Candidate For Cancer Therapeutics Research Articles

Synthesis, antiproliferative and apoptosis induction potential activities of novel bis(indolyl)hydrazide-hydrazone derivatives.

In recent years, indole-indazolyl hydrazide-hydrazone derivatives with strong cell growth inhibition and apoptosis induction characteristics are being strongly screened for their cancer chemo-preventive potential. In the present study, N-methyl and N,N-dimethyl bis(indolyl)hydrazide-hydrazone analog derivatives were designed, synthesized and allowed to evaluate for their anti-proliferative and apoptosis induction potential against cervical (HeLa), breast (MCF-7 and MDA-MB-231) and lung (A549) cancer cell lines relative to normal HEK293 cells. The MTT assay in conjunction with mitochondrial potential assays and the trypan blue dye exclusion were employed to ascertain the effects of the derivatives on the cancer cells. Further, mechanistic studies were conducted on compound 14a to understand the biochemical mechanisms and functional interactions with various signaling pathways triggered in HeLa and MCF-7 cells. Compound 14a induced apoptosis via caspase independent pathway through the participation of mitogen-activated protein kinases (MAPK) such as extracellular signal related kinase (ERK) and p38 as well as p53 pathways. It originates the activation of pro-apoptotic proteins such as Bak and Mcl-1s and also strongly induced the generation of reactive oxygen species. In downstream signaling pathway, activated p53 protein interacted with MAPK pathways, including SAPK/c-Jun N-terminal protein kinase (JNK), p38 and ERK kinases resulting in apoptotic cell death. The involvement of MAPK cascades such as p38, ERK and p38 on compound 14a induced apoptotic cell death was evidenced by the fact that the inclusion of specific inhibitors of p38, ERK1/2 and JNK MAPK (SB2035809, PD98059 and SP600125) prevented the compound 14a towards induced apoptosis. The results clearly showed that MAP kinase cascades were crucial for apoptotic response in compound 14a induced cellular killing and were dependent on p53 activity. Based on the results, compound 14a was identified as a promising candidate for cancer therapeutics and these findings furnish a basis for further in vivo experiments on anti-proliferative activity.

N-(3-oxo-acyl) homoserine lactone induced germ cell apoptosis and suppressed the over-activated RAS/MAPK tumorigenesis via mitochondrial-dependent ROS in C. elegans.

As a quorum-sensing molecule for bacteria-bacteria communication, N-(3-oxododecanoyl)-homoserine lactone (C12) has been found to possess pro-apoptotic activities in various cell culture models. However, the detailed mechanism of how this important signaling molecule function in the cells of live animals still remains largely unclear. In this study, we systematically investigated the mechanism for C12-mediated apoptosis and studied its anti-tumor effect in Caenorhabditis elegans (C. elegans). Our data demonstrated that C12 increased C. elegans germ cell apoptosis, by triggering mitochondrial outer membrane permeabilization (MOMP) and elevating the reactive oxygen species (ROS) level. Importantly, C12-induced ROS increased the expression of genes critical for DNA damage response (hus-1, clk-2 and cep-1) and genes involved in p38 and JNK/MAPK signaling pathway (nsy-1, sek-1, pmk-1, mkk-4 and jnk-1). Furthermore, C12 failed to induce germ cell apoptosis in animals lacking the expression of each of those genes. Finally, in a C. elegans tumor-like symptom model, C12 significantly suppressed tumor growth through inhibiting the expression of RAS/MAPK pathway genes (let-23/EGFR, let-60/RAS, lin-45/RAF, mek-2/MEK and mpk-1/MAPK). Overall, our results indicate that DNA damage response and MAPK activation triggered by mitochondrial ROS play important roles in C12-induced apoptotic signaling in C. elegans, and RAS/MAPK suppression is involved in the tumor inhibition effect of C12. This study provides in vivo evidence that C12 is a potential candidate for cancer therapeutics by exerting its pro-apoptotic and anti-tumor effects via elevating mitochondria-dependent ROS production.

