Synthesis, biological evaluation, and physicochemical property assessment of 4-substituted 2-phenylaminoquinazolines as Mer tyrosine kinase inhibitors.

Current results identified 4-substituted 2-phenylaminoquinazoline compounds as novel Mer tyrosine kinase (Mer TK) inhibitors with a new scaffold. Twenty-one 2,4-disubstituted quinazolines (series 4-7) were designed, synthesized, and evaluated against Mer TK and a panel of human tumor cell lines aimed at exploring new Mer TK inhibitors as novel potential antitumor agents. A new lead, 4b, was discovered with a good balance between high potency (IC50 0.68µM) in the Mer TK assay and antiproliferative activity against MV4-11 (GI50 8.54µM), as well as other human tumor cell lines (GI50<20µM), and a desirable druglike property profile with low logP value (2.54) and high aqueous solubility (95.6µg/mL). Molecular modeling elucidated an expected binding mode of 4b with Mer TK and necessary interactions between them, thus supporting the hypothesis that Mer TK might be a biologic target of this kind of new active compound.

Anticancer activity of tuftsin-derived T peptide in postoperative residual tumors.

Immune adjuvants have been used in cancer biotherapies to stimulate immune response to tumor cells. Despite their potential as anticancer reagents, there are several impediments to their use in clinical applications. In this study, we aim to modify the existing tuftsin structure and evaluate its antitumor activity in preclinical models. We synthesized a novel tuftsin derivative, namely, the T peptide (TP), by linking four tuftsin peptides, which showed enhanced stability in vivo. We then evaluated its anticancer activity in a postoperative residual tumor model in mice, where we surgically removed most of the primary tumor from the host, a procedure mimicking clinically postoperative patients. Despite the limited effect in intact solid tumors, TP strongly inhibited relapsed growth of residual tumors in postsurgical mice. Surgical resection of tumors accelerated residual tumor growth, but TP slowed down this process significantly. Interestingly, TP showed similar effects in human xenograft residual models. As an immunomodulator, TP could synergize the functions of macrophages, thus inhibiting the growth of cocultured tumor cells in vitro. Furthermore, TP could shift the macrophages to the tumor-suppressive M1 type and mobilize them to produce elevated cytotoxic TNF-α and NO. As a result, TP effectively prolonged the survival time of tumor-resected mice. Using the postoperative residual tumor models, we provide a body of evidence showing the antitumor activity of TP, which causes no obvious toxicity. Our study highlights the potential of TP as a postoperative adjuvant in cancer therapies.

Exploration of the protection of riboflavin laurate on oral mucositis induced by chemotherapy or radiotherapy at the cellular level: what is the leading contributor?

Oral or gastrointestinal mucositis is a frequent phenomenon in cancer patients receiving chemotherapy or radiotherapy. In addition, several clinical investigations have demonstrated in recent years that riboflavin laurate has the potential to protect the patients from the disease induced by chemotherapy or radiotherapy. In our studies, it is observed that riboflavin laurate can ameliorate either chemotherapy- or radiotherapy-induced toxicities on Helf cells, and the effect is greater than that of riboflavin. In addition, riboflavin laurate is able to transport through the Caco-2 cell monolayer as the prototype, indicating the protective effects may be produced by the prototype of riboflavin laurate, rather than simply by the released riboflavin.

[Corrigendum] Fully human VEGFR2 monoclonal antibody BC001 attenuates tumor angiogenesis and inhibits tumor growth.

Subsequently to the publication of the above paper, an interested reader drew to the authors' attention that, for the scratch wound assay experiments shown in Fig. 1 on p. 2413, the panels showing the '0 h' experiments for the respective incubations with VEGF or BC001 were apparently identical. The authors were able to re­examine their original data files, and realized that this figure had been inadverently assembled incorrectly. The revised version of Fig. 1, containing the correct data for the '0 h / BC001' panel, is shown below. Note that the revisions made to this figure do not affect the overall conclusions reported in the paper. The authors are grateful to the Editor of International Journal of Oncology for allowing them the opportunity to publish this Corrigendum, and apologize to the readership for any inconvenience caused. [International Journal of Oncology 45: 2411­2420, 2014; DOI: 10.3892/ijo.2014.2690].

