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Invasion is a complex process controlled by secretion and activation of proteases, alteration of integrin levels and GSL (glycosphingolipid) patterns. Differential organization of GSLs with specific membrane proteins and signal transducers in GEMs (GSL-enriched microdomains), initiates signalling events to modify cellular phenotype. Although the GSL monosialyl-Gb5 has been linked with invasion, its functional role in invasion is poorly described and understood. To investigate this problem, we induced the invasion of human breast cancer cells and subsequently explored the underlying mechanism. In the present study, the invasion of human MCF-7 breast cancer cells is highly dependent on clustering of monosialyl-Gb5, and the subsequent activation of monosialyl-Gb5-associated focal adhesion kinase and cSrc in GEM leading to the downstream activation of extracellular-signal-regulated kinase (ERK). As a result, we observed increased expression levels and activity of matrix metalloproteinases-2 and -9, which correlated with decreased expression of integrins α1 and β1. Together these results suggest that the organization of crucial molecules in GEMs of MCF-7 cells is critical for their invasive properties.

Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase known to promote cell migration and invasiveness. Overexpression and increased activity of FAK are closely associated with metastatic breast tumors and are linked to poor prognosis. This study discovered an inverse correlation between FAK activity and migratory and invasive behavior. We show decreased phosphorylation levels of FAK at tyrosine residues 397 and 861, and most prominently at Y407, in the more invasive Hs578Ts(i)8 subclone of the Hs578T breast cancer progression model. There is limited information available on FAK Y407, and here we demonstrate its presence in triple-negative breast cancer (TNBC) cell lines. Furthermore, our studies propose that localization of FAK Y407, rather than FAK expression and overall FAK Y407 phosphorylation levels, is crucial for the control of cell motility. FAK Y407 is found extensively at the cell periphery in focal adhesion-like structures at each end of actin stress fibers and organized with integrin αVβ5 receptors, linking the αVβ5 integrin-mediated migratory behavior of Hs578Ts(i)8 cells to FAK Y407. These data suggest that subcellular localization, next to expression and activity levels, are important for understanding TNBC progression. Such an approach opens new avenues for further studies and may provide novel insight for the classification of TNBC and facilitate the discovery of effective biomarkers for diagnosis and therapy of TNBC.

Two human mammary carcinoma cell variants, MCF-7/AZ and MCF-7/6, show the same composition in their glycosphingolipid-enriched microdomain (GEM) with regard to globo-series structures Gb3, Gb4, Gb5, monosialyl-Gb5, GM2, and cSrc and FAK. Both variants are non-invasive into collagen gel layer, and showed similar motility in wound migration assay. Whereas invasiveness and motility of MCF-7/AZ cells were enhanced greatly by treatment with mAb RM1 directed to monosialyl-Gb5, the same RM1 treatment had no effect on MCF-7/6. cSrc and FAK of MCF-7/AZ, but not MCF-7/6, were activated by RM1 treatment. Thus, malignancy of MCF-7 is highly dependent on monosialyl-Gb5, and its activation of cSrc and FAK in GEM.

Complex interplays among proteins, lipids and carbohydrates can alter the phenotype and are suggested to have a crucial role in tumour metastasis. Our previous studies indicated that a complex of the GSLs (glycosphingolipids), AsGM1 (asialo-GM1), which lacks α2,3-linked sialic acid, and α2β1 integrin receptors is responsible for the metastatic behaviour of C4-2B prostate cancer cells. Herein, we identified and addressed the functional significance of changes in sialylation during prostate cancer progression. We observed an increase in α2,3-linked sialic acid residues on α2 subunits of α2β1 integrin receptors, correlating with increased gene expression of α2,3-STs (sialyltransferases), particularly ST3GAL3. Cell surface α2,3-sialylation of α2 subunits was required for the integrin α2β1-dependent cell adhesion to collagen type I and the same α2,3-linked sialic acid residues on the integrin receptor were responsible for the interaction with the carbohydrate moiety of AsGM1, explaining the complex formation between AsGM1 and α2β1 integrin receptors. These results provide novel insights into the role of sialic acids in the organization and function of important membrane components in invasion and metastatic processes.

