Intravenous delivery of anti-RhoA small interfering RNA loaded in nanoparticles of chitosan in mice: safety and efficacy in xenografted aggressive breast cancer

Overexpression of RhoA in cancer indicates a poor prognosis, because of increased tumor cell proliferation and invasion and tumor angiogenesis. We showed previously that anti-RhoA small interfering RNA (siRNA) inhibited aggressive breast cancer more effectively than conventional blockers of Rho-mediated signaling pathways. This study reports the efficacy and lack of toxicity of intravenously administered encapsulated anti-RhoA siRNA in chitosan-coated polyisohexylcyanoacrylate (PIHCA) nanoparticles in xenografted aggressive breast cancers (MDA-MB-231). The siRNA was administered every 3 days at a dose of 150 or 1500 microg/kg body weight in nude mice. This treatment inhibited the growth of tumors by 90% in the 150-microg group and by even more in the 1500-microg group. Necrotic areas were observed in tumors from animals treated with anti-RhoA siRNA at 1500 microg/kg, resulting from angiogenesis inhibition. In addition, this therapy was found to be devoid of toxic effects, as evidenced by similarities between control and treated animals for the following parameters: body weight gain; biochemical markers of hepatic, renal, and pancreatic function; and macroscopic appearance of organs after 30 days of treatment. Because of its efficacy and the absence of toxicity, it is suggested that this strategy of anti-RhoA siRNA holds significant promise for the treatment of aggressive cancers.     

抑癌基因IL-24在乳腺癌细胞中的表达及意义

目的研究hIL-24（人白细胞介素24）对MDA-MB-231乳腺癌细胞的生长抑制作用。方法将pDC316-hIL-24-EGFP转染人乳腺癌MDA-MB-231细胞,用RT-PCR法、Western blot法检测IL-24基因在肿瘤细胞中的表达。结晶紫染色法检测IL-24基因的表达对乳腺癌细胞的生长抑制。RT-PCR检测侵袭、转移相关基因的转录。ELISA法检测VEGF、Ang-1的分泌水平。结果 IL-24基因可在MDA-MB-231细胞中成功转录及表达,并对乳腺癌细胞增殖有明显抑制作用。能明显下调MMP-2、MMP-9的转录。VEGF、Ang-1表达下降。结论 IL-24对乳腺癌细胞有明显的抑制生长、侵袭转移、血管生成的作用。     

BRMS1对乳腺癌细胞生物学特性影响的体外研究

为探讨乳腺癌转移抑制基因1（BRMS1）对乳腺癌细胞生物学特性影响,利用已建立的高表达BRMS1基因的乳腺癌细胞MDA-MB-231,研究了BRMS1基因对MDA-MB-231细胞的增殖活性、浸润迁移能力、侵袭能力、细胞周期和凋亡的影响.结果 表明：BRMS1基因通过乳腺癌MDA-MB-231细胞的侵袭能力而抑制肿瘤细胞的转移能力;而对MDA-MB-231细胞增殖活性、浸润迁移能力以及细胞周期和凋亡无影响.     

Ras同源类似物小干扰RNA对乳腺癌细胞恶性表型的影响

目的观察下调Ras同源类似物E(RhoE)表达对人乳腺癌细胞231生物学行为的影响。方法蛋白质印迹技术检测小干扰RNA(siRNA)转染前后RhoE在乳腺癌细胞231中的表达;RhoE siRNA的细胞转染用lipofectamine 2000脂质体法;Cell Counting Kit-8检测转染细胞及对照细胞的增殖变化;损伤刮擦试验和体外侵袭实验(Transwell小室)分别检测转染细胞及对照细胞的迁移与侵袭能力。结果 RhoE在乳腺癌细胞231中的表达较高;成功转染RhoE siRNA的乳腺癌细胞,蛋白质印迹显示RhoE的表达被明显的抑制;RhoE的表达被抑制后对乳腺癌细胞的增殖、迁移和侵袭有着明显的促进作用。结论下调RhoE表达能够明显促进乳腺癌细胞的增殖﹑迁移和侵袭,RhoE可能在乳腺癌的发生发展中起着重要作用。     

