Effect of cyanide ions (CN-) extracted from cassava (Manihotesculenta Crantz) on Alveolar Epithelial Cells (A549 cells).

Cassava (Manihotesculenta Crantz) is one of the most important root crops in tropical countries. It is a major source of cyanogenic glycosides viz. linamarin and lotaustralin, and these on breakdown liberate HCN and ketone. Cassava cyanide extract (CCE) from cassava leaves and tuber rinds were formulated as a biopesticide against certain borer insect pests of horticultural crops. Adenocarcinomic human alveolar basal epithelial cells (A549) were treated with three different concentrations (100, 200, 400 ppm) of CCE. The MTT and NRU assays showed dose-dependent cytotoxicity. The DCFH-DA assay does not show any free radical scavenging activity, whereas the NRR assay showed a reduction in the nitrile radicals with an increase in the concentration of the bioactive compound. A negative correlation was found between the concentration of the bioactive principles and mitochondrial and lysosomal functions. Various cellular assays demonstrated the cellular response of the CCE, and it was found that at higher concentration (400 ppm), the CCE exert a significant necrotic cell death rather than apoptosis. The results of the study indicated that the CCE have a remarkable tendency of anti-proliferative ability.

Downregulation of NKG2DLs by TGF-ß in human lung cancer cells.

BACKGROUND: Transforming growth factor beta (TGF-ß) is a typical immuno-inhibitory cytokine and highly secreted by lung cancer cells. It was supposed that its immunosuppressive effects to NK cell might be related with the altered expression of activating and inhibitory molecules in lung cancer cells. In this study, we examined the expression of NKG2DLs, PD-L1 and PD-L2 in lung cancer cells after treatment of TGF-ß and a TGF-ß inhibitor, Galunisertib (LY2157299). RESULTS: TGF-ß reduced the level of surface proteins of five NKG2DLs without altered transcription levels in lung cancer cells. Galunisertib reversed the effect of TGF-ß on the expression of NKG2DLs. Since MMP inhibitors, MMPi III and MMP2 inhibitor I, restored the reduced expression of NKG2DLs after treatment of TGF-ß, it was thought that TGF-ß induced the expression of MMP2 which facilitated the shedding of the NKG2DLs in cancer cells. However, the expression of PD-L1, L2 were not changed by treatment with TGF-ß or Galunisertib. CONCLUSIONS: Therefore, inhibition of TGF-ß might reverse the immunosuppressive status on immune cells and restore NK cell mediated anticancer immune responses by upregulation of NKG2DLs in cancer cells.

One- and Two-Photon Phototherapeutic Effects of RuII Polypyridine Complexes in the Hypoxic Centre of Large Multicellular Tumor Spheroids and Tumor-Bearing Mice*.

During the last decades, photodynamic therapy (PDT), an approved medical technique, has received increasing attention to treat certain types of cancer. Despite recent improvements, the treatment of large tumors remains a major clinical challenge due to the low ability of the photosensitizer (PS) to penetrate a 3D cellular architecture and the low oxygen concentrations present in the tumor center. To mimic the conditions found in clinical tumors, exceptionally large 3D multicellular tumor spheroids (MCTSs) with a diameter of 800 µm were used in this work to test a series of new RuII polypyridine complexes as one-photon and two-photon PSs. These metal complexes were found to fully penetrate the 3D cellular architecture and to generate singlet oxygen in the hypoxic center upon light irradiation. While having no observed dark toxicity, the lead compound of this study showed an impressive phototoxicity upon clinically relevant one-photon (595 nm) or two-photon (800 nm) excitation with a full eradication of the hypoxic center of the MCTSs. Importantly, this efficacy was also demonstrated on mice bearing an adenocarcinomic human alveolar basal epithelial tumor.

Hyaluronan decoration of milk exosomes directs tumor-specific delivery of doxorubicin.

