RAFT polymerization mediated core–shell supramolecular assembly of PEGMA-co-stearic acid block co-polymer for efficient anticancer drug delivery

In this work, core–shell supramolecular assembly polymeric nano-architectures containing hydrophilic and hydrophobic segments were synthesized via reversible addition fragmentation chain transfer (RAFT) polymerization. Herein, polyethylene glycol methyl ether methacrylate (PEGMA), and stearic acid were used to synthesize the poly(PEGMA) homopolymer and stearyl ethyl methacrylate (SEMA), respectively. Then, PEGMA and SEMA were polymerized through controlled RAFT polymerization to obtain the final diblock copolymer, poly(PEGMA-co-SEMA) (BCP). Model anticancer drug, doxorubicin (DOX) was loaded on BCPs. Interestingly, efficient DOX release was observed at acidic pH, similar to the cancerous environment pH level. Significant cellular uptake of DOX loaded BCP50 (BCP50-DOX) was observed in MDA-MB-231 triple negative breast cancer cells and resulted in a 35 fold increase in anticancer activity against MDA MB-231 cells compared to free DOX. Scanning electron microscopy (SEM) imaging confirmed the apoptosis mediated cellular death. These core–shell supramolecular assembly polymeric nano-architectures may be an efficient anti-cancer drug delivery system in the future.

In this work, core–shell supramolecular assembly polymeric nano-architectures containing hydrophilic and hydrophobic segments were synthesized via reversible addition fragmentation chain transfer (RAFT) polymerization.     

Development of a pH-sensitive functionalized metal organic framework: in vitro study for simultaneous delivery of doxorubicin and cyclophosphamide in breast cancer

Exploration of an efficient dual-drug based nanocarrier with high drug loading capacity, specific targeting properties, and long-term stability is highly desirable in cancer therapy. Metal–organic frameworks (MOFs) have proven to be a promising class of drug carriers due to their high porosity, crystalline properties with defined structure information, and their potential for further functionalization. To enhance the drug efficacy as well as to overcome the burst effect of drugs, here we synthesized a pH responsive folic acid (FA) and graphene oxide (GO) decorated zeolitical imidazolate frameworks-8 (GO–FA/ZIF-8), for targeted delivery of doxorubicin (DOX) and cyclophosphamide (CP), simultaneously. In this system, DOX molecules were encapsulated in the pores of ZIF-8 during in situ synthesis of ZIF-8 and CP molecules have been captured by the GO surface via hydrogen bonding and π–π interactions as well. Furthermore, the resulting pH-responsive nanocarrier (DOX@ZIF-8/GO–FA/CP) showed in vitro sustained release characteristics (76% of DOX and 80% of CP) by cleavage of chemical bonding and disruption of the MOFs structure under acidic condition (at pH 5.6). Moreover, DOX@ZIF-8/GO–FA/CP has synergistic cytotoxic effects as compared to the combination of both the drugs without ZIF-8/GO–FA when treating MCF-7 and MDA-MB-231 breast cancer cell lines (with a combination index of 0.29 and 0.75 for MCF-7 and MDA-MB-231 cell-lines, respectively). Hence this system can be applied as an effective platform for smart dual drug delivery in breast cancer treatment through its remarkable manageable multidrug release.

Exploration of an efficient dual-drug based nanocarrier with high drug loading capacity, specific targeting properties, and long-term stability is highly desirable in cancer therapy.     

Liposomes co-delivery system of doxorubicin and astragaloside IV co-modified by folate ligand and octa-arginine polypeptide for anti-breast cancer

Doxorubicin (DOX) is one of the core drugs in triple-negative breast cancer (TNBC) chemotherapy, but its resistance has severely limited its clinical application. Our previous study found that astragaloside IV (AS-IV) has a good reversal effect on doxorubicin resistance. In order to encapsulate DOX and AS-IV simultaneously, a new liposome-targeted co-delivery system co-modified by the folate ligand (FA) and octa-arginine polypeptide (R8) (FA-R8-LPs, for short) was prepared. In this co-delivery system, R8 not only served as a bond connecting the FA to the liposome, but also played the role of cell penetrating peptides (CPPs). This design effectively increased the tumor targeting and cellular uptake capacity of liposomes. The results of the cytotoxicity test indicated that FA-R8-LPs significantly inhibited the proliferation of the DOX resistant cell line MDA-MB-231/DOX in vitro. In nude mice tumor models inoculated with MDA-MB-231/DOX cells, FA-R8-LPs significantly inhibited tumor growth, and overcame doxorubicin resistance, exhibiting excellent antitumor effects. This study demonstrates that liposome-targeted co-delivery systems based on FA and R8 double modifying may provide a new and effective strategy for the treatment of TNBC, which is of great significance for drug combination.

To more efficiently co-deliver DOX and AS-IV, R8 was used as a “connecting bridge” to connect FA with cholesterol. A new liposome-targeted co-delivery system, co-modified by FA and R8 (FA-R8-LPs, for short), was prepared.     

Targeting self-assembled F127-peptide polymer with pH sensitivity for release of anticancer drugs

The treatment of breast cancer mainly relies on chemotherapy drugs, which present significant side effects. The most typical example is the cardiotoxicity and bone marrow suppression associated with doxorubicin (DOX). Therefore, this drug is not the first choice in clinical treatment. We designed ATN-FFPFF-ATN, a new targeted antitumor drug carrier, polymerized from phenylalanine dipeptide (FF), ATN-161 peptide, and Pluronic® F-127. The peptide and Pluronic® F-127 are linked with acetal and are, therefore, acid-sensitive. As cancer can reduce pH through complex mechanisms and subsequently maintain acid ambience, our vehicle can smartly unravel at a peculiar position, through which the drug can specifically accumulate inside the tumor. ATN-161 is a protein ligand of integrin α₅β₁, which is highly expressed on the surface of some breast cancer cells. This targeting peptide sequence can play a role in the selective delivery of DOX to tumor cells. The DOX-carrying vector was able to significantly inhibit cell proliferation and promote cell apoptosis in MDA-MB-231 cells. Based on these results, ATN-FFPFF-ATN with pH response is a promising vehicle for DOX delivery.

