGA effectively slowed down the release of DOX from nanoparticles, and decreased the lipolysis of lipid NPs in simulated gastrointestinal medium. The cell study showed that DOX-loaded lipid NPs and DOX-NPs/CS/ γ-PGA remarkably enhanced the cell uptake in comparison with DOX solution. The DOX-NPs/CS/γ-PGA significantly improved oral absorption of DOX compared with DOX-loaded lipid NPs. The C(max), t(max) were 0.76 ± 0.25 μg·m L(-1) and 0.5 h, respectively; AUC(0-24 h) was 3.02 folds and the relative bioavailability was 302.46% with DOX solution as reference. The stability of lipid NPs against lipolysis and drug release were significantly improved by layer-by-layer assembling, leading to an improved oral absorption.

To enhance the glucose sensitivity and self-regulated release of insulin, biobased capsules with glucose-responsive and competitive properties were fabricated based on poly(γ-glutamic acid) (γ-PGA) and chitosan oligosaccharide (CS) polyelectrolytes. First, poly(γ-glutamic acid)-g-3-aminophenylboronic acid) (γ-PGA-g-APBA) and galactosylated chitosan oligosaccharide (GC) were synthesized by grafting APBA and lactobionic acid (LA) to γ-PGA and CS, respectively. The (γ-PGA-g-APBA/GC)5 capsules were then prepared by layer-by-layer (LBL) assembly of γ-PGA-g-APBA and GC via electrostatic interaction. The size and morphology of the particles and capsules were investigated by DLS, SEM, and TEM. The size of the (γ-PGA-g-APBA/GC)5 capsules increased with increasing glucose concentration due to the swelling of the capsules. The capsules could be dissociated at high glucose concentration due to the breaking of the cross-linking bonds between APBA and LA by the competitive reaction of APBA with glucose. The encapsulated insulin was able to undergo self-regulated release from the capsules depending on the glucose level and APBA composition. The amount of insulin release increased with incubation in higher glucose concentration and decreased with higher APBA composition. Moreover, the on-off regulation of insulin release from the (γ-PGA-g-APBA/GC)5 capsules could be triggered with a synchronizing and variation of the external glucose concentration, whereas the capsules without the LA functional groups did not show the on-off regulated release. Furthermore, the (γ-PGA-g-APBA/GC)5 capsules are biocompatible. These (γ-PGA-g-APBA/GC)5 with good stability, glucose response, and controlled insulin delivery are expected to be used for future applications to glucose-triggered insulin delivery.

Efficient oral administration of anticancer agents requires a nanocarrier to long survive in the stomach, effectively penetrate across the small intestine, tightly retain the drug during bloodstream and quickly release drug in tumor cells. Herein a kind of dual pH-sensitive polyelectrolyte complex nanoparticles (CNPs) was developed by employing electrostatic interaction between positively charged chitosan (CS) and negative poly (L-glutamic acid) grafted polyethylene glycol-doxorubicin conjugate nanoparticles (PG-g-PEG-hyd-DOX NPs) with acid-labile hydrazone linkages. The obtained NPs and CNPs were characterized for their morphology, particle size, ζ-potential, pH-sensitivity under the simulated physiological conditions, drug release, as well as in vivo antitumor activity and biodistribution. The results indicated that CNPs can remain intact structure in pH range from 3.0 to 6.5. After detaching CS layer due to the pH-induced deprotonation with increasing pH to 7.4 in the mucus layer of the small intestine, the inner NPs would be released and effectively absorbed into blood circulation via opening the tight junctions by CS. PG-g-PEG-hyd-DOX NPs with demonstrated long-circulating properties can be accumulated in the tumor via EPR effect and dump the drug within tumor cells by acid-cleavage of hydrazone bonds between PG-g-PEG and DOX, achieving high therapeutic efficacy and low systemic toxicity. These results suggest that the design presented here, combining the functions of the gastrointestinal pH-sensitive electrostatic complex and intracellular acid-sensitive macromolecular prodrugs NPs, can sequentially overcome the biological barriers of oral anticancer drug delivery, which thus provides a promising nanomedicine platform for oral chemotherapy., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Diabetes mellitus is a high prevalence and one of the most severe and lethal diseases in the world. Insulin is commonly used to treat diabetes in order to give patients a better life condition. However, due