Milk exosomes (mExo), similar to cell-derived exosomes, are emerging as promising nanocarriers for delivery of therapeutic molecules such as chemical drugs and siRNA, due to the excellent biocompatibility and low-cost production from bovine milk. However, additional modification remains required to apply milk exosomes for tumor-specific drug delivery. Here, we attempted to develop a novel strategy for directing doxorubicin (Dox)-loaded mExo to CD44-overexpressing tumor cells. Hyaluronan (HA), a CD44-specific ligand, was functionalized with an amphiphilic molecule DSPE-PEG2000, which enabled the spontaneous decoration of Dox-loaded mExo with HA onto the phospholipid bilayer. The obtained nanocarrier HA-mExo-Dox was shown to be able to selectively deliver Dox into cells with over-expressed CD44 instead of control cells and trigger the notable tumor cells death in the in vitro analysis. This study demonstrates the potential use of HA-displaying mExo for tumor cell-specific drug delivery and this strategy should prove to be feasible for targeted cancer therapy.

Tumor-targeting multi-shelled hollow nanospheres as drug loading platforms for imaging-guided combinational cancer therapy.

In this work, we developed multi-shelled hollow nanospheres [RGD@am-ZnO@CuO@Au@DOX HNSs] as multifunctional therapeutic agents to achieve effective and targeted Zn2+/Cu2+ therapy, induced drug delivery under low pH/red-light conditions, and enhanced phototherapy under single red-light. The photothermal and photodynamic performance of am-ZnO@CuO@Au HNSs was enhanced relative to that of am-ZnO nanoparticles (NPs) or am-ZnO@CuO HNSs by utilizing the resonance energy transfer process and broad red-light absorption. The pH-sensitive am-ZnO@CuO@Au HNSs were dissolved to Zn2+/Cu2+ in the acidic endosomes/lysosomes of cancer cells, resulting in a cancer cell killing effect. The release performance of doxorubicin (DOX) from RGD@am-ZnO@CuO@Au@DOX HNSs was evaluated under low pH and red-light-irradiated conditions, and targeting of HNSs was confirmed by dual-modal imaging (magnetic resonance/fluorescence) of the tumor area. Moreover, in vivo synergistic therapy using RGD@am-ZnO@CuO@Au@DOX HNSs was further evaluated in mice bearing human pulmonary adenocarcinoma (A549) cells, achieving a remarkable synergistic antitumor effect superior to that obtained by monotherapy. This study validated that RGD@am-ZnO@CuO@Au@DOX HNSs can be a promising candidate for efficient postoperative cancer therapy.

Metformin attenuates adhesion between cancer and endothelial cells in chronic hyperglycemia by recovery of the endothelial glycocalyx barrier.

BACKGROUND: Epidemiologic studies suggest that diabetes is associated with an increased risk of cancer. Concurrently, clinical trials have shown that metformin, which is a first-line antidiabetic drug, displays anticancer activity. The underlying mechanisms for these effects are, however, still not well recognized. METHODS: Methods based on atomic force microscopy (AFM) were used to directly evaluate the influence of metformin on the nanomechanical and adhesive properties of endothelial and cancer cells in chronic hyperglycemia. AFM single-cell force spectroscopy (SCFS) was used to measure the total adhesion force and the work of detachment between EA.hy926 endothelial cells and A549 lung carcinoma cells. Nanoindentation with a spherical AFM probe provided information about the nanomechanical properties of cells, particularly the length and grafting density of the glycocalyx layer. Fluorescence imaging was used for glycocalyx visualization and monitoring of E-selectin and ICAM-1 expression. RESULTS: SCFS demonstrated that metformin attenuates adhesive interactions between EA.hy926 endothelial cells and A549 lung carcinoma cells in chronic hyperglycemia. Nanoindentation experiments, confirmed by confocal microscopy imaging, revealed metformin-induced recovery of endothelial glycocalyx length and density. The recovery of endothelial glycocalyx was correlated with a decrease in the surface expression of E-selectin and ICAM-1. CONCLUSION: Our results identify metformin-induced endothelial glycocalyx restoration as a key factor responsible for the attenuation of adhesion between EA.hy926 endothelial cells and A549 lung carcinoma cells. GENERAL SIGNIFICANCE: Metformin-induced glycocalyx restoration and the resulting attenuation of adhesive interactions between the endothelium and cancer cells may account for the antimetastatic properties of this drug.

Rational design of some substituted phenyl azanediyl (bis) methylene phosphonic acid derivatives as potential anticancer agents and imaging probes: Computational inputs, chemical synthesis, radiolabeling, biodistribution and gamma scintigraphy.

