C5a receptor antagonism coming of age for vascular pathology.

The Food and Drug Administration recently approved the new drug avacopan for a relatively rare disease, anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis. Avacopan is an antagonist of receptor-1 for anaphylatoxin C5a (C5aR1) that is the first one to meet all expectations of an orally bioavailable drug. Pharmacological effects of C5a on vascular tissue are reviewed; they are essentially indirect, via resident or infiltrating leukocytes, and largely mediated by vasoconstrictor prostanoids that are potentially thrombogenic. The in vivo acute neutropenic effect of C5a and various responses of isolated neutrophils to the peptide have been exploited in the preclinical development of avacopan, but not the prominent hemodynamic responses. Possible clinical risks and extension of therapeutic C5aR1 blockade are discussed. Therapeutic intervention on the blood-derived peptide C5a and on its G protein coupled receptor for specific forms of vascular injury contrasts with other current research approaches in vascular pathology, such as investigating the roles of cytokines and intracellular signaling.

Quick and simple detection technique to assess the binding of antimicrotubule agents to the colchicine-binding site.

Development of antimitotic binding to the colchicine-binding site for the treatment of cancer is rapidly expanding. Numerous antimicrotubule agents are prepared every year, and the determination of their binding affinity to tubulin requires the use of purified tubulins and radiolabeled ligands. Such a procedure is costly and time-consuming and therefore is limited to the most promising candidates. Here, we report a quick and inexpensive method that requires only usual laboratory resources to assess the binding of antimicrotubules to colchicine-binding site. The method is based on the ability of N,N'-ethylene-bis(iodoacetamide) (EBI) to crosslink in living cells the cysteine residues at position 239 and 354 of ß-tubulin, residues which are involved in the colchicine-binding site. The ß-tubulin adduct formed by EBI is easily detectable by Western blot as a second immunoreacting band of ß-tubulin that migrates faster than ß-tubulin. The occupancy of colchicine-binding site by pertinent antimitotics inhibits the formation of the EBI: ß-tubulin adduct, resulting in an assay that allows the screening of new molecules targeting this binding site.

ASK1-P38 pathway is important for anoikis induced by microtubule-targeting aryl chloroethylureas.

PURPOSE: We investigated the involvement of MAPK signaling in the cell death mechanisms of classical microtubule interfering agents (MIA) and aryl-3-(2-chloroethyl)ureas (CEU) acting as antimitotics, along with CEU that don't affect directly microtubules (non-MIA CEU). METHODS: To ascertain the activated signaling pathway profile of MIA and non-MIA CEU, Western blot, immunoprecipitation and transfection experiments were performed. RESULTS: Non-MIA CEU do not activate p38, as opposed to MIA, and the extent of ERK and JNK activation is lower than in response to MIA. The effect of MIA and non-MIA CEU on focal adhesion associated protein was also studied; MIA were shown to induce focal adhesion dismantlement associated with a sustained increase in paxillin phosphorylation and FAK cleavage, as opposed to non-MIA CEU. In addition, bcl-2 phosphorylation and AKT cleavage, induced by all MIA tested, was not observed in response to non-MIA CEU further emphasizing the differential cell death mechanisms induced by MIA and non-MIA CEU. Pharmacologic and genetic approaches emphasize that the ASK1-p38 pathway activation contributes to the cytotoxic mechanism of MIA, in contrast to non-MIA CEU. ASK1-p38 is important for increased paxillin phosphorylation and FAK cleavage, suggesting that ASK-1-p38 is an upstream event of FA structure dismantlement induced by MIA. Moreover, the endogen inhibitor of ASK-1, thioredoxin, is released from ASK-1 in response to MIA as opposed to non-MIA CEU. CONCLUSION: Our study supports that ASK1-p38 activation is an important signaling event, induced by MIA, which impairs focal adhesion structure and induces anchorage-dependent apoptosis or anoikis.

Chloroethyl urea derivatives block tumour growth and thioredoxin-1 nuclear translocation.