1-O-octadecyl-2-O-methyl-glycerophosphocholine (ET-18-OMe) is an analogue of the naturally occurring 2-lysophosphatidylcholine belonging to the class of antitumor lipids. Previously, we demonstrated that ET-18-OMe modulates cell-cell adhesion of human breast cancer MCF-7 cells. In the present study, we tested the effect of ET-18-OMe on adhesion, invasion and localisation of episialin and E-cadherin in MCF-7/AZ cells expressing a functional E-cadherin/catenin complex. The MCF-7/6 human breast cancer cells were used as negative control since their E-cadherin/catenin complex is functional in cells grown on solid substrate but not in suspension. The function of E-cadherin, a calcium-dependent transmembrane cell-cell adhesion and signal-transducing molecule, is disturbed in invasive cancers by mutation, loss of mRNA stability, proteolytic degradation, tyrosine phosphorylation of associated proteins and large cell-associated proteoglycans or mucin-like molecules such as episialin. Episialin, also called MUC1, is an anti-adhesion molecule that by its large number of glycosylated tandem repeats can sterically hinder the adhesive properties of other glycoproteins. ET-18-OMe inhibited the E-cadherin functions of MCF-7/AZ cells as measured by inhibition of fast and slow aggregation and by the induction of collagen invasion. These effects were enhanced by MB2, an antibody against E-cadherin and blocked by monoclonal antibodies (MAbs) 214D4 or M8 against episialin. ET-18-OMe had no influence on tyrosine phosphorylation of beta-catenin and the E-cadherin/catenin complex remained intact. Transcription, translation, protein turnover and cell surface localisation of episialin were not altered. ET-18-OMe induced finger-like extensions with clustering of episialin together with E-cadherin and carcinoembryonic antigen but not with occludin. In cells in suspension, ET-18-OMe caused a shift in the flow-cytometric profile of episialin toward a lower intensity for MCF-7/AZ cells. In contrast with MCF-7/AZ cells, the adhesion-deficient and noninvasive MCF-7/6 cells showed neither morphotypic changes nor induction of aggregation nor invasion in collagen I upon treatment with ET-18-OMe. Co-localisation of episialin with E-cadherin was rarely observed. We conclude that in the human breast cancer cells MCF-7/AZ, E-cadherin and episialin are key molecular players in the regulation of promotion and suppression of cell-cell adhesion and invasion.

A series of ethacrynic acid analogues, lacking the alpha,beta-unsaturated carbonyl unit, was synthesized and subsequently evaluated for their ability to inhibit the migration of human breast cancer cells, MCF-7/AZ. Several of the analogues were already active in the low micromolar range, whereas ethacrynic acid itself shows no potential to inhibit the migration of these cancer cells. Preliminary studies show that the presence of one or more methoxy groups at the phenyl ring of ethacrynic acid is important in order for the ethacrynic acid analogues to demonstrate an inhibitory effect on the migration.

On the basis of the detection of expressed sequence tag ('EST') similar to the rat N-acetylgalactosamine alpha2,6-sialyltransferase (ST6GalNAc) III cDNA, we have identified a novel member of the human ST6GalNAc family. We have isolated a cDNA clone containing an open reading frame that codes for a type II membrane protein of 302 amino acids with a seven-amino-acid cytoplasmic domain, an 18-amino-acid transmembrane domain and the smallest described catalytic domain of 277 amino acids. This predicted sialyltransferase sequence is similar to the rat ST6GalNAc III (46.6%), but was found to be even more similar to the recently reported mouse ST6GalNAc IV (88.1%) on the basis of amino acid sequence identity. Northern-blot analysis showed that the newly identified gene is expressed constitutively in various adult human tissues as a 2.2kb transcript, but was also found to be expressed at lower levels in brain, heart and skeletal muscle as a 2.5kb transcript. Expression of the hST6GalNAc IV gene was investigated by reverse transcription PCR in various human cancer cells, and was found to be present in the majority of cell types with the exception of the carcinoma cell line T47D and pro-monocyte THP cells. The transient expression in COS-7 cells of the full-length cDNA led to the production of an active enzyme sharing the acceptor specificity of the ST6GalNAc family towards Neu5Ac alpha 2-3Gal beta 1-3GalNAc alpha-O-R (where 'R' denotes H, benzyl, or a peptidic chain). Detailed analysis in vitro of substrate specificity revealed that the enzyme required the trisaccharide Neu5Ac alpha 2-3Gal beta1-3GalNAc found on O-glycans and arylglycosides. In addition, we have clarified the genomic organization of ST6GalNAc IV gene.