Peptidomimetic Src/Pretubulin Inhibitor KX-01 Alone and in Combination with Paclitaxel Suppresses Growth, Metastasis in Human ER/PR/HER2-Negative Tumor Xenografts

Src kinase is elevated in breast tumors that are ER/PR negative and do not overexpress HER2, but clinical trials with Src inhibitors have shown little activity. The present study evaluated preclinical efficacy of a novel peptidomimetic compound, KX-01 (KX2-391), that exhibits dual action as an Src and pretubulin inhibitor. KX-01 was evaluated as a single-agent and in combination with paclitaxel in MDA-MB-231, MDA-MB-157, and MDA-MB-468 human ER/PR/HER2-negative breast cancer cells. Treatments were evaluated by growth/apoptosis, isobologram analysis, migration/invasion assays, tumor xenograft volume, metastasis, and measurement of Src, focal adhesion kinase (FAK), microtubules, Ki67, and microvessel density. KX-01 inhibited cell growth in vitro and in combination with paclitaxel resulted in synergistic growth inhibition. KX-01 resulted in a dose-dependent inhibition of MDA-MB-231 and MDA-MB-157 tumor xenografts (1 and 5 mg/kg, twice daily). KX-01 inhibited activity of Src and downstream mediator FAK in tumors that was coincident with reduced proliferation and angiogenesis and increased apoptosis. KX01 also resulted in microtubule disruption in tumors. Combination of KX-01 with paclitaxel resulted in significant regression of MDA-MB-231 tumors and reduced metastasis to mouse lung and liver. KX-01 is a potently active Src/pretubulin inhibitor that inhibits breast tumor growth and metastasis. As ER/PR/HER2-negative patients are candidates for paclitaxel therapy, combination with KX-01 may potentiate antitumor efficacy in management of this aggressive breast cancer subtype. Mol Cancer Ther; 11(9); 1936-47. ?2012 AACR.     

Stathmin表达水平与乳腺癌细胞侵袭能力相关性研究

目的 探讨乳腺癌细胞MDA-MB-231和MCF-7中Stathmin基因表达水平与细胞生长、黏附、侵袭等生物学行为之间的关系,为进一步研究乳腺癌转移机制奠定实验基础。方法 应用RT-PCR和Western Blot方法检测MDA-MB-231和MCF-7细胞中Stathmin基因的表达水平,同时利用细胞增殖试验、细胞黏附试验和细胞侵袭试验检测MDA-MB-231和MCF-7细胞的生长、黏附、侵袭能力,分析Stathmin表达与细胞的生长、黏附、侵袭能力之间的关系。结果 RT-PCR和Western Blot检测结果显示,Stathmin基因在MDA-MB-231和MCF-7细胞中表达均高于正常对照细胞(F=10.173,P<0.05),且MDA-MB-231细胞中的表达水平明显高于MCF-7细胞中的表达水平(t=4.562,P<0.05)。而MDA-MB-231细胞在生长、黏附和侵袭能力方面均强于MCF-7细胞(P<0.05)。结论 Stathmin表达水平高的乳腺癌细胞相应的生长、黏附、侵袭能力较强,Stathmin表达水平与细胞侵袭能力密切相关。     

Caprin-1 is a novel microRNA-223 target for regulating the proliferation and invasion of human breast cancer cells