Novel antitumor compound optimized from natural saponin Albiziabioside A induced caspase-dependent apoptosis and ferroptosis as a p53 activator through the mitochondrial pathway

It is highly desirable to activation p53 function with small-molecule compounds for colon cancer therapy. Triterpene saponin has been characterized with the favorable selectivity and safety profiles. However, the application of triterpene saponin as cancer chemotherapy drugs was hampered primarily by moderate anticancer potency and the lack the mechanism of action. In this study, we synthesized a series of Albiziabioside A derivatives and evaluated the antitumor activity both in vitro and in vivo. Compounds D13 possessed strong inhibitory activity against HCT116 cells with IC50 values of 5.19 μM. More importantly, compound D13 had a favorable selectivity and was efficacious against MDR cancer cells. Moreover, compound D13 could induce apoptosis and ferroptosis through the mitochondrial pathway as a p53 activator. In addition, compound D13 significantly suppressed tumorigenesis without inducing toxicity in normal organs in vivo. Collectively, this study provides a clinically relevant argument for considering triterpene saponin derivatives D13 as potential cancer therapeutic candidates with enhanced activity, acceptable safety and novel mechanisms of action. To the best of our knowledge, this compound is the first drug candidate which can induce apoptosis and ferroptosis as a p53 activator.

ING4 expressing oncolytic vaccinia virus promotes anti-tumor efficiency and synergizes with gemcitabine in pancreatic cancer.

With no effective treatments available for most pancreatic cancer patients, pancreatic cancer continues to be one of the most difficult malignancies to treat. Oncolytic virus mediated-gene therapy has exhibited ubiquitous antitumor potential. In this study, we constructed a novel oncolytic vaccinia virus harboring the inhibitor of growth family member 4 gene (VV-ING4) to investigate its therapeutic efficacy alone or in combination with gemcitabine against pancreatic cancer cells in vitro and in vivo. ING4 expression was determined via quantitative real-time polymerase chain reaction (qPCR) and western blot. The cytotoxicity of VV-ING4 was measured using a cell proliferation assay. Both flow cytometry and western blot were applied to analyze the cell cycle and apoptosis. Furthermore, the combination inhibitory effect of VV-ING4 and gemcitabine was assessed using Chou-Talalay analysis in vitro and a BLAB/c mice model in vivo. We found that VV-ING4 significantly increases ING4 expression, displayed greater cytotoxic efficiency, and induced pancreatic cancer cell apoptosis and G2/M phase arrest. Additionally, the combination of VV-ING4 and gemcitabine synergistically effect in vitro and in vivo. Taken together, our data implicate VV-ING4 as a conceivable pancreatic cancer therapeutic candidate alone or in combination with gemcitabine.

Abstract 3073: Potential predictive biomarkers of clinical responses for a novel CDC7-selective inhibitor TAK-931

