Bile acid-conjugated chondroitin sulfate A-based nanoparticles for tumor-targeted anticancer drug delivery

Chondroitin sulfate A-deoxycholic acid (CSA-DOCA)-based nanoparticles (NPs) were produced for tumor-targeted delivery of doxorubicin (DOX). The hydrophobic deoxycholic acid (DOCA) derivative was conjugated to the hydrophilic chondroitin sulfate A (CSA) backbone via amide bond formation, and the structure was confirmed by ¹H-nuclear magnetic resonance (NMR) analysis. Loading the DOX to the CSA-DOCA NPs resulted in NPs with an approximately 230 nm mean diameter, narrow size distribution, negative zeta potential, and relatively high drug encapsulation efficiency (up to 85%). The release of DOX from the NPs exhibited sustained and pH-dependent release profiles. The cellular uptake of DOX from the CSA-DOCA NPs in CD44 receptor-positive human breast adenocarcinoma MDA-MB-231 cells was reduced when co-treated with free CSA, indicating the interaction between CSA and the CD44 receptor. The lower IC₅₀ value of DOX from the CSA-DOCA NPs compared to the DOX solution was also probably due to this interaction. Moreover, the ability of the developed NPs to target tumors could be inferred from the in vivo and ex vivo near-infrared fluorescence (NIRF) imaging results in the MDA-MB-231 tumor-xenografted mouse model. Both passive and active strategies appear to have contributed to the in vivo tumor targetability of the CSA-DOCA NPs. Therefore, these CSA-DOCA NPs could further be developed into a theranostic nanoplatform for CD44 receptor-positive cancers.     

CD44-tropic polymeric nanocarrier for breast cancer targeted rapamycin chemotherapy

In contrast with the conventional targeting of nanoparticles to cancer cells with antibody or peptide conjugates, a hyaluronic acid (HA) matrix nanoparticle with intrinsic-CD44-tropism was developed to deliver rapamycin for localized CD44-positive breast cancer treatment. Rapamycin was chemically conjugated to the particle surface via a novel sustained-release linker, 3-amino-4-methoxy-benzoic acid. The release of the drug from the HA nanoparticle was improved by 42-fold compared to HA-temsirolimus in buffered saline. In CD44-positive MDA-MB-468 cells, using HA as drug delivery carrier, the cell viability was significantly decreased compared to free rapamycin and CD44-blocked controls. Rat pharmacokinetics showed that the area under the curve of HA nanoparticle formulation was 2.96-fold greater than that of the free drug, and the concomitant total body clearance was 8.82-fold slower. Moreover, in immunocompetent BALB/c mice bearing CD44-positive 4T1.2neu breast cancer, the rapamycin-loaded HA particles significantly improved animal survival, suppressed tumor growth and reduced the prevalence of lung metastasis.From the Clinical EditorThis study demonstrates increased efficiency of rapamycin delivery and consequential treatment effects in a breast cancer model by hyaluronic acid - L-rapamycin conjugates with intrinsic tropism for CD44-positive cells.     

Positive hyaluronan/PEI/DNA complexes as a target-specific intracellular delivery to malignant breast cancer

In the present study, The PEI/DNA (PD) complexes was first prepared with positive surface charge under a suitable N/P ratio of 10. The redundant positive charge was partially and excessively shielded by a polysaccharide, hyaluronic acid (HA), in aqueous solution. The HA/PEI/DNA ternary complexes were characterized by assessing the zeta potential and size, then transferred to MDA-MB-435, MDA-MB-231, and MCF-7 cell lines with different amounts of HA-specific CD44 receptors on the surface. Consequently, The transfection efficiency of all the prepared complexes show a little increased to MCF-7 (low CD44 level) while a large increased to MDA-MB-231 and MDA-MB-435 cells (high CD44 level) with adding HA. Also, when HA:PEI charge ratio was 7.5%, the ternary complexes show the highest transfection efficiency. The prepared ternary complexes exhibited increased 2–13-fold fluorescence intensity and lower cell toxicity compared to the PD (N/P, 10). These results indicated that the positive HA/PEI/DNA ternary complexes (HA:PEI charge ratio, 7.5%) can target malignant breast cancer cells with high CD44 level and might be a promising candidate vector for gene therapy.     