A cDNA clone encoding a human Galbeta1-3GalNAc alpha2, 6-sialyltransferase (designated hST6GalNAc II) was identified employing the PCR with degenerated primers to the sialylmotifs, followed by BLAST analysis of databanks. This sialyltransferase sequence is similar to that of previously cloned ST6GalNAc II (chicken and mouse) and shows the sialylmotifs that are present in all eukaryotic members of the sialyltransferase gene family. The predicted amino acid sequence encodes a putative type II transmembrane protein as found for other eukaryotic sialyltransferases and shows significant similarity to chicken (56. 8% identity) and mouse (74.6% identity) enzymes. Expression of a secreted form of hST6GalNAc II in COS-7 cells showed that the gene product had Galbeta1-3GalNAc (sialyl to GalNAc) alpha2, 6-sialyltransferase activity. In vitro analysis of substrate specificity revealed that the enzyme required a peptide aglycone fraction to be active and used both Galbeta1-3GalNAc and Neu5Acalpha2-3Galbeta1-3GalNAc as acceptor substrates. Northern analysis revealed a restricted expression pattern of two hST6GalNAc II transcripts, a 2.0 kb mRNA found mainly in skeletal muscle, heart and kidney and a 1.8 kb mRNA found in placenta, lung and leukocytes. No transcriptional expression was detected in brain, thymus or spleen. Transcriptional expression of the ST6GalNAc II gene was followed in various human cell lines and found to be expressed in almost all cell types with notable exceptions for several myeloid and lymphoid cell lines.

We have investigated the role of sialylation on cell-cell adhesion mediated by E-cadherin. Two MCF-7 human breast cancer cell variants were studied: MCF-7/AZ cells showed a spontaneous cell-cell adhesion in the fast and slow aggregation assay, whereas the adhesion deficient MCF-7/6 cell variant failed to form larger aggregates, suggesting that E-cadherin was not functional under the conditions of both assays. We measured the sialyltransferase activities using Galβ1-3GalNAcα-O-benzyl and Galβ1- 4GlcNAcα-O-benzyl as acceptor substrates as well as mRNA levels of four sialyltransferases, ST3Gal I, ST3Gal III, ST3Gal IV, ST6Gal I, using multiplex RT-PCR in MCF-7 cell variants. The α2-6 and α2-3 sialylation of E-cadherin was investigated by immuno-blot using Sambucus nigra agglutinin and Maackia amurensis agglutinin. Compared to the adhesion-proficient MCF-7/AZ cells, the adhesion-deficient MCF-7/6 cell line apparently lacks ST6Gal I mRNA, has a lower ST3Gal I mRNA, a lower ST3Gal I sialyltransferase activity, and no α2-3 linked sialic acid moieties on E-cadherin. The potential anti-cancer drug 1-O-octadecyl-2-O-methylglycero-3-phosphocholine (ET-18-OMe, 48 h, 25 μg/ml) belonging to the class of alkyllysophospholipids restored the E-cadherin function in the adhesion-deficient MCF-7/6 cells as evidenced by an increased aggregation. ET-18-OMe caused loss of ST6Gal I mRNA in MCF-7/AZ cells but no changes of sialyltransferase activities or sialic acid moieties on E-cadherin could be observed. We conclude that Ca2+-dependent, E-cadherin-specific homotypic adhesion of MCF-7/AZ or MCF-7/6 cells treated with ET-18-OMe was not affected by sialylation of E-cadherin.