MicroRNAs (miRNAs) are 21–22 nucleotides regulatory small non-coding RNAs that inhibit gene expression by binding to complementary sequences especially the 3’ untranslated region (3’UTR) of mRNA. One miRNA can target many messenger RNAs, leading to a complex metabolic network. Previous studies have shown that miRNA-223 regulates migration and invasion of tumor cells and targets cytoplasmic activation/proliferation-associated protein-1 (Caprin-1). In the present study, we detected the expression of miRNA-223 and Caprin-1 in MCF-7, T-47D and MDA-MB-231 cancer cell lines, and MCF-10A normal breast cell line, and analyzed the role of miRNA-223 in Caprin-1-induced proliferation and invasion of human breast cancer cells. We found that miRNA-223 expression levels are significantly lower in MCF-7, T-47D and MDA-MB-231 cancer cells than in MCF-10A normal breast cells, while Caprin-1 expression is higher in cancer cells than in normal breast cells. The most malignant cancer cell line MDA-MB-231 has the lowest expression of miR-223, but the highest expression of Caprin-1. Further, we found that miR-223 targets the 3’UTR of Caprin-1 miRNA and down-regulates the expression of Caprin-1. We also found that over-expression of Caprin-1 can promote the proliferation and the invasion of breast cancer cells, but miRNA-223 can inhibit the proliferation and the invasion. miRNA-223-induced inhibition can be reversed by ectopic over-expression of Caprin-1. These findings suggest that miR-223 may suppress the proliferation and invasion of cancer cells by directly targeting Caprin-1. Our study also indicates that expression levels of miR-223 and Caprin-1 can be used to predict the state of cancer in breast cancer patient.     

Amino-terminal fragment of urokinase inhibits tumor cell invasion in vitro and in vivo: respective contribution of the urokinase plasminogen activator receptor-dependent or -independent pathway

The urokinase plasminogen activator (uPA) is implicated in both cancer cell invasion and angiogenesis. It can interact with a specific receptor (uPAR) via the epidermal growth factor (EGF)-like domain in the urokinase amino-terminal fragment (ATF) in a species-specific manner. Our previous studies showed that adenovirusmediated delivery of murine ATF (AdmATF) suppressed human tumor growth in mouse models, by inhibiting murine angiogenesis. However, we cannot exclude its putative inhibitory action on human cancer cell invasion through a uPAR-independent pathway. To further investigate the mechanisms of ATF, we constructed another adenovirus, AdhmATF, expressing humanized murine ATF (hmATF). hmATF binds to human uPAR but not to murine uPAR. We compared the antagonist effect of both AdmATF and AdhmATF on human and murine cancer cells. In vitro, the supernatant from AdhmATF-infected cells repressed 79% of membrane-associated uPA activity on human MDA-MB-231 cells, whereas that from AdmATF-infected cells repressed 35% of membrane-associated uPA activity. On murine LLC cells, the supernatant from AdhmATF-infected cells inhibited 29% of cell surface uPA activity, whereas that from AdmATF-infected cells inhibited 74% of cell surface uPA activity. Similar results were obtained in a cell invasion assay. In vivo, intratumoral injection of the adenoviruses into LLC tumors on day 24 postinjection induced lower but significant tumor growth suppression by AdhmATF (tumor volume was 1185 +/- 128 mm3), whereas suppression by AdmATF was greater (407 +/- 147 mm3). In the MDA-MB-231 tumor model, on day 52 postinjection, tumor size was 187 +/- 47 mm3 in the AdhmATF-treated group and 468 +/- 65 mm3 in the AdmATF-treated group. The LLC and MDA-MB- 231 cell lines transfected by mATF or hmATF genes showed growth inhibition In vivo equivalent to the results obtained by adenovirus treatment. These results demonstrate the strong anticancer activity of ATF even when its uPAR-binding affinity has been suppressed, and indicate that ATF exerts an antitumor effect via dual mechanisms: essentially through targeting the uPA-uPAR system via the EGF-like domain and partially through targeting a uPAR-independent interaction via the kringle domain.     