Production and radioprotective effects of pyrroloquinoline quinone.

Pyrroloquinoline quinone (PQQ) was produced by fermentation of the Methylovorus sp. MP688 strain and purified by ion-exchange chromatography, crystallization and recrystallization. The yield of PQQ reached approximately 125 mg/L and highly pure PQQ was obtained. To determine the optimum dose of PQQ for radioprotection, three doses (2 mg/kg, 4 mg/kg, 8 mg/kg) of PQQ were orally administrated to the experimental animals subjected to a lethal dose of 8.0 Gy in survival test. Survival of mice in the irradiation + PQQ (4 mg/kg) group was found to be significantly higher in comparison with the irradiation and irradiation + nilestriol (10 mg/kg) groups. The numbers of hematocytes and bone marrow cells were measured for 21 days after sublethal 4 Gy gamma-ray irradiation with per os of 4 mg/kg of PQQ. The recovery of white blood cells, reticulocytes and bone marrow cells in the irradiation + PQQ group was faster than that in the irradiation group. Furthermore, the recovery of bone marrow cell in the irradiation + PQQ group was superior to that in irradiation + nilestriol group. Our results clearly indicate favourable effects on survival under higher lethal radiation doses and the ability of pyrroloquinoline quinine to enhance haemopoietic recovery after sublethal radiation exposure.

A Modified Surgical Ventricular Reconstruction in Post-infarction Mice Persistently Alleviates Heart Failure and Improves Cardiac Regeneration.

Objectives: Large ventricular aneurysm secondary to myocardial infarction (MI) results in severe heart failure (HF) and limits the effectiveness of regeneration therapy, which can be improved by surgical ventricular reconstruction (SVR). However, the conventional SVR procedures do not yield optimal long-term outcome in post-MI rodents. We hypothesized that a modified SVR procedure without aggressive purse string suture would persistently alleviate HF and improve cardiac regeneration in post-MI mice. Methods: Adult male C57 mice were subjected to MI or sham surgery. Four weeks later, mice with MI underwent SVR or 2nd open-chest operation alone. SVR was performed by plicating the aneurysm with a single diagonal linear suture from the upper left ventricle (LV) to the right side of the apex. Cardiac remodeling, heart function and myocardial regeneration were evaluated. Results: Three weeks after SVR, the scar area, LV volume, and heart weight/body weight ratio were significantly smaller, while LV ejection fraction, the maximum rising and descending rates of LV pressure, LV contractility and global myocardial strain were significantly higher in SVR group than in SVR-control group. The inhibitory effects of SVR on LV remodeling and HF persisted for at least eight-week. SVR group exhibited improved cardiac regeneration, as reflected by more Ki67-, Aurora B- and PH3-positive cardiomyocytes and a higher vessel density around the plication area of the infarcted LV. Conclusions: SVR with a single linear suture results in a significant and sustained reduction in LV volume and improvement in both LV systolic and diastolic function as well as cardiac regeneration.

Microbial transformation of epothilone A by Aspergillus niger AS 3.739.

The microbiological transformation of epothilone A (1) by Aspergillus niger AS 3.739 afforded four main metabolites. Their structures were elucidated by (1)H, (13)C NMR and HSQC, COSY, HMBC, and NOESY spectra as trans-12,13-hydroxylated epothilone A (2), cis-12,13-hydroxylated epothilone A (3), trans-12,15-epoxidated epothilone A (4), and cis-12,15-epoxidated epothilone A (5). All four compounds were firstly found based on their stereochemistry. These new compounds displayed cytotoxicity against human breast carcinoma cells MCF-7 with IC(50) 9.88 microg/ml of 2, 2.52 microg/ml of 3, 9.88 microg/ml of 4, and 5.68 microg/ml of 5.

Differential sensitivity of colorectal cancer cell lines to artesunate is associated with expression of beta-catenin and E-cadherin.