The gas-chromatographic determination of hexadecylphosphocholine (HePC), an experimental antitumor agent of the alkyllysophospholipid group, in Caco-2T cell culture and cell culture media is described. The Caco-2T cells were treated with HePC at a concentration of 40 micrograms/ml (9.8 microM) and the uptake of the drug into the cells (calculated per milligram protein) was measured after 48 h culture (37 degrees C, 10% CO2). Also, a reversibility test for another 48 h was carried out in which the retention of the drug was measured. The toxicity of HePC on Caco-2T cells in viability assays was determined. Before the capillary gas-chromatographic determination, sample cleanup was performed by solid-phase extraction (SPE) on a weak cation-exchange column of the CBA (carboxylic acid) type. For quantitation, racemic 1-O-octadecyl-2-O-methylglycero-3-phosphorylcholine (ET-18-OMe) was added as internal standard, followed by derivatization with trimethylsilylbromide. The results showed that HePC taken up by the cells during 48 h of treatment was still detectable 48 h after removal of the drug from the medium.

4H-Pyrano-[2,3-b]naphthoquinone is a structural motif commonly found in natural products manifesting anticancer activities. As part of a program aimed at structural simplification of bioactive natural products utilizing multicomponent synthetic processes, we developed a compound library based on this heterocyclic scaffold. We found that several library members displayed low micromolar antiproliferative activity and induced apoptosis in human cancer cells. Selected compounds showed promising activity against cancer cell lines resistant to proapoptotic stimuli, demonstrating their potential in treating cancers with dismal prognoses.

Structural simplification of an antimitotic natural product podophyllotoxin with mimetic heterocyclic scaffolds constructed using multicomponent reactions led to the identification of compounds exhibiting low nanomolar antiproliferative and apoptosis-inducing properties. The most potent compounds were found in the dihydropyridopyrazole, dihydropyridonaphthalene, dihydropyridoindole and dihydropyridopyrimidine scaffold series. Biochemical mechanistic studies performed with dihydropyridopyrazole compounds showed that these heterocycles inhibit in vitro tubulin polymerization and disrupt the formation of mitotic spindles in dividing cells at low nanomolar concentrations, in a manner similar to podophyllotoxin itself. Separation of a racemic dihydropyridonaphthalene into individual enantiomers demonstrated that only the optical antipode matching the absolute configuration of podophyllotoxin possessed potent anticancer activity. Computer modeling, performed using the podophyllotoxin binding site on beta-tubulin, provided a theoretical understanding of these successful experimental findings.

After the initial discovery of antiproliferative and apoptosis-inducing properties of a camptothecin-inspired pentacycle based on a 1H-indeno[2′,1′:5,6]dihydropyrido[2,3-d]pyrimidine scaffold, a library of its analogues as well as their oxidized planar counterparts were prepared utilizing a practical multicomponent synthetic protocol. The synthesized compounds exhibited submicromolar to low micromolar antiproliferative potencies toward a panel of human cancer cell lines. Biochemical experiments are consistent with the dihydropyridine library members undergoing intracellular oxidation to the corresponding planar pyridines, which then inhibit topoisomerase II activity, leading to inhibition of proliferation and cell death. Because of facile synthetic preparation and promising antitopoisomerase activity, both the dihydropyridine and planar pyridine-based compounds represent a convenient starting point for anticancer drug discovery.

A series of ethacrynic acid analogues, lacking the α,β-unsaturated carbonyl unit, was synthesized and subsequently evaluated for their ability to inhibit the migration of human breast cancer cells, Hs578Ts(i)8 as well as of human prostate cancer cells, C4-2B. These cell lines provide a good model system to study migration and invasion, since they represent metastatic cancer. Our studies show that ethacrynic acid analogues with methyl substituents at the aromatic ring demonstrate no inhibitory effect on the migration of both cancer cell lines, whereas a precursor in the synthesis of these ethacrynic acid analogues (II-1, a para-acylated m-cresol) is an excellent inhibitor of the migration of both cancer cell lines.