to bioavailability problems, the most common route of insulin administration is the subcutaneous route, which may present patients compliance problems to treatment. The oral administration is thus considered the most convenient alternative to deliver insulin, but it faces important challenges. The low stability of insulin in the gastrointestinal tract and low intestinal permeation, are problems to overcome. Therefore, the encapsulation of insulin into polymer-based nanoparticles is presented as a good strategy to improve insulin oral bioavailability. In the last years, different strategies and polymers have been used to encapsulate insulin and deliver it orally. Polymers with distinct properties from natural or synthetic sources have been used to achieve this aim, and among them may be found chitosan, dextran, alginate, poly(γ-glutamic acid), hyaluronic acid, poly(lactic acid), poly(lactide-co-glycolic acid), polycaprolactone (PCL), acrylic polymers and polyallylamine. Promising studies have been developed and positive results were obtained, but there is not a polymeric-based nanoparticle system to deliver insulin orally available in the market yet. There is also a lack of long term toxicity studies about the safety of the developed carriers. Thus, the aims of this review are first to provide a deep understanding on the oral delivery of insulin and the possible routes for its uptake, and then to overview the evolution of this field in the last years of research of insulin-loaded polymer-based nanoparticles in the academic and industrial fields. Toxicity concerns of the discussed nanocarriers are also addressed., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Since the legendary 1964 article of Folkman and Long entitled "The use of silicone rubber as a carrier for prolonged drug therapy" the role of polymers in controlled drug delivery has come a long way. Today it is evident that polymers play a crucial if not the prime role in this field. The latest boost owes to the interest in drug delivery for the purpose of tissue engineering in regenerative medicine. The focus of this commentary is on a selection of general and personal observations that are characteristic for the current state of polymer therapeutics and carriers. It briefly highlights selected examples for the long march of synthetic polymer-drug conjugates from bench to bedside, comments on the ambivalence of selected polymers as inert excipients versus biological response modifiers, and on the yet unsolved dilemma of cationic polymers for the delivery of nucleic acid therapeutics. Further subjects are the complex design of multifunctional polymeric carriers including recent concepts towards functional supramolecular polymers, as well as observations on stimuli-sensitive polymers and the currently ongoing trend towards natural and naturally-derived biopolymers. The final topic is the discovery and early development of a novel type of biodegradable polyesters for parenteral use. Altogether, it is not the basic and applied research in polymer therapeutics and carriers, but the translational process that is the key hurdle to proceed towards an authoritative approval of new polymer therapeutics and carriers., (Copyright © 2015 Elsevier B.V. All rights reserved.)

There is a need to develop new and innovative polymer carriers to be used as drug delivery systems and/or imaging agents owing to the fact that there is no universal polymeric system that can be used in the treatment of all diseases. Additionally, limitations with existing systems, such as a lack of biodegradability and biocompatibility, inevitably lead to side effects and poor patient compliance. New polymer therapeutics based on amino acids are excellent candidates for drug delivery, as they do not suffer from these limitations. This article reports on a simple yet powerful methodology for the synthesis of 3-arm star-shaped polyglutamic acid with well-defined structures, precise molecular weights (MW), and low polydispersity (Đ = <1.3). These were synthesized by ring-opening polymerization (ROP) of N-carboxyanhydrides (NCA) in a divergent method from novel multifunctional initiators. Herein, their exhaustive physicochemical characterization is presented. Furthermore, preliminary in vitro evaluation in selected cell models, and exhaustive in vivo biodistribution and pharmacokinetics, highlighted the advantages of these branched systems when compared with their linear counterparts in terms of cell uptake enhancement and prolonged plasma half-life.