Bisphosphonates are widely used for treatment of osteoporosis. Recently, they have been reported to be effective anticancer agents. In this work, we designed some substituted phenyl (azanediyl) bis (methylene phosphonic acid) to be tested for their anticancer effect. Both molecular docking and dynamics studies were used to select the top ranked highly scored compounds. The selected hits showed potential in vitro anticancer effect against some cell lines. Biodistribution pattern and gamma scintigraphy were conducted to the most effective derivative (BMBP) after radiolabeling with 99mTc. Results of biodistribution and scintigraphic imaging of 99mTc-BMBP in tumor bearing mice showed a notable tumor affinity, and confirmed the targeting affinity of BMBP to the tumor tissues. As a conclusion, BMBP could act as potential anticancer agent and imaging probe.

Targeted Modification of the Cationic Anticancer Peptide HPRP-A1 with iRGD To Improve Specificity, Penetration, and Tumor-Tissue Accumulation.

The chimeric peptide HPRP-A1-iRGD, composed of a chemically conjugated tumor-homing/penetration domain (iRGD) and a cationic anticancer peptide domain (HPRP-A1), was used to study the effect of targeted modification to enhance the peptide's specificity, penetration, and tumor accumulation ability. The iRGD domain exhibits tumor-targeting and tumor-penetrating activities by specifically binding to the neuropilin-1 receptor. Acting as a homing/penetration domain, iRGD contributed to enhancing the tumor selectivity, permeability, and targeting of HPRP-A1 by targeted receptor dependence. As the anticancer active domain, HPRP-A1 kills cancer cells by disrupting the cell membrane and inducing apoptosis. The in vitro membrane selectivity toward cancer cells, such as A549 and MDA-MB-23, and human umbilical vein endothelial cells (HUVECs), normal cells, the penetrability assessment in the A549 3D multiple cell sphere model, and the in vivo tumor-tissue accumulation test in the A549 xenograft model indicated that HPRP-A1-iRGD exhibited significant increases in the selectivity toward membranes that highly express NRP-1, the penetration distance in 3D multiple cell spheres, and the accumulation in tumor tissues after intravenous injection, compared with HPRP-A1 alone. The mechanism of the enhanced targeting ability of HPRP-A1-iRGD was demonstrated by the pull-down assay and biolayer interferometry test, which indicated that the chimeric peptide could specifically bind to the neuropilin-1 protein with high affinity. We believe that chemical conjugation with iRGD to increase the specificity, penetration, and tumor-tissue accumulation of HPRP-A1 is an effective and promising approach for the targeted modification of peptides as anticancer therapeutics.

Assessment of in vitro particle dosimetry models at the single cell and particle level by scanning electron microscopy.

BACKGROUND: Particokinetic models are important to predict the effective cellular dose, which is key to understanding the interactions of particles with biological systems. For the reliable establishment of dose-response curves in, e.g., the field of pharmacology and toxicology, mostly the In vitro Sedimentation, Diffusion and Dosimetry (ISDD) and Distorted Grid (DG) models have been employed. Here, we used high resolution scanning electron microscopy to quantify deposited numbers of particles on cellular and intercellular surfaces and compare experimental findings with results predicted by the ISDD and DG models. RESULTS: Exposure of human lung epithelial A549 cells to various concentrations of differently sized silica particles (100, 200 and 500 nm) revealed a remarkably higher dose deposited on intercellular regions compared to cellular surfaces. The ISDD and DG models correctly predicted the areal densities of particles in the intercellular space when a high adsorption ("stickiness") to the surface was emulated. In contrast, the lower dose on cells was accurately inferred by the DG model in the case of "non-sticky" boundary conditions. Finally, the presence of cells seemed to enhance particle deposition, as aerial densities on cell-free substrates were clearly reduced. CONCLUSIONS: Our results further validate the use of particokinetic models but also demonstrate their limitations, specifically, with respect to the spatial distribution of particles on heterogeneous surfaces. Consideration of surface properties with respect to adhesion and desorption should advance modelling approaches to ultimately predict the cellular dose with higher precision.

Crystal Structures and Cytotoxicity of ent-Kaurane-Type Diterpenoids from Two Aspilia Species.