Abstract Cell division cycle 7 (CDC7) is a serine/threonine kinase, which plays important roles in initiation of DNA replication by phosphorylating MCM2. Kinase activity of CDC7 is controlled by its binding protein DBF4 in a cell-cycle dependent manner. Here we developed a potent CDC7 inhibitor TAK-931 (IC50< 0.3 nM) as a cancer therapeutic drug candidate, which exhibits a time-dependent ATP-competitive kinetics to its ATP-binding pocket. The selectivity studies using the 308 kinases revealed >120-fold selectivity of TAK-931 for CDC7 kinase inhibition compared to other kinase inhibitions. Treatment with TAK-931 suppressed the cellular MCM2 phosphorylation at Ser40, resulting in a delayed S phase progression, checkpoint activation, apoptosis, and potent growth suppression in various cancer cell lines. Furthermore, oral administration of TAK-931 as a single agent caused a significant antitumor activity in multiple xenograft models which include both cell line-based xenografts and patient-derived xenograft (PDX) models. These results demonstrate that TAK-931 is a highly potent and selective inhibitor of CDC7 kinase, and causes a potent antiproliferation both in vitro and in vivo studies using various cancer cells. Next, to identify potential predictive biomarkers to guide patient-selection strategies, in vitro cell panel screening of TAK-931 using was tested for its ability to antiproliferation in 246 cell lines, which includes both solid and hematological cancer cells. TAK-931 inhibited proliferation of multiple cancer cell lines, with mean concentration producing a half-maximal response (EC50) values ranging from 29.1 nM to > 30 μM (median = 554.5 nM). While the wide range of TAK-931 antiproliferative spectrum was observed, neither doubling speed nor CDC7 expression profile did predict tTAK-931 sensitivity in cancer cell lines. A correlative study of the tumor genetic mutations in relation to antiproliferative activity that KRAS mutant cancer cells were more sensitive to TAK-931 compared to KRAS non-mutant cell lines (p<0.05). We also confirmed this KRAS-associated antiproliferative effect of TAK-931 in the SW48 isogenic cell lines of KRAS mutations (G12V, G13D). The ectopic expressions of G12V- and G13D-KRAS mutations increased the TAK-931 sensitivity compared to the KRAS-wild parental SW48 cell line. Given that KRAS mutations are frequently detected in clinical pancreatic tumors, we next conducted in vivo efficacy studies using pancreatic PDX models. Consistent to our hypothesis from in vitro studies, KRAS-mutant pancreatic PDX tumors were more sensitive to TAK-931 than the KRAS-wild pancreatic tumors; all KRAS-mutant pancreatic PDX models we tested exhibited >60% TGI. Our findings suggest that the KRAS-mutant pancreatic tumors could be the potential candidate for the TAK-931 target indication. Citation Format: Kenichi Iwai, Tadahiro Nambu, Osamu Kurasawa, Noriko Uchiyama, Ryo Dairiki, Yukiko Yamamoto, Satoru Nishizawa, Mengkun Zhang, Yuko Ishii, Huifeng Niu, Akihiro Ohashi. Potential predictive biomarkers of clinical responses for a novel CDC7-selective inhibitor TAK-931 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3073. doi:10.1158/1538-7445.AM2017-3073

Specific humoral and cellular immune responses in cancer patients undergoing chronic immunization with a VEGF-based therapeutic vaccine

CIGB-247 is a cancer therapeutic vaccine, based on recombinant modified human vascular endothelial growth factor (VEGF) as antigen, in combination with the adjuvant VSSP, a bacterially-derived adjuvant. The vaccine have demonstrated efficacy in several murine malignancy models. These studies supported the rationale for a phase I clinical trial where safety, tolerance, and immunogenicity of CIGB-247 was studied in patients with advanced solid tumors at three antigen dose level. Surviving individuals of this clinical trial were eligible to receive off-trial voluntary re-immunizations. The present work is focus in the immunological follow up of these patients after approximately three years of immunizations, without additional oncological treatments. Long term vaccination was feasible and safe. Our results indicated that after sustained vaccination most of the patients conserved their seroconversion status. The specific anti-VEGF IgG titer diminished, but in all the cases keeps values up from the pre-vaccination levels. Continued vaccination was also important to produce a gradual shift in the anti-VEGF IgG response from IgG1 to Ig4. Outstanding, our results indicated that long-term off-trial vaccination could be associated with the maintaining of one reserve of antibodies able to interfere with the VEGF/Receptor interaction and the production of IFNγ secretion in CD8+ cells. The results derived from the study of this series of patients suggest that long term therapeutic vaccination is a feasible strategy, and highlight the importance of continuing the clinical development program of this novel cancer therapeutic vaccine candidate. We also highlight the future clinical applications of CIGB-247 in cancer and explain knowledge gaps that future studies may address.Registration number and name of trial registry: RPCEC00000102. Cuban Public Clinical Trial Registry (WHO accepted Primary Registry). Available from: http://registroclinico.sld.cu/.

In vivo immunotherapy of lung cancer using cross-species reactive vascular endothelial growth factor nanobodies.