Despite considerable progress being made in breast cancer therapy, the complete eradication of highly aggressive triple negative breast cancer (TNBC) remains a notable challenge today. We herein report on the fabrication of novel gold nanorods (GNRs) with covalent cisplatin-polypeptide wrapping and folic acid (FA) conjugation (FA-GNR@Pt) for the targeted photothermal (PT) therapy and chemotherapy of TNBC. The FA-GNR@Pt hybrid nanoparticles are designed to integrate the photothermal conversion property of GNRs, the superior biocompatibility of polypeptide poly(l-glutamic acid) (PGA), the chemotoxicity of cisplatin, and the tumor targeting ability of FA into one single nanoplatform. In combination with localized near infrared (NIR) laser illumination, the resulting FA-GNR@Pt hybrid nanoparticles are able to significantly inhibit the growth of the TNBC tumor when administered systemically. In particular, they can extensively suppress the dissemination of TNBC cells from the primary tumor to the lung by eliminating the peripheral tumor blood vessels. Collectively, our studies demonstrate that the combined PT therapy and chemotherapy using cisplatin-loaded GNRs with FA conjugation might imply a promising strategy for targeted treatment of TNBC.

Incomplete insertion is a common problem associated with polymer microneedles (MNs) that results in a limited drug delivery efficiency and wastage of valuable medication. This paper presents a fully insertable MN system that is composed of poly-γ-glutamic acid (γ-PGA) MNs and polyvinyl alcohol (PVA)/polyvinyl pyrrolidone (PVP) supporting structures. The PVA/PVP supporting structures were designed to provide an extended length for counteracting skin deformation during insertion and mechanical strength for fully inserting the MNs into the skin. When inserted into the skin, both the supporting structures and MNs can be dissolved in the skin within 4min, thus quickly releasing the entire drug load from the MNs. To evaluate the feasibility and reproducibility of using the proposed system for treating diabetes, we administered insulin-loaded MNs to diabetic rats once daily for 2days. The results indicated that the hypoglycemic effect in the rats receiving insulin-loaded MNs was comparable to that observed in rats receiving subcutaneous insulin injections. The relative pharmacological availability and relative bioavailability of the insulin were in the range of 90-97%, indicating that the released insulin retained its pharmacological activity. We observed no significant differences in the plasma insulin concentration profiles between the first and second administrations, confirming the stability and accuracy of using the proposed MN system for insulin delivery. These results indicated that the γ-PGA MNs containing the supporting structure design enable complete and efficient delivery of encapsulated bioactive molecules and have great potential for the relatively rapid and convenient transdermal delivery of protein drugs., Statement of Significance: Incomplete insertion of microneedles largely limits drug delivery efficiency and wastage of valuable medication. To address this problem, we developed a fully insertable poly-glutamic acid microneedles with a supporting structure design to ensure complete and efficient delivery of encapsulated drugs. The supporting structures were designed to provide an extended length for counteracting skin compressive deformation during puncture and mechanical strength for fully inserting the microneedles into the skin. When inserted into the skin, both the supporting structures and microneedles can be dissolved in the skin within 4min, thus quickly releasing the entire drug load. This study demonstrated that the proposed microneedle system featuring this unique design allows more convenient and efficient self-administration of drugs into the skin., (Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Poly(γ-glutamic acid)-based targeted drug delivery system (PAMCN) targeting transmembrane metalloprotease aminopeptidase-N (APN/CD13) was fabricated and evaluated for the enhancement of targeting efficiency and cytotoxicity. The cisplatin (CDDP) loading content of PAMCN was about 36 ± 5% and PAMCN showed a sustainable release profile with a half-maximal release time (t1/2) of 23 h. The average size of PAMCN was 132 ± 18 nm determined by light scattering (LS) and 158 ± 67 nm by atomic force microscopy (AFM). Flow cytometry and fluorescence microscope analysis showed that the drug carrier (PAMN) could specifically bind to human umbilical vein endothelial cells (HUVEC). PAMCN enhanced the efficacy of CDDP to HUVEC cells with the half maximal inhibitory concentration (IC50) value decreased to 90.83 ± 33.00 μg/ml comparing with free CDDP treatment and showed less tube formation amounts (p < 0.01) than free CDDP in matrigel angiogenesis inhibition assay in vitro. In vivo toxicity experiment indicated that the survival rate of KM mice in PAMCN group was 100% and PAMCN reduced the hepatic and renal toxicity significantly compared to free CDDP group. Therefore, this novel drug delivery system presents a promising potential for antiangiogenic chemotherapy.

