Sensitization of ovarian cancer cells to cisplatin by gold nanoparticles

Recently we reported that gold nanoparticles (AuNPs) inhibit ovarian tumor growth and metastasis in mice by reversing epithelial-mesenchymal transition (EMT). Since EMT is known to confer drug resistance to cancer cells, we wanted to investigate whether anti-EMT property of AuNP could be utilized to sensitize ovarian cancer cells to cisplatin. Herein, we report that AuNPs prevent cisplatin-induced acquired chemoresistance and stemness in ovarian cancer cells and sensitize them to cisplatin. AuNPs inhibit cisplatin induced EMT, decrease the side population cells and key stem cell markers such as ALDH1, CD44, CD133, Sox2, MDR1 and ABCG2 in ovarian cancer cells. Mechanistically, AuNPs prevent cisplatin-induced activation of Akt and NF-κB signaling axis in ovarian cancer cells that are critical for EMT, stem cell maintenance and drug resistance. In vivo, AuNPs sensitize orthotopically implanted ovarian tumor to a low dose of cisplatin and significantly inhibit tumor growth via facilitated delivery of both AuNP and cisplatin. These findings suggest that by depleting stem cell pools and inhibiting key molecular pathways gold nanoparticles sensitize ovarian cancer cells to cisplatin and may be used in combination to inhibit tumor growth and metastasis in ovarian cancer.     

The expression pattern of PFKFB3 enzyme distinguishes between induced-pluripotent stem cells and cancer stem cells

Induced pluripotent stem cells (iPS) have become crucial in medicine and biology. Several studies indicate their phenotypic similarities with cancer stem cells (CSCs) and a propensity to form tumors. Thus it is desirable to identify a trait which differentiates iPS populations and CSCs. Searching for such a feature, in this work we compare the restriction (R) point-governed regulation of cell cycle progression in different cell types (iPS, cancer, CSC and normal cells) based on the expression profile of 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase3 (PFKFB3) and phosphofructokinase (PFK1). Our study reveals that PFKFB3 and PFK1 expression allows discrimination between iPS and CSCs. Moreover, cancer and iPS cells, when cultured under hypoxic conditions, alter their expression level of PFKFB3 and PFK1 to resemble those in CSCs. We also observed cell type-related differences in response to inhibition of PFKFB3. This possibility to distinguish CSC from iPS cells or non-stem cancer cells by PFKB3 and PFK1 expression improves the outlook for clinical application of stem cell-based therapies and for more precise detection of CSCs.     

上皮源性卵巢癌干细胞的研究进展

肿瘤干细胞(cancer stem cells,CSCs)是肿瘤组织内具有自我更新能力以及无限增殖和多向分化潜能的一群细胞,对化疗药物耐受.卵巢癌干细胞(epithelial ovarian cancer stem cells,EOCSCs)在卵巢癌的发生发展、侵袭转移和耐药复发过程中都起到了重要作用.传统的肿瘤细胞减灭术联合顺铂、紫杉醇全身化疗对减小肿瘤体积,缓解临床症状具有一定的作用,但治疗后残留的EOCSCs能够短时间内重建肿瘤组织,是卵巢癌复发和难治的根本原因.深入了解EOCSCs的生物学特性,探索EOCSCs的发生发展机制,研究针对EOCSCs的靶向治疗药物,是抗肿瘤治疗的关键.     

Cisplatin and TRAIL enhance breast cancer stem cell death

Triple negative breast cancer (TNBC) has increased recurrence and poor survival, despite a high response rate to neoadjuvant chemotherapy. The aim of this study was to determine whether current drug treatment(s) eliminates bulk of tumor cells, but it has a minimal effect on cancer stem cells (CSCs) leading to tumor recurrence. We studied the effects of PARP inhibitors (AZD2281 and BSI-201), paclitaxel, docetaxel, cisplatin and cisplatin plus TRAIL on CSCs derived from CRL-2335 and MDA-MB-468 TNBC cells in?vitro. The in?vitro data indicate that cisplatin plus TRIAL treatment was most effective in eliminating CSCs compared to PARP inhibitors, cisplatin, paclitaxel and docetaxel. Treatment with cisplatin plus TRAIL also inhibits Wnt-1 signaling and its downstream target, β-catenin, phospho β-catenin, cyclin D1, increased apoptosis, reduced proliferation and mammosphere formation. Inhibition of Wnt-1 by siRNA significantly reduced the ability of CSCs to form mammospheres compared to control. However, maximum effect was seen in cisplatin plus TRAIL-treated cells. Taken together the data suggest that cisplatin plus TRAIL treatment has the potential of providing a new strategy for improving the therapeutic outcome in TNBC patients.     