Objective(s):Lung cancer is the main leading cause of cancer death worldwide. Angiogenesis is the main step in proliferation and spreading of tumor cells. Targeting vascular endothelial growth factor (VEGF) is an effective approach for inhibition of cancer angiogenesis. Nanobodies (NBs) are a novel class of antibodies derived from the camel. Unique characteristics of Nbs like their small size and good penetration to tumor tissues makes them promising tools in drug development. Development of NBs targeting both human and mouse VEGF is required for understanding their in vivo functions. Therefore, development of cross-species reactive anti-VEGF Nbs for immunotherapy of lung cancer was the main aim of the current study.Materials and Methods:Here we developed NBs from Camelus dromedarius library with high specificity and binding affinity to both human and mouse VEGF. In vitro and In vivo function of developed NB was evaluated on human endothelial cells and lung epithelial tumor cells (TC-1).Results:A nanobody showed the highest affinity to human and mouse VEGF and potently inhibited VEGF in the ELISA experiment. Anti-VEGF NBs significantly inhibited in vitro human endothelial cell migration through blockade of VEGF (P=0.045). Anti-VEGF NBs also significantly inhibited in vivo TC-1 growth in a dose-dependent manner (P=0.001) and resulted in higher survival rate in the nanobody treated groupConclusion:These findings demonstrate the potential of anti-VEGF NBs in tumor growth inhibition and are promising as novel cancer therapeutic candidate.

MicroRNA-379-5p plays a tumor-suppressive role in human bladder cancer growth and metastasis by directly targeting MDM2.

Bladder cancer is the second most common urological malignancy in the US and is the most frequently diagnosed urological malignancy in China. An increasing amount of evidence indicates that microRNAs perform extremely important functions in many biological processes related to the formation and progression of cancers, including bladder cancer. Previous studies have reported that microRNA‑379-5p (miR-379-5p) is involved in tumour initiation and development in human cancers. However, the expression pattern, biological functions and the underlying mechanisms of miR-379-5p in bladder cancer remain unknown. The present study demonstrated that the expression levels of miR‑379-5p in bladder cancer tissues and cell lines were lower than the levels in adjacent normal tissues and the human bladder epithelial immortalized SV-HUC-1 cell line. Restoration of the expression of miR-379-5p inhibited bladder cancer cell proliferation, migration and invasion. Mouse double minute2 (MDM2) was identified as a direct target gene of miR-379-5p. Furthermore, similar to miR-379-5p overexpression in bladder cancer cells, inhibition of MDM2 exerted tumor-suppressive effects. Rescue experiments showed that upregulation of MDM2 reversed the inhibitory effects of miR-379-5p on bladder cancer cell proliferation, migration and invasion. MDM2 was highly expressed and inversely correlated with miR-379-5p expression in bladder cancer tissues. These findings suggest that the miR-379-5p/MDM2 pathway plays an important role in bladder cancer and could serve as a potential candidate for bladder cancer therapeutics.

MicroRNA-138 inhibits proliferation, migration and invasion through targeting hTERT in cervical cancer.

A growing body of evidence suggests that microRNA-138 (miR-138) functions as a tumor suppressor, and is involved in tumor initiation, development and metastasis in certain types of human cancers. However, the function and underlying molecular mechanism of miR-138 in cervical cancer remains unclear. Therefore, the purpose of the present study was to investigate the clinical significance of miR-138 expression in cervical cancer, and to evaluate its role and underlying mechanisms in cervical cancer. The present study indicated that miR-138 expression was significantly downregulated in cervical cancer tissues and cell lines, and that the low miR-138 expression was negatively associated with advanced FIGO stage and lymph node metastasis (P<0.01). Functional analyses indicated that the overexpression of miR-138 in cervical cancer cells inhibited cell proliferation, migration and invasion, induced cell apoptosis in vitro, and suppressed tumor growth in a nude mice model. Luciferase reporter assays confirmed that human telomerase reverse transcriptase was a novel target gene of miR-138. The findings of the present study suggested that miR-138 could be a potential candidate for cervical cancer therapeutics.

2-Azetidinones: Synthesis and biological evaluation as potential anti-breast cancer agents

A series of twenty-five 2-azitidinone (β-lactam) derivatives were synthesized and evaluated for anti-cancer properties against breast cancer, MCF-7 and MDA-MB-231. These β-lactam derivatives depicted significant cytotoxicity in cancer cell lines but not in normal human mammary epithelial cells, MEpiC. Interestingly, derivatives of 2-bromo ethyl acrylonitrile (19w) exhibited - potent anti-proliferative activity with IC50, 5.79 ± 0.01 μM in MCF-7 and 6.86 ± 0.009 μM in MDA-MB-231. In addition, an increased expression of pro-apoptotic genes (p53, Bax, Bid) as well as decreased mRNA expression of cyclins D1, E and Cdk 2, 6 along with cell cycle arrest at G1 phase was observed. 19w treatment has shown higher percentage of Annexin-positive cells indicating induction of apoptosis. Further, docking studies confirmed an interaction between 19w and ATP-binding catalytic site of AKT1. Mechanistically, 19w depicted dose-dependent decrease in phosphorylation of AKT and GSK-3β and significant decrease in AKT kinase activity. In conclusion, β-lactam derivative 19w is a potential anti-breast cancer therapeutic candidate targeting cell survival pathway (AKT/GSK3β).