Among the most common nanoparticulate systems, the polymeric nanocarriers have a number of key benefits, which give a great choice of delivery platforms. Nevertheless, polymeric nanoparticles possess some limitations that include use of toxic solvents in the production process, polymer degradation, drug leakage outside the diseased tissue, and polymer cytotoxicity. The combination of polymers of biological and synthetic origin is an appealing modern strategy for the production of novel nanocarriers with unprecedented properties. Proteins' interface can play an important role in determining bioactivity and toxicity and gives perspective for future development of the polymer-based nanoparticles. The design of hybrid constructs composed of synthetic polymer and biological molecules such as proteins can be considered as a straightforward tool to integrate a broad spectrum of properties and biofunctions into a single device. This review discusses hybrid protein-synthetic polymer nanoparticles with different structures and levels in complexity and functionality, in view of their applications as drug delivery systems., (© 2015 Elsevier Inc. All rights reserved.)

Polymers are widely applied to drug delivery systems because polymers are generally excreted from the body more slowly than small molecules. Subcutaneous injection is one plausible means of administration. In this study, the in vivo behaviors of subcutaneously injected polymers, linear poly(glutamic acid) (Poly-Glu), acetylated dendrimer (Ac-den) and collagen peptide-conjugated dendrimer (CP-den), were investigated. Single photon emission computed tomography (SPECT) imaging was used to noninvasively monitor the in vivo behaviors. Diethylenetriaminepentaacetic acid (DTPA) was conjugated to these polymers, which were labeled with radioactive (111)In. These (111)In-DTPA-bearing polymers (Poly-Glu-DTPA, Ac-den-DTPA and CP-den-DTPA) and unconjugated DTPA were subcutaneously injected into tumor-bearing mice, which were subjected to SPECT imaging. These (111)In-DTPA-bearing polymers were largely retained at the injection site for at least 1 day, whereas the unconjugated DTPA was rapidly cleared from the whole body through excretion. Poly-Glu-DTPA and Ac-den-DTPA were partly accumulated in the kidney (and the liver), but the CP-den-DTPA was not. However, these (111)In-DTPA-bearing polymers were accumulated in the liver and the kidney following intravenous administration. These results indicate that the subcutaneously injected polymers did not largely gain substantial access to the systemic circulation, which is useful for a depot of drug around the injection site., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Targeted drug delivery strategies have shown great potential in solving some problems of chemotherapy, such as non-selectivity and severe side effects, thus enhancing the anti-tumor efficiency of chemotherapeutic agents. In this work, we have prepared a novel nanoparticle consisted of amphiphilic poly(γ-glutamic acid-maleimide-co-L-lactide)-1,2-dipalmitoylsn-glycero-3-phosphoethanolamine (γ-PGA-MAL-PLA-DPPE) copolymer decorated with transferrin (Tf), which can specifically deliver anti-cancer drug paclitaxel (PTX) to the tumor cells for targeting chemotherapy. These nanoparticles (NPs) have preferable particle size, high encapsulation efficiency and a pH-dependent release profile. As expected, The Tf modification mediate specific targeting to nasopharyngeal carcinoma (C666-1) cells and human cervical carcinoma (Hela) cells with the transferrin receptor (TfR) overexpressed and enhance cellular uptake of the NPs, as demonstrated by flow cytometry and confocal microscopy assays. In vitro cytotoxicity studies reveal that the NPs have excellent biocompatibility, and the presence of Tf enhance the activity of PTX to the targeted cells. All these results prove that Tf modified γ-PGA-MAL-PLA-DPPE NPs could facilitate the tumor-specific therapy. Therefore, such a targeting drug delivery system provides significant advances toward cancer therapy., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The oral route is a convenient and commonly employed way for drug delivery. However, therapeutic proteins have poor bioavailability upon oral administration due to the impermeable barrier from intestinal epithelial tight junction (TJ). Moreover, the pH