Multidrug resistance (MDR) remains a significant impediment to chemotherapy during cancer therapy. In this study, the amphiphilic biomaterials PEI-TOS and HA-QU were synthesized to self-assemble into PEI-TOS/HA-QU core-shell micelles for the targeted codelivery of paclitaxel (PTX) and quercetin (QU) to alleviate multidrug drug resistance and enhance therapeutic efficacy. The PTX-loaded micelles possessed a uniform particle size (167.60 ± 8.185 nm), stable negative charge (-19.13 ± 0.321 mV), and pH-responsive drug release with good compatibility. The drug-loaded micelles increased the chemosensitivity of MDR tumor cells (MDA-MB-231/MDR1) to PTX and activated mitochondria-dependent apoptotic pathways (the IC50 was 2.22-fold lower than that of PTX alone). Moreover, PEI-TOS/HA-QU micelles increased the cellular uptake of lipophilic antitumor drugs by downregulating P-gp expression in MDA-MB-231/MDR1 cells. Compared with Taxol, PTX-loaded PEI-TOS/HA-QU micelles presented excellent antitumor efficacy in tumor-bearing mice, with an average tumor size that was 3.7-fold lower than that of the control group. The drug-loaded formulation showed low in vitro / in vivo toxicity and better tumor accumulation than the free drug, which led to a high tumor inhibition rate of 80.56% and considerable biocompatibility. This work describes a new platform for the codelivery of lipophilic anticancer drugs and natural active ingredients such as PTX and QU for the treatment of MDR cancer cells.

The therapeutic efficacy of chemotherapy in many types of hematological malignancies and solid tumors is dramatically hindered by multidrug resistance (MDR). This work presents a combination strategy of pretreatment of MDA-MB-231/MDR1 cells with quercetin (QU) followed by doxorubicin (DOX) to overcome MDR, which can be delivered by mixed micelles composed of the reduction-sensitive hyaluronic acid-based conjugate and d-α-tocopheryl poly(ethylene glycol) 1000 succinate. The combination strategy can enhance the cytotoxicity of DOX on MDA-MB-231/MDR1 cells by increasing intracellular DOX accumulation and facilitating DOX-induced apoptosis. The probable MDR reversal mechanisms are that the pretreatment cells with QU-loaded mixed micelles downregulate P-glycoprotein expression to decrease DOX efflux as well as initiate mitochondria-dependent apoptotic pathways to accelerate DOX-induced apoptosis. In addition, this combination strategy can not only potentiate in vivo tumor-targeting efficiency but also enhance the antitumor effect in MDA-MB-231/MDR1-bearing nude mice without toxicity or side effects. This research suggests that the co-administration of natural compounds and chemotherapeutic drugs could be an effective strategy to overcome tumor MDR, which deserves further exploration.

目的 比较乙醇、丙二醇、混合醇(乙醇-丙二醇2:8)3种柔软剂对制备光甘草定醇质体(GLA-ES)的影响,为难溶性药物的醇质体研究提供柔软剂选择依据.方法 分别选用乙醇、丙二醇、混合醇(乙醇-丙二醇2∶8)作为柔软剂,采用注入-超声结合法制备GLA-ES,并进行不同GLA-ES的形态、粒径、Zeta电位、包封率、稳定性和体外释药等考察.以黑色素瘤B 16-OVA细胞内酪氨酸酶活性评价GLA-ES抑制黑色素合成能力,采用铁氰化钾还原力实验评估其抗氧化效果,并使用人表皮HaCaT细胞和大鼠皮肤进行初步安全性评价.结果 所得GLA-ES为黄色半透明液体,具有囊泡状磷脂双分子层结构;光甘草定乙醇醇质体(GLA-Et-ES)、丙二醇醇质体(GLA-PG-ES)和混合醇醇质体(GLA-MA-ES)的平均粒径分别为(34.24±0.29)、(62.31±1.66)、(41.20±1.13) nm,电位分别为(-41.0±1.8)、(-32.9±0.2)、(-35.8±1.6) mV,包封率分别为(91.47±2.39)％、(87.33±1.31)％、(91.39±3.59)％,在4℃放置20 d较稳定;其体外释药行为均符合Higuchi方程,具有一定缓释效果.与光甘草定混悬液相比,GLA-Et-ES、GLA-PG-ES和GLA-MA-ES对黑色素瘤B16-OVA细胞的酪氨酸酶活性抑制率分别提高了38.07％、19.58％、40.42％;铁氰化钾还原力实验结果也显示GLA-ES有较强的体外抗氧化作用;GLA-ES对正常细胞几乎没有毒性,对大鼠皮肤无刺激性.结论 3种柔软剂均可制备GLA-ES,其抑制黑色素生成、抗氧化作用增强,安全性良好,为进一步开发美白护肤产品或医药外用制剂奠定基础.对于光甘草定这类难溶性药物,选择混合醇(乙醇-丙二醇2∶8)作为柔软剂,所制备的醇质体的包封率、稳定性优于单用乙醇或丙二醇.