CD44‐Targeted Docetaxel Conjugate for Cancer Cells and Cancer Stem‐Like Cells: A Novel Hyaluronic Acid‐Based Drug Delivery System

A CD44‐targeted macromolecular conjugate of docetaxel was prepared via a pH‐sensitive linkage to hyaluronic acid and was characterized using NMR, gel permeation chromatography, and differential scanning calorimetry. The conjugated species were further evaluated in terms of drug release, cytotoxicity, cellular uptake, cell cycle inhibition, and subacute toxicity in mice. Cellular microscopic studies revealed that CD44‐expressing cells including MCF‐7 cancer stem cells and MDA‐MB‐231 metastatic breast cancer cells had internalized the conjugates via a selective receptor‐mediated mechanism, leading to cell cycle arrest in the G2/M phase. Hyaluronic acid–docetaxel conjugates showed specific toxicity only in CD44‐expressing cells in vitro, along with a decreased risk of neutropenia and dose‐dependent mortality in vivo. Hyaluronic acid–drug conjugates represent a promising and efficient platform for solubilization of sparingly soluble molecules as well as active and selective targeted delivery to cancer cells and cancer stem cells.     

Uptake and Cytotoxicity of Docetaxel-Loaded Hyaluronic Acid-Grafted Oily Core Nanocapsules in MDA-MB 231 Cancer Cells

Purpose

It is hypothesized that docetaxel (Doc)-loaded hyaluronic acid (HA)-polyethylene glycol/poly(ε-caprolactone)-grafted oily core nanocapsules (NCs) can enhance the drug cytotoxicity and uptake in CD44 expressing breast cancer (BC) cells (MDA-MB 231).

Methods

NCs were prepared, optimized and characterized by dynamic light scattering, transmission electron microscopy (TEM), and powder X-ray diffraction (PXRD). In vitro cytotoxicity tests [MTS, level of reactive oxygen species (ROS) and level of reduced glutathione (GSH)] were performed in BC cells. The contribution of CD44 to the NCs cellular uptake was elucidated using an anti CD44 antibody blockage and a CD44 negative NIH3T3 cell line.

Results

The optimum formulation of Doc-loaded HA oily core NCs had respective mean diameter, polydispersity, and drug encapsulation efficiency of 224.18 nm, 0.32, and 60.38%. The NCs appeared spherical with low drug crystallinity, while the drug release data fitted to first order equation. Compared to that of ungrafted NCs, the cytotoxicity of Doc-loaded HA-grafted NCs was significantly enhanced (p<0.05). A decrease of the intracellular level of ROS was reversely correlated with that of GSH. Interestingly, the cellular internalization of HA-grafted NCs mediated CD44 was dramatically enhanced (3 to 4-fold) with respect to the absence of specific biomarker or targeting ligand.

Conclusions

The use of HA-grafted NCs enhanced the selective drug payload, cytotoxicity and uptake in MDA-MB 231 cells. Therefore, it could be a promising template for safe and effective delivery of Doc and similar chemotherapeutic agents in cancer cells.     