of the small intestine varies among different regions of the intestinal tract where digestion and absorption occur at different levels. In this study, a tunable dual-emitting and pH-responsive nanocarrier that can alter the fluorescent color and emission intensity in response to pH changes and can trigger the opening of intestinal epithelial TJ at different levels were developed from chitosan-N-arginine and poly(γ-glutamic acid)-taurine conjugates. As pH increased from 6.0 to 8.0, the binding affinity of the oppositely charged polyions decreased, whereas the ratio of the intensity of the donor-to-acceptor emission intensity (ID/IA) increased by 27-fold. The fluorescent and pH-responsive nanocarrier was able to monitor the pH change of intestinal environment and to control the release of an anti-angiogenic protein in response to the pH gradient. The nanocarrier triggered the opening of intestinal epithelial TJ and consequently enhanced the permeation of the released protein through the intestinal epithelial barrier model (Caco-2 cell monolayer) to inhibit tube formation of human umbilical vein endothelial cells.

The bioactive polymer poly(L-glutamic acid)n-b-poly(D, L-lactic acid)m was synthesized and used to form doxorubicin-loaded hybrid polymeric micelles to treat melanoma. These polymers exhibited pH-responsive changes in conformation, which controlled the diverse functionalities of the micelles. During circulation, poly(L-glutamic acid)n-b-poly(D, L-lactic acid)m protected Tat peptides on the micelles from proteolysis. Under tumor-acidic conditions, polymers with shorter poly(l-glutamic acid) blocks underwent a conformational change to form channels that accelerated the release of doxorubicin. The conformational change also exposed the Tat peptides to tumor cells, thereby promoting cellular internalization of the micelles. Enhanced cellular uptake of the micelles induced significant apoptosis of A375 melanoma cells in tumor-acidic conditions. In vivo studies demonstrated that the micelles with shorter poly(L-glutamic acid) blocks could effectively accumulate in tumor tissues, suppress tumor growth and help maintain the body weight of tumor-bearing mice. However, micelles with longer poly(l-glutamic acid) blocks did not undergo a conformational change under acidic conditions and performed poorly in both in vitro and in vivo evaluations. Our work provides a strategy for applying bioactive polymers to the rational construction of pH-responsive delivery systems for solid tumors and lends insight into possible conformational effects on the bioactivity of drug carriers., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Hydroxyapatite (HAP), a highly specific component of bone tissue, is the main target in order to impart osteotropicity. Bone targeted nanoparticles can increase the strength of the interaction with HAP through multivalency and thus constitute a valuable strategy in the therapeutics of skeletal diseases. PBLG10k-b-PEG6k-alendronate nanoparticles (~ 75 nm) were prepared by a simple nanoprecipitation method. The calcium affinity (KCa(+2)=1.8 × 10(4)M(-1)) of these nanoparticles was evaluated using isothermal titration calorimetry. The multivalent interaction with HAP surfaces (KHAP) was studied by fluorescence and was estimated to be 1.1 × 10(10)M(-1), which is more than 4000 times stronger than the reported monovalent interaction between alendronate and HAP surfaces. Molecular modeling suggests that the number of binding sites available at the HAP surface is in large excess than what is required for the whole surface coverage by alendronate decorated nanoparticles. The lower calcium affinity of these nanoparticles than for HAP allows calcium bound nanoparticles to interact with HAP, which yields a deeper understanding of bone targeted carriers and could potentially improve their bone targeting properties., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Highly viscous mucus poses a big challenge for the delivery of particulates carrying therapeutics to patients with cystic fibrosis. In this study, surface modifying DNase I loaded particles using different excipients to achieve better lung deposition, higher enzyme stability or better biological activity had been exploited. For the purpose, controlled release microparticles (MP) were prepared by co-spray drying DNase I with the