Three different extract conditions (aqueous, EtOH and EtOAc) of four different parts (bracts, leaves, roots and stems) of the plant Anemopsis californica (A. californica) were evaluated for their effect on the growth and migration of human colon cancer cells, HCT-8, and the breast cancer cell lines Hs 578T and MCF-7/AZ. Our aim was to identify potential anticancer activity in crude A. californica extracts, given that this plant is used by Native Americans to treat a variety of diseases, including cancer. Our results demonstrated that for each of the cell lines tested, the majority of ethyl acetate extracts of all the plant parts are more toxic than the aqueous and ethanol extracts. Furthermore, significant growth inhibitory activity against the three cell lines was found for the ethyl acetate extract of the roots, while the aqueous extract of the roots influenced the migratory capacity of the three cell lines. This study provides evidence for the anticancer properties of A. californica when extracted in water and ethyl acetate, and supports the importance for further purification of the crude extracts and isolation of potential new anticancer compounds through bio-guided fractionation.

The mechanisms of invasion and metastasis are poorly understood. Our previous studies demonstrated that cancer cell invasion may result from reorganization of membrane molecules, thereby initiating signaling pathways. To increase our understanding on how cancer cells govern metastases we studied the established LNCaP prostate cancer progression model. Herein we show that the bone metastatic derivative cell line, C4-2B, displays changes in adhesion to collagen type I and invasion into collagen type I. Moreover, we found that these changes were concomitant with activation of the FAK/src/paxillin/Rac/JNK signaling pathway and increased activity of matrix metalloproteinases (MMPs)-2 and -9. Inhibition of src and JNK resulted in inhibition of adhesion and invasion, and deactivation of the signaling molecules in the identified pathway as well as reduced activity of MMPs. Additionally, we found a pivotal role for the integrin alpha2 subunit since lateral redistribution and clustering were responsible for activation of the downstream signaling and function blocking of the integrin alpha2 subunit resulted in poor adhesion and inhibition of invasion. In conclusion, our results suggest that invasion of prostate cancer cells can be ascribed to reorganization and clustering of integrin alpha2 subunits, resulting in activation of associated FAK/src/paxillin/Rac/JNK, leading to increased activity of MMPs and thus invasion.

Twenty-nine Amaryllidaceae alkaloids and their derivatives belonging to the five most common groups, including lycorine, lycorenine, tazettine, crinine, and narciclasine types, were evaluated for antiproliferative, apoptosis-inducing, and anti-invasive activities in vitro. The antiproliferative properties of each test compound are in agreement with those reported in the literature, while the high potency of amarbellisine is reported for the first time. It was also found that with the exception of ungeremine, amarbellisine, and hippeastrine, the antiproliferative effect of the potent compounds is apoptosis mediated. Thus, apoptosis in Jurkat cells was triggered by narciclasine, narciclasine tetraacetate, C10b-R-hydroxypancratistatin, cis-dihydronarciclasine, trans-dihydronarciclasine, lycorine, 1-O-acetyllycorine, lycorine-2-one, pseudolycorine, and haemanthamine. With the exception of narciclasine, lycorine, and haemanthamine, the apoptosis-inducing properties of these compounds are reported for the first time. The collagen type I invasion assay revealed potent anti-invasive properties associated with N-methyllycorine iodide, hippeastrine, clivimine, buphanamine, and narciclasine tetraacetate, all of which were tested at non-toxic concentrations. The anti-invasive activity of buphanamine is particularly promising because this alkaloid is not toxic to cells even at much higher doses. This work has resulted in the identification of several novel leads for anticancer drug design.

Diversely substituted 2-pyrrolines have been prepared by a novel multicomponent process involving a reaction of various N-(aryl- and alkylsulfonamido)-acetophenones with aldehydes and malononitrile. While the reaction is highly regioselective, it is not stereoselective, generating a mixture of cis and trans 2-pyrrolines. A number of analogs from both cis and trans 2-pyrroline libraries were found to have antiproliferative activity in human cancer cell lines.