Hsa-miR-145对人三阴性乳腺癌细胞侵袭和迁移的影响

目的 探讨Hsa-miR-145对人三阴性乳腺癌细胞增殖、侵袭及迁移的影响,并初步分析其影响三阴性乳腺癌细胞增殖、侵袭及迁移的可能机制.方法 运用脂质体介导的转染方法将miR-145阻遏物(miR-145 inhibitors)转染人三阴性乳腺癌细胞株MDA-MB-231,以inhibitor negative control(inhibitor NC)作为阴性对照,通过MTT法和Transwell侵袭实验检测细胞的增殖能力和侵袭力;采用Transwell迁移实验及划痕试验检测细胞的迁移能力;利用生物信息学方法预测miR-145的靶基因,并对其靶基因进行基因功能分析.结果 (1)miR-145 inhibitors组细胞增殖活性明显高于inhibitor NC组(P<0.05);(2)划痕后miR-145 inhibitors组细胞迁移能力比inhibitor NC组明显增强(P<0.05);(3)Transwell侵袭及迁移实验均显示转染miR-145 inhibitors后,MDA-MB-231细胞的侵袭及迁移能力明显增强(P<0.01,P<0.05);(4)生物信息学方法预测miR-145的靶基因中,部分发挥了促进细胞增殖、侵袭及迁移的生物学功能.结论 (1)miR-145对人三阴性乳腺癌细胞的增殖、侵袭及迁移能力可能存在负性调控作用;(2)miR-145可能通过多种靶基因发挥其对肿瘤的调控作用.     

Targeting the Akt/mammalian target of rapamycin pathway for radiosensitization of breast cancer

The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is known to be activated by radiation. The mammalian target of rapamycin (mTOR) is downstream of Akt, and we investigated the effects of radiation on Akt/mTOR signaling in breast cancer cell models. RAD001 (everolimus), a potent derivative of the mTOR inhibitor rapamycin, was used to study the effects of mTOR inhibition, as the role of mTOR inhibition in enhancing radiation remains unexplored. RAD001 decreased clonogenic cell survival in both breast cancer cell lines MDA-MB-231 and MCF-7, although the effect is greater in MDA-MB-231 cells. Irradiation induced Akt and mTOR signaling, and this signaling is attenuated by RAD001. The radiation-induced signaling activation is mediated by PI3K because inhibition of PI3K with LY294002 inhibited the increase in downstream mTOR signaling. Additionally, caspase-dependent apoptosis is an important mechanism of cell death when RAD001 is combined with 3 Gy radiation, as shown by induction of caspase-3 cleavage. An increase in G(2)-M cell cycle arrest was seen in the combination treatment group when compared with controls, suggesting that cell cycle arrest may have been a contributing factor in the increased radiosensitization seen in this study. We conclude that RAD001 attenuates radiation-induced prosurvival Akt/mTOR signaling and enhances the cytotoxic effects of radiation in breast cancer cell models, showing promise as a method of radiosensitization of breast cancer.     

miR-203调控NEDD9抑制乳腺癌细胞MDA-MB-231侵袭及迁移作用机制研究

目的 :研究mi R-203抑制乳腺癌MDA-MB-231细胞NEDD9蛋白的作用对细胞侵袭和迁移的影响。方法:mi R-203脂质体转染MDA-MB-231细胞,通过细胞划痕实验和Transwell实验观察细胞侵袭和迁移能力的变化;通过蛋白印迹检测mi R-203对NEDD9表达的调控作用,并用sensor reporter确定mi R-203的靶标位点;最后,通过细胞"拯救"实验研究相关蛋白表达量的变化,用免疫荧光-激光共聚焦显微镜观察细胞片状伪足和黏着斑的改变,探究mi R-203对MDA-MB-231细胞侵袭和迁移能力的调节机制。结果:细胞划痕实验和Transwell实验显示,mi R-203抑制了MDA-MB-231细胞的侵袭和迁移能力;通过生物信息学网站预测mi R-203的靶基因是NEDD9;蛋白印迹和sensor reporter检测结果显示,mi R-203通过与NEDD9 3′-UTR结合,下调NEDD9;共转染si NEDD9和mi R-203的"拯救"实验后的细胞划痕实验、蛋白印迹和免疫荧光-激光共聚焦结果显示,mi R-203抑制NEDD9表达导致激活态Rac1-GTP减少,致使细胞运动状态改变,侵袭迁移能力减弱,而si NEDD9干涉作用能"拯救"mi R-203对MDA-MB-231细胞迁移能力的抑制。结论 :mi R-203通过降解NEDD9,下调激活的Rac1水平,导致乳腺癌细胞MDA-MB-231片状伪足和黏着斑的减少或消失,从而抑制细胞的侵袭及迁移。     