Artesunate, a remarkable antimalarial agent, also reveals profound cytotoxic activity. In the present investigation, we compared the anticancer effects of artesunate on three colorectal cancer cell lines and analyzed the relationship between drug sensitivity and malignant phenotype of the tumor cells. The findings are as follows: poorly-differentiated was colorectal cancer cell line CLY showing nuclear beta-catenin accumulation and loss of E-cadherin; moderately-differentiated were Lovo cells with cytoplasmic distribution of the two proteins; and well-differentiated were HT-29 cells with membranous localization of them. Also, both in vitro and in vivo, poorly- or moderately-differentiated CLY and Lovo were more susceptible to artesunate treatment than well-differentiated HT-29. Furthermore, the sensitive response of CLY and Lovo to artesunate was associated with membranous translocation of beta-catenin and increased expression of E-cadherin, which indicated the inhibition of hyperactive Wnt signaling pathway and the reversion of the epithelial to mesenchymal transition, respectively. Due to the vital roles of Wnt pathway and the epithelial to mesenchymal transition in tumor differentiation, we thought artesunate could promote the re-differentiation and apoptosis of colorectal cancer cells by inhibition of hyperactive Wnt pathway and reversion of the epithelial to mesenchymal transition.

CT­1042, a novel anticancer agent, exhibits effects by activating p53 and inhibiting survivin.

A novel small molecular compound, 4­ethyl­8­fluoro­hydroxy­9­methoxy­11­methyl­1,12­dihydro­4H­2­oxa­6,12a­diaza­dibenzo[b,h]fluorene­3,13­dione (CT­1042) exhibits potent antitumor activity against many tumor cells in vitro. However, the effects and underlying mechanisms of CT­1042 in non­small cell lung cancer (NSCLC) remain unclear. The present study was designed to determine the anticancer properties and underlying molecular mechanisms of CT­1042 in NCI­H460 NSCLC cells. A thiazolyl blue tetrazolium bromide assay (MTT) was performed to evaluate cell viability and flow cytometry was used to analyze apoptosis, mitochondrial membrane potential (MMP) and cell cycle. Real­time quantitative PCR and western blotting were conducted to determine relative mRNA and protein levels. A tumor xenograft experiment was performed to investigate the effects of CT­1042 on tumor growth in vivo. CT­1042 markedly inhibited the proliferation of twelve cancer cell lines, decreased MMP in subject cells and increased caspase­3 activity. Cell cycle analysis indicated that CT­1042 delayed the cell cycle progression during the G2/M phase in a dose­dependent manner. In addition, CT­1042 induced mitochondrial­mediated apoptosis by activating p53 and Bax, as well as inhibiting Bcl­2 and survivin. Finally, CT­1042 significantly suppressed NCI­H460 xenograft tumor growth in vivo, with low systemic toxicity. Collectively, these results revealed that CT­1042 has significant lung anticancer properties.

Novel proapoptotic agent SM-1 enhances the inhibitory effect of 5-fluorouracil on colorectal cancer cells in vitro and in vivo.

5-Fluorouracil (5-FU) is one of the most important agents used to treat colorectal cancer. However, the therapeutic effect of 5-FU on colon cancer is limited. SM-1 is a novel type of proapoptotic agent that directly activates procaspase-3 to caspase-3, leading to apoptosis in human cancer cells. The aim of the present study was to evaluate the antitumor effects of 5-FU in combination with SM-1. The human colorectal cancer cell lines HCT116 and LoVo were cultured in the presence of SM-1 and 5-FU. The combination of SM-1 and 5-FU treatment exhibited increased proliferation inhibitory effects compared with 5-FU treatment alone in HCT116 and LoVo cells, as determined using an MTT assay. SM-1 significantly decreased the half-maximal inhibitory concentration of 5-FU from 8.07±0.49 to 2.55±0.41 µmol/l in HCT116 cells, and from 7.90±0.98 to 3.14±0.81 µmol/l in LoVo cells. Similarly, the apoptotic activity was increased to 47.95 and 35.19% in HCT116 and LoVo cells, respectively, as determined using Annexin V/propidium iodide staining and flow cytometry. The combination of SM-1 and 5-FU treatment led to significantly increased caspase-3 activity compared with either compound alone. The reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis revealed the downregulation of B-cell lymphoma 2 and Survivin, and the upregulation of apoptosis regulator Bcl-2-associated X protein and cleaved poly (ADP-ribose) polymerase in HCT116 and LoVo cells. In addition, RT-qPCR identified downregulation of X-linked inhibitor of apoptosis protein mRNA. 5-FU and SM-1 treatment in combination increased tumor proliferation inhibition in HCT116 and LoVo xenograft mouse models of colorectal cancer, compared with SM-1 or 5-FU treatment alone. SM-1 significantly enhanced the antitumor activity of 5-FU in colorectal cancer. These improved effects were due to increased activity of the apoptotic signaling pathway.

