Role of p38 MAPK in enhanced human cancer cells killing by the combination of aspirin and ABT‐737

Regular use of aspirin after diagnosis is associated with longer survival among patients with mutated‐PIK3CA colorectal cancer, but not among patients with wild‐type PIK3CA cancer. In this study, we showed that clinically achievable concentrations of aspirin and ABT‐737 in combination could induce a synergistic growth arrest in several human PIK3CA wild‐type cancer cells. In addition, our results also demonstrated that long‐term combination treatment with aspirin and ABT‐737 could synergistically induce apoptosis both in A549 and H1299 cells. In the meanwhile, short‐term aspirin plus ABT‐737 combination treatment induced a greater autophagic response than did either drug alone and the combination‐induced autophagy switched from a cytoprotective signal to a death‐promoting signal. Furthermore, we showed that p38 acted as a switch between two different types of cell death (autophagy and apoptosis) induced by aspirin plus ABT‐737. Moreover, the increased anti‐cancer efficacy of aspirin combined with ABT‐737 was further validated in a human lung cancer A549 xenograft model. We hope that this synergy may contribute to failure of aspirin cancer therapy and ultimately lead to efficacious regimens for cancer therapy.     

Stochastic elimination of cancer cells

Tissues of multicellular organisms consist of stem cells and differentiated cells. Stem cells divide to produce new stem cells or differentiated cells. Differentiated cells divide to produce new differentiated cells. We show that such a tissue design can reduce the rate of fixation of mutations that increase the net proliferation rate of cells. It has, however, no consequence for the rate of fixation of neutral mutations. We calculate the optimum relative abundance of stem cells that minimizes the rate of generating cancer cells. There is a critical fraction of stem cell divisions that is required for a stochastic elimination ('wash out') of cancer cells.     

Synergistic cytotoxicity of perifosine and ABT-737 to colon cancer cells

An acidic environment and hypoxia within the tumour are hallmarks of cancer that contribute to cell resistance to therapy. Deregulation of the PI3K/Akt pathway is common in colon cancer. Numerous Akt-targeted therapies are being developed, the activity of Akt-inhibitors is, however, strongly pH-dependent. Combination therapy thus represents an opportunity to increase their efficacy. In this study, the cytotoxicity of the Akt inhibitor perifosine and the Bcl-2/Bcl-xL inhibitor ABT-737 was tested in colon cancer HT-29 and HCT-116 cells cultured in monolayer or in the form of spheroids. The efficacy of single drugs and their combination was analysed in different tumour-specific environments including acidosis and hypoxia using a series of viability assays. Changes in protein content and distribution were determined by immunoblotting and a “peeling analysis” of immunohistochemical signals. While the cytotoxicity of single agents was influenced by the tumour-specific microenvironment, perifosine and ABT-737 in combination synergistically induced apoptosis in cells cultured in both 2D and 3D independently on pH and oxygen level. Thus, the combined therapy of perifosine and ABT-737 could be considered as a potential treatment strategy for colon cancer.     

Platinum-based combination nanomedicines for cancer therapy

The relatively low success rate of cancer nanomedicines has raised debate on the roles of the enhanced permeability and retention (EPR) effect in enhancing drug delivery to tumors and improving therapeutic efficacy. In this review, we highlight new strategies beyond the EPR effect for enhancing nanoparticle delivery to tumors. We discuss the roles of transcellular extravasation, receptor-mediated pathways, and protein corona interactions on nanoparticle deposition in tumors. We summarize recent progress in platinum-based combination nanomedicines containing multiple chemotherapeutics with synergistic anticancer mechanisms and multiple anticancer therapies with novel mechanisms to enhance drug delivery and antitumor activities. We also highlight future opportunities in platinum-based combination nanomedicines and key hurdles for the translation of these combination nanomedicines into the clinic.     

Synergistic cytotoxicity of perifosine and ABT‐737 to colon cancer cells

An acidic environment and hypoxia within the tumour are hallmarks of cancer that contribute to cell resistance to therapy. Deregulation of the PI3K/Akt pathway is common in colon cancer. Numerous Akt‐targeted therapies are being developed, the activity of Akt‐inhibitors is, however, strongly pH‐dependent. Combination therapy thus represents an opportunity to increase their efficacy. In this study, the cytotoxicity of the Akt inhibitor perifosine and the Bcl‐2/Bcl‐xL inhibitor ABT‐737 was tested in colon cancer HT‐29 and HCT‐116 cells cultured in monolayer or in the form of spheroids. The efficacy of single drugs and their combination was analysed in different tumour‐specific environments including acidosis and hypoxia using a series of viability assays. Changes in protein content and distribution were determined by immunoblotting and a “peeling analysis” of immunohistochemical signals. While the cytotoxicity of single agents was influenced by the tumour‐specific microenvironment, perifosine and ABT‐737 in combination synergistically induced apoptosis in cells cultured in both 2D and 3D independently on pH and oxygen level. Thus, the combined therapy of perifosine and ABT‐737 could be considered as a potential treatment strategy for colon cancer.     