Aryl chloroethyl ureas (CEUs) are new protein alkylating agents exhibiting anticancer activity both in vitro and in vivo. We report herein that 14C-labeled CEU derivatives, designated CEU-025 and CEU-027, covalently bind to thioredoxin-1 (TRX1). Covalent binding of these molecules slightly decreases the disulfide-reducing activity of recombinant TRX1, when compared with the effect of strong thioalkylating agents such as N-ethylmaleimide. Moreover, site-directed mutagenesis and diamide competition assays demonstrated that TRX1 cysteinyl residues are not the prime targets of CEUs. CEU-025 abrogates the nuclear translocation of TRX1 in human cancer cells. In addition, we show that CEU-025 can block TRX1 nuclear translocation induced by cisplatin. Unexpectedly, pretreatment with sublethal CEU-025 concentrations that block TRX1 nuclear translocation protected the cells against cisplatin cytotoxicity. Overexpression of TRX1 in HT1080 fibrosarcoma cells attenuated CEU-025 cytotoxicity, while its suppression using TRX1-specific siRNA increased the effects of CEU-025, suggesting that loss of function of TRX1 is involved, at least in part, in the cytotoxic activity of CEU-025. These results suggest that CEU-025 and CEU-027 exhibit anticancer activity through a novel, unique mechanism of action. The importance of TRX1 and the dependence of the cytotoxicity of CEU-025 and CEU-027 on TRX1 intracellular localization are also discussed.

Mechanism of action of N-phenyl-N'-(2-chloroethyl)ureas in the colchicine-binding site at the interface between alpha- and beta-tubulin.

Computational tools such as CoMSIA and CoMFA models reported in a recent study revealed the structure-activity relationships ruling the interactions occurring between hydrophobic N-phenyl-N'-(2-chloroethyl)ureas (CEU) and the colchicine-binding site (C-BS) on beta(II)-tubulin. Here, we describe the mechanisms involved in the covalent binding of three subsets of CEU derivatives to the C-BS. The FlexiDock experiments confirmed that the interaction of non-covalent portions of the CEU auxophore moiety of CEU is involved in the binding of the drug to the C-BS facilitate the nucleophilic attack of Glu-beta198 rather than Cys-beta239. In addition, these studies suggest that Cys-beta239 together with Asn-alpha99, Ser-alpha176, Thr-alpha177, Leu-beta246, Asn-beta247, Ala-beta248, Lys-beta252 and Asn-beta256 are implicated in the stabilization of a C-BS-CEU complex prior to the acylation of Glu-beta198 by CEU. Our molecular models propose the formation of a stabilized C-BS-CEU complex before the completion of the Glu-beta198 acylation; acylation triggering conformational changes of beta-tubulin, microtubule depolymerization and anoikis. The computational models presented here might be useful to the design of selective and more potent C-BS inhibitors. Of interest, in vivo acylation of acidic amino acid residues by xenobiotics is an unusual reaction and may open new approaches for the design of irreversible protein inhibitors such as tubulin.

Tumor cells expressing tissue factor influence the migration of smooth muscle cells in a catalytic activity-dependent way.

The expression of tissue factor (TF) in tumors reportedly exacerbates the aggressiveness of several types of cancers. The shedding of TF-containing membrane particles is believed to influence the ability of tumors to expand and metastasize, and these microparticles may also be harmful in the onset of disseminated intravascular coagulation in specific cancers. Furthermore, the intracellular signaling that is elicited after the formation of the TF / coagulation factor VIIa complex at the cell membrane modulates the activity of adhesion molecules and mitogen-activated protein (MAP) kinases. To evaluate whether TF overexpression in tumor cells modulates its shedding and neighboring stromal cells by its catalytic or intracellular activity, TF-GFP (green fluorescent protein) and a tailless form (TFDeltaC-GFP) were stably expressed in the rat Morris hepatoma and human HT1080 fibrosarcoma cell lines. Both TF proteins were efficiently produced by tumor cells and functionally active, and their clotting activity could be blocked by the active site-inhibited factor VIIa (ASIS). TF-expressing tumorigenic cells produced a soluble factor that increased the migration of arterial smooth muscle cells in vitro. This effect was abrogated by ASIS and the PAR-1 receptor antagonist ATAP-2, showing that it is dependent on the proteolytic activity of the TF ligand factor VIIa and the thrombin-activated cell membrane receptor. We propose that TF-containing microparticles that are released in the culture medium by tumor cells influence the migratory behavior of neighboring stromal cells, thus aiding the cancer cell's tumorigenic potential.