Sulforaphane induces cell type-specific apoptosis in human breast cancer cell lines

Sulforaphane, an isothiocyanate found in cruciferous vegetables, has been shown to induce phase 2 detoxication enzymes and inhibit the growth of chemically induced mammary tumors in rats, although the exact mechanisms of action of sulforaphane are not understood. In this study, we evaluated the effects of sulforaphane on cell growth and death in several human breast cancer cell lines and examined the hypothesis that sulforaphane acts as a histone deacetylase (HDAC) inhibitor in these cell lines. Sulforaphane treatment inhibited cell growth, induced a G(2)-M cell cycle block, increased expression of cyclin B1, and induced oligonucleosomal DNA fragmentation in the four human breast cancer cell lines examined, MDA-MB-231, MDA-MB-468, MCF-7, and T47D cells. Activation of apoptosis by sulforaphane in MDA-MB-231 cells seemed to be initiated through induction of Fas ligand, which resulted in activation of caspase-8, caspase-3, and poly(ADP-ribose) polymerase, whereas apoptosis in the other breast cancer cell lines was initiated by decreased Bcl-2 expression, release of cytochrome c into the cytosol, activation of caspase-9 and caspase-3, but not caspase-8, and poly(ADP-ribose) polymerase cleavage. Sulforaphane inhibited HDAC activity and decreased the expression of estrogen receptor-alpha, epidermal growth factor receptor, and human epidermal growth factor receptor-2 in each cell line, although no change in the acetylation of H3 or H4 was seen. These data suggest that sulforaphane inhibits cell growth, activates apoptosis, inhibits HDAC activity, and decreases the expression of key proteins involved in breast cancer proliferation in human breast cancer cells. These results support testing sulforaphane in vivo and warrant future studies examining the clinical potential of sulforaphane in human breast cancer.     

TMPRSS4通过上皮间质转化促进乳腺癌细胞的增殖、侵袭和转移

目的通过检测Ⅱ型跨膜丝氨酸蛋白酶4（TMPRSS4）在乳腺癌细胞中的表达情况,分析其在乳腺癌细胞增殖、侵袭和转移过程中的作用及其与上皮间质转化（EMT）的关系。 方法采用实时荧光定量PCR（qRT-PCR）和Western blot法检测5种不同乳腺癌细胞系中TMPRSS4 mRNA和蛋白水平的表达情况。将过表达质粒转染至乳腺癌细胞中,采用qRT-PCR方法检测转染效率,并通过MTS和EdU细胞增殖实验、Transwell和Matrigel细胞迁移和侵袭实验,研究过表达TMPRSS4对乳腺癌细胞增殖、侵袭和迁移的作用。采用qRT-PCR和Western blot法检测过表达TMPRSS4后细胞EMT相关基因E-cadherin、Vimentin、Claudin-1、Slug、ZEB1的表达变化。 结果TMPRSS4在乳腺癌MDA-MB-468和MDA-MB-231细胞系中高表达。MTS实验显示TMPRSS4过表达组乳腺癌细胞MDA-MB-468和MDA-MB-231的增殖能力与阴性对照组相比明显增加,差异具有统计学意义（P=0.039和0.038）,EdU实验进一步证实过表达TMPRSS4促进乳腺癌细胞的增殖,MDA-MB-468细胞和MDA-MB-231细胞的增殖率与阴性对照组比较差异具有统计学意义（P=0.001和0.008）。Transwell实验显示,TMPRSS4过表达组MDA-MB-468细胞和MDA-MB-231细胞的迁移能力与阴性对照组比较明显提高,差异具有统计学意义（p均=0.001）。Matrigel实验显示,TMPRSS4过表达组MDA-MB-468细胞和MDA-MB-231细胞的侵袭浸润能力与阴性对照组比较明显提高,差异具有统计学意义（P=0.012和0.000）。与阴性对照组比较,过表达TMPRSS4后,EMT相关基因包括上皮性标记E-cadherin和Claudin-1的表达均显著下降,而间质标记Vimentin和Slug的表达均明显提高,差异具有统计学意义（P分别=0.024,0.003,0.002和0.012）。 结论TMPRSS4参与了乳腺癌EMT的发生过程,并通过EMT促进乳腺癌细胞的增殖、侵袭和转移。     