Ethnotraditional use of plant-derived natural products plays a significant role in the discovery and development of potential medicinal agents. Plants of the genus Taraxacum, commonly known as dandelions, have a history of use in Chinese, Arabian and Native American traditional medicine, to treat a variety of diseases including cancer. To date, however, very few studies have been reported on the anti-carcinogenic activity of Taraxacum officinale (TO). In the present study, three aqueous extracts were prepared from the mature leaves, flowers and roots, and investigated on tumor progression related processes such as proliferation and invasion. Our results show that the crude extract of dandelion leaf (DLE) decreased the growth of MCF-7/AZ breast cancer cells in an ERK-dependent manner, whereas the aqueous extracts of dandelion flower (DFE) and root (DRE) had no effect on the growth of either cell line. Furthermore, DRE was found to block invasion of MCF-7/AZ breast cancer cells while DLE blocked the invasion of LNCaP prostate cancer cells, into collagen type I. Inhibition of invasion was further evidenced by decreased phosphorylation levels of FAK and src as well as reduced activities of matrix metalloproteinases, MMP-2 and MMP-9. This study provides new scientific data on TO and suggests that TO extracts or individual components present in the extracts may be of value as novel anti-cancer agents.

Pyrano[3,2- c]pyridone and pyrano[3,2- c]quinolone structural motifs are commonly found in alkaloids manifesting diverse biological activities. As part of a program aimed at structural simplification of bioactive natural products utilizing multicomponent synthetic processes, we developed compound libraries based on these privileged heterocyclic scaffolds. The selected library members display low nanomolar antiproliferative activity and induce apoptosis in human cancer cell lines. Mechanistic studies reveal that these compounds induce cell cycle arrest in the G2/M phase and block in vitro tubulin polymerization. Because of the successful clinical use of microtubule-targeting agents, these heterocyclic libraries are expected to provide promising new leads in anticancer drug design.

Diversely substituted 2–pyrrolines have been prepared by a novel multicomponent process involving a reaction of various N–(aryl– and alkylsulfonamido)–acetophenones with aldehydes and malononitrile. While the reaction is highly regioselective, it is not stereoselective, generating a mixture of cis and trans 2–pyrrolines. A number of analogues from both cis and trans 2–pyrroline libraries were found to have antiproliferative activity in human cancer cell lines.

The ether lipid 1-O-octadecyl-2-O-methyl-3-glycero-phosphocholine (ET-18-OMe) inhibits cell-cell adhesion and induces invasiveness of breast cancer cells. Previously, we showed that a loss of cell-cell adhesion was due to sterical hindrance of E-cadherin by the anti-adhesive properties of the cell surface mucin episialin. Here, we demonstrated that the ether lipid ET-18-OMe induced the translocation of E-cadherin and episialin to membrane microdomains, enriched in glycosphingolipids, known to be involved in cell-cell adhesion and cell signaling. In addition, it was found that E-cadherin and clusters of episialin colocalized and associated with the glycosphingolipid, MSGb5, upon treatment with ET-18-OMe. Together, these results suggest that ET-18-OMe inhibits cell-cell adhesion by inducing the translocation of E-cadherin and episialin into MSGb5-enriched membrane microdomains, which leads to clustering and colocalization of the pro-adhesive E-cadherin and the anti-adhesive episialin thereby inhibiting cell-cell adhesion.