New soft alkylating agents with enhanced cytotoxicity against cancer cells resistant to chemotherapeutics and hypoxia.

Chloroethylureas (CEU) are soft alkylating agents that covalently bind to beta-tubulin (betaTAC) and affect microtubule polymerization dynamics. Herein, we report the identification of a CEU subset and its corresponding oxazolines, which induce cell growth inhibition, apoptosis, and microtubule disruption without alkylating beta-tubulin (N-betaTAC). Both betaTAC and N-betaTAC trigger the collapse of mitochondrial potential (DeltaPsi(m)) and modulate reactive oxygen species levels, following activation of intrinsic caspase-8 and caspase-9. Experiments using human fibrosarcoma HT1080 respiratory-deficient cells (rho(0)) and uncoupler of the mitochondrial respiratory chain (MRC) showed that betaTAC and N-betaTAC impaired the MRC. rho(0) cells displayed an increased sensitivity toward N-betaTAC as compared with rho(+) cells but, in contrast, were resistant to betaTAC or classic chemotherapeutics, such as paclitaxel. Oxazoline-195 (OXA-195), an N-betaTAC derivative, triggered massive swelling of isolated mitochondria. This effect was insensitive to cyclosporin A and to Bcl-2 addition. In contrast, adenine nucleotide translocator (ANT) antagonists, bongkrekic acid or atractyloside, diminished swelling induced by OXA-195. The antiproliferative activities of the N-betaTACs CEU-025 and OXA-152 were markedly decreased in the presence of atractyloside. Conversely, pretreatment with cyclosporin A enhanced growth inhibition induced by betaTAC and N-betaTAC. One of the proteins alkylated by N-betaTAC was identified as the voltage-dependent anion channel isoform-1, an ANT partner. Our results suggest that betaTAC and N-betaTAC, despite their common ability to affect the microtubule network, trigger different cytotoxic mechanisms in cancer cells. The role of mitochondria in these mechanisms and the potential of N-betaTAC as a new therapeutic approach for targeting hypoxia-resistant cells are discussed.

Cycloalkyl-substituted aryl chloroethylureas inhibiting cell cycle progression in G0/G1 phase and thioredoxin-1 nuclear translocation.

1-(2-Chloroethyl)-3-(4-cyclohexylphenyl)urea (cHCEU) has been shown to abrogate the presence of thioredoxin-1 into the nucleus through its selective covalent alkylation. In the present letter we have evaluated the structure-activity relationships of the substituents at positions 3 and 4 of the phenyl ring of cHCEU derivatives on cell cycle progression and thioredoxin-1 nuclear translocation. Active CEU derivatives exhibited GI(50) ranging from 1.9 to 49muM on breast carcinoma MCF-7, skin melanoma M21, and colon carcinoma HT-29 cells. On one hand, compounds 1, 2, 9c, 10c, 13, and 14 arrested the cell cycle in G(2)/M phase while CEUs 3, 4, 5c, 6c, 11c, and 12c blocked the cell division in G(0)/G(1) phase. On the other hand, CEUs 2-4, 5c, 7c, 8c, 11c, and 12c abrogated the translocation of thioredoxin-1 while the other CEU derivatives were inactive in that respect. Our results suggest that CEU substituted on the phenyl ring at position 3 or 4 by lower cycloalkyl or cycloalkoxy groups arrest cell progression in G(0)/G(1) phase through mechanism of action different from their antimicrotubule counterparts, presumably via thioredoxin-1 alkylation and modulation of its activity. The mechanism of action of these new molecules is still undetermined. However, the significant accumulation of cells in G(0)/G(1) phase suggests that these molecules may act similarly to known chemopreventive agents against cancers. In addition, the inhibition of Trx-1 nuclear localization also suggests the abrogation of an important chemoresistance mechanism towards a variety of chemotherapeutic agents.

Aromatic 2-chloroethyl urea derivatives and bioisosteres. Part 2: Cytocidal activity and effects on the nuclear translocation of thioredoxin-1, and the cell cycle progression.