The treatment of breast cancer mainly relies on chemotherapy drugs, which present significant side effects.     

Niclosamide encapsulated polymeric nanocarriers for targeted cancer therapy

Localized cancer rates are on an upsurge, severely affecting mankind across the globe. Timely diagnosis and adopting appropriate treatment strategies could improve the quality of life significantly reducing the mortality and morbidity rates. Recently, nanotherapeutics has precipitously shown increased efficacy for controlling abnormal tissue growth in certain sites in the body, among which ligand functionalized nanoparticles (NP) have caught much attention for improved survival statistics via active targeting. Our focus was to repurpose the antihelminthic drug, niclosamide (NIC), which could aid in inhibiting the abnormal growth of cells restricted to a specific region. The work here presents a one-pot synthesis of niclosamide encapsulated, hyaluronic acid functionalized core–shell nanocarriers [(NIC-PLGA NP)HA] for active targeting of localized cancer. The synthesized nanocarriers were found to possess spherical morphology with mean size of 150.8 ± 9 nm and zeta potential of −24.9 ± 7.21 mV. The encapsulation efficiency was found to be 79.19 ± 0.16% with a loading efficiency of 7.19 ± 0.01%. The nanohybrids exhibited extreme cytocompatibility upon testing with MDA-MB-231 and L929 cell lines. The rate of cancer cell elimination was approximately 85% with targeted cell imaging results being highly convincing. [(NIC-PLGA NP)HA] demonstrates increased cellular uptake leading to a hike in reactive oxygen species (ROS) generation, combating tumour cells aiding in the localized treatment of cancer and associated therapy.

Localized binding of nanoparticulate formulation, actively targeting the receptors present on the cell surface.     

Novel non-mulberry silk fibroin nanoparticles with enhanced activity as potential candidate in nanocarrier mediated delivery system

Silk fibroin (SF) is well known for its excellent biocompatible properties facilitating its application in the field of biomedical engineering through different biomaterial fabrications in the recent era. Here in this study, novel nanoparticles from non-mulberry SF of Antheraea assamensis were fabricated, characterized and evaluated for its applicability as nanocarrier. Fabricated nanoparticles were initially compared with prevailing SF nanoparticles from Bombyx mori. Fabricated A. assamensis silk fibroin nanoparticles (AA-SFNps) were found to be lesser in size (80–300 nm in diameter) than B. mori silk fibroin nanoparticles (BM-SFNps) (120–500 nm in diameter). When checked for stability, AA-SFNps were found to be more stable than BM-SFNps in biological media. FTIR and XRD studies revealed persistence of structural properties even after fabrication. TGA and DSC studies showed AA-SFNps to be thermally more stable than BM-SFNps without any cytotoxicity (MTT assay). On loading with model drug Doxorubicin hydrochloride (DOX), AA-SFNps exhibited an encapsulation efficiency of 94.47% with 11.81% loading of the anticancer drug. Cumulative release study revealed highest percentage release of DOX (42.1 ± 0.4%) at pH 5.2 on day 7 in comparison to pH 7.4 and 8.0. Sustained release profile of the DOX loaded AA-SFNps (AA-SFNps-DOX) was clearly reflected and it was found to be highly cytotoxic against triple negative MDA-MB-231 cells in comparison to free DOX at different time points. Overall, this study showed the efficacy of the AA-SFNps as a nanocarrier for future drug delivery applications.

Novel Antheraea assamensis silk fibroin nanoparticles (AA-SFNps) exhibiting enhanced activity as doxorubicin hydrochloride (DOX) loaded nanocarriers for future drug delivery applications.     

P1c peptide decorated liposome targeting αvβ3-expressing tumor cells in vitro and in vivo

Integrin αvβ3 is a promising target for integrin-rich tumor and neovascular. In the present study, we prepared a doxorubicin (DOX)-loaded liposome of which the surface was decorated with PEG and a novel αvβ3 targeting peptide of P1c. The in vitro targeting efficiency was evaluated in αvβ3-positive (U87MG) and -negative (MCF-7) tumor cells by flow cytometry and laser confocal scanning microscopy. The in vivo therapeutic effects were evaluated in the glioblastoma U87MG-tumor bearing mouse model. The results indicated that the prepared liposomes showed mean sizes of 131.2 and 128.4 nm in diameter for P1c-modified targeting liposomes (P1c-DOXL) and non-targeting liposomes (DOXL), respectively. The DOX encapsulation efficiencies were more than 95% in both types of liposomes. The conjugation ratio for P1c decoration was 66.8%. The flow cytometry and confocal laser-scanning microscopy experiments consistently showed that the intracellular fluorescence intensity of the P1c-modified targeted liposome group was stronger than that of the non-targeted liposome group (P < 0.05) in U87MG cells. In vivo results revealed that compared with DOX or DOXL treatment, P1c-DOXL dramatically reduced tumor growth (P < 0.05) and tumor angiogenesis while much lower hepatotoxicity was observed. P1c-modified targeting liposome exhibited sustained release, enhancing the antitumor effect of DOX through targeting tumor cells and neovascular where integrin αvβ3 was overexpressed. The results indicated that P1c might be promising for active targeting delivery in cancer therapy.

A novel peptide of P1c decorated liposomes targets an integrin αvβ3 expressed tumor.     