结肠癌靶向米托蒽醌脂质体的制备及其抗肿瘤活性研究

目的:比较透明质酸(HA)修饰的脂质体(HA-LP)与普通脂质体(LP)的抗转移及抗肿瘤活性。方法:采用薄膜水化法制备空白脂质体,通过透明质酸上的羧基与膜材中DSPE的氨基反应将HA连接在脂质体上,运用硫酸铵梯度法包载模型药物米托蒽醌;通过透射电镜表征脂质体形态;免疫荧光法检测结肠癌细胞(CT26 cells)中透明质酸受体CD44表达情况;高内涵扫描仪检测结肠癌细胞对脂质体的摄取情况;划痕实验考察2组脂质体抑制细胞转移的能力;体内抗肿瘤实验考察2组脂质体的抑瘤效果。结果:所制备的脂质体形态均一,包封率均大于95%。免疫荧光结果显示,结肠癌细胞中CD44受体高度表达。相同的药物浓度下,HA-LP相较于未修饰的脂质体具有明显增加的细胞摄取。划痕实验结果表明,HA可能与受体CD44存在某种相互作用,抑制了结肠癌细胞的转移,表现出HA-LP具有更强的抗转移活性。体内抗肿瘤结果显示,HA-LP对小鼠荷结肠癌实体瘤的抑瘤率为62.1%,显著高于普通脂质体组(46.6%,P<0.05)。结论:HA-LP与细胞膜上的受体CD44作用,显著增加了其细胞摄取,并抑制了结肠癌细胞的转移,体内抑瘤结果同样显示,具有肿瘤靶向功能的HA-LP具有更强的抑制肿瘤生长的能力。     

索拉非尼联合多柔比星对人乳腺癌MDA-MB-231细胞和人肺癌A549细胞抑制作用的研究

目的观察索拉非尼联合多柔比星对人乳腺癌MDA-MB-231细胞和人肺癌A549细胞的抑制作用。方法不同浓度索拉非尼联合多柔比星处理人乳腺癌MDA-MB-231细胞和人肺癌A549细胞。①以CCK-8法检测处理24、48和72 h后MDA-MB-231和A549的抑制率;②以细胞划痕试验观察受试细胞的迁移能力;③以印迹试验检测处理24 h后MDAMB-231细胞内P-ERK和Bcl-2的表达情况。结果索拉非尼联合多柔比星能显著抑制人肺癌A549细胞和人乳腺癌MDAMB-231细胞的体外增殖,且具时间依赖性;人乳腺癌MDA-MB-231细胞经不同浓度索拉非尼联合多柔比星处理24 h,P-ERK和Bcl-2蛋白的表达明显下调并呈剂量依赖性。结论索拉非尼联合多柔比星具有抑制人乳腺癌MDA-MB-231细胞和人肺癌A549细胞的增殖,诱导细胞凋亡的作用。     

槐耳颗粒联合DC-CIK细胞对乳腺癌MDA-MB-231干细胞体内外杀伤作用研究

目的 探究白屈菜红碱（CHE）对腺样囊性癌细胞（ACC2）生长的抑制作用及机制。方法 利用CCK8法、EdU法、Hoechst33342/PI双染色法、试剂盒法检测CHE对ACC2细胞活力、细胞增殖、细胞凋亡和活性氧（ROS）水平的影响;通过Western blotting技术检测CHE对Cleaved-Caspase 3、PARP、NF-κB、p-JNK、p-p38蛋白表达的影响;利用斑马鱼移植瘤模型检测CHE对斑马鱼体内ACC2细胞生长的抑制作用。结果 CCK-8结果显示：与对照组比较,2、3、4、5、6、7、8、9、10 μmol/L的CHE显著降低ACC2细胞的存活率（P<0.05、0.01）,且呈浓度相关性; ROS检测结果显示：与对照组比较,5、8 μmol/L的CHE导致ACC2细胞内的ROS水平显著上升（P<0.05、0.01）; EdU增殖检测结果表明：与对照组比较,5、8 μmol/L的CHE致使ACC2细胞的增殖能力显著下降（P<0.01）;Hoechst/PI染色结果显示：与对照组比较,CHE 5、8 μmol/L组ACC2细胞凋亡率显著上升（P<0.01）。抗氧化剂N-乙酰半胱氨酸（NAC）显著抑制CHE诱导的ROS水平升高、细胞凋亡增加（P<0.01）;Western blotting结果显示：2、5、8 μmol/L的CHE能够显著上调Cleaved-Caspase 3、PARP、NF-κB蛋白的表达（P<0.01）,且呈现浓度相关性,5、8 μmol/L的CHE能够显著上调p-JNK的蛋白表达（P<0.01）,8 μmol/L的CHE能够显著上调p-p38的蛋白表达（P<0.01）;NAC显著降低由CHE导致的Cleaved-Caspase 3、PARP、NF-κB、p-JNK、p-p38蛋白表达增加（P<0.01）,5、8 μmol/L CHE能够有效抑制斑马鱼体内肿瘤的生长（P<0.01）。结论 体外及斑马鱼移植瘤模型证明,CHE可以有效抑制ACC2细胞生长,其机制与提高细胞ROS水平,上调NF-κB、p-JNK、p-p38表达,从而抑制细胞增殖、诱导细胞凋亡相关。     