Pharmaceutical cocrystals are a promising technology that can be used to improve the solubility of poor aqueous compounds. The objective of this study was to systematically investigate the solubility of myricetin (MYR) cocrystals, including their kinetic solubility, thermodynamic solubility, and intrinsic dissolution rate (IDR). The effects of pH, surfactant, ion concentration, and coformers on the cocrystal solubility were evaluated. Furthermore, single crystal structures of MYR, myricetin–isonicotinamide (MYR–INM) and myricetin–caffeine (MYR–CAF) cocrystals were analyzed to discuss the possible reasons for the enhancement of cocrystal solubility from the perspective of the spatial structure. The results indicated that the kinetic solubility of MYR cocrystals was modulated by pH and cocrystal coformer (CCF) ionization in buffer solution, while it primarily depended on the CCF solubility in pure water. In addition, the solubility of MYR cocrystals was increased in a concentration dependent fashion by the surfactant or ion concentration. The thermodynamic solubility of MYR–INM (1:3) cocrystals decreased with the increases of the pH value of the dissolution media. The IDR of MYR cocrystals was faster than that of MYR in the same medium and extremely fast in pH 4.5 buffer. The improved solubility of MYR cocrystals was probably related to the alternate arrangements of MYR and INM/CAF molecules and increased intermolecular distance. The present study provides some references to investigate the solubility behavior of pharmaceutical cocrystals.

BACKGROUNDKruppel-like factors (KLFs) are involved in various biological processes; emerging studies have indicated that KLF9 plays a critical role in regulating tumorigenesis. The role of KLF9 in prostate cancer (PCa), however, has not yet been investigated. METHODSThe expression of KLF members, AKT- and apoptosis-related proteins were analyzed by Western blot or qRT-PCR. Tet-On inducible KLF9 expression was established for the evaluation of the effects of KLF9 on cell proliferation, apoptosis, and xenograft tumor growth in nude mice. Cell cycle and apoptosis were determined by flow cytometry. RESULTSKLF9 was induced in a time-dependent manner in flutamide-caused apoptosis, and knockdown of KLF9 significantly decreased flutamide-induced growth inhibition and apoptosis in LNCaP cells. The levels of KLF9 were relatively lower in PCa cell lines, particularly in androgen-independent cell lines compared with those in nontumorous prostate epithelial cell lines. Overexpression of KLF9 dramatically suppressed cell proliferation and caused cell cycle arrest in the G2/M phase and cell apoptosis in the androgen-independent cell lines, PC3 and DU145. Intriguingly, KLF9 expression severely suppressed the activation of AKT and its downstream targets. AKT reactivation partially rescued the KLF9-mediated inhibitory effects on the proliferation of PCa cells. More importantly, we found that KLF9 overexpression efficiently inhibited the xenograft tumor growth of PCa cells. CONCLUSIONSThese data collectively showing that KLF9 substantially inhibits AKT activation and abrogates tumor growth of PCa cells, suggest the potential of either genetic or pharmacological activation of KLF9 in the therapeutic treatment of castration-resistant PCa. Prostate 74:946-958, 2014. (c) 2014 Wiley Periodicals, Inc.