Mouse induced pluripotent stem cell microenvironment generates epithelial-mesenchymal transition in mouse Lewis lung cancer cells

Induced pluripotent stem (iPS) cells may be a powerful tool in regenerative medicine, but their potential tumorigenicity is a significant challenge for the clinical use of iPS cells. Previously, we succeeded in converting miPS cells into cancer stem cells (CSCs) under the conditions of tumor microenvironment. Both stem cells and tumor cells are profoundly influenced by bi-directional communication with their respective microenvironment, which dictates cell fate determination and behavior. The microenvironment derived from iPS cells has not been well studied. In this paper, we have investigated the effects of secreted factors from Nanog-mouse iPS (miPS) cells on mouse Lewis lung cancer (LLC) cells that are found in the conditioned media. The results demonstrated that miPS cells secrete factors that can convert the epithelia phenotype of LLC cells to a mesenchymal phenotype, and that can promote tumorigenisity, migration and invasion. Furthermore, LLC cells that have been exposed to miPS conditioned medium became resistant to apoptosis. These various biological effects suggest that the miPS microenvironment contain factors that can promote an epithelial-mesenchymal transition (EMT) through an active Snail-MMP axis or by suppressing differentiation in LLC cells.     

Tissue transglutaminase protects epithelial ovarian cancer cells from cisplatin-induced apoptosis by promoting cell survival signaling

Tissue transglutaminase (TG2), an enzyme involved in protein cross-linking and overexpressed in ovarian tumors, has antiapoptotic effects in cancer cells and may play a role in response to chemotherapy. In this study, we investigated the role of TG2 in the sensitivity of ovarian cancer cells to cisplatin. By using stable knockdown and overexpression strategies, we demonstrate that the level of expression of TG2 regulates apoptosis induced by cisplatin in SKOV3 and OV-90 ovarian cancer cells. Interestingly, not only TG2 knockdown but also a TG2 enzymatic inhibitor (KCC009) sensitized SKOV3 cells to cisplatin. To understand the mechanism by which TG2 exerts its antiapoptotic role, we examined the effects of protein kinase B (Akt) and nuclear factor-kappa B (NF-kappaB), two survival pathways commonly involved in development of drug resistance. Overexpression of the constitutively active p65 subunit of NF-kappaB, but not constitutively active Akt, rescued cells with diminished TG2 expression from cisplatin-induced apoptosis. This implicates activation of NF-kappaB as the main cisplatin resistance mechanism downstream of TG2. Indeed, NF-kappaB activity is decreased and the level of the inhibitory subunit I kappaB alpha is increased in ovarian cancer cells engineered to express diminished levels of TG2 or treated with the enzymatic inhibitor, KCC009. Our data show that TG2 prevents apoptosis induced by cisplatin by activating the NF-kappaB survival pathway in ovarian cancer cells.     

Cisplatin inhibits paclitaxel-induced apoptosis in cisplatin-resistant ovarian cancer cell lines: possible explanation for failure of combination therapy.