Let-7a mimic attenuates CCL18 induced breast cancer cell metastasis through Lin 28 pathway

BackgroundMicroRNAs are believed to influence breast cancer cell tumorgenicity by interacting with the production of tumor associated macrophages. At this stage, this hypothesis lacks sufficient empirical evidence. Our study is an investigation of the effects of let-7a on the function of human breast cancer cell lines that had undergone chemokine ligand 18 (CCL18) stimulation.MethodsTwo breast cancer cell lines MDA-MB-231 and MCF-7 were transfected with let-7a mimics with or without CCL18 simulation. The expression level of let-7a was evaluated with qRT-PCR. Our study examined cell proliferation, migration and cell cycles following let-7a treatment. The predicted target of let-7a was identified and confirmed in vitro by a dual luciferase reporter system. The associations between let-7a, CCL18 and target gene expression were evaluated using RT-PCR and the Western blotting method.ResultsThe downregulated expression level of let-7a was observed in both breast cancer cell lines. When compared to the control and CCL18 stimulation groups, cell proliferation and migration in MDA-MB-231 and MCF-7 cells were significantly inhibited by let-7a. Furthermore, the cell cycle was dramatically blocked at the G2/M phase. The luciferase reporter identified Lin28 as the direct binding target of let-7a in both breast cancer cell lines.ConclusionUpregulation of let-7a carries the potential to reverse CCL18 induced cell proliferation and migration alteration in breast cancer cells by regulating Lin28 expression. Our results provided evidence which suggests the use of let-7a as a therapeutic agent in the treatment of breast cancer.     

Tamoxifen-resistant human breast cancer cell growth: inhibition by thioridazine, pimozide and the calmodulin antagonist, W-13.

Estrogen receptor (ER)-negative human breast cancer cell lines (MDA-MB-231 and MDA-MB-435) and ER-positive derivatives of the MCF-7 cell line selected for growth in the presence of antiestrogens (LY2 and RR) were used as in vitro models of tamoxifen-resistant human breast cancer in this study. The sensitivity of the tamoxifen-sensitive (MCF-7) and tamoxifen-resistant human breast cancer cell growth to two noncytotoxic neuroleptic drugs, pimozide and thioridazine, and the anticalmodulin agent, W-13, were compared. Inhibition of cell growth was measured as a decrease in cell number following a 72-h incubation with drug. Growth of the ER-negative cell lines MDA-MB-231 and MDA-MB-435 was inhibited by all three drugs. The average Ki values in these two lines were 6.3 and 3.8 microM for pimozide and 4.1 and 15 microM for thioridazine, respectively. Both ER-negative cell lines were more sensitive than MCF-7 cells to growth inhibition by W-13. MCF-7 cells selected for antiestrogen resistance were sensitive to growth inhibition by W-13 and thioridazine (LY2, average Ki = 10.4 microM; RR, average Ki = 5.2 microM). LY2 and RR cells were resistant to pimozide except when treated with estradiol (Ki = 4.6 and 7.9 microM, respectively). Pimozide, thioridazine and W-13 all exerted different effects on the distribution of human breast cancer cells within the cell cycle, suggesting that each drug may utilize a distinct pathway for inhibition of cell growth. We conclude that all three drugs are potential noncytotoxic alternatives to tamoxifen for the treatment of tamoxifen-resistant human breast cancer.     