Recently, a subset of N-phenyl-N'-(2-chloroethyl)ureas (CEU) was found abrogating the nuclear translocation of thioredoxin-1 and arresting the cell cycle in G(0)/G(1) phase. Several derivatives were prepared to assess their effect on cell cycle progression and on the intracellular location of Trx-1. Compounds 1-20, 21-40, and 41-60 exhibited GI(50) between 1 and 80 microM. Immunocytochemistry analysis showed compounds 4, 6, 8, 10, 11, 23, 24, 26-31, 34, 37, 41, 44, 46-51, 53, 56, and 57 inhibiting the nuclear translocation of Trx-1. Our results suggest that increasing the electrophilic character of these molecules might enhance the antiproliferative activity at the expense of the selectivity toward thioredoxin-1 and the G(0)/G(1) phase arrest.

Selective alkylation of beta(II)-tubulin and thioredoxin-1 by structurally related subsets of aryl chloroethylureas leading to either anti-microtubules or redox modulating agents.

Aryl chloroethylureas (CEUs) are potent anti-neoplastic agents alkylating specific intracellular proteins such as beta(II)-tubulin. Recently we have identified a new subset of CEU derived from compound 36 that alkylates thioredoxin isoform 1 (Trx-1), inhibits the nuclear translocation of Trx-1, and favors the accumulation of cells in G(0)/G(1) phase. We have evaluated the effects of various substituents and their position on the aromatic ring of a series of derivatives of 36 on (i) the anti-proliferative activity, (ii) the cell cycle progression, (iii) the nuclear translocation of Trx-1, and (iv) their covalent binding to beta-tubulin. The same experiments were performed on representative CEU derivatives where the 2-chloroethyl amino moiety is replaced by either an ethyl, a 2-aminooxazolinyl or a 2-chloroacetyl group. On one hand, our results suggest that CEUs substituted on the phenyl ring at position 3 or 4 by cycloalkyl and substituted cycloalkyl or cycloalkoxy groups inhibit the nuclear translocation of Trx-1 and arrest the cell cycle progression in G(0)/G(1). On the other hand, CEUs substituted by a fused aromatic ring, an aliphatic chain, or a fused aliphatic ring are alkylating beta(II)-tubulin but not Trx-1. Beside the expected inactivity of the ethylurea derivatives, none of the modification to the electrophilic moiety led to cross-selectivity of the drugs toward beta-tubulin but increased the anti-proliferative activity and resulted in mitigated effects on Trx-1 translocation.

Covalent grafting of fibronectin onto plasma-treated PTFE: influence of the conjugation strategy on fibronectin biological activity.

Surface coating of synthetic materials is often considered to improve biomedical devices biocompatibility. In this study, we covalently bound fibronectin (FN) onto ammonia plasma-treated PTFE via two crosslinkers, namely glutaric anhydride (GA) and sulfosuccinimidyl-4-(p-maleimidophenyl)butyrate (sulfo-SMPB). With respect to clean PTFE, cell adhesion increased markedly on both FN grafted surfaces, although it was twice higher on PTFE-GA-FN than on PTFE-SMPB-FN. ELISA experiments performed with a polyclonal antibody revealed that the amount of FN is identical on both surfaces while monoclonal antibody specific to the RGD binding site clearly demonstrated a greater availability when FN is surface grafted through GA. These results provide evidence of a variation in protein conformation correlated with the surface conjugation strategy.

Antiangiogenic and antitumoral activity of phenyl-3-(2-chloroethyl)ureas: a class of soft alkylating agents disrupting microtubules that are unaffected by cell adhesion-mediated drug resistance.