Soluble VEGF receptor 1 (sFLT1) induces non-apoptotic death in ovarian and colorectal cancer cells.

Soluble Vascular Endothelial Growth Factor Receptor 1 (sVEGFR1/sFLT1) is an angiogenesis inhibitor that competes with angiogenic factors such as VEGF and Placental Growth Factor (PlGF). Imbalances of VEGF and sFLT1 levels can cause pathological conditions such as tumour growth or preeclampsia. We observed direct damage caused by sFLT1 in tumour cells. We exposed several kinds of cells derived from ovarian and colorectal cancers as well as HEK293T cells to sFLT1 in two ways, transfection and exogenous application. The cell morphology and an LDH assay revealed cytotoxicity. Additional experiments were performed to clarify how sFLT1 injured cells. In this study, non-apoptotic cell damage was found to be induced by sFLT1. Moreover, sFLT1 showed an anti-tumour effect in a mouse model of ovarian cancer. Our results suggest that sFLT1 has potential as a cancer therapeutic candidate.

Identification of a Small Molecule That Overcomes HdmX-Mediated Suppression of p53.

Inactivation of the p53 tumor suppressor by mutation or overexpression of negative regulators occurs frequently in cancer. As p53 plays a key role in regulating proliferation or apoptosis in response to DNA-damaging chemotherapies, strategies aimed at reactivating p53 are increasingly being sought. Strategies to reactivate wild-type p53 include the use of small molecules capable of releasing wild-type p53 from key, cellular negative regulators, such as Hdm2 and HdmX. Derivatives of the Hdm2 antagonist Nutlin-3 are in clinical trials. However, Nutlin-3 specifically disrupts Hdm2-p53, leaving tumors harboring high levels of HdmX resistant to Nutlin-3 treatment. Here, we identify CTX1, a novel small molecule that overcomes HdmX-mediated p53 repression. CTX1 binds directly to HdmX to prevent p53-HdmX complex formation, resulting in the rapid induction of p53 in a DNA damage-independent manner. Treatment of a panel of cancer cells with CTX1 induced apoptosis or suppressed proliferation and, importantly, CTX1 demonstrates promising activity as a single agent in a mouse model of circulating primary human leukemia. CTX1 is a small molecule HdmX inhibitor that demonstrates promise as a cancer therapeutic candidate. Mol Cancer Ther; 15(4); 574-82. ©2016 AACR.

N-Hydroxyphthalimide exhibits antitumor activity by suppressing mTOR signaling pathway in BT-20 and LoVo cells.

BackgroundN-Hydroxyphthalimide (NHPI), an important chemical raw material, was found to have potent and selective anti-proliferative effect on human breast carcinoma BT-20 cells, human colon adenocarcinoma LoVo and HT-29 cells during our screening for anticancer compounds. The purpose of this study is to assess the antitumor efficacy of NHPI in vitro and in vivo and to explore the underlying antitumor mechanism.MethodsCell cytotoxicity of NHPI was evaluated using MTS assay and cell morphological analysis. After NHPI treatment, cell cycle, apoptosis and mitochondrial membrane potential were analyzed using flow cytometer. The subcellular localization of eukaryotic initiation factor 4E (eIF4E) was analyzed by immunofluorescence assay. The antitumor efficacy of NHPI in vivo was tested in BT-20 xenografts. The underlying antitumor mechanisms of NHPI in vitro and in vivo were investigated with western blot analysis in NHPI-treated cancer cells and tumor tissues. Statistical significance was determined using Student’s t-test.ResultsIn vitro, NHPI selectively inhibited the proliferation and induced G2/M phase arrest in BT-20 and LoVo cells, which was attributed to the inhibition of cyclin B1 and cdc2 expressions. Furthermore, NHPI induced apoptosis via mitochondrial pathway. Of note, NHPI effectively inhibited mammalian target of rapamycin (mTOR) complex 1 (mTORC1) and mTOR complex 2 (mTORC2) signaling, and overcame the feedback activation of Akt and extracellular signal-regulated kinase (ERK) caused by mTORC1 inhibition in BT-20 and LoVo cells. In vivo, NHPI inhibited tumor growth and suppressed mTORC1 and mTORC2 signaling in BT-20 xenografts with no obvious toxicity.ConclusionsWe found for the first time that NHPI displayed antitumor activity which is associated with the inhibition of mTOR signaling pathway. Our findings suggest that NHPI may be developed as a promising candidate for cancer therapeutics by targeting mTOR signaling pathway and as such warrants further exploration.