A phytochemical investigation of the roots of Aspilia plurisetaled to the isolation of ent-kaurane-type diterpenoids and additional phytochemicals (1⁻23). The structures of the isolated compounds were elucidated based on Nuclear Magnetic Resonance (NMR) spectroscopic and mass spectrometric analyses. The absolute configurations of seven of the ent-kaurane-type diterpenoids (3⁻6, 6b, 7 and 8) were determined by single crystal X-ray diffraction studies. Eleven of the compounds were also isolated from the roots and the aerial parts of Aspilia mossambicensis. The literature NMR assignments for compounds 1 and 5 were revised. In a cytotoxicity assay, 12α-methoxy-ent-kaur-9(11),16-dien-19-oic acid (1) (IC50 = 27.3 ± 1.9 µM) and 9ß-hydroxy-15α-angeloyloxy-ent-kaur-16-en-19-oic acid (3) (IC50 = 24.7 ± 2.8 µM) were the most cytotoxic against the hepatocellular carcinoma (Hep-G2) cell line, while 15α-angeloyloxy-16ß,17-epoxy-ent-kauran-19-oic acid (5) (IC50 = 30.7 ± 1.7 µM) was the most cytotoxic against adenocarcinomic human alveolar basal epithelial (A549) cells.

Novel dihydroartemisinin derivative DHA-37 induces autophagic cell death through upregulation of HMGB1 in A549 cells.

Dihydroartemisinin (DHA) and its analogs are reported to possess selective anticancer activity. Here, we reported a novel DHA derivative DHA-37 that exhibited more potent anticancer activity on the cells tested. Distinct from DHA-induced apoptosis, DHA-37 triggered excessive autophagic cell death, and became the main contributor to DHA-37-induced A549 cell death. Incubation of the cells with DHA-37 but not DHA produced increased dots distribution of GFP-LC3 and expression ratio of LC3-II/LC3-I, and enhanced the formation of autophagic vacuoles as revealed by TEM. Treatment with the autophagy inhibitor 3-MA, LY294002, or chloroquine could reverse DHA-37-induced cell death. In addition, DHA-37-induced cell death was associated significantly with the increased expression of HMGB1, and knockdown of HMGB1 could reverse DHA-37-induced cell death. More importantly, the elevated HMGB1 expression induced autophagy through the activation of the MAPK signal but not PI3K-AKT-mTOR pathway. In addition, DHA-37 also showed a wonderful performance in A549 xenograft mice model. These findings suggest that HMGB1 as a target candidate for apoptosis-resistant cancer treatment and artemisinin-based drugs could be used in inducing autophagic cell death.

Turning Toxicants into Safe Therapeutic Drugs: Cytolytic Peptide-Photosensitizer Assemblies for Optimized In Vivo Delivery of Melittin.

Melittin (MEL) is recognized as a highly potent therapeutic peptide for treating various human diseases including cancer. However, the clinical applications of MEL are largely restricted by its severe hemolytic activity and nonspecific cytotoxicity. Here, it is found that MEL can form a stable supramolecular nanocomplex of ≈60 nm with the photosensitizer chlorin e6 (Ce6), which after hyaluronic acid (HA) coating can achieve robust, safe, and imaging-guided tumor ablation. The as-designed nanocomplex (denoted as MEL/Ce6@HA) shows largely reduced hemolysis and selective cytolytic activity toward cancer cells. Upon laser irradiation, the loaded photosensitive Ce6 can synergistically facilitate the membrane-lytic efficiency of melittin and greatly increase the tumor penetration depth of the complexes in multicellular tumor spheroids. In vivo experiments reveal that MEL/Ce6@HA can realize enhanced tumor accumulation, reduced liver deposition, and rapid body clearance, which are beneficial for highly efficient and safe chemo-photodynamic dual therapy. This work develops a unique supramolecular strategy for optimized in vivo delivery of melittin and may have implications for the development of peptide-based theranostics.

Organometallic Gold(III) Complexes Similar to Tetrahydroisoquinoline Induce ER-Stress-Mediated Apoptosis and Pro-Death Autophagy in A549 Cancer Cells.