Tandem microfluidic chip isolation of prostate and breast cancer cells from simulated liquid biopsies using CD71 as an affinity ligand

The use of blood as a liquid biopsy provides a minimally invasive and less traumatic approach for initial cancer screens as well as patient monitoring. However, current clinical protocols require a priori knowledge of cancer type for liquid biopsy analyses. Previously, we proposed the use of the human transferrin 1 receptor protein (CD71) as a universal capture target for cancer cells analyses. In this study we have attempted to identify the lowest limit of detection for circulating tumor cells of prostate (PC-3) and breast cancers (MDA-MB-231) using CD71. We used a novel high-throughput herringbone chip design which could extract PC-3 cells at 34 ± 5% purity and MDA-MB-231 cells at 43 ± 35% purity when spiked to lysed blood at 0.1%. MDA-MB-231 cell spiked samples showed higher standard deviation, but the system captured 55 ± 16 cells, which is a sufficient number of cells for subsequent analyses. Further, this herringbone chip design has been shown to be compatible with an erythrocyte lysis chip we have described in previous studies. This circuit was capable of capturing 510 ± 120 cells with a purity of 82 ± 14% using <7 μL of a whole blood sample spiked with 10% MDA-MB-231 cells. Using an erythrocyte lysis circuit eliminates the need for human intervention for target cell enrichment, thereby reducing cell loss and sample contamination. We have shown that, when used with the high-throughput herringbone chip CD71 has the capacity to sensitively detect rare target cells for routine low-cost cancer screens.

The use of blood as a liquid biopsy provides a minimally invasive and less traumatic approach for initial cancer screens as well as patient monitoring.     

Synergic effect of doxorubicin release and two-photon irradiation of Mn2+-doped Prussian blue nanoparticles on cancer therapy

We demonstrate here that Mn²⁺-doped Prussian blue nanoparticles of ca. 55 nm loaded with doxorubicin may be used as efficient therapeutic agents for combined photothermal and chemo-therapy of cancer cells with a synergic effect under two photon irradiation.

Mn²⁺-doped Prussian blue nanoparticles loaded with doxorubicin present high efficiency for combined photothermal and chemotherapy of cancer cells with a synergic effect under two-photon irradiation.