Systems Perturbation Analysis of a Large-Scale Signal Transduction Model Reveals Potentially Influential Candidates for Cancer Therapeutics.

Dysregulation in signal transduction pathways can lead to a variety of complex disorders, including cancer. Computational approaches such as network analysis are important tools to understand system dynamics as well as to identify critical components that could be further explored as therapeutic targets. Here, we performed perturbation analysis of a large-scale signal transduction model in extracellular environments that stimulate cell death, growth, motility, and quiescence. Each of the model’s components was perturbed under both loss-of-function and gain-of-function mutations. Using 1,300 simulations under both types of perturbations across various extracellular conditions, we identified the most and least influential components based on the magnitude of their influence on the rest of the system. Based on the premise that the most influential components might serve as better drug targets, we characterized them for biological functions, housekeeping genes, essential genes, and druggable proteins. The most influential components under all environmental conditions were enriched with several biological processes. The inositol pathway was found as most influential under inactivating perturbations, whereas the kinase and small lung cancer pathways were identified as the most influential under activating perturbations. The most influential components were enriched with essential genes and druggable proteins. Moreover, known cancer drug targets were also classified in influential components based on the affected components in the network. Additionally, the systemic perturbation analysis of the model revealed a network motif of most influential components which affect each other. Furthermore, our analysis predicted novel combinations of cancer drug targets with various effects on other most influential components. We found that the combinatorial perturbation consisting of PI3K inactivation and overactivation of IP3R1 can lead to increased activity levels of apoptosis-related components and tumor-suppressor genes, suggesting that this combinatorial perturbation may lead to a better target for decreasing cell proliferation and inducing apoptosis. Finally, our approach shows a potential to identify and prioritize therapeutic targets through systemic perturbation analysis of large-scale computational models of signal transduction. Although some components of the presented computational results have been validated against independent gene expression data sets, more laboratory experiments are warranted to more comprehensively validate the presented results.

Nitroxide radical containing nanoparticle as a potential candidate for redox therapeutics in breast cancer