Aiming to immune elimination of ovarian cancer stem cells

Ovarian cancer accounts for only 3% of all cancers in women, but it causes more deaths than any other gynecologic cancer. Treatment with chemotherapy and cytoreductive surgery shows a good response to the therapy. However, in a large proportion of the patients the tumor grows back within a few years. Cancer stem cells, that are less responsive to these treatments, are blamed for this recurrence of disease. Immune therapy either cellular or humoral is a novel concept to treat cancer. It is based on the notice that immune cells invade the tumor. However, the tumor invest heavily to escape from immune elimination by recruiting several immune suppressive mechanisms. These processes are normally in place to limit excessive immune activation and prevent autoimmune phenomena. Here, we discuss current knowledge about the immune (suppressive) status in ovarian cancer. Moreover, we discuss the immunological targets of ovarian cancer stem cells.     

Methylseleninic acid potentiates multiple types of cancer cells to ABT-737-induced apoptosis by targeting Mcl-1 and Bad

ABT-737, a novel small molecule inhibitor of Bcl-2 family proteins, holds great promise to complement current cancer therapies. However many types of solid cancer cells are resistant to ABT-737. One practical approach to improve its therapeutic efficacy is to combine with the agents that can overcome such resistance to restore the sensitivity. In the present study, a second-generation selenium compound methylseleninic acid (MSeA) synergistically sensitized MDA-MB-231 human breast cancer cells, HT-29 human colon cancer cells and DU145 human prostate cancer cells to apoptosis induction by ABT-737, as evidenced by greater than additive enhancement of Annexin V/FITC positive (apoptotic) cells and activation of multiple caspases and PARP cleavage. Mechanistic investigation demonstrated that MSeA significantly decreased basal Mcl-1 expression and ABT-737-induced Mcl-1 expression. Knocking down of Mcl-1 with RNAi approach supported the functional significance of this molecular target. More importantly, we identified inactivation of Bad by phosphorylation on ser-136 and ser-112 as a novel mechanism involved in ABT-737 resistance, which can be overcome by combining with MSeA. In addition, we found that expression of Bax was required for the efficient execution of synergistic sensitization. Our findings, for the first time, provide a strong mechanistic rationale for developing MSeA as a novel sensitizing agent of ABT-737.     

Efficient photodynamic therapy of cancer using chemotherapeutic porphyrin-ruthenium metalla-cubes

Two cationic octanuclear metalla-cubes [Ru(8)(η(6)-C(6)H(5)Me)(8)(tpp-H2)(2)(dhbq)(4)](8+) and [Ru(8)(η(6)-p-iPrC(6)H(4)Me)(8)(tpp-H2)(2)(dhbq)(4)](8+) were prepared and evaluated as dual photosensitizers and chemotherapeutics in cancer cells. In the dark, the complexes presented high cytotoxicity towards only melanoma and ovarian cancer cells. However, the complexes exhibited good phototoxicities toward all cancer cells (1μM concentration, LD(50)=2-7J/cm(2)), thus suggesting a dual synergistic effect with good properties of both the arene ruthenium chemotherapeutics and the porphyrin photosensitizers.     

Efficient production of uridine 5'-diphospho-N-acetylglucosamine by the combination of three recombinant enzymes and yeast cells

Uridine 5'-diphospho N-acetylglucosamine (UDP-GlcNAc) is an important nucleotide sugar in the biochemistry of all living organisms, and it is an important substrate in the synthesis of oligosaccharides. In the present work, three bioactive enzymes, namely, glucokinase (YqgR), GlcNAc-phosphate mutase (Agm1), and N-acetylglucosamine-1-phosphate uridyltransferase (GlmU), were produced effectively as soluble form in recombinant Escherichia coli. These three enzymes and dried yeast together were used to construct a multistep enzymatic system, which could produce UDP-GlcNAc efficiently with N-acetylglucosamine (GlcNAc) as the substrate. After the optimization of various reaction conditions, 31.5 mMUDP-GlcNAc was produced from 50 mMGlcNAc and 50 mMUMP.     