polymer poly-lactic-co-glycolic acid (PLGA) and the biocompatible lipid surfactant 1,2-dipalmitoyl-Sn-phosphatidyl choline (DPPC) using various hydrophilic excipients. The effect of the included modifiers on the particle morphology, size, zeta potential as well as enzyme encapsulation efficiency, biological activity and release had been evaluated. Powder aerosolisation performance and particle phagocytosis by murine macrophages were also investigated. The results showed that more than 80% of enzyme activity was recovered after MP preparation and that selected surface modifiers greatly increased the enzyme encapsulation efficiency. The particle morphology was greatly modified altering in turn the powders inhalation indices where dextran, ovalbumin and chitosan hydrochloride increased considerably the respirable fraction compared to the normal hydrophilic carriers lactose and PVP. Despite of the improved aerosolisation caused by chitosan hydrochloride, yet retardation of chitosan coated particles in artificial mucus samples discouraged its application. On the other hand, dextran and polyanions enhanced DNase I effect in reducing cystic fibrosis mucus viscosity. DPPC proved good ability to reduce particles phagocytic uptake even in the presence of the selected adjuvants. The prepared MP systems were biocompatible with lung epithelial cells. To conclude, controlled release DNase I loaded PLGA-MP with high inhalation indices and enhanced mucolytic activity on CF sputum could be obtained by surface modifying the particles with PGA or dextran., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Background: Bombesin has been used to target Bombesin receptor, a growth receptor, which is over-expressed in many cancers, including prostate cancer. Polymer-anti-neoplastic-drug-conjugates (PDC) were also developed to reduce non-specific toxicity and increase tumor toxicity utilizing the enhanced permeability and retention effect, benefitting treatment of large tumors with well-established vasculature., Purpose: If PDCs were delivered by targeted delivery to cancer cells, tumor toxicity would be enhanced and non-specific toxicity decreased., Methods: Cardiocyte toxicity was assessed in H9c2 cardiocytes with doxorubicin (Dox) or N-terminal DTPA-modified-Doxorubicin-loaded-polyglutamic acid polymers (D-Dox-PGA). Therapeutic efficacy of targeted D-Dox-PGA after pretargeting with Bombesin-conjugated anti-DTPA-antibody Bispecific Complexes (Bom-BiSpCx) was compared to that of Dox in PC3 cells. Bom-BiSpCx was generated by thioether bond between Bombesin to Anti-DTPA antibody., Results: D-Dox-PGA was demonstrated to have less cardiocyte toxicity (IC50 = 20 µg/ml) than free Dox (1.55 µg/ml, p < 0.001). However, after pre-targeting of human prostate cancer PC3 cells with Bom-BiSpCx and targeting with D-Dox-PGA, IC50 (13.2 µg/ml) was about two times less than that of Dox (28.5 µg/ml, p < 0.0001)., Discussion: Targeted delivery of PDCs having lower cardiocyte toxicity enabled higher efficiency cancer cell therapy., Conclusion: This study may allow development of very efficient targeted prostate cancer pro-drug therapy.

Polymer nanogels have gained considerable attention as a potential platform for drug delivery applications. Here we describe the design and synthesis of novel polypeptide-based nanogels with hydrophobic moieties in the cross-linked ionic cores. Diblock copolymer, poly(ethylene glycol)-b-poly(L-glutamic acid), hydrophobically modified with L-phenylalanine methyl ester moieties was used for controlled template synthesis of nanogels with small size (ca. 70 nm in diameter) and narrow particle size distribution. Steady-state and time-resolved fluorescence studies using coumarin C153 indicated the existence of hydrophobic domains in the ionic cores of the nanogels. Stable doxorubicin-loaded nanogels were prepared at high drug capacity (30 w/w%). We show that nanogels are enzymatically-degradable leading to accelerated drug release under simulated lysosomal acidic pH. Furthermore, we demonstrate that the nanogel-based formulation of doxorubicin is well tolerated and exhibit an improved antitumor activity compared to a free doxorubicin in an ovarian tumor xenograft mouse model. Our results signify the point to a potential of these biodegradable nanogels as attractive carriers for delivery of chemotherapeutics.