WX-132-18B, a novel microtubule inhibitor, exhibits promising anti-tumor effects.

Cancer drug researchers have been seeking microtubule-inhibiting agents (MIAs) with higher bioactivity and lower toxicity than currently marketed drugs. WX-132-18B, a novel structural compound synthesized at our institute, specifically bound to the colchicine-binding site on tubulin rather than the vinblastine site, and concentration-dependently reduced microtubule content via depolymerization. It exhibited the same cellular phenotypic profiles as the classic MIAs (colchicine, vincristine, and taxol), including inducing cell cycle arrest at the G2/M phase, triggering tumor cell apoptosis, promoting nuclear membrane permeability, reducing mitochondrial membrane potential, and disrupting the redox system balance. Importantly, WX-132-18B displayed more potent in vitro bioactivity (IC50 0.45-0.99 nM) than did the classic MIAs; it inhibited the proliferation of human umbilical vein endothelial cells and seven types of human tumor cells, especially the taxol-resistant breast cancer cells MX-1/T. WX-132-18B also dose-dependently inhibited mice sarcoma, human lung, and gastric cancer xenograft tumors and the formation of tumor blood vessels in mice. In conclusion, WX-132-18B is a novel microtubule-depolymerizing agent that selectively acts on the colchicine-binding site of tubulin and exerts potent in vitro and in vivo anti-tumor effects. These characteristics, along with its anti-angiogenesis and anti-drug resistance properties, make WX-132-18B a promising anti-tumor drug candidate.

Preclinical pharmacodynamic evaluation of antibiotic nitroxoline for anticancer drug repurposing.

The established urinary antibiotic nitroxoline has recently regained considerable attention, due to its potent activities in inhibiting angiogenesis, inducing apoptosis and blocking cancer cell invasion. These features make nitroxoline an excellent candidate for anticancer drug repurposing. To rapidly advance nitroxoline repurposing into clinical trials, the present study performed systemic preclinical pharmacodynamic evaluation of its anticancer activity, including a methyl thiazolyl tetrazolium assay in vitro and an orthotopic urological tumor assay in vivo. The current study determined that nitroxoline exhibits dose-dependent anti-cancer activity in vitro and in urological tumor orthotopic mouse models. In addition, it was demonstrated that the routine nitroxoline administration regimen used for urinary tract infections was effective and sufficient for urological cancer treatment, and 2 to 4-fold higher doses resulted in obvious enhancement of anticancer efficacy without corresponding increases in toxicity. Furthermore, nitroxoline sulfate, one of the most common metabolites of nitroxoline in the urine, effectively inhibited cancer cell proliferation. This finding increases the feasibility of nitroxoline repurposing for urological cancer treatment. Due to the excellent anticancer activity demonstrated in the present study, and its well-known safety profile and pharmacokinetic properties, nitroxoline has been approved to enter into a phase II clinical trial in China for non-muscle invasive bladder cancer treatment (registration no. CTR20131716).

Antitumor Effects of Orally and Intraperitoneally Administered Chitosan Oligosaccharides (COSs) on S180-Bearing/Residual Mouse.