Platinum compounds sensitize ovarian carcinoma cells to ABT-737 by modulation of the Mcl-1/Noxa axis

Ovarian cancer is the leading cause of death from gynecological cancer. The anti-apoptotic protein Bcl-x_L is frequently overexpressed in ovarian carcinoma which correlates with chemotherapy resistance. It has been demonstrated that Bcl-x_L cooperates with another anti-apoptotic protein, Mcl-1, to protect ovarian cancer cells against apoptosis, and that their concomitant inhibition induces massive cell death. Here, we examined the interest of ABT-737, a potent BH3-mimetic molecule targeting Bcl-x_L, both alone and in combination with Mcl-1 modulators, in ovarian cancer cell lines. As a single agent, ABT-737 was ineffective at promoting cell death in the four cell lines we tested in vitro. However, the specific inhibition of Mcl-1 by siRNA dramatically increased the sensitivity of chemoresistant cells to ABT-737. Platinum compounds also sensitize to ABT-737 by dose-dependently decreasing Mcl-1 expression or by increasing the expression of pro-apoptotic BH3-only proteins Noxa and, to a lower extent, Bim. Furthermore, we demonstrated that Noxa accumulation was involved in apoptosis occurring in response to the combination of ABT-737 and platinum compounds, since cells were protected from apoptosis by its silencing. Moreover, the combination was also highly cytotoxic ex vivo in sliced SKOV3 tumor nodes. However we observed in these slices a strong basal expression of Noxa and apoptotic cell death in response to ABT-737 alone. Therefore, we have revealed that the modulation of the Mcl-1/Noxa axis by platinum compounds results in a strong sensitization of chemoresistant ovarian carcinoma cells to ABT-737, which could constitute a promising therapeutic in these cancers.     

Independent prognostication and therapy monitoring of breast cancer patients by DNA/RNA typing of minimal residual cancer cells

Clinical relevance, purification techniques and molecular characterization of minimal residual cancer cells (MRCC) is a controversial topic in the literature. An analytical concept including a novel isolation procedure and a panel of tests for DNA and RNA typing of MRCCs is described and clinically evaluated in this paper. The purification procedure exploiting the physical characteristics of MRCCs shows superior performance leading to > 50% pure and viable tumor cells. Proof of the presence and purity of MRCCs in an isolated sample is given by multiparametric DNA typing (amplifications, mutations, losses of heterozygosity). On the basis of the proven presence of MRCCs tumor-relevant mRNAs can be adequately analyzed by normalized quantitative real-time RT-PCR. The molecular characterization of MRCCs isolated from blood of breast cancer patients could have a strong clinical impact on prognostication, drug targeting and therapy monitoring.     

Simvastatin-romidepsin combination kills bladder cancer cells synergistically

The HMG-CoA reductase inhibitor simvastatin activates AMP-activated protein kinase (AMPK) and thereby induces histone acetylation. We postulated that combining simvastatin with the histone deacetylase (HDAC) inhibitor romidepsin would kill bladder cancer cells by inducing histone acetylation cooperatively. The combination of romidepsin and simvastatin induced robust apoptosis and killed bladder cancer cells synergistically. In murine subcutaneous tumor models using MBT-2 cells, a 15-day treatment with 0.5 mg/kg romidepsin and 15 mg/kg simvastatin was well tolerated and inhibited tumor growth significantly. Mechanistically, the combination induced histone acetylation by activating AMPK. The combination also decreased the expression of HDACs, thus further promoting histone acetylation. This AMPK activation was essential for the combination's action because compound C, an AMPK inhibitor, suppressed the combination-induced histone acetylation and the combination's ability to induce apoptosis. We also found that the combination increased the expression of peroxisome proliferator-activated receptor (PPAR) γ, leading to reactive oxygen species production. Furthermore, the combination induced endoplasmic reticulum (ER) stress and this ER stress was shown to be associated with increased AMPK expression and histone acetylation, thus playing an important role in the combination's action. Our study also suggests there is a positive feedback cycle between ER stress induction and PPARγ expression.     

Efficient microbial elimination of methanol inhibition for Naproxen resolution by a lipase

To eliminate methanol inhibition of the activity of a lipase, methanotrophic bacteria, which can convert methanol into water and CO₂, were introduced to the reaction of enantioselective hydrolysis of Naproxen methyl ester catalysed by lipase from Candida rugosa. Both the activity and stability of lipase were improved by the removal of methanol by the bacteria.     