Redox-responsive SCMs based on amphiphilic PBLG-b-dextran with good biocompatibility are synthesized and used for efficient intracellular drug delivery. The molecular structures and SCMs characteristics are characterized by (1) H NMR, FT-IR, TEM, and DLS. The hydrodynamic radius of SCMs increases gradually in PBS due to the cleavage of disulfide bond in micellar shell caused by the presence of GSH. The encapsulation efficiency and release kinetics of DOX are investigated. The fastest DOX release is observed under intracellular-mimicking reductive environments. An MTT assay demonstrates that DOX-loaded SCMs show higher cellular proliferation inhibition against GSH-OEt pretreated HeLa and HepG2 than that of the non-pretreated and BSO-pretreated ones., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Micelles for mucosal immunity: A mucosal vaccine system based on γ-PGA nanomicelles and viral antigens was synthesized. The intranasal administration of the vaccine system induces a high immune response both in the humoral and cellular immunity (see picture)., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

In this work, Taylor dispersion analysis and capillary electrophoresis were used to characterize the size and charge of polymeric drug delivery nanogels based on polyglutamate chains grafted with hydrophobic groups of vitamin E. The hydrophobic vitamin E groups self-associate in water to form small hydrophobic nanodomains that can incorporate small drugs or therapeutic proteins. Taylor dispersion analysis is well suited to determine the weight average hydrodynamic radius of nanomaterials and to get information on the size polydispersity of polymeric samples. The effective charge was determined either from electrophoretic mobility and hydrodynamic radius using electrophoretic modeling (three different approaches were compared), or by indirect UV detection in capillary electrophoresis. The influence of vitamin E hydrophobicity on the polymer effective charge has been studied. The presence of vitamin E leads to a drastic decrease in polymer effective charge in comparison to non-modified polyglutamate. Finally, the electrophoretic behavior of polyglutamate backbone grafted with hydrophobic vitamin E (pGVE) nanogels according to the ionic strength was investigated using the recently proposed slope plot approach. It was deduced that the pGVE nanogels behave electrophoretically as polyelectrolytes which is in good agreement with the high water content of the nanogels.

This research is aimed to develop a nano-sized supramolecular micelle delivery system of cis-dichlorodiammine platinum (II) (CDDP) in order to achieve the passive tumor targeting. Firstly, star-shaped poly (γ-benzyl-L-glutamate) was synthesized by the ring-opening polymerization of γ-benzyl-L-glutamate-N-carboxyanhydride initiated with per-6-amino-β-cyclodextrin. After removal of benzyl groups, β-cyclodextrin based seven-armed poly (L-glutamic acid) (β-CD-7PLGA) was obtained. β-CD-7PLGA/CDDP complexes were prepared by the complex reaction between the carboxylic groups of β-CD-7PLGA and CDDP. Further inclusion of β-CD-7PLGA/CDDP complexes with adamantine terminated mPEG (mPEG-Ad) gave CDDP supramolecular micelles (mPEG-Ad@β-CD-7PLGA/CDDP). The formation of mPEG-Ad@β-CD-7PLGA/CDDP supramolecular micelles was confirmed by fluorescence spectrophotoscopy and particle size measurements. All the micelles showed spherical shape, and their sizes increased from 100 to 135 nm with the increase of PLGA arm molecular weight. mPEG-Ad@CD-7PLGA/CDDP micelles showed sustained drug release profiles over 50h in PBS. Compared with CDDP, mPEG-Ad@β-CD-7PLGA/CDDP supramolecular micelles showed essential decreased cytotoxicity to KB cells, suggesting their great potential as the delivery carriers of CDDP., (Copyright © 2013 Elsevier B.V. All rights reserved.)