Improved delivery of miR-1296 loaded cationic nanoliposomes for effective suppression of triple negative breast cancer

Nowadays, microRNA is considered an attractive strategy for the effective treatment of cancer. A significant delivery of microRNA for cancer therapy remains a significant obstacle to target cancer cells. The restoring microRNA-1296 (miR-1296) has immense therapeutic efficacy in triple-negative breast cancer (TNBC). TNBC is an aggressive subtype of breast tumors with the progression of malignant transformation. This study aimed to develop a cationic nanoliposome that can serve as a miR-1296 carrier and studied its efficiency in TNBC. The efficacy of miR-1296 liposomes was evaluated on its apoptotic effect, cellular uptake, and potential chemotherapy sensitization in the TNBC cell line (MDA-MB-231). For in vitro viability study, the apoptotic effect was performed to validate protein expression using Alamar blue kit and western blot. The transfection of miR-1296 into TNBC cells was also investigated using cisplatin as a TNBC resistance drug. The fluorescent miR-1296-cy3 liposome was used for cellular uptake study. The miR-liposome was successfully prepared with a particle size of 123.6 ± 1.3 nm and encapsulation efficiency of 94.33%. A dose of 0.5 uM has significantly reduced the viability of MDA-MB-231 to be 33.45%±5.29 (P < 0.001). This result was validated by down-expression of CCND1, and PARP1, the miR-1296 receptor, and apoptosis marker. The image of the miR-1296-cy3 liposome showed cytoplasmic intracellular localization. It was found high sensitization of TNBC cell line for miR-1296 liposome compared to cisplatin (P < 0.001). Future in vivo research may answer questions concerning safety and stability. This study demonstrates that miR-191 liposomes may have promising clinical applications for TNBC therapy.     

Multifunctional Tumor-Targeting Nanocarriers Based on Hyaluronic Acid-Mediated and pH-Sensitive Properties for Efficient Delivery of Docetaxel

Purpose

The objective of this work was to develop a multifunctional tumor-targeting nanocarrier based on the mechanism of CD44-mediated endocytosis and pH-induced drug release to improve the therapeutic efficacy of docetaxel (DTX).

Methods

Hyaluronic acid-coated docetaxel-loaded cholesteryl hemisuccinate vesicles (HA-CHEMS vesicles) were prepared. The physiochemical properties and pH-dependent drug release of HA-CHEMS vesicles were evaluated. The HA-CHEMS vesicles were investigated for CD44-mediated internalization and in vitro cell viability using MCF-7,A549 and L929 cells.In addition,tissue distribution as well as antitumor efficacy was also evaluated in MCF-7 tumor-bearing mouse model.