Photothermal therapy (PTT) is a promising strategy for the therapeutic treatment of tumours, which consists in “burning” cancer cells by laser irradiation at low energy wavelengths in the presence of an NIR photo-adsorbing agent.^1,2 In comparison to conventional cancer treatments, it offers several advantages, such as high specificity, deep penetration, lower tumour recurrence, minimal invasiveness and low toxicity to normal tissues.^3,4 Due to the fact that the treatment efficiency highly depends on the outstanding capacity of the PTT agent to convert light into heat, numerous materials have recently been reported, such as organic dyes^5,6 or various inorganic nano-objects, including metallic nanostructures (Au nanorods or nanoshells, Ag, Pd), carbon based materials (nanotubes, spheres, graphene oxide), transition metal dichalcogenide nanoparticles, metal oxide nano-objects or up-conversion nanocrystals.^7–16Among these, Prussian blue (PB) nano-objects have been developed as interesting PTT agents since they benefit from many advantages, such as: (i) a controlled size ranging from a few to hundreds of nanometers, (ii) easy surface functionalization, (iii) excellent absorption properties in the NIR spectral domain due to an Fe²⁺ to Fe³⁺ intervalence charge transfer band ranging from 650 to 900 nm, (iv) high stability under irradiation, and (v) the fact that bulk PB is validated by the FDA under the brand name Radiogardase® as an antidote for human beings exposed to radioactive Cs⁺.¹⁷ It has been shown in vitro that PB nanoparticles upon single photon excitation (SPE) (at 808 nm) can convert the laser irradiation into thermal energy contributing to an important temperature increase.^18,19 Their efficiency depends on the nanoparticles'' concentration, their chemical composition, the laser power, the mode of irradiation and the irradiation time. Moreover, it has been demonstrated that nano-sized PB presents a higher efficiency in comparison with Au nanorods and a better photothermal stability than organic dyes used as conventional photothermal agents.^19,20 PTT effect of PB nanoparticles has also been demonstrated in vivo through their intra-tumoral injection that causes significant tumour necrosis 24 h after NIR irradiation in comparison with non-treated mice.¹⁸ Nevertheless, the complete eradication of cancer cells using solely PTT treatment is rather difficult due to the suboptimal laser energy in deep tissues related to light scattering and absorption effects, a limited light penetration and the insufficient tumour target specificity of PTT agents. Some of these drawbacks may be circumvented using two photon excitation (TPE), which permits to increase the penetration depth and laser focalization allowing selective and efficient destruction of the targeted cancer cells with less damage to healthy tissues.^21–23 In this line of thought, we reported recently Mn²⁺-doped PB nanoparticles of ca. 70 nm as efficient multifunctional PTT agents permitting to eradicate 97% of cancer cells 24 h after irradiation under TPE during 10 min, which may also be followed by Magnetic Resonance Imaging (MRI).²⁴ However, the comparison between the efficiency of the single and two-photon excitations on the same nano-objects has never been investigated.Another possibility to increase treatment efficiency and thus decrease the concentration of the PTT agent consists in the association of PTT with a drug or gene delivery to develop a personalized cancer therapy.^25–29 Several works report on the design of PB nanoparticles wrapped by gelatine or PEGylated lipids containing the incorporated anticancer drug^1,25,30–32 or conjugated with pDNA³³ or antigen specific cytotoxic lymphocytes.³⁴ In these cases, an improvement in cancer cell mortality or in the efficiency of gene transfection under NIR irradiation has been observed. But the synergic effect of PTT and chemotherapy has been clearly demonstrated only for hollow PB nanoparticles of 236 nm loaded with DOX presenting a therapeutic efficacy increase from 13.6 to 51.2% in comparison with the non-loaded nanoparticles.²⁰Herein, we investigate the loading of Mn²⁺-doped PB nanoparticles of ca. 55 nm with DOX by experimental means joined with a theoretical modelling and demonstrate that such systems may be used as efficient agents for combined PTT and chemotherapy, able to eradicate almost all cancer cells (91%) under TPE due to a synergic effect with a relatively low concentration (50 μg mL⁻¹). Moreover, we compare for the first time the efficiency of the nano-objects with TPE and SPE and investigate their cytotoxicity. The Mn²⁺ doped PB nanoparticles have been chosen for this study due to their multifunctionality as MRI contrast agents and PTT therapeutic agent, since the presence of Mn²⁺ ion in such amount does not impact the PTT efficiency.²⁴Mn²⁺-doped PB nanoparticles Na_0.38Mn_0.12Fe[Fe(CN)₆]_0.911 have been obtained by using the previously described procedure involving the controlled self-assembling reaction between Na₄[Fe(CN)₆]·10H₂O, FeCl₃·6H₂O and MnCl₂·6H₂O in water. The drug loading was performed by suspending 1 in an aqueous DOX solution for 24 h under stirring in the dark. The loading capacity of DOX of 7.6 wt% (ratio between the adsorbed DOX weight and the total weight of the loaded nanoparticles) was determined by UV-Vis absorption spectroscopy (see Experimental part and Fig. S1, ESI^†). The obtained nanoparticles Na_0.28Mn_0.12Fe[Fe(CN)₆]_0.88@DOX_0.041@DOX present typical stretching vibrations of the bridging cyanide group ν(Fe^II–CN–Fe^III) at 2080 cm⁻¹ in the FTIR spectra, as the pristine nanoparticles 1 with an appearance of small intensity bands fingerprints of DOX in the 800–1500 cm⁻¹ frequency range (Fig. S2, ESI^†). Their Powder X-ray diffraction pattern shows the typical fcc structure with a lattice parameter a = 10.0 Å (Fig. S3, ESI^†). The obtained zeta potential (negative value) of −20.8 mV is slightly lower than for the pristine nanoparticles 1 (zeta potential of −35.7 mV), that suggests a surface modification after DOX loading. Transmission Electronic Microscopy (TEM) images for 1@DOX reveal typical cubic nanoparticles with a mean size of 55 ± 13 nm (Fig. 1a, S4 and S5, ESI^†). A slight bathochromic shift from 725 to 757 nm can be observed in the absorption spectra for 1@DOX in comparison with 1 (Fig. S6^†). Yet, due to the relatively small quantity of the adsorbed DOX, its characteristic band at 480 nm is not visible on the spectra of nanoparticles (Fig. S6, ESI^†).Open in a separate windowFig. 1(a) TEM image for 1@DOX; (b) snapshot illustrating the PB network and the density of DOX inside from Monte Carlo simulations (red points = density of DOX presence; green points = density of presence of Na⁺).In order to provide an insight about the nature of interactions between DOX and the PB network, Monte-Carlo calculations using classical force field have been performed (see ESI^† for details^35–37). The size of DOX molecules of 16 Å × 9 Å × 8 Å is clearly larger in comparison with the size of the classical pores of the PB network generated by the presence of both, tetrahedral sites and cyanometallates'' monovacancies, except in the case of double lacunas, in which DOX molecules can be fixed (Fig. 1b, S7, ESI^†). The results of the Monte-Carlo simulations indicate that the saturation of doxorubicin molecules inside the pores is equal to 5.0 wt% (related to 7.6 wt% found experimentally). The larger experimental loading suggests that DOX is situated within the pores of the network (accessible double lacunas) and also at the surface of the nanoparticles attached by physisorption, as suggested by the change in the zeta potential. But the presence of covalent bonds between the nanoparticles and amino and OH groups cannot be excluded.The DOX release experiments performed for 1@DOX in phosphate buffered saline (PBS) show a slow and quasi-linear drug release during the first 4 hours and a more gradual behaviour resulting in a 10.3% release of the adsorbed DOX after 48 h (Fig. S8^†).In order to evaluate the safety of the nanoparticles, an in vitro cytotoxicity study has been carried out on human breast adenocarcinoma MDA-MB-231 cell line, which is treated with nanoparticles 1 and 1@DOX at a concentration of 50 μg mL⁻¹ and cell viability was determined after 1, 2 and 3 days. Results showed that 1 are non-toxic, as indicated by the increase of the percentage of cell viability with time, a similar behaviour as for untreated cells, which suggests their biocompatibility (Fig. S9, ESI^†).^1,38,39 In contrast, cell death was observed in cells treated with 1@DOX, where cell viability decreases during the treatment (40% after 1 day, 35% after 2 days, and 32% after 3 days) due to DOX release.Chemo-PTT ability of the nanoparticles 1@DOX to kill the living cancer MDA-MB-231 cells has been investigated by using both, TPE and SPE lasers and compared with the efficiency of nanoparticles 1 and free DOX at a concentration of 6 mg mL⁻¹. The latter is comparable with the amount of loaded DOX into nanoparticles 1@DOX.In the case of pulsed TPE at 808 nm (3.7 W, 5% of total laser power) for 10 min and after 24 h of irradiation, the cells treated with 50 μg mL⁻¹ of 1@DOX show 91 ± 7% of cell death, while it was 76 ± 12% in the case of 1 and 24 ± 17% for cells treated with free DOX (Fig. 2a). These results were analysed by using CompuSyn software (ComboSyn, Inc)⁴⁰ in order to determine the synergic effect of combined PTT and drug. The calculated combination index (CI) value of 0.4 confirms the presence of an important synergistic effect (the synergic effect is operational for CI value lower than 1, while the additive and antagonist effects occur for values equal or greater than 1, respectively). The cell death is highlighted by confocal imaging on green fluorescent living cells as shown in Fig. 2b. The safety of TPE laser was confirmed on untreated cells by the increase in the number of green fluorescent spots signifying their important multiplication 24 h after irradiation. On the contrary, a decrease or an almost completely disappearance of green fluorescent nuclei was observed 24 h after irradiation in cells treated with free DOX, 1 or 1@DOX, confirming cell death. The synergic effect is certainly due to the important temperature elevation around each nanoparticle permitting the efficient DOX liberation.Open in a separate windowFig. 2(a) Cell counting (%) of living MDA-MB-231 cells treated with nanoparticles 1@DOX and 1 at 50 μg mL⁻¹ concentration and free doxorubicin before irradiation, immediately after irradiation and 24 h after irradiation with a TPE laser at 808 nm (3.7 W, 5% of total laser power) for 10 min. Data are presented as (mean ± SEM), n = 3. (b) Fluorescence imaging of living MDA-MB-231 cells treated with 50 μg mL⁻¹ concentration of 1, 1@DOX and free doxorubicin (6 mg mL⁻¹) before irradiation, immediately and 24 h after irradiation with TPE laser at 808 nm (3.7 W, 5% of total laser power) for 10 min.The increase of nanoparticles concentration to 100 μg mL⁻¹ did not hardly impact the efficiency of the treatment. In fact, 95 ± 4%, 89 ± 6% and 34 ± 1% of cell death were found for 1@DOX, 1 and free DOX, respectively. This fact may be explained by the restricted amount of nanoparticles uptake at higher concentration. The effect of the concentration of 1@DOX on cells mortality is shown on Fig. S10.^†The obtained results with the TPE laser have been compared with the effect obtained under the SPE laser by using the same concentration of nanoparticles. For this, cells were irradiated at 808 nm (2.5 W cm⁻²) during 30 min (10 min of irradiation did not provide any cells'' mortality). Data showed that only 35 ± 5% of cell death was found with 1@DOX 24 h after irradiation (Fig. 3 and Fig. S11^†) clearly demonstrating the advantage of TPE over SPE in our experimental conditions. However, the cancer cells continue to die and reached a death level of 55 ± 10% and 77 ± 3% after 48 h and 72 h of irradiation, respectively (Fig. 3a). Fluorescent images for live (green colour) and dead cells (red colour due to propidium iodide staining) confirm this result (Fig. 3b and S12^†). Note that the obtained results on SPE are comparable with the previously published ones on other PB nanoparticles containing doxorubicin.^1,25,30,32Open in a separate windowFig. 3(a) Cell counting (%) of living MDA-MB-231 cells treated with nanoparticles 1@DOX and 1 at 50 μg mL⁻¹ concentration before irradiation and 24 h, 48 h, and 72 h after irradiation with SPE laser at 808 nm (2.5 W cm⁻²) for 30 min. Data are presented as (mean ± SEM) n = 3. Inset: enlargement of figure for 1@DOX + NIR; (b) fluorescence imaging of living MDA-MB-231 cells treated with 50 μg mL⁻¹ nanoparticles'' concentration after 72 h of irradiation with SPE at 808 nm (2.5 W cm⁻²) for 30 min. Cells were treated with propidium iodide (PI) for cell death detection (shown in red).In summary, we demonstrated that Mn²⁺-doped PB nanoparticles of ca. 55 nm can be easily loaded with doxorubicin drug (7.6 wt%). We established for the first time by using theoretical modelling that the latter mainly enters into the network''s pores and a minor amount is also attached on the surface of the nanoparticles. A synergic effect of the PTT and DOX release is observed upon the treatment of MDA-MB-231 cancer cells with a relatively low concentration of nanoparticles (50 μg mL⁻¹) under TPE laser inducing their almost total eradication. These results are much improved in comparison to the ones obtained by combined DOX release and PTT under an SPE laser taking the same concentration of nanoparticles, which provides lower cell mortality even 72 h after irradiation. This strategy provides an interesting opportunity towards multifunctional nanomedicines with combined therapy and imaging.     