Chitosan oligosaccharides (COSs) are hydrolysate mixture of chitin and possess various biomedical properties, such as antimicrobial, immunoenhancing, and antitumor effects. Antiproliferation activity of COS and commercially available samples was compared in the terms of A549 and HCT-116 cells. Ten tumor cells were used to estimate cytotoxicity of COS. Although there were some researches on the antitumor effects of COS, we highlighted the in vivo antitumor activities of COS administrated orally and intraperitoneally on S180-bearing/residual mouse. Results turned out that in vitro IC50 values of COS were 48.6 ± 7.0 to 1329.9 ± 93.4 µg/mL against 10 different tumor cell lines. Then, the in vivo experiments proved that the inhibition rate was high up to 58.5%. Significant cell death and necrosis were observed in COS-treated groups by histological analysis. COS stimulated the mRNA expression of tumor necrosis factor alpha. In summary, COS may be considered promising candidate as antitumor functional food or pharmaceutic adjuvant in oncotherapy, especially for patients after surgical resection.

Micheliolide derivative DMAMCL inhibits glioma cell growth in vitro and in vivo.

BACKGROUND: There is no highly effective chemotherapy for malignant gliomas to date. We found that dimethylaminomicheliolide (DMAMCL), a selective inhibitor of acute myeloid leukemia (AML) stem/progenitor cells, inhibited the growth of glioma cells. METHODS: The distribution of DMAMCL in brain was analyzed by an ultraperformance liquid chromatography-mass spectrometry (UPLC-MS/MS) system. The anti-tumor evaluations of DMAMCL in vitro were performed by MTT, FACS and RT-PCR. In vivo, the mixture of C6 cells and matrigel was injected into caudatum, and the anti-tumor activity of DMAMCL was evaluated by tumor growth and rat survival. The toxicity of DMAMCL was evaluated by body weight, daily food intake, hematological or serum biochemical analyses, and histological appearance of tissues. RESULTS: The IC50 values of DMAMCL against the C6 and U-87MG cell lines in vitro were 27.18 ± 1.89 µM and 20.58 ± 1.61 µM, respectively. DAMMCL down-regulated the anti-apoptosis gene Bcl-2 and increased apoptosis in C6 and U-87MG cells in a dose-dependent manner. In a C6 rat tumor model, daily administration of DMAMCL for 21 days reduced the burden of C6 tumors by 60% to 88% compared to controls, and more than doubled the mean lifespan of tumor-bearing rats. Distribution analysis showed that the DMAMCL concentration was higher in the brain than in plasma. Evaluations for toxicity revealed that oral administration of DMAMCL at 200 or 300 mg/kg once a day for 21 days did not result in toxicity. CONCLUSIONS: These results suggest that DMAMCL is highly promising for the treatment of glioma.

Fully human VEGFR2 monoclonal antibody BC001 attenuates tumor angiogenesis and inhibits tumor growth.

The critical role of VEGFR2 in tumor neovascularization and progression has allowed the design of clinically beneficial therapies based on it. Here we show that BC001, a new fully human anti-VEGFR2 monoclonal antibody, inhibits VEGF-stimulated endothelial cell migration, tube formation, and effectively suppressed the transdifferentiation of cancer stem cells into endothelial cells in vitro. Since BC001 exhibited no activity against the mouse VEGFR2 and mouse based study was required to confirm its efficacy in vivo, BC101, the mouse analogue of BC001, was developed. BC101 significantly attenuated angiogenesis according to Matrigel plug assay and resulted in ~80% growth inhibition of mouse B16F10 homograft tumors relative to vehicle control. Similarly, human analogue BC001 suppressed the growth of human xenograft tumors HCT116 and BGC823. Furthermore, immunohistochemical results showed reduced expression of CD31, VEGFR2 and Ki-67, as well as increased expression of Caspase 3 in BC001-treated tumor, which indicated BC001 was able to significantly decrease microvessel density, suppress proliferation and promote apoptosis. These results demonstrate the fully human VEGFR2 monoclonal antibody BC001 can work as an effective inhibitor of tumor angiogenesis and tumor growth both in vitro and in vivo.

Preparation, characterization, and pharmacodynamics of thermosensitive liposomes containing docetaxel.