Results

The particle size and zeta potential of HA-CHEMS vesicles were 131.4?±?6.2 nm and ?13.3?±?0.04 mV,respectively. The in vitro drug release results demonstrated a pH-responsive drug release under different pH conditions. In vitro cell viability tests suggested that the encapsulation of DTX in HA-CHEMS vesicles led to more than 51.6-fold and 46.3-fold improved growth inhibition in MCF-7 and A549 cell lines,respectively compared to Taxotere®. From the cell uptake studies,the coumarin 6-loaded HA-CHEMS vesicles enhanced intracellular fluorescent intensity in the CD44-overexpressing cell line (MCF-7). Biodistribution studies revealed selective accumulation of HA-CHEMS vesicles in the MCF-7 bearing BalB/c nude mice as a result of passive accumulation and active targeting (CD44-mediated endocytosis). Compared to Taxotere®,HA-CHEMS vesicles exhibited higher antitumor activity by reducing tumor volume (P?<?0.05) and drug toxicity,demonstrating the success of the multifunctional targeting delivery.

Conclusions

This work corresponds to the preparation of a multifunctional tumor-targeted delivery system. Our investigation shows that hyaluronan-bearing docetaxel-loaded cholesteryl hemisuccinate vesicles (HA-CHEMS vesicles) is a highly promising therapeutic system,leading to tumor regression after intravenous administration without visible toxicity.     

Docetaxel-loaded PLGA nanoparticles to increase pharmacological sensitivity in MDA-MB-231 and MCF-7 breast cancer cells

This study aimed to develop docetaxel (DTX) loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (DTX-NPs) and to evaluate the different pharmacological sensitivity of NPs to MCF-7 and MDA-MB-231 breast cancer cells. NPs containing DTX or coumarin-6 were prepared by the nanoprecipitation method using PLGA as a polymer and d-α-tocopherol polyethylene glycol 1000 succinate (TPGS) as a surfactant. The physicochemical properties of NPs were characterized. In vitro anticancer effect and cellular uptake were evaluated in breast cancer cells. The particle size and zeta potential of the DTX-NPs were 160.5 ± 3.0 nm and –26.7 ± 0.46 mV, respectively. The encapsulation efficiency and drug loading were 81.3 ± 1.85% and 10.6 ± 0.24%, respectively. The in vitro release of DTX from the DTX-NPs was sustained at pH 7.4 containing 0.5% Tween 80. The viability of MDA-MB-231 and MCF-7 cells with DTX-NPs was 37.5 ± 0.5% and 30.3 ± 1.13%, respectively. The IC₅₀ values of DTX-NPs were 3.92- and 6.75-fold lower than that of DTX for MDA-MB-231 cells and MCF-7 cells, respectively. The cellular uptake of coumarin-6-loaded PLGA-NPs in MCF-7 cells was significantly higher than that in MDA-MB-231 cells. The pharmacological sensitivity in breast cancer cells was higher on MCF-7 cells than on MDA-MB-231 cells. In conclusion, we successfully developed DTX-NPs that showed a great potential for the controlled release of DTX. DTX-NPs are an effective formulation for improving anticancer effect in breast cancer cells.     

Targeting Effect of Betulinic Acid Liposome Modified by Hyaluronic Acid on Hepatoma Cells In Vitro

Betulinic acid (BA) is a natural pentacyclic triterpenoid with broad-spectrum anticancer activity, which has great development potential as an anti-cancer drug. In this study, a novel hyaluronic acid (HA)-modified BA liposome (BA-L) was developed for use in targeted liver cancer therapy. The size, polymer dispersity index (PDI), zeta potential, and entrapment efficiency were measured. Cell viability, cell migration and clonogenicity, cellular uptake, immunohistochemistry of CD44, and protein expression of ROCK1/IP3/RAS were also investigated. BA, BA-L, and HA-BA-L had no inhibitory effect on the activity of LO2 normal hepatocytes, but they inhibited the proliferation of HepG2 and SMMC-7721 cells in a dose- and time-dependent manner, with HA-BA-L exhibiting the most prominent inhibitory effect. Compared with the BA-L group, the expression of CD44 in HepG2 cells in the HA-BA-L group was decreased. The results of WB showed that BA, BA-L, and HA-BA-L downregulated the expression of ROCK1, IP3, and RAS in HepG2 cells, and the expression level in the HA-BA-L group was significantly decreased. The easily prepared HA-BA-L was demonstrated to be an excellent CD44-mediated intracellular delivery system capable of targeting effects. Further mechanistic research revealed that the inhibition of HA-BA-L on HepG2 cells may be mediated by blocking the ROCK1/IP3/RAS signaling pathways.     