Agents inducing both apoptosis and autophagic death can be effective chemotherapeutic drugs. In our present work, we synthesized two organometallic gold(III) complexes harboring C^N ligands that structurally resemble tetrahydroisoquinoline (THIQ): Cyc-Au-1 (AuL1Cl2, L1 = 3,4-dimethoxyphenethylamine) and Cyc-Au-2 (AuL2Cl2, L2 = methylenedioxyphenethylamine). In screening their in vitro activity, we found both gold complexes exhibited lower toxicity, lower resistance factors, and better anticancer activity than those of cisplatin. The organometallic gold(III) complexes accumulate in mitochondria and induce elevated ROS and an ER stress response through mitochondrial dysfunction. These effects ultimately result in simultaneous apoptosis and autophagy. Importantly, compared to cisplatin, Cyc-Au-2 exhibits lower toxicity and better anticancer activity in a murine tumor model. To the best of our knowledge, Cyc-Au-2 is the first organometallic Au(III) compound that induces apoptosis and autophagic death. On the basis of our results, we believe Cyc-Au-2 to be a promising anticancer agent or lead compound for further anticancer drug development.

Co-administration of iRGD with peptide HPRP-A1 to improve anticancer activity and membrane penetrability.

To improve the specificity and penetration of anticancer peptides against tumors, in this study, we examined the effects of co-administration of the membrane-active peptide HPRP-A1 and the tumor homing/penetrating peptide iRGD. iRGD peptide is widely recognized as an efficient cell membrane penetration peptide targeting to αvß3 integrins and neuropilin-1 (NRP-1) receptors, which show high expression in many tumor cells. The anticancer activity, cancer specificity and penetration activity in vitro and in vivo of the co-administered peptides were examined on 2D monolayer cells, 3D multi-cellular spheroids (MCS) and xenograft nude mice. Co-administration of iRGD and HPRP-A1 exhibited stronger anticancer activity and tumor specificity against A549 non-small cell lung cancer cells with NRP-1 receptor overexpression compared with HPRP-A1 alone. A549 cells showed uptake of the peptide combination and destruction of the integrity of the cell membrane, as well as adherence to the mitochondrial net, resulting in induction of apoptosis by a caspase-dependent pathway. The iRGD peptide dramatically increased the penetration depth of HPRP-A1 on A549 MCS and anticancer efficacy in an A549 xenograft mouse model. Our results suggest that the co-administration strategy of anticancer and penetrating peptides could be a potential therapeutic approach for cancer treatment in clinical practice.

Paired Phase II Studies of Erlotinib/Bevacizumab for Advanced Bronchioloalveolar Carcinoma or Never Smokers With Advanced Non-Small-cell Lung Cancer: SWOG S0635 and S0636 Trials.

BACKGROUND: Before mutation testing of the epidermal growth factor receptor (EGFR) gene was recognized as highly associated with the activity of EGFR tyrosine kinase inhibitors (TKIs), clinically defined patient populations with bronchioloalveolar carcinoma (BAC) and never smokers were identified as likely to benefit from EGFR TKIs. From preclinical and clinical data suggesting potentially improved efficacy with a combination of an EGFR TKI and the antiangiogenic agent bevacizumab, the Southwestern Oncology Group (SWOG) initiated paired phase II trials to evaluate the combination of erlotinib/bevacizumab in patients with advanced BAC (SWOG S0635) or never smokers with advanced lung adenocarcinoma (SWOG S0636). MATERIALS AND METHODS: Eligible patients with BAC or adenocarcinoma with BAC features (SWOG S0635) or never smokers with advanced lung adenocarcinoma (SWOG S0636) received erlotinib 150 mg/day with bevacizumab 15 mg/kg until progression or prohibitive toxicity. Never smokers with BAC were preferentially enrolled to SWOG S0636. The primary endpoint for both trials was overall survival. RESULTS: A total of 84 patients were enrolled in the SWOG S0635 trial and 85 in the SWOG S0636 trial. The objective response rate was 22% (3% complete response) in the SWOG S0635 trial and 50% (38% confirmed; 3% complete response) in the SWOG S0636 trial. The median progression-free survival was 5 and 7.4 months in the S0635 and S0636 trials, respectively. The median overall survival was 21 and 29.8 months, respectively. Toxicity consisted mainly of rash and diarrhea in both trials. CONCLUSION: Although the field has moved toward molecular, rather than clinical, selection of patients as optimal candidates for EGFR TKI therapy, these results support the hypothesis that a subset of patients in whom erlotinib is particularly active could receive an incremental benefit from the addition of bevacizumab.