Increased src tyrosine kinase expression and activity has been associated with colon cancer cell invasion and survival. Several signaling pathways are involved in the oncogenic activation of src during the adenoma to carcinoma progression and cellular invasion. In the present study, the synthetic ether lipid analog ET-18-OMe was shown to promote invasion of HCT-8/S11 colon cancer cells into collagen type I through the concomitant activation of src by phosphorylation at Tyr416 (5-30 min) in alpha1-integrin immunoprecipitates containing the integrin binding proteins talin and paxillin, as well as the phoshorylated and activated forms of focal adhesion kinase (FAK) at Tyr397 (a FAK kinase activation signal), Tyr576 and Tyr861. This was associated with the lateral redistribution of alpha1-integrins in focal aggregates and persistent activation of the p130Cas/JNK pathways at 5-30 min, with the subsequent induction and activation of the matrix metalloproteinases MMP-2 and MMP-9 (2-12 h). These activated molecular scaffolds and signaling cascades were not observed in immunoprecipitates of alpha2- and beta1-integrins, and tetraspanin CD9, an invasion and metastasis suppressor linked to integrins and FAK signaling. Our data demonstrate that the lateral redistribution and clustering of alpha1-integrins results in the recruitment of the FAK/src motility-promoting signaling complex involved in cancer cell invasion. Disruption of this proinvasive pathway was accomplished by the dominant negative mutant of src (K295R, kinase dead), src pharmacological inhibitor (PP1) and alpha1-integrin function blocking antibodies. These findings support the notion that the alpha1-integrin- and src-dependent signalosome is a relevant therapeutic target against tumor progression in colon cancer patients.

Dynamic crosstalk between cell adhesion molecules, extracellular matrix and soluble informative factors is essential for cancer cell migration and invasion. Here, we investigated the mechanisms by which the E-cadherin/catenin complex and alpha v integrin can modulate insulin-like growth factor-I (IGF-I)-induced cell migration. Human colon mucosa, human colon cancer cell lines, HT29-D4 and HCT-8 derivatives that differ in their expression of alpha-catenin, were used as models. Interactions between E-cadherin, alpha v integrin and IGF-I receptor (IGF-IR) were analyzed by coimmunoprecipitation and immunolocalization experiments. The impact of these interactions on cell mobility was determined by haptotaxis assays. We report that alpha v integrin, E-cadherin and IGF-IR form a ternary complex in both cultured cancer cells and human normal colonic mucosa. alpha-Catenin regulates the scaffolding of this complex. IGF-IR ligation by IGF-I induces the disruption of the complex and the relocalization of alpha v integrin from cell-cell contacts to focal contact sites. This perturbation is correlated with the observed increase in cell migration. These results suggest that regulation of the alpha v integrin/E-cadherin/IGF-IR scaffolding is essential for the modulation of cell mobility. Its alteration could be of major importance to sustain alterations in cell adhesion that occur during cancer cell invasion and metastasis.

Plants used in folklore medicine continue to be an important source of discovery and development of novel therapeutic agents. In the present study, we determined the effects of crude aqueous extracts of a panel of medicinal plants on the growth and invasion of cancer cells. Our results showed that extracts of L. tridentata (Creosote Bush) and J. communis L. (Juniper Berry) significantly decreased the growth of MCF-7/AZ breast cancer cells. The latter as well as A. californica (Yerba Mansa) inhibited invasion into the collagen type I gel layer. Furthermore, the phosphorylation levels of extracellular signal-regulated kinase 1 and 2 (ERK1/2) decreased when the cells were exposed to aqueous extracts of L. tridentata, J. communis L. and A. californica. This study provides original scientific data on the anticancer activity of selected aqueous medicinal plant extracts used in traditional medicine.

A new type of photosensitizers used in photodynamic therapy, which is based on photon upconverting nanoparticles, is reported. These photosensitizers are excitable with infrared irradiation, which has several times larger tissue penetration depth than the currently available ones. They are brought close to the target cancer cells through antigen-antibody interaction with good specificity and versatility. The design is also flexible in that various photosensitive molecules can be potentially adopted into the design. Results from in vitro experiments demonstrate their promise of becoming the next generation photodynamic therapy drugs.

A three-component condensation of 5-amino-3-methylpyrazole, tetronic acid, and various aromatic, heteroaromatic, and aliphatic aldehydes leads to the formation of dihydropyridopyrazole analogues of a cytotoxic lignan podophyllotoxin. This new heterocyclic scaffold-based library allows a drastic reduction of the structural complexity of the natural product with the retention of its potent cytotoxic properties. Similarly to podophyllotoxin, the presented analogues induce apoptosis in Jurkat cells.