Combination chemotherapy using paclitaxel with a platinum-based regimen is currently the standard first-line therapy for ovarian cancer after surgical cytoreduction. Whereas cisplatin-paclitaxel combination chemotherapy has shown significant efficacy over previous drug combinations in ovarian cancer, 20-30% of patients fail to respond to this combination. These patients are deemed cisplatin-paclitaxel resistant, although it is unclear whether the tumors are resistant to one or both drugs. Because the options available to ovarian cancer patients for second-line therapy are limited, and knowing that mechanistic differences exist between cisplatin and paclitaxel, we assessed the efficacy of combination drug therapy on cisplatin-resistant (cisplatinR) ovarian cancer cells. We found that paclitaxel induced apoptosis in cisplatinR cells as well as in the cisplatin-sensitive parental cell lines. In cisplatinR C-13 cells, the concomitant addition of cisplatin blocked paclitaxel-induced apoptosis as determined by DNA fragmentation assays, fluorescence microscopy, and flow cytometry. Paclitaxel-induced multimininucleation was also inhibited when the cells were exposed sequentially to paclitaxel and then cisplatin. Cisplatin did not block paclitaxel-induced stabilization of microtubules or prevent paclitaxel-induced loss of Bcl-2 expression in cisplatinR cells. Conversely, paclitaxel did not inhibit p53 protein accumulation by cisplatin. These results suggest that cisplatin blocks paclitaxel-induced apoptosis at a point downstream of Bcl-2 degradation and independent of microtubule stabilization. Our research shows that cisplatin can inhibit the effectiveness of paclitaxel in cispatinR cell lines. Therefore, the establishment of a clinical protocol to evaluate the efficacy of paclitaxel alone versus another second-line regimen in patients with cisplatin-paclitaxel-resistant ovarian cancer is warranted.     

Mdm2 antagonists induce apoptosis and synergize with cisplatin overcoming chemoresistance in TP53 wild‐type ovarian cancer cells

Ovarian cancer (OVCa) is the leading cause of death from gynecological malignancies. Although treatment for advanced OVCa has improved with the introduction of taxane–platinum chemotherapy, the majority of patients will develop resistance to the treatment, leading to poor prognosis. One of the causes of chemoresistance is the reduced ability to undergo apoptosis. Cisplatin is a genotoxic drug that leads cells to apoptosis through the activation of the p53 pathway. Defective signaling in this pathway compromises p53 function, and thus cisplatin does not induce apoptosis. A new group of nongenotoxic small molecules called Nutlins have been developed to inhibit p53‐Mdm2 binding, inducing apoptosis in chemoresistant tumors through the activation of the p53 pathway. The wild‐type p53 cisplatin‐resistant ovarian cancer cell‐line A2780cis was used to test the effect of Nutlin‐3a (Nut3a) on apoptosis response. The results showed that Nut3a synergized with cisplatin, inducing cell‐cycle arrest in G2/M and potentiating apoptotic cell death. Increased apoptosis was also induced in wild‐type TP53 primary OVCa cultures by double cisplatin–Nut3a treatment. In conclusion, Nut3a appears to sensitize chemoresistant OVCa cells to cisplatin, inducing apoptosis. As increased response was generalized in primary tumors, this cisplatin–Nut3a combination could be useful for the treatment of patients harboring wild‐type TP53 who do not respond to standard chemotherapy.     

PGE2/EP4 antagonism enhances tumor chemosensitivity by inducing extracellular vesicle‐mediated clearance of cancer stem cells

Cells expressing mesenchymal/basal phenotypes in tumors have been associated with stem cell properties. Cancer stem cells (CSCs) are often resistant to conventional chemotherapy. We explored overcoming mesenchymal CSC resistance to chemotherapeutic agents. Our goal was to reduce CSC numbers in vivo, in conjunction with chemotherapy, to reduce tumor burden. Analysis of clinical samples demonstrated that COX‐2/PGE₂/EP₄ signaling is elevated in basal‐like and chemoresistant breast carcinoma and is correlated with survival and relapse of breast cancer. EP₄ antagonism elicts a striking shift of breast cancer cells from a mesenchymal/CSC state to a more epithelial non‐CSC state. The transition was mediated by EP₄ antagonist‐induced extracellular vesicles [(EVs)/exosomes] which removed CSC markers, mesenchymal markers, integrins, and drug efflux transporters from the CSCs. In addition, EP₄ antagonism‐induced CSC EVs/exosomes can convert tumor epithelial/non‐CSCs to mesenchymal/CSCs able to give rise to tumors and to promote tumor cell dissemination. Because of its ability to induce a CSC‐to‐non‐CSC transition, EP₄ antagonist treatment in vivo reduced the numbers of CSCs within tumors and increased tumor chemosensitivity. EP₄ antagonist treatment enhances tumor response to chemotherapy by reducing the numbers of chemotherapy‐resistant CSCs available to repopulate the tumor. EP₄ antagonism can collaborate with conventional chemotherapy to reduce tumor burden.     