Celecoxib normalizes the tumor microenvironment and enhances small nanotherapeutics delivery to A549 tumors in nude mice.

Barriers presented by the tumor microenvironment including the abnormal tumor vasculature and interstitial matrix invariably lead to heterogeneous distribution of nanotherapeutics. Inspired by the close association between cyclooxygenase-2 (COX-2) and tumor-associated angiogenesis, as well as tumor matrix formation, we proposed that tumor microenvironment normalization by COX-2 inhibitors might improve the distribution and efficacy of nanotherapeutics for solid tumors. The present study represents the first time that celecoxib, a special COX-2 inhibitor widely used in clinics, was explored to normalize the tumor microenvironment and to improve tumor nanotherapeutics delivery using a human-derived A549 tumor xenograft as the solid tumor model. Immunofluorescence staining of tumor slices demonstrated that oral celecoxib treatment at a dose of 200 mg/kg for two weeks successfully normalized the tumor microenvironment, including tumor-associated fibroblast reduction, fibronectin bundle disruption, tumor vessel normalization, and tumor perfusion improvement. Furthermore, it also significantly enhanced the in vivo accumulation and deep penetration of 22-nm micelles rather than 100-nm nanoparticles in tumor tissues by in vivo imaging and distribution experiments and improved the therapeutic efficacy of paclitaxel-loaded micelles in tumor xenograft-bearing mouse models in the pharmacodynamics experiment. As celecoxib is widely and safely used in clinics, our findings may have great potential in clinics to improve solid tumor treatment.

Polyphenolic Polymersomes of Temperature-Sensitive Poly(N-vinylcaprolactam)-block-Poly(N-vinylpyrrolidone) for Anticancer Therapy.

We report a versatile synthesis for polyphenolic polymersomes of controlled submicron (<500 nm) size for intracellular delivery of high and low molecular weight compounds. The nanoparticles are synthesized by stabilizing the vesicular morphology of thermally responsive poly(N-vinylcaprolactam)n-b-poly(N-vinylpyrrolidone)m (PVCLn-PVPONm) diblock copolymers with tannic acid (TA), a hydrolyzable polyphenol, via hydrogen bonding at a temperature above the copolymer's lower critical solution temperature (LCST). The PVCL179-PVPONm diblock copolymers are produced by controlled reversible addition-fragmentation chain transfer (RAFT) polymerization of PVPON using PVCL as a macro-chain transfer agent. The size of the TA-locked (PVCL179-PVPONm) polymersomes at room temperature and upon temperature variations are controlled by the PVPON chain length and TA:PVPON molar unit ratio. The particle diameter decreases from 1000 to 950, 770, and 250 nm with increasing PVPON chain length (m = 107, 166, 205, 234), and it further decreases to 710, 460, 290, and 190 nm, respectively, upon hydrogen bonding with TA at 50 °C. Lowering the solution temperature to 25 °C results in a slight size increase for vesicles with longer PVPON. We also show that TA-locked polymersomes can encapsulate and store the anticancer drug doxorubicin (DOX) and higher molecular weight fluorescein isothiocyanate (FITC)-dextran in a physiologically relevant pH and temperature range. Encapsulated DOX is released in the nuclei of human alveolar adenocarcinoma tumor cells after 6 h incubation via biodegradation of the TA shell with the cytotoxicity of DOX-loaded polymersomes being concentration-dependent. Our approach offers biocompatible and intracellular degradable nanovesicles of controllable size for delivery of a variety of encapsulated materials. Considering the particle monodispersity, high loading capacity, and a facile two-step aqueous assembly based on the reversible temperature-responsiveness of PVCL, these polymeric vesicles have significant potential as novel drug nanocarriers and provide a new perspective for fundamental studies on thermo-triggered polymer assemblies in solutions.

Trichopeptides A and B, trichocyclodipeptides A-C, new peptides from the ascomycete fungus Stagonospora trichophoricola.