Dual targeted and pH-responsive gold nanorods with improved chemotherapy and photothermal ablation for synergistic cancer treatment

Cancer is considered to be one of the leading causes of morbidity and mortality worldwide. A multifunctional nanosystem based on gold nanorods (GNRs) has demonstrated the potential to enhance therapeutic performance. In this research, dual-targeted pH-responsive GNRs for synergistic cancer treatment were developed and investigated. The GNRs could target angiogenic endothelial cells in the tumor region using αvβ3-mediated recognition and subsequently facilitate its specific binding to tumor cells mediated via recognition of the folate receptor, which could accumulate precisely at the tumor site. Doxorubicin (DOX) was loaded on to the surface of GNRs via a pH-sensitive hydrazone (hz) bond, which could effectively control the drug release by responding to the tumor acidic microenvironment. In vitro, the FA/RGD-DOX-hz-GNRs showed higher tumor specificity and killing ability under near-infrared irradiation. Furthermore, in B16-F10 xenograft tumor-bearing mice, FA/RGD-DOX-hz-GNRs produced the optimal tumor therapeutic efficacy by antagonizing angiogenesis, inhibiting cell proliferation and causing necrosis. Therefore, the strategy of integration of a photothermal effect, chemotherapy and a molecular active targeting based double-targeting mode appeared advantageous over chemotherapy or a photothermal therapy alone.