Event Abstract Back to Event Nitroxide radical containing nanoparticle as a potential candidate for redox therapeutics in breast cancer Babita Shashni1 and Yukio Nagasaki1, 2, 3 1 Graduate School of Pure and Applied Sciences, University of Tsukuba, Department of Material Science, Japan 2 Graduate School of Comprehensive Human Sciences, University of Tsukuba, Master’s School of Medical Sciences, Japan 3 National Institute for Materials Science (NIMS), Satellite Laboratory, International Center for Materials Nanoarchitectonics (WPI-MANA), Japan Introduction: Elevated basal level of reactive oxygen species (ROS) is a hallmark for cancer cell phenotype. ROS, which is considered to be an oncogenic, has been reported to play a critical role in cancer development, progression and metastasis. Therefore ROS manipulating strategies in cancers may hold a great potential for cancer therapy. Henceforth, in this line we focused our study to investigate the therapeutic efficacy of our pH sensitive and non-sensitive ROS scavenging redox nanoparticle (RNP) [1],[2] (Figure 1) in breast cancer by specific inhibition of ROS regulated proteins associated with cancer metastasis and angiogenesis. Figure 1: Chemical structure representation of a) RNP(N): pH-sensitive and b) RNP(O): non-pH sensitive Materials and Methods: Therapeutic efficacy of RNPs was evaluated by in vitro and in vivo assays. In vitro anti-proliferative, -metastatic and -angiogenic activities were confirmed by cell viability, migration and endothelial migration and tube cell formation assays, respectively, whereas in in vivo the efficacy was investigated in breast cancer mice models by biochemical assays. The mechanisms of anti -metastatic and -angiogenic activities were investigated by expression studies. Results: We confirmed that RNPs significantly restricts proliferation, migration and angiogenesis in breast cancer. Furthermore, we also confirmed RNP directed suppression of ROS regulated migration (NF-kβ and metallo matrix proteases (MMPs)) and angiogenesis (vascular endothelial growth factor (VEGF)) promoting proteins. Anti-metastatic tendency was also confirmed in in vivo breast cancer mice models. Discussion: ROS plays a critical role in cancer cell survival. Considering the dependence of cancer cells on ROS-induced molecular signaling pathways, ROS can thus represent a vulnerable specific target for redox cancer therapeutics. In this study we specifically inhibited the ROS induced metastasis and angiogenesis promoting characteristics of breast cancer cells by down-regulating transcriptional factor NF-kB, MMPs and VEGF (Figure 2). This preclinical evaluation study will thus establish RNP as a potential candidate for breast cancer therapeutics. Figure 2: Schematic representation of mechanism of RNP directed restriction of metastasis and angiogenesis in breast cancer Conclusion: We provide the first molecular evidence of anti-metastatic and anti-angiogenic activities of RNP. This report thus strengthens the role of ROS scavenging nanoparticles as potential candidates for breast cancer therapeutics.

Smac mimetic with TNF-α targets Pim-1 isoforms and reactive oxygen species production to abrogate transformation from blebbishields.

Cancer cells are capable of sphere formation (transformation) through reactive oxygen species (ROS) and glycolysis shift. Transformation is linked to tumorigenesis and therapy resistance, hence targeting regulators of ROS and glycolysis is important for cancer therapeutic candidates. Here, we demonstrate that Smac mimetic AZ58 in combination with tumour necrosis factor-α (TNF-α) was able to inhibit the production of ROS, inhibit glycolysis through Pim-1 kinase-mediated Ser-112 phosphorylation of BAD, and increase depolarization of mitochondria. We also identified mitochondrial isoforms of Pim-1 kinase that were targeted for degradation by AZ58 in combination with TNF-α or AZ58 in combination with Fas ligand (FasL) plus cycloheximide (CHX) through caspase-3 to block transformation. Our study demonstrates that Smac mimetic in combination with TNF-α is an ideal candidate to target Pim-1 expression, inhibit ROS production and to block transformation from blebbishields.

Phytocannabinoids for Cancer Therapeutics: Recent Updates and Future Prospects.

Phytocannabinoids (pCBs) are lipid-soluble phytochemicals present in the plant, Cannabis sativa L. and non-cannabis plants which have a long history in recreation and traditional medicine. The plant and the constituents isolated were central in the discovery of the endocannabinoid system (ECS), the most new target for drug discovery. The ECS includes two G-protein-coupled receptors; the cannabinoid receptors-1 and -2 (CB1 and CB2) for marijuana's psychoactive principle Δ(9)-tetrahydrocannabinol (Δ(9)-THC), their endogenous small lipid ligands; namely anandamide (AEA) and 2-arachidonoylglycerol (2-AG), also known as endocannabinoids and the enzymes for endocannabinoid biosynthesis and degradation such as fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). The ECS has been suggested as a pro-homeostatic and pleiotropic signaling system activated in a time- and tissue-specific way during pathological conditions including cancer. Targeting the CB1 receptors becomes a concern because of adverse psychotropic reactions. Hence, targeting the CB2 receptors or the endocannabinoid metabolizing enzymes by pCBs obtained from plants lacking psychotropic adverse reactions has garnered interest in drug discovery. These pCBs derived from plants appear safe and effective with a wider access and availability. In the recent years, several pCBs derived other than non-cannabinoid plants have been reported to bind to and functionally interact with cannabinoid receptors and appear promising candidate for drug development including cancer therapeutics. Several of them also targets the endocannabinoid metabolizing enzymes that control endocannabinoid levels. In this article, we summarize and critically discuss the updates and future prospects of the pCBs as novel and promising candidates for cancer therapeutics.