Caffeic Acid Phenethyl Ester (CAPE) derived from propolis, a honeybee product, inhibits growth of breast cancer stem cells

Cancer stem cells (CSC) are chemoresistant and implicated in tumor recurrence, metastasis and high patient mortality; thus substances impairing CSC activity, could be invaluable as novel cancer therapeutics. We previously showed that CAPE (caffeic acid phenethyl ester), a component of propolis, a honeybee product, inhibits growth of MDA-MB-231 (MDA-231) cells, mdr gene expression, NF-κB, EGFR, and VEGF. We hypothesized that CAPE also acts by interfering with CSC-mediated effects. We isolated breast CSC (bCSC) from MDA-231 cells, a model of human triple-negative breast cancer, and mouse xenografts. bCSC grow as mammospheres (MMS) and when dissociated into single cells, form MMS again, a sign of self-renewal. bCSC exhibited the characteristic CD44(+)/CD24(-/low) phenotype and generated progenitors in the presence of serum, a CSC trait responsible for regenerating tumor mass. CAPE caused dose-dependent bCSC self-renewal inhibition and progenitor formation. Clonal growth on soft agar was inhibited dose-dependently, but apoptosis was not induced as determined by Annexin-V/PI assay. Instead, bCSC were noted to significantly progress from a quiescent cell cycle state in G0/G1 (82%), S phase (12%) to a cycling state with an increase in S phase (41%) and subsequent decrease in G0/G1 (54%). Treatment of bCSC with CAPE (4.5-days) decreased CD44 levels by 95%, while another cell population containing 10-100-fold lower CD44 content concurrently increased. Results suggest that CAPE causes pronounced changes in bCSC characteristics manifested by inhibition of self renewal, progenitor formation, clonal growth in soft agar, and concurrent significant decrease in CD44 content, all signs of decreased malignancy potential.     

Ultrasound-enhanced chemotherapy of drug-resistant breast cancer tumors by micellar-encapsulated Paclitaxel

Background and aim

Currently, delivery of the poorly water-soluble chemotherapeutic agent paclitaxel is associated with a substantial array of systemic toxicities and results in low-efficiency tumor treatment. A novel on-demand delivery system based on paclitaxel encapsulated in polymeric micelles in conjunction with triggered release of the drug by local ultrasonic irradiation of the tumor was evaluated in vitro and in vivo using a drug-resistant MCF7/ADmt breast cancer human cell line.

Method

The effects of local ultrasonic tumor irradiation on cellular proliferation and intracellular drug uptake were compared for a developmental micellar paclitaxel formulation (SYP-PM) and a currently available clinical intravenous formulation of paclitaxel.

Results

Without ultrasound, the uptake of paclitaxel from the micellar formulation was significantly lower than that from the clinical formulation, which is advantageous for preventing unwanted drug interactions with healthy tissues in vivo. When micellar encapsulation was combined with ultrasonically triggered release, drug uptake from micellar paclitaxel was increased more than 20-fold and cellular proliferation was inhibited by nearly 90%. Without ultrasound, the clinical formulation of paclitaxel and SYP-PM manifested low efficacy in vivo, whereas injections of SYP-PM combined with ultrasound resulted in complete tumor resolution.

Conclusion

The ability of micellar-encapsulated paclitaxel to exert a significant cytotoxic effect only when subjected to ultrasound proves promising for the development of a tumor-targeted ultrasound-enhanced paclitaxel delivery system for clinical application. This treatment modality could be successfully used for the therapy of both drug-sensitive and drug-resistant tumors. The major advantages of a micellar formulation of paclitaxel combined with local tumor sonication are the aqueous base of the drug formulation, reduced systemic toxicity, potential for tumor targeting, and on-demand delivery of drug to tumor cells.     