Overexpression of DDB2 enhances the sensitivity of human ovarian cancer cells to cisplatin by augmenting cellular apoptosis

Cisplatin is one of the most widely used anticancer agents, displaying activity against a wide variety of tumors. However, development of drug resistance presents a challenging barrier to successful cancer treatment by cisplatin. To understand the mechanism of cisplatin resistance, we investigated the role of damaged DNA binding protein complex subunit 2 (DDB2) in cisplatin‐induced cytotoxicity and apoptosis. We show that DDB2 is not required for the repair of cisplatin‐induced DNA damage, but can be induced by cisplatin treatment. DDB2‐deficient noncancer cells exhibit enhanced resistance to cell growth inhibition and apoptosis induced by cisplatin than cells with fully restored DDB2 function. Moreover, DDB2 expression in cisplatin‐resistant ovarian cancer cell line CP70 and MCP2 was lower than their cisplatin‐sensitive parental A2780 cells. Overexpression of DDB2 sensitized CP70 cells to cisplatin‐induced cytotoxicity and apoptosis via activation of the caspase pathway and downregulation of antiapoptotic Bcl‐2 protein. Further analysis indicates that the overexpression of DDB2 in CP70 cells downregulates Bcl‐2 expression through decreasing Bcl‐2 mRNA level. These results suggest that ovarian cancer cells containing high level of DDB2 become susceptible to cisplatin by undergoing enhanced apoptosis.     

Prostate cancer stem cells and their potential roles in metastasis

Most solid tumors have now been reported to contain stem cell-like cells called cancer stem cells (CSCs). These cells are endowed with high tumorigenic capacity and may be the cells that drive tumor formation, maintain tumor homeostasis, and mediate tumor metastasis. Since these self-renewing cancer cells may be the sole population to develop a primary tumor, it is predicted that CSCs may also represent the lethal seeds of metastasis, as supported by a flurry of recent studies on the relationship between CSCs and metastasis. Herein, we summarize current knowledge of stem/progenitor cells and CSCs, especially in the context of normal human prostate and prostate cancer. We further update the recently gained knowledge on the involvement of CSCs in metastasis. We also discuss the fundamental influence of tumor microenvironment on the manifestation of CSCs and metastasis. Finally, we discuss the clinical implication of CSC-based therapy.     

Programmed cell death 4 enhances chemosensitivity of ovarian cancer cells by activating death receptor pathway in vitro and in vivo

Chemoresistance is a major cause of treatment failure in ovarian cancer. Therefore, it is necessary to explore alternative therapeutic methods to overcome drug resistance for ovarian cancer treatment. We previously reported that programmed cell death 4 (PDCD4), a tumor suppressor, significantly suppresses the malignant phenotype of ovarian cancer cells and its lost or low expression in ovarian cancer is associated with unfavorable prognosis of patients. Here we show that PDCD4 improves the sensitivity of ovarian cancer cells to platinum‐based chemotherapy. Overexpression of PDCD4 enhanced chemosensitivity in SKOV3 and CAOV3 cells with low levels of PDCD4, whereas knockdown of PDCD4 reduced chemosensitivity in OVCAR3 cells with high levels of PDCD4. Subsequently, the combination of enforced PDCD4 expression with cisplatin treatment significantly suppressed ovarian tumor growth in a xenograft animal model. The PDCD4 effect appears to be specific for cisplatin and carboplatin, not affecting cyclophosphamide, etoposide, or paclitaxel. Mechanistically, PDCD4 significantly increased cisplatin‐induced cleavage of caspase‐3 and caspase‐8, but had only a slight impact on caspase‐9 cleavage and the expression of Bax and Bcl‐2 in vitro and in vivo. A specific caspase‐8 inhibitor, Z‐ITED‐FMK, attenuated cisplatin‐induced apoptosis in PDCD4‐overexpressing ovarian cancer cells. Taken together, our results indicate that PDCD4 enhances cisplatin‐induced apoptosis by mainly activating the death receptor pathway, and PDCD4 gene transfer in combination with cisplatin therapy may break the resistance of ovarian cancer cells to chemotherapy. (Cancer Sci 2010)     