Inhibition of angiogenesis in human endothelial cell using VEGF specific nanobody

Angiogenesis is an important step in tumor development and metastasis. Vascular endothelial growth factor (VEGF) plays an important role in progression of angiogenesis. VEGF121 and VEGF165 are the most relative forms of VEGF family which contain the full biological activity. Nanobodies derived from camelidae are the smallest biding site of antigen. Unique characteristic of nanobodies make them as a useful candidate for research. In this report, we describe the isolation of VEGF specific nanobodies from dromedaries immunized with purified VEGF antigen using phage display. Four clones that showed the highest signal value in ELISA experiment were selected and expressed as a His-tagged fusion protein. Four selected nanobodies were reacted strongly to VEGF in cross-reactivity assay. The binding affinity of selected nanobodies named Nb22, Nb23, Nb35 and Nb42 were differed from 0.1 to 60nM. The nanobodies inhibited endothelial cell proliferation or tube formation in response of VEGF in a dose-dependent manner. These results indicate the potential of nanobodies in inhibition of VEGF and represent a promising candidate for cancer research and therapeutics.

The tetravalent anti-DR5 antibody without cross-linking direct induces apoptosis of cancer cells

Tumor necrosis factor-related apoptosis-inducing ligand can induce apoptosis in many tumor cell lines. This apoptotic effect is mediated by interaction of TRAIL and its receptors, which include Death Receptor 4 (DR4) and Death Receptor 5 (DR5). Some antibodies to DR4 or DR5 do not have anti-tumor ability without cross-linking but exhibit anti-tumor ability in the presence of a cross-linking reagent. Here, we suggest that the tetravalent anti-DR5 antibody can induce apoptosis of cancer cells independent of cross-linking reagent. The single-chain variable fragment of the anti-DR5 antibody, HSA (human serum albumin) – p53 gene, comprising residues 490–513 of HSA and the tetramerization domain of human p53 were assembled into the tetravalent antibody by an overlapping PCR. Results of size exclusion HPLC indicated that the purified protein exhibited a major peak (tetramer) and a minor peak (dimer). MTT assay demonstrated the tetravalent antibody without cross-linking could inhibit survival of Jurkat and EC9706 cells in a dose-dependent manner while the monovalent antibody could not inhibit survival of Jurkat and EC9706 cells. IC50 of Jurkat cell was 3.2 mg/L and IC50 of EC9706 cell was 3.9 mg/L. Furthermore, the Annexin V/PI assay and the Hoechst 33258 staining showed that the tetravalent antibody could efficiently induce apoptosis of Jurkat and EC9706 cells. Therefore, the tetravalent anti-DR5 antibody can act as a direct agonistic antibody, and initiate efficient apoptotic independent of cross-linking reagent. Thus, the tetravalent anti-DR5 antibody will be a new kind of candidate for potential cancer therapeutics.

Expression of nicotinamide phosphoribosyltransferase-influenced genes predicts recurrence-free survival in lung and breast cancers.

Nicotinamide phosphoribosyltransferase (NAMPT) is a rate-limiting enzyme in the salvage pathway of nicotinamide adenine dinucleotide biosynthesis. NAMPT protein is a secreted plasma biomarker in inflammation and in cancer. The NAMPT enzymatic inhibitor, FK866, acts as an inducer of apoptosis and is a cancer therapeutic candidate, however, little is known regarding the influence of NAMPT on cancer biological mechanisms or on the prognosis of human cancers. We interrogated known microarray data sets to define NAMPT knockdown-influenced gene expression to demonstrate that reduced NAMPT expression strongly dysregulates cancer biology signaling pathways. Comparisons of gene expression datasets of four cancer types generated a N39 molecular signature exhibiting consistent dysregulated expression in multiple cancer tissues. The N39 signature provides a significant and independent prognostic tool of human recurrence-free survival in lung and breast cancers. Despite the absence of clear elucidation of molecular mechanisms, this study validates NAMPT as a novel “oncogene” with a central role in carcinogenesis. Furthermore, the N39 signature provides a potentially useful tool for prediction of recurrence-free survival in lung and breast cancer and validates NAMPT as a novel and effective therapeutic target in cancer.