Trichopeptides A (1) and B (2), new linear tetrapeptide and tripeptide, respectively, and three new diketopiperazines trichocyclodipeptides A-C (3-5) were isolated from the fermentation of the ascomycete fungus Stagonospora trichophoricola, a fungus isolated from the soil sample surrounding the fruiting body of Ophiocordyceps sinensis in Maqin Country, Qinghai Province, People's Republic of China. Their structures were primarily elucidated by interpretation of NMR and MS experiments. The absolute configurations of 1-5 were assigned through Marfey's method on their acid hydrolyzates. Compound 3 showed antifungal activity against Candida albicans with the IC50 and MIC values of 22 and 90 µg ml-1, respectively.

Jolkinolide A and Jolkinolide B Inhibit Proliferation of A549 Cells and Activity of Human Umbilical Vein Endothelial Cells.

BACKGROUND Jolkinolide A (JA) and Jolkinolide B (JB) are diterpenoids extracted from the roots of Euphorbia fischeriana Steud and have been shown to have anti-tumor activity. However, their effects on the ability of tumor cells to invade blood vessels and metastasize remain largely unknown. Investigations into the effects of JA and JB on the angiogenesis of tumor tissues may facilitate the identification of new natural drugs with anti-tumor growth and metastasis activities. MATERIAL AND METHODS We used different concentrations of JA and JB (20 µg/ml, 40 µg/ml, 60 µg/ml, 80 µg/ml, and 100 µg/ml) to stimulate A549 cells and then studied the effects on the growth and metastasis of lung cancers. In addition, we used conditional media from A549 cells (A549-CM) stimulated by either JA or JB in different concentrations to culture human umbilical vein endothelial cells (HUVECs). RESULTS We found that both JA and JB significantly inhibited the Akt-STAT3-mTOR signaling pathway and reduced the expression of VEGF in A549 cells, but JB exhibited more significant inhibitory effects than JA. The JB-stimulated A549 cell conditional media had a greater inhibitory effect on the proliferation and migration of HUVECs than did the conditional media of JA-stimulated A549 cells. This effect gradually increased with increasing concentrations of either type of Jolkinolide. CONCLUSIONS Our results suggest that JA and JB inhibited VEGF expression in A549 cells through the inhibition of the Akt-STAT3-mTOR signaling pathway, and directly inhibited the proliferation and migration of HUVECs. These findings are of great significance for the development of new plant-derived chemotherapy agents for the treatment of cancer.

Phase I Results from a Study of Crizotinib in Combination with Erlotinib in Patients with Advanced Nonsquamous Non-Small Cell Lung Cancer.

INTRODUCTION: This phase I trial was conducted to determine the safety, maximum tolerated dose (MTD)/recommended phase II dose, and efficacy of crizotinib plus erlotinib in patients with advanced NSCLC. METHODS: Patients with NSCLC and an Eastern Cooperative Oncology Group performance status of 0 to 2 after failure of one or two prior chemotherapy regimens were eligible. Erlotinib, 100 mg, was given continuously once daily starting between day -14 and -7; crizotinib, 200 mg twice daily (dose level 1) or 150 mg twice daily (dose level -1), was added continuously beginning on day 1 of treatment cycle 1. Potential pharmacokinetic interactions between crizotinib and erlotinib were evaluated. RESULTS: Twenty-seven patients received treatment; 26 received crizotinib plus erlotinib. Frequent adverse events were diarrhea, rash, decreased appetite, and fatigue. Dose-limiting toxicities were dehydration, diarrhea, dry eye, dysphagia, dyspepsia, esophagitis and vomiting. The MTD was crizotinib, 150 mg twice daily, with erlotinib, 100 mg once daily. Crizotinib increased the erlotinib area under the concentration-time curve 1.5-fold (dose level -1) and 1.8-fold (dose level 1). The plasma level of crizotinib appeared to be unaffected by coadministration of erlotinib. Two patients whose tumors harbored activating EGFR mutations achieved confirmed partial responses, one at each crizotinib dose level. CONCLUSIONS: The MTD of the combination of crizotinib and erlotinib in patients with advanced NSCLC was crizotinib, 150 mg twice daily, with erlotinib, 100 mg once daily, which is less than the approved dose of either agent. The phase II portion of the study was not initiated.