A dual-targeted pH-responsive GNR for synergistic cancer treatment was developed and investigated, which demonstrated the desired potential for enhancing therapeutic performance.     

In vivo targeting of breast cancer with a vasculature-specific GQDs/hMSN nanoplatform

According to our previous experiment, graphene quantum dots capped in hollow mesoporous silica nanoparticles, denoted as GQDs@hMSN, and its conjugates exhibited great potential for medical applications due to their commendable biocompatibility. Due to the fluorescence and structural stability, and enormous porosity, polyethylene glycol (PEG) modified GQDs@hMSN (GQDs@hMSN-PEG) is a good candidate in a drug carrying and delivery system. However, the goal of targeted drug delivery couldn''t be achieved simply by utilizing the enhanced permeability and retention (EPR) effect of tumors. In this study, GQDs@hMSN-PEG was further functionalized with vascular endothelial growth factor antibodies (VEGF Abs) for VEGF targeting of breast tumors. Doxorubicin (DOX) was loaded into GQDs@hMSN-VEGF Abs with a drug loading capacity of 0.80 mg DOX per mg GQDs@hMSN. With GQDs as the fluorescent source, GQDs@hMSN-VEGF Abs demonstrated strong fluorescence intensity in VEGF-positive cells. Results from in vitro and in vivo targeting experiments indicated that GQDs@hMSN-VEGF Abs had high specificity on tumor vasculature, and it could be used as an image-guidable, tumor-selective delivery nanoplatform for breast cancer.

According to our previous experiment, graphene quantum dots capped in hollow mesoporous silica nanoparticles, denoted as GQDs@hMSN, and its conjugates exhibited great potential for medical applications due to their commendable biocompatibility.     

Mitochondria-targeted half-sandwich iridium(iii)-Cp*-arylimidazophenanthroline complexes as antiproliferative and bioimaging agents against triple negative breast cancer cells MDA-MB-468

To reduce the side effects of marketed cancer drugs against triple negative breast cancer cells we have reported mitochondria targeting half-sandwich iridium(iii)-Cp*-arylimidazophenanthroline complexes for MDA-MB-468 cell therapy and diagnosis. Out of five Ir(iii) complexes (IrL1–IrL5), [iridium(iii)-Cp*-2-(naphthalen-1-yl)-1H-imidazo[4,5-f][1,10]phenanthroline]PF₆ (IrL1) has exhibited the best cytoselectivity against MDA-MB-468 cells compared to normal HaCaT cells along with excellent binding efficacy with DNA as well as serum albumin. The subcellular localization study of the complex revealed the localization of the compound in cytoplasm thereby pointing to a possible mitochondrial localization and consequent mitochondrial dysfunction via MMP alteration and ROS generation. Moreover, the IrL1 complex facilitated a substantial G₁ phase cell-cycle arrest of MDA-MB-468 cells at the highest tested concentration of 5 μM. The study verdicts support the prospective therapeutic potential of the IrL1 complex in the treatment and eradication of triple negative breast cancer cells. These results validate that these types of scaffolds will be fairly able to exert great potential for tumor diagnosis as well as therapy in the near future.

Mitochondria targeting half-sandwich Iridium(iii)-Cp*-arylimidazophenanthroline complexes have been developed for MDA-MB-468 cell therapy and diagnosis.     

Lecithin–chitosan–TPGS nanoparticles as nanocarriers of (−)-epicatechin enhanced its anticancer activity in breast cancer cells

Natural compounds such as (−)-epicatechin show a variety of biological properties including anticancer activity. Nonetheless, (−)-epicatechin''s therapeutic application is limited due to its low water solubility and sensitivity to oxygen and light. Additionally, previous studies have reported that the encapsulation of flavonoids in nanoparticles might generate stable deliverable forms, which improves the availability and solubility of the bioactive compounds. The aims of this study were to generate (−)-epicatechin-loaded lecithin–chitosan nanoparticles (EC-LCT-NPs) by molecular self-assembly and to assess their cytotoxic potential against breast cancer cells. Various parameters were measured to characterize the EC-LCT-NPs including size, polydispersity index (PdI), zeta potential, morphology and entrapment efficiency. The results showed that the mean particle size of the EC-CLT-NPs was 159 ± 2.23 nm (PdI, 0.189), and the loading and entrapment efficiencies of (−)-epicatechin were 3.42 ± 0.85% and 56.1 ± 3.9%, respectively. The cytotoxic effect of the EC-CLT-NPs was greater than that of free (−)-epicatechin on breast cancer cell lines (MCF-7, MDA-MB-231, MDA-MB-436 and SK-Br3). Indeed, EC-LCT-NPs showed an IC₅₀ that was four-fold lower (85 μM) than free (−)-epicatechin (350 μM) and showed selectivity to cancerous cells. This study demonstrated that encapsulating (−)-epicatechin into lecithin–chitosan nanoparticles opens new options for breast cancer treatment.

Natural compounds such as (−)-epicatechin show a variety of biological properties including anticancer activity.     

LyP-1 Conjugated Nanoparticles for Magnetic Resonance Imaging of Triple Negative Breast Cancer

Purpose

Triple-negative breast cancer (TNBC) does not express estrogen receptor, progesterone receptor, or Her2/neu. Both diagnosis and treatment of TNBC remain a clinical challenge. LyP-1 is a cyclic 9 amino acid peptide that can bind to breast cancer cells. The goal of this study was to design and characterize LyP-1 conjugated to fluorescent iron oxide nanoparticles (LyP-1-Fe₃O₄-Cy5.5) as a contrast agent for improved and specific magnetic resonance imaging (MRI) in a preclinical model of TNBC.