磷脂酶C-γ1对乳腺癌细胞增殖、粘附及迁移的作用

摘要：目的：探讨磷脂酶C-γ1 (phospholipase C-γ1,PLC-γ1)对乳腺癌细胞生长、粘附和迁移等生物学行为的影响,揭示乳腺癌发生的分子机制。 方法：在乳腺癌细胞株MDA-MB-231中加入PLC-γ1抑制剂（U-73122）,用MTT试验、划痕试验、细胞粘附及趋化运动试验检测细胞生物学行为的变化,western blot检测抑制PLC-γ1后的信号传导通路中整合素β1和PKCζ分子磷酸化激活的情况。 结果：PLC-γ1被U-73122抑制后MDA-MB-231细胞的增殖、粘附和迁移运动能力下降,差异均有统计学意义（P<0.05）。western blot结果表明,细胞粘附相关整合素β1的磷酸化激活水平降低;趋化运动相关PKCζ磷酸化水平降低。 结论：PLC-γ1参与调控乳腺癌细胞增殖、粘附和迁移运动,可能是乳腺癌分子诊断和靶向治疗的有效靶点。     

Liposomal delivery enhances short-chain ceramide-induced apoptosis of breast cancer cells

It is therapeutically desirable to effectively deliver ceramide, an antimitogenic and proapoptotic lipid second messenger, to transformed cell types. However, the targeted delivery of cell-permeable ceramide analogs, including C6-ceramide, to cells may be impeded by the hydrophobicity of these bioactive lipids, resulting in reduced efficacy. The objective of this study is to develop and optimize liposomal vehicles to augment ceramide delivery to a breast adenocarcinoma cell line. We designed conventional, cationic, and pegylated drug release vesicles to efficaciously deliver ceramide to MDA-MB-231 breast adenocarcinoma cells. In vitro pharmacokinetic analysis demonstrated that liposomal ceramide delivery resulted in significantly greater accumulation of ceramide in MDA-MB-231 cells. Ceramide-formulated liposomes significantly inhibited MDA-MB-231 cell proliferation as compared with nonliposomal administration of ceramide. Ceramide-induced apoptosis correlated with the pharmacokinetic profile and the diminished proliferation in this highly aggressive, metastatic cell line. Liposomal ceramide formulations inhibited phosphorylated Akt levels and stimulated caspase-3/7 activity more effectively than nonliposomal ceramide, events consistent with apoptosis. Together, these results indicate that bioactive ceramide analogs can be incorporated into conventional, cationic, or pegylated liposomal vehicles for improved drug delivery and release.     

Aryl hydrocarbon receptor agonists induce microRNA-335 expression and inhibit lung metastasis of estrogen receptor negative breast cancer cells

The aryl hydrocarbon receptor (AHR) was initially identified as a receptor that bound 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related environmental toxicants; however, there is increasing evidence that the AHR is an important new drug target for treating multiple diseases including breast cancer. Treatment of estrogen receptor (ER)-negative MDA-MB-231 and BT474 breast cancer cells with TCDD or the selective AHR modulator 6-methyl-1,3,-trichlorodibenzofuran (MCDF) inhibited breast cancer cell invasion in a Boyden chamber assay. These results were similar to those previously reported for the antimetastic microRNA-335 (miR-335). Both TCDD and MCDF induced miR-335 in MDA-MB-231 and BT474 cells and this was accompanied by downregulation of SOX4, a miR-335-regulated (inhibited) gene. The effects of TCDD and MCDF on miR-335 and SOX4 expression and interactions of miR-335 with the 3'-UTR target sequence in the SOX4 gene were all inhibited in cells transfected with an oligonucleotide (iAHR) that knocks down the AHR, thus confirming AHR-miR-335 interactions. MCDF (40 mg/kg/d) also inhibited lung metastasis of MDA-MB-231 cells in a tail vein injection model, showing that the AHR is a potential new target for treating patients with ER-negative breast cancer, a disease where treatment options and their effectiveness are limited.