肿瘤转移靶向肽修饰的阿霉素脂质体对高转移性乳腺癌细胞的靶向特异性研究

肿瘤转移日渐成为肿瘤治疗的重要靶标。本研究采用肿瘤转移靶向肽（TMT）与脂质材料（PEG-DSPE）偶联获得靶向化合物（TMT-PEG-DSPE）,用以构建靶向阿霉素脂质体（TMT-LS-DOX）。结果表明,TMT-LS-DOX呈现出良好的药剂学性质。选用高转移性乳腺癌细胞（MDA-MB-435S和MDA-MB-231）对该转移特异性递送系统进行评价,采用非转移性乳腺癌细胞（MCF-7）作为对照。游离TMT多肽浓度达100μg/mL时仍未显示出细胞毒性。与MCF-7相比,MDA-MB-435S及MDA-MB-231细胞对TMT-LS-DOX摄取增加,并经受体竞争性实验证明该促进作用由TMT介导。因此,TMT修饰的纳米载体可能成为增加化疗药物对高转移性乳腺癌特异性的一种新策略。     

miR-200c/PAI-2 promotes the progression of triple negative breast cancer via M1/M2 polarization induction of macrophage

PurposeTo investigate the effect of miR-200c/PAI-2 on macrophage polarization into M2-type TAMs in TNBC.Methods and materialsPAI-2 expression in MDA-MB-231^con, MDA-MB-231^miR-200ab and MDA-MB-231^miR-200c breast cancer cells was evaluated by RT-PCR and immunofluorescence (IF), while the expression of the TAM marker F4/80 and the M2-type TAM marker CD206 in MDA-MB-231^con, MDA-MB-231^miR-200c and MDA-MB-231^{miR-200c/siPAI-2} mouse lung metastatic tumor tissues was examined with immunohistochemistry (IHC). The effects of RAW264.7 cells on MDA-MB-231^con, MDA-MB-231^miR-200c and MDA-MB-231^{miR-200c/siPAI-2} were examined by transwell co-culture. CD206 expression in RAW264.7 cells were confirmed by immunostaining. The level of PAI-2 and IL-10 in the co-culture supernatants were assessed using ELISA.Results1. RT-PCR and IF analysis showed that PAI-2 was upregulated in MDA-MB-231^miR-200c cells. 2. IHC assays analysis showed that the numbers of F4/80 and CD206 positive cells were increased in MDA-MB-231^miR-200c tumor tissues, while in MDA-MB-231^{miR-200c/siPAI-2} tumor tissues were decreased. 3. Transwell co-culture assays analysis showed that MDA-MB-231^miR-200c cells significantly promoted the cell migration ability compared with the control group, while knockdown PAI-2 significantly inhibited the cell migration ability (P < 0.05). 4. Transwell co-culture and immunostaining assays analysis showed that overexpression miR-200c in MDA-MB-231 cell line increased the CD206 expression in RAW264.7 cells, while knockdown PAI-2 decreased. 5. ELISA assays analysis showed that miR-200c-mediated MDA-MB-231 cells significantly increased the secretion of PAI-2 and IL-10, while decreased the secretion of PAI-2 and IL-10 in MDA-MB-231 ^{miR-200c/siPAI-2} cells.ConclusionsmiR-200c promotes the malignant progressions of TNBC by PAI-2 upregulation and M2 phenotype macrophages polarization.     