Efficient detection of PrPSc (263K) in human plasma.

The potential contamination of human blood or plasma with prions, such as variant Creuftzfelt-Jacob disease (vCJD), is becoming a serious problem. In this study, we established a Western blot-based detection method for PrP(Sc) (263K) spiked in plasma. Although plasma contains a large amount of protein, specific detection of a small amount of 263K in plasma could be specifically detected only after ultra-centrifugation followed by heat-denaturation of plasma proteins included in the resulting precipitate, before the digestion with proteinase K.     

In silico drug combination discovery for personalized cancer therapy

Background

Drug combination therapy, which is considered as an alternative to single drug therapy, can potentially reduce resistance and toxicity, and have synergistic efficacy. As drug combination therapies are widely used in the clinic for hypertension, asthma, and AIDS, they have also been proposed for the treatment of cancer. However, it is difficult to select and experimentally evaluate effective combinations because not only is the number of cancer drug combinations extremely large but also the effectiveness of drug combinations varies depending on the genetic variation of cancer patients. A computational approach that prioritizes the best drug combinations considering the genetic information of a cancer patient is necessary to reduce the search space.

Results

We propose an in-silico method for personalized drug combination therapy discovery. We predict the synergy between two drugs and a cell line using genomic information, targets of drugs, and pharmacological information. We calculate and predict the synergy scores of 583 drug combinations for 31 cancer cell lines. For feature dimension reduction, we select the mutations or expression levels of the genes in cancer-related pathways. We also used various machine learning models. Extremely Randomized Trees (ERT), a tree-based ensemble model, achieved the best performance in the synergy score prediction regression task. The correlation coefficient between the synergy scores predicted by ERT and the actual observations is 0.738. To compare with an existing drug combination synergy classification model, we reformulate the problem as a binary classification problem by thresholding the synergy scores. ERT achieved an F1 score of 0.954 when synergy scores of 20 and -20 were used as the threshold, which is 8.7% higher than that obtained by the state-of-the-art baseline model. Moreover, the model correctly predicts the most synergistic combination, from approximately 100 candidate drug combinations, as the top choice for 15 out of the 31 cell lines. For 28 out of the 31 cell lines, the model predicts the most synergistic combination in the top 10 of approximately 100 candidate drug combinations. Finally, we analyze the results, generate synergistic rules using the features, and validate the rules through the literature survey.

Conclusion

Using various types of genomic information of cancer cell lines, targets of drugs, and pharmacological information, a drug combination synergy prediction pipeline is proposed. The pipeline regresses the synergy level between two drugs and a cell line as well as classifies if there exists synergy or antagonism between them. Discovering new drug combinations by our pipeline may improve personalized cancer therapy.

     

Gemcitabine/cannabinoid combination triggers autophagy in pancreatic cancer cells through a ROS-mediated mechanism

Gemcitabine (GEM, 2′,2′-difluorodeoxycytidine) is currently used in advanced pancreatic adenocarcinoma, with a response rate of < 20%. The purpose of our work was to improve GEM activity by addition of cannabinoids. Here, we show that GEM induces both cannabinoid receptor-1 (CB1) and cannabinoid receptor-2 (CB2) receptors by an NF-κB-dependent mechanism and that its association with cannabinoids synergistically inhibits pancreatic adenocarcinoma cell growth and increases reactive oxygen species (ROS) induced by single treatments. The antiproliferative synergism is prevented by the radical scavenger N-acetyl--cysteine and by the specific NF-κB inhibitor BAY 11-7085, demonstrating that the induction of ROS by GEM/cannabinoids and of NF-κB by GEM is required for this effect. In addition, we report that neither apoptotic nor cytostatic mechanisms are responsible for the synergistic cell growth inhibition, which is strictly associated with the enhancement of endoplasmic reticulum stress and autophagic cell death. Noteworthy, the antiproliferative synergism is stronger in GEM-resistant pancreatic cancer cell lines compared with GEM-sensitive pancreatic cancer cell lines. The combined treatment strongly inhibits growth of human pancreatic tumor cells xenografted in nude mice without apparent toxic effects. These findings support a key role of the ROS-dependent activation of an autophagic program in the synergistic growth inhibition induced by GEM/cannabinoid combination in human pancreatic cancer cells.