A novel thermosensitive liposome (TL) containing docetaxel (DTX) was designed to enhance DTX-targeted delivery and antitumor effect. TL loading DTX (DTX-TL) were prepared by thin film hydration. The mean particle size of the liposomes was about 100 nm, and the drug entrapment efficiency was more than 95%. The phase transition temperature of liposomes was about 42 °C. In vitro drug release showed that drug released at 37 °C was obviously less than that at 42 °C. For in vivo experiments, the human breast tumor model was established by subcutaneous xenotransplantation on nude mice; liposomes and injection containing DTX were injected i.v. in nude mice, followed by exposure of the tumors to hyperthermia (HT) for 30 min after administration. The tumor inhibition ratio of DTX-TL group was significantly higher than the normal injection group. Combining TL with HT enhanced the delivery of DTX and thereby its antitumor effects. The liposomes reported in this paper could potentially produce viable clinical strategies for improved targeting and delivery of DTX for the treatment of breast cancer.

Optimization of 4-(N-cycloamino)phenylquinazolines as a novel class of tubulin-polymerization inhibitors targeting the colchicine site.

The 6-methoxy-1,2,3,4-tetrahydroquinoline moiety in prior leads 2-chloro- and 2-methyl-4-(6-methoxy-3,4-dihydroquinolin-1(2H)-yl)quinazoline (1a and 1b) was modified to produce 4-(N-cycloamino)quinazolines (4a-c and 5a-m). The new compounds were evaluated in cytotoxicity and tubulin inhibition assays, resulting in the discovery of new tubulin-polymerization inhibitors. 7-Methoxy-4-(2-methylquinazolin-4-yl)-3,4-dihydroquinoxalin- 2(1H)-one (5f), the most potent compound, exhibited high in vitro cytotoxic activity (GI50 1.9-3.2 nM), significant potency against tubulin assembly (IC50 0.77 µM), and substantial inhibition of colchicine binding (99% at 5 µM). In mechanism studies, 5f caused cell arrest in G2/M phase, disrupted microtubule formation, and competed mostly at the colchicine site on tubulin. Compound 5f and N-methylated analogue 5g were evaluated in nude mouse MCF7 xenograft models to validate their antitumor activity. Compound 5g displayed significant in vivo activity (tumor inhibitory rate 51%) at a dose of 4 mg/kg without obvious toxicity, whereas 5f unexpectedly resulted in toxicity and death at the same dose.

Preclinical evaluation of herpes simplex virus armed with granulocyte-macrophage colony-stimulating factor in pancreatic carcinoma.

AIM: To investigate the therapeutic efficacy and mechanisms of action of oncolytic-herpes-simplex-virus encoding granulocyte-macrophage colony-stimulating factor (HSV(GM-CSF)) in pancreatic carcinoma. METHODS: Tumor blocks were homogenized in a sterile grinder in saline. The homogenate was injected into the right armpit of each mouse. After vaccination, the mice were randomly assigned into four groups: a control group, a high dose HSV(GM-CSF) group [1 × 107 plaque forming units (pfu)/tumor], a medium dose HSV(GM-CSF) group (5 × 106 pfu/tumor) and a low dose HSV(GM-CSF) group (5 × 105 pfu/tumor). After initiation of drug administration, body weights and tumor diameters were measured every 3 d. Fifteen days later, after decapitation of the animal by cervical dislocation, each tumor was isolated, weighed and stored in 10% formaldehyde solution. The drug effectiveness was evaluated according to the weight, volume and relative volume change of each tumor. Furthermore, GM-CSF protein levels in serum were assayed by enzyme-linked immunosorbent assays at 1, 2, 3 and 4 d after injection of HSV(GM-CSF). RESULTS: Injection of the recombinant mouse HSV encoding GM-CSF resulted in a significant reduction in tumor growth compared to the control group, and dose-dependent effects were observed: the relative tumor proliferation rates of the low dose, medium dose and high dose groups on 15 d after injection were 45.5%, 55.2% and 65.5%, respectively. The inhibition rates of the tumor weights of the low, middle, and high dose groups were 41.4%, 46.7% and 50.5%, respectively. Furthermore, the production of GM-CSF was significantly increased in the mice infected with HSV(GM-CSF). The increase in the GM-CSF level was more pronounced in the high dose group compared to the other two dose groups. CONCLUSION: Our study provides the first evidence that HSV(GM-CSF) could inhibit the growth of pancreatic cancer. The enhanced GM-CSF expression might be responsible for the phenomenon.