Therapeutic strategies targeting cancer stem cells

Increasing studies have demonstrated a small proportion of cancer stem cells (CSCs) exist in the cancer cell population. CSCs have powerful self-renewal capacity and tumor-initiating ability and are resistant to chemotherapy and radiation. Conventional anticancer therapies kill the rapidly proliferating bulk cancer cells but spare the relatively quiescent CSCs, which cause cancer recurrence. So it is necessary to develop therapeutic strategies acting specifically on CSCs. In recent years, studies have shown that therapeutic agents such as metformin, salinomycin, DECA-14, rapamycin, oncostatin M (OSM), some natural compounds, oncolytic viruses, microRNAs, cell signaling pathway inhibitors, TNF-related apoptosis inducing ligand (TRAIL), interferon (IFN), telomerase inhibitors, all-trans retinoic acid (ATRA) and monoclonal antibodies can suppress the self-renewal of CSCs in vitro and in vivo. A combination of these agents and conventional chemotherapy drugs can significantly inhibit tumor growth, metastasis and recurrence. These strategies targeting CSCs may bring new hopes to cancer therapy.     

SOX8 regulates cancer stem‐like properties and cisplatin‐induced EMT in tongue squamous cell carcinoma by acting on the Wnt/β‐catenin pathway

A sub‐population of chemoresistant cells exhibits biological properties similar to cancer stem cells (CSCs), and these cells are believed to be a main cause for tumor relapse and metastasis. In our study, we explored the role of SOX8 and its molecular mechanism in the regulation of the stemness properties and the epithelial mesenchymal transition (EMT) of cisplatin‐resistant tongue squamous cell carcinoma (TSCC) cells. We found that SOX8 was upregulated in cisplatin‐resistant TSCC cells, which displayed CSC‐like properties and exhibited EMT. SOX8 was also overexpressed in chemoresistant patients with TSCC and was associated with higher lymph node metastasis, advanced tumor stage and shorter overall survival. Stable knockdown of SOX8 in cisplatin‐resistant TSCC cells inhibited chemoresistance, tumorsphere formation, and EMT. The Wnt/β‐catenin pathway mediated the cancer stem‐like properties in cisplatin‐resistant TSCC cells. Further studies showed that the transfection of active β‐catenin in SOX8 stable‐knockdown cells partly rescued the SOX8 silencing‐induced repression of stem‐like features and chemoresistance. Through chromatin immunoprecipitation and luciferase assays, we observed that SOX8 bound to the promoter region of Frizzled‐7 (FZD7) and induced the FZD7‐mediated activation of the Wnt/β‐catenin pathway. In summary, SOX8 confers chemoresistance and stemness properties and mediates EMT processes in chemoresistant TSCC via the FZD7‐mediated Wnt/β‐catenin pathway.     

小细胞肺癌干细胞信号通路的研究进展

小细胞肺癌是具有高度侵袭性的肺肿瘤,其主要临床特征是化疗有效率高但易在短时间内复发转移,这一特点可能与肿瘤干细胞的存在有关。肿瘤干细胞被认为是恶性肿瘤发生发展、耐药、复发及转移的根源。目前多认为肿瘤干细胞与正常干细胞有着相同的信号通路,如Hedgehog、Notch、Wnt等通路。本文就这几条信号通路在小细胞肺癌干细胞中所起的作用以及针对这几条信号通路治疗药物的研究进展和可能的信号通路交互作用等方面进行综述。     

Epithelial-mesenchymal transition,cancer stem cells and treatment resistance

Breast cancer relapse, in a large number of patients, after initial response to standard of care therapy warrants development of novel therapies against recurrent and metastatic cancer. Cancer stem cells (CSCs), present in breast tumors while being intrinsically resistant to conventional therapy, have the ability to self renew and cause tumor recurrence. The residual tumors after therapy, with dramatic enrichment of the CSCs, have all the hallmarks of epithelial- mesenchymal transition (EMT). This review will focus on the link between EMT, CSCs and treatment resistance, since a better understanding of these interactions will allow us to effectively target the residual population after therapy.