Cell penetrating peptides functionalized gambogic acid-nanostructured lipid carrier for cancer treatment

Tumor-targeted delivery is considered a crucial component of current anticancer drug development and is the best approach to increase the efficacy and reduce the toxicity. Nanomedicine, particularly ligand-based nanoparticles have shown a great potential for active targeting of tumor. Cell penetrating peptide is one of the promising ligands in a targeted cancer therapy. In this study, the gambogic acid-loaded nanostructured lipid carrier (GA-NLC) was modified with two kinds of cell penetrating peptides (cRGD and RGERPPR). The GA-NLC was prepared by emulsification and solvent evaporation method and coupled with cRGD, RGERPPR, and combination cRGD and RGERPPR to form GA-NLC-cRGD, GA-NLC-RGE, and GA-NLC-cRGD/RGE, respectively. The formulations were characterized by their particle size and morphology, zeta potential, encapsulation efficiency, and differential scanning calorimetry. In vitro cytotoxicity and cellular uptake study of the formulations were performed against breast cancer cell (MDA-MB-231). Furthermore, in vivo biodistribution and antitumor activity of the formulations were determined by in vivo imaging and in tumor-bearing nude mice, respectively. The result of in vitro cytotoxicity study showed that GA-NLC-RGE exhibited a significantly higher cytotoxicity on MDA-MB-231 as compared with GA-NLC and GA-Sol. Similarly, RGE-Cou-6-NLC showed remarkably higher uptake by the cells than other NLCs over the incubation period. The in vivo imaging study has demonstrated that among the formulations, the RGE-decorated DiR-NLC were more accumulated in the tumor site. The in vivo antitumor activity revealed that RGE-GA-NLC inhibits the tumor growth more efficiently than other formulations. In conclusion, RGERPPR has a potential as an effective carrier in targeting drug delivery of anticancer agents.     

Selective delivery of PLXDC1 small interfering RNA to endothelial cells for anti-angiogenesis tumor therapy using CD44-targeted chitosan nanoparticles for epithelial ovarian cancer

Angiogenesis plays an essential role in the growth and metastasis of tumor cells, and the modulation of angiogenesis can be an effective approach for cancer therapy. We focused on silencing the angiogenic gene PLXDC1 as an important factor for anti-angiogenesis tumor therapy. Herein, we developed PLXDC1 small interfering siRNA (siRNA)-incorporated chitosan nanoparticle (CH-NP/siRNA) coated with hyaluronic acid (HA) to target the CD44 receptor on tumor endothelial cells. This study aimed to improve targeted delivery and enhance therapeutic efficacy for tumor anti-angiogenesis. The HA-CH-NP/siRNA was 200?±?10?nm in size with a zeta potential of 26.4?mV. The loading efficiency of siRNA to the HA-CH-NP/siRNA was up to 60%. The selective binding of HA-CH-NP/siRNA to CD44-positive tumor endothelial cells increased by 2.1-fold compared with that of the CD44 nontargeted CH-NP/siRNA. PLXDC1 silencing by the HA-CH-NP/siRNA significantly inhibited tumor growth in A2780 tumor-bearing mice compared with that in the control group (p?p?p?p?相似文献   

A combination of LightOn gene expression system and tumor microenvironment-responsive nanoparticle delivery system for targeted breast cancer therapy

A light-switchable transgene system called LightOn gene expression system could regulate gene expression with a high on/off ratio under blue light, and have great potential for spatiotemporally controllable gene expression. We developed a nanoparticle drug delivery system (NDDS) to achieve tumor microenvironment-responsive and targeted delivery of diphtheria toxin A (DTA) fragment-encoded plasmids to tumor sites. The expression of DTA was induced by exposure to blue light. Nanoparticles composed of polyethylenimine and vitamin E succinate linked by a disulfide bond, and PEGylated hyaluronic acid modified with RGD peptide, accumulated in tumor tissues and were actively internalized into 4T1 cells via dual targeting to CD44 and α_vβ₃ receptors. The LightOn gene expression system was able to control target protein expression through regulation of the intensity or duration of blue light exposure. In vitro studies showed that light-induced DTA expression reduced 4T1 cell viability and induced apoptosis. Furthermore, the LightOn gene expression system enabled spatiotemporal control of the expression of DTA in a mouse 4T1 tumor xenograft model, which resulted in excellent antitumor effects, reduced tumor angiogenesis, and no systemic toxicity. The combination of the LightOn gene expression system and NDDS may be an effective strategy for treatment of breast cancer.