Procedures

The binding of LyP-1-Fe₃O₄-Cy5.5 to MDA-MB-231 TNBC cells was evaluated and compared to scrambled peptide bio-conjugated to iron oxide nanoparticles (Ctlpep-Fe₃O₄-Cy5.5) as a negative control. Following the in vitro study, the MDA-MB-231 cells were injected into mammary glands of nude mice. Mice were divided into two groups: control group received Ctlpep- Fe₃O₄-Cy5.5 and LyP-1 group received LyP-1-Fe₃O₄-Cy5.5 (tail vein injection at 2 mg/kg of Fe₃O₄). Mice were imaged with an in vivo fluorescence imager and a 9.4 T MRI system at various time points after contrast agent injection. The T2 relaxation time was measured to observe accumulation of the contrast agent in breast tumor and muscle for both targeted and non-targeted contrast agents.

Results

Immunofluorescence revealed dense binding of the LyP-1-Fe₃O₄-Cy5.5 contrast agent to MDA-MB-231 cells; while little appreciable binding was observed to the scrambled negative control (Ctlpep-Fe₃O₄-Cy5.5). Optical imaging performed in tumor-bearing mice showed increased fluorescent signal in mammary gland of animals injected by LyP-1-Fe₃O₄-Cy5.5 but not Ctlpep- Fe₃O₄-Cy5.5. The results were confirmed ex vivo by the 2.6-fold increase of fluorescent signal from LyP-1-Fe₃O₄-Cy5.5 in extracted tumors when compared to the negative control. In MR imaging studies, there was a statistically significant (P < 0.01) difference in normalized T2 between healthy breast and tumor tissue at 1, 2, and 24 h post injection of the LyP-1-Fe₃O₄-Cy5.5. In animals injected with LyP-1-Fe₃O₄, distinct ring-like structures were observed with clear contrast between the tumor core and rim.

Conclusion

The results demonstrate that LyP-1-Fe₃O₄ significantly improves MRI contrast of TNBC, hence has the potential to be exploited for the specific delivery of cancer therapeutics.

     

LyP-1标记超顺磁性氧化铁的制备及体外成像

目的 探讨LyP-1标记超顺磁性氧化铁(SPIO)与乳腺癌细胞结合进行体外成像的可行性。方法 以共沉淀法制备SPIO,用LyP-1与3-氨丙基三甲氧基硅烷(APTMS)包被的SPIO耦联,以乳腺癌MDA-MB-231细胞株为研究对象,设立LyP-1-SPIO组、竞争组、SPIO组和对照组,普鲁士蓝染色,评价不同组中铁颗粒在细胞中的分布情况,四甲基偶氮唑盐(MTT)比色法检测细胞活性并观察其体外MR成像效果。结果 成功制备SPIO。 LyP-1-SPIO组有较多铁颗粒进入细胞内,竞争组和SPIO组仅有少量铁颗粒进入细胞中,对照组细胞中无铁颗粒。MTT比色法检测结果显示不同作用时间LyP-1-SPIO和SPIO对细胞的生长均无显著影响;体外MR成像提示LyP-1-SPIO能够显著增强阴性对比效果。结论 LyP-1-SPIO对MDA-MB-231细胞具有较好的靶向作用,可用于对肿瘤细胞的影像学诊断。     

Thermosensitive star polymer pompons with a core–arm structure as thermo-responsive controlled release drug carriers

In contrast with traditional chemotherapy, controlled drug delivery systems provide many advantages. Herein, a thermosensitive star polymer pompon with a core–arm structure was synthesized using a grafting-on method as a thermo-responsive controlled release drug carrier. Single-chain cyclized/knotted poly tetra(ethylene glycol) diacrylate (polyTEGDA) was used as the hydrophobic core, and thermosensitive linear poly(N-isopropylacrylamide-co-N-methylolacrylamide) (poly(NIPAM-co-NMA)) was selected as the hydrophilic arm. Below or above its lower critical solution temperature (LCST), the linear poly(NIPAM-co-NMA) grafted onto the polyTEGDA core adopted a stretched or curled status, respectively, then the drug could be loaded in or extruded out. The LCST of star polyTEGDA-b-poly(NIPAM-co-NMA) was adjusted to slightly above body temperature (37 °C). The antitumor drug doxorubicin (DOX) was successfully loaded into the pompons with a high loading capacity of 19.45%. The cumulative release of DOX from loaded pompons in vitro for 72 hours was 71% and 20.7% at 42 °C and 37 °C, respectively, indicating that the excellent temperature-controlled release characteristics result from the unique thermo-responsive extrusion effect. Moreover, DOX loaded polyTEGDA-b-poly(NIPAM-co-NMA) pompons achieved better antitumor ability against ovarian carcinoma SKOV3 cells at 42 °C compared with that at 37 °C. These results suggest that star polyTEGDA-b-poly(NIPAM-co-NMA) pompons have considerable promise as thermo-responsive controlled drug delivery carriers.

Herein, a thermosensitive star polymer pompon with a core–arm structure was synthesized using a grafting-on method as a thermo-responsive controlled release drug carrier.     

Synthesis and anticancer studies of Michael adducts and Heck arylation products of sesquiterpene lactones,zaluzanin D and zaluzanin C from Vernonia arborea

Sesquiterpene lactones containing α-methylene-γ-lactones, zaluzanin D 1 and zaluzanin C 2 were isolated from the leaves of Vernonia arborea. Several diverse Michael adducts (3–22) and Heck arylation analogs (23–34) of 1 have been synthesized by reacting with various amines and aryl iodides, respectively and were assayed for their in vitro anticancer activities against human breast cancer cell lines MCF7 and MDA-MB-231. Among all the synthesized analogs, Michael adducts 9 and 10 showed better anticancer activities as compared to 1. However, among these compounds, only 10 has minimal cytotoxic effect on normal breast epithelial MCF10A cells. Our detailed mechanistic studies reveal that compounds 9 and 10 execute their antiproliferative activity through induction of apoptosis and thereby inhibit the cancer cells proliferation and compound 10 could be a lead compound for designing potential anti-cancer compound.