The development of new anticancer agents with lower toxicity, higher therapeutic index, and weaker tendency to induce resistant phenotypes in tumor cells is a continuous challenge for the scientific community. Toward that end, we showed previously that a new class of soft alkylating agents designed as phenyl-3-(2-chloroethyl)ureas (CEUs) inhibits tumor cell growth in vitro and that their efficiency is not altered by clinically relevant mechanisms of resistance such as overexpression of multidrug resistance proteins, increase in intracellular concentration of glutathione and/or glutathione S-transferase activity, alteration of topoisomerase II, and increased DNA repair. Mechanistic studies have showed recently that the cytotoxic activity of several CEUs was mainly related to the disruption of microtubules. Here, we present results supporting our assumption that 4-tert-butyl-[3-(2-chloroethyl)ureido]phenyl (tBCEU) (and its bioisosteric derivative 4-iodo-[3-(2-chloroethyl)ureido]phenyl (ICEU) are potent antimicrotubule agents both in vitro and in vivo. They covalently bind to beta-tubulin, leading to a microtubule depolymerization phenotype, consequently disrupting the actin cytoskeleton and altering the nuclear morphology. Accordingly, tBCEU and ICEU also inhibited the migration and proliferation of endothelial and tumor cells in vitro in a dose-dependent manner. It is noteworthy that ICEU efficiently blocked angiogenesis and tumor growth in three distinct animal models: (a) the Matrigel plug angiogenesis assay; (b) the CT-26 tumor growth assay in mice; and (c) the chick chorioallantoic membrane tumor assay. In addition, we present evidence that CEU cytotoxicity is unaffected by additional resistance mechanisms impeding tumor response to DNA alkylating agents such as cisplatin, namely the cell adhesion mediated-drug resistance mechanism, which failed to influence the cytocidal activity of CEUs. On the basis of the apparent innocuousness of CEUs, on their ability to circumvent many classical and recently described tumor cell resistance mechanisms, and on their specific biodistribution to organs of the gastrointestinal tract, our results suggest that CEUs represent a promising new class of anticancer agents.

MMP-2 expression in uveal melanoma: differential activation status dictated by the cellular environment.

PURPOSE: MMPs are recognized to play a major role in tumor progression and metastasis of many forms of cancers. The purpose of this study was to compare the expression and activity of MMP-2 in uveal melanoma cell lines grown either in vitro on plastic culture plates or in vivo as tumors produced in chick embryos. METHODS: The chick chorioallantoic membrane (CAM) model was used to evaluate the tumorigenic potential of uveal melanoma cell lines derived either from the primary uveal melanoma tumor isolated from three different patients (cell lines SP6.5, SP8.0, and TP31) or from a metastatic lesion derived from the liver of a patient diagnosed with uveal melanoma (cell line H79). The presence of MMP-2 in the vicinity of the tumor cells was determined by immunofluorescence analyses. Gelatin zymography was used for the detection of latent and activated forms of MMP-2 in uveal melanoma cell lines when grown in vitro on plastic, or in the solid tumors these cell lines produced in vivo on the CAM of the chick embryo. The gelatinase activity was quantified by densitometric analyses and the active/(active+pro-form) ratio was calculated as the MMP-2 activation ratio. Western blot analyses were performed to confirm the zymographic profile. RESULTS: Only the inactive form of MMP-2 was expressed and secreted in vitro by all uveal melanoma cell lines, higher levels being found for the liver-derived H79 cell line whereas SP8.0 only expressed MMP-2 to a very low level. On the other hand, all solid tumors produced in the CAM from these cell lines expressed and secreted, although to varying levels (SP6.5 and SP8.0, TP31 and H79), primarily the active form of MMP-2. Gelatinolytic activities of active MMP-2 were significantly higher in uveal melanoma tissues than in the non-neoplastic CAM, as revealed by the measurement of the activation ratio. The immunolocalization of MMP-2 revealed that all cell lines were MMP-2-positive although a reduced and more diffuse staining was observed for H79 and SP6.5 than in SP8.0 and TP31 cells. CONCLUSIONS: These results suggest the activation of proMMP-2 as an important event in the process of uveal melanoma progression. An elevated active to inactive MMP-2 ratio in the tumor environment of uveal melanoma suggests that a potential MMP-2 activity could be related to the progression of this type of cancer.

Role of the extracellular matrix proteins in the resistance of SP6.5 uveal melanoma cells toward cisplatin.