Strategies for peroxisome proliferator-activated receptor (PPAR) activation or survivin inhibition have potential for cancer therapy. However, whether the combination of these two approaches can be developed as a rational regimen with enhanced efficiency in the inhibition of tumor cells remains to be determined. In this study, the combinatory effect of PPAR-γ agonist and survivin inhibition on bladder cancer cells was investigated. T24 and 5637 cells were treated with 15d-PGJ(2) to determine whether 15d-PGJ(2) had an inhibitory effect. Cell viability and proliferation were analyzed and efficiency of survivin siRNAs was assessed using western blot analysis. The results showed that, in the human bladder cancer cell lines T24 and 5637, the natural PPAR-γ ligand 15d-PGJ(2) significantly decreased cell proliferation and loci formation. The increase in the proportion of apoptotic cells was observed in the cells 48 h after 15d-PGJ(2) treatment. Furthermore, 15d-PGJ(2) substantially inhibited the levels of stemness-related genes in these cells. The ability of sphere formation was markedly suppressed in the cells treated with 15d-PGJ(2). More importantly, the downregulation of survivin with siRNAs significantly enhanced the 15d-PGJ(2)-mediated induction of cell apoptosis and inhibition of sphere formation. Accordingly, we also found that survivin inhibition significantly enhanced 15d-PGJ(2)-induced production of reactive oxygen species (ROS) in bladder cancer cells. Taken together, these findings suggest that the combination of 15d-PGJ(2) and survivin inhibition play a potentially role in the therapeutical manipulation of bladder cancer.

Krüppel-like factor 9 (KLF9) is known to be a tumor suppressor gene in colorectal tumors and glioblastoma; however, the functional status and significance of KLF9 in hepatocellular carcinoma (HCC) is unclear. We report here that KLF9 is downregulated in HCC tissues. Restoration of KLF9 significantly inhibited growth and caused apoptosis in SK-Hep1 and HepG2 cells. We found that KLF9 positively regulated p53 levels by directly binding to GC boxes within the proximal region of the p53 promoter. Moreover, in the presence of cycloheximide, KLF9 significantly increased p53 stability in HCC cells. Remarkably, ectopic expression of KLF9 was sufficient to delay the onset of tumors and to promote regression of the established tumors in vivo, suggesting that KLF9 plays a critical role in HCC development and that pharmacological or genetic activation of KLF9 may have potential in the treatment of HCC.

The generation of CD 4 + F oxp3 + T reg cells in the thymus is crucial for immune homeostasis and self‐tolerance. Recent studies have shown T reg‐cell plasticity when T h‐related transcriptional factors and cytokines are present. However, the mechanisms that maintain the stability of T reg cells are poorly understood. Here, using mice with a T ‐cell‐specific deletion of the transforming growth factor‐β receptor 2 ( T gfbr2 −/− mice), we identify the restriction of AKT activation as a key event for the control of T reg‐cell stability in T h1 inflammation. AKT regulation was evident in thymic CD 4 + F oxp3 + T reg cells before they egressed to peripheral tissues. CD 4 + F oxp3 + thymocytes from mice with the T gfbr2 deletion expressed high levels of CXCR 3 and T ‐bet, and produced IFN ‐γ and TNF ‐α. Thymic T gfbr2 −/− T reg cells also showed an increase in the activation of AKT pathway. Enhanced AKT activity induced the expression of IFN ‐γ both in natural and inducible T reg cells. Inhibition of AKT activity markedly attenuated the expression of IFN ‐γ and TNF ‐α in thymic T gfbr2 −/− T reg cells in vivo. In addition, mixed bone marrow transplantation showed that TGF ‐β signaling maintained T reg‐cell stability in an intrinsic manner. Our results demonstrate that AKT hyperactivation contributes to the conversion of T reg cells to a T h1 phenotype.

Emerging evidence suggests that tumor-initiating cells (TICs) are the most malignant cell subpopulation in tumors because of their resistance to chemotherapy or radiation treatment. Targeting TICs may be a key innovation for cancer treatment. In this study, we found that PPARγ agonists inhibited the cancer stem cell-like phenotype and attenuated tumor growth of human hepatocellular carcinoma (HCC) cells. Reactive oxygen species (ROS) initiated by NOX2 upregulation were partially responsible for the inhibitory effects mediated by PPARγ agonists. However, PPARγ agonist-mediated ROS production significantly activated AKT, which in turn promoted TIC survival by limiting ROS generation. Inhibition of AKT, by either pharmacological inhibitors or AKT siRNA, significantly enhanced PPARγ agonist-mediated inhibition of cell proliferation and stem cell-like properties in HCC cells. Importantly, in nude mice inoculated with HCC Huh7 cells, we demonstrated a synergistic inhibitory effect of the PPARγ agonist rosiglitazone and the AKT inhibitor triciribine on tumor growth. In conclusion, we observed a negative feedback loop between oxidative stress and AKT hyperactivation in PPARγ agonist-mediated suppressive effects on HCCs. Combinatory application of an AKT inhibitor and a PPARγ agonist may provide a new strategy for inhibition of stem cell-like properties in HCCs and treatment of liver cancer.