Sesquiterpene lactones containing α-methylene-γ-lactones, zaluzanin D 1 and zaluzanin C 2 were isolated from the leaves of Vernonia arborea.     

Active targeting drug-gold nanorod hybrid nanoparticles for amplifying photoacoustic signal and enhancing anticancer efficacy

The development of theranostic nanomaterials with limited side effects and increased therapeutic efficacy is a promising approach for cancer imaging and therapy. In the present study, the development of a multifunctional metal–organic hybrid nanoparticle (NP) with enhanced photoacoustic (PA) imaging performance able to be actively uptaken by cancer cells for synergistic chemo-photothermal cancer therapy was reported. The theranostic NP was composed through the coordination effect between an ultrasmall gold nanorod (AuNR), a thick coating layer of the organic near-infrared dye IR780, and the anticancer drug doxorubicin (DOX), named AuNR@IR780/DOX-RGD-PEG. In addition, the theranostic NP surface was conjugated with targeting ligand RGD and a protective PEG shell, where the PEG played a role in concealing or exposing the RGD for specific targeting of the NPs to the cancer cells. The theranostic NP demonstrated a greatly enhanced PA imaging signal compared to AuNR or IR780, due to the fact that the electromagnetic field of the AuNR increased the light absorption efficiency of the IR780 coating based on the theoretical simulation results. Furthermore, the “Trojan-horse” active targeting strategy not only increased the uptake of NPs by tumor cells, but also decreased the non-specific uptake by healthy cells, thus limiting the side effects. This study developed a smart theranostic NP for enhanced cancer PA imaging and specific cancer therapy.

We developed a multifunctional metal–organic hybrid nanoparticle with enhanced photoacoustic imaging performance and specific chemo-photothermal cancer therapy.     

Effects of pore size on in vitro and in vivo anticancer efficacies of mesoporous silica nanoparticles

Mesoporous silica nanoparticles (MSN) have been widely applied for drug delivery systems. To investigate the effects of pore size on anticancer efficacies, MSN with different pore sizes but similar particle sizes and surface charges were synthesized via a microemulsion method. The pore structures of MSN were characterized by transmission electron microscopy (TEM), small-angle X-ray scattering (SAXS), and N₂ adsorption–desorption isotherms. Doxorubicin loaded MSN (DOX/MSN) were prepared and the minimum drug loading capacity was detected in DOX/MSN with a pore size of 2.3 nm (DOX/MSN2). DOX/MSN with a pore size of 8.2 nm (DOX/MSN8) showed a comparable drug loading amount in comparison with ones with a pore size of 5.4 nm (DOX/MSN5). In vitro drug release profiles showed that DOX/MSN5 could release DOX quickly and completely. Compared with DOX/MSN2 and DOX/MSN8, DOX/MSN5 showed the higher cellular uptake and nucleic concentration of DOX in QGY-7703 cells, which led to efficient cell-apoptosis induction and anti-proliferation effect, and thus the optimal in vivo anticancer activities. Taken together, these results highlighted the importance of pore size in anticancer efficacies, which would serve as a guideline in the rational design of MSN for cancer therapy.

MSN with suitable pore sizes achieved an outstanding performance for in vitro and in vivo antitumor efficacies.     

Chemical constituents from Carica papaya Linn. leaves as potential cytotoxic,EGFRwt and aromatase (CYP19A) inhibitors; a study supported by molecular docking

The phytochemical investigation of the hydromethanolic extract of Carica papaya Linn. leaves (Caricaceae) resulted in the isolation and characterization of ten compounds, namely; carpaine (1), methyl gallate (2), loliolide (3), rutin (4), clitorin (5), kaempferol-3-O-neohesperidoside (6), isoquercetin (7), nicotiflorin (8) and isorhamnetin-3-O-β-d-glucopyranoside (9). The compounds 2, 3, 5–7 and 9 were isolated for the first time from the genus Carica. An in vitro breast cancer cytotoxicity study was evaluated with an MCF-7 cell line using the MTT assay. Methyl gallate and clitorin demonstrated the most potent cytotoxic activities with an IC₅₀ of 1.11 ± 0.06 and 2.47 ± 0.14 μM, respectively. Moreover, methyl gallate and nicotiflorin exhibited potential EGFR^wt kinase inhibition activities with an IC₅₀ of 37.3 ± 1.9 and 41.08 ± 2.1 nM, respectively, compared with the positive control erlotinib (IC₅₀ = 35.94 ± 1.8 nM). On the other hand, clitorin and nicotiflorin displayed the strongest aromatase kinase inhibition activities with an IC₅₀ of 77.41 ± 4.53 and 92.84 ± 5.44 nM, respectively. Clitorin was comparable to the efficacy of the standard drug letrozole (IC₅₀ = 77.72 ± 4.55). Additionally, molecular docking simulations of the isolated compounds to EGFR and human placental aromatase cytochrome P450 (CYP19A1) were evaluated. Methyl gallate linked with the EGFR receptor through hydrogen bonding with a pose score of −4.5287 kcal mol⁻¹ and RMSD value of 1.69 Å. Clitorin showed the strongest interaction with aromatase (CYP19A1) for the breast cancer receptor with a posing score of −14.2074 and RMSD value of 1.56 Å. Compounds (1–3) possessed a good bioavailability score with a 0.55 value.

The phytochemical investigation of the hydromethanolic extract of Carica papaya Linn. leaves (Caricaceae) resulted in the isolation and characterization of ten compounds.