Native American medicinal plants are traditionally used to prevent and treat a variety of diseases, including cancer. These herbal preparations are alleged to have many biological activities, such as stimulation or suppression of immune responses and antiproliferative effects on cancer cells. In the present study, we investigated the effects of aqueous and ethanol extracts from two Native American plants, Ligusticum porteri (Osha) and Anemopsis californica (Yerba Manza), on the growth of human MCF-7/AZ breast and HCT8/E11 colon cancer cells. The aqueous and ethanol extracts from A. californica potently inhibited growth of MCF-7/AZ in a concentration-dependent manner, whereas the growth of HCT8/E11 was unaltered. Extracts from L. porteri showed no activity on either cell line. In addition, we observed that the extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) activities were markedly decreased when exposed to both extracts from A. californica. These results suggest that the growth inhibitory effect of A. californica in breast cancer cells is ERK-mediated.

Recombinant human α(1→3/4)-fucosyltransferases (FucT-III) expressed in CHO cells or baculovirus-infected insect cells, immobilized on Ni2+-agarose through a 6His tag, exhibit a marked stability, which was exploited in the synthesis of Lewis a and Lewis x trisaccharides.

A multicomponent reaction of indane-1,3-dione, an aldehyde and an amine-containing aromatic compound leading to the formation of indenopyridine-based heterocyclic medicinal scaffolds has been investigated. It was found that the yields significantly improve when oxygen gas is bubbled through the reaction mixture, facilitating the oxidation of the intermediate dihydropyridine-containing compounds to their aromatic counterparts. Investigation of the reaction scope revealed that formaldehyde, as well as various aliphatic, aromatic and heteroaromatic aldehydes, works well as the aldehyde component. In addition, substituted anilines and diverse aminoheterocycles can be utilized in this process as the amine-containing component. Preliminary biological evaluation of the synthesized library identified a pyrimidine-based polycycle, which rivals the anticancer drug etoposide in its toxicity and apoptosis inducing properties toward a human T-cell leukemia cell line.

Our previous work has shown that long-term treatment of mucus-secreting HT-29 cells with 1-benzyl-2-acetamido-2-deoxy-alpha-D-galactopyranoside (GalNAcalpha-O-bn), a competitive inhibitor of O-glycosylation, induced several phenotypic changes, in particular a blockade in the secretion of mucins, which are extensively O-glycosylated glycoproteins. Here, we have analyzed the effects of GalNAcalpha-O-bn upon the intracellular trafficking of basolateral and apical membrane glycoproteins at the cellular and biochemical levels in two types of cells, HT-29 G(-) and Caco-2, differentiated into an enterocyte-like phenotype. In HT-29 G(-) cells, but not in Caco-2 cells, DPP-IV and CD44 failed to be targeted to the apical or basolateral membrane, respectively, and accumulated inside intracytoplasmic vesicles together with GalNAcalpha-O-bn metabolites. We observed a strong inhibition of alpha2,3-sialylation of glycoproteins in HT-29 G(-) cells correlated to the high expression of alpha2,3-sialyltransferases ST3Gal I and ST3Gal IV. In these cells, DPP-IV and CD44 lost the sialic acid residue substituting the O-linked core 1 structure Galbeta1-3GalNAc (T-antigen). In contrast, sialylation was not modified in Caco-2 cells, but a decrease of alpha1,2-fucosylation was observed, in correlation with the high expression of alpha1,2-fucosyltransferases Fuc-TI and Fuc-TII. In conclusion, in HT-29 G(-) cells, GalNAcalpha-O-bn induces a specific cellular phenotype, which is morphologically characterized by the formation of numerous intracellular vesicles, in which are accumulated defectively sialylated O-glycosylproteins originally targeted to basolateral or apical membranes, and GalNAcalpha-O-bn metabolites.