Uveal melanoma is the most frequent primary intraocular tumor in the adult population. This malignancy has a high mortality rate and responds poorly to existing chemotherapy. Recently, the tumor environment has been found to exert a profound influence on drug response through cell interaction with components from the extracellular matrix (ECM). In the present study, we investigated whether individual components from the ECM may affect cell survival and/or cell death induced by the cytotoxic agent cisplatin on the SP6.5 uveal melanoma cell line. Tumor cells were shown by immunofluorescence analyses to be surrounded by the ECM proteins fibronectin (FN), type IV collagen (CIV) and laminin (LM), both at the primary and metastatic sites. Binding of SP6.5 cells to FN, LM and CIV is primarily dictated by the expression of membrane bound integrins from the beta1 family as revealed by cell adhesion assays conducted on ECM-coated culture plates. Analysis of cell death by flow cytometry demonstrated that culturing SP6.5 cells in the presence of FN, CIV and LM, substantially reduced the percentage of cells undergoing apoptosis after cisplatin treatment when compared with those seeded on a non-permissive matrix. These results suggest that adhesion of the SP6.5 uveal melanoma cells to the ECM proteins FN, CIV and LM might therefore confer resistance to the chemotherapeutic agent cisplatin. The cellular resistance induced by the ECM proteins toward cisplatin could explain in part the local recurrence of metastasis derived from uveal melanoma often observed clinically after chemotherapy.

Effect of reproductive hormones on ovarian epithelial tumors: II. Effect on angiogenic activity.

Menstrual cycle activity predisposes to ovarian epithelial tumors based on numerous epidemiological studies. We showed that the hormones involved in menstrual cycle regulation modulate cell cycle activity in these tumors in an accompanying paper. We investigated whether such hormones could also influence angiogenesis, an important determinant of tumor progression, in the same tumors. Treatment with progesterone (P4) stimulated VEGF protein secretion in 4 of 5 ovarian carcinoma cell lines examined. Northern blot analyses performed in MCV50 cells showed that this effect was accompanied by increased VEGF mRNA levels. P4 also stimulated VEGF promoter activity in these cells. Estradiol (E2) showed a similar, but substantially smaller effect on VEGF secretion which was additive to that of P4. Conditioned medium from P4-treated cells strongly stimulated angiogenesis on chicken chorio-allantoic membranes. Incubating the conditioned medium with a neutralizing anti-VEGF antibody, but not with non-specific immunoglobulins abolished this effect. Angiogenic activity was not altered by treatment of the membranes with P4 directly. We conclude that P4 can stimulate angiogenic activity via induction of VEGF secretion in some ovarian epithelial tumors. Therapeutic use of progestins may be most effective when administered in combination with an anti-angiogenic agent, at least against a subset of ovarian carcinomas.

Plasminogen activator inhibitor-1 in tumor growth, angiogenesis and vascular remodeling.

Plasminogen activator inhibitor-1 (PAI-1) is the principal inhibitor of urokinase type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA), and as such is thought to play an important role in the regulation of extracellular matrix remodeling. In blood, PAI-1 is bound to the adhesion protein vitronectin and is associated with vitronectin in fibrin clots and the provisional matrix. Elevated levels of PAI-1 are associated with atherosclerosis and an increased thrombotic tendency, while PAI-1 deficiency leads to increased fibrinolysis and bleeding. PAI-1 is also elevated in many solid tumors and is associated with a poor prognosis in cancer. PAI-1 has been shown to be a potent regulator of both vascular cell migration in vitro and of angiogenesis and tumor growth in vivo. PAI-1 can both promote and inhibit tumor growth and angiogenesis. Low concentrations of PAI-1 can stimulate tumor angiogenesis while treatment of animals with high doses of PAI-1 inhibits angiogenesis and tumor growth. Hence, PAI-1 appears to have a multifunctional role in regulating the migratory and fibrinolytic activity of vascular cells, and this, in turn, may help to explain the many varied actions of PAI-1.

Kinin receptors: functional aspects.

Two types of receptors (B1R, B2R) for kinins are defined in mammalian species. Comparative experiments involving recombinant fusion proteins consisting of rabbit B1R or B2R fused to GFP-related proteins are exploited to study the regulation of the response to kinins at the receptor level. The following points will be briefly reviewed and supported by some novel data. (1) The constitutive B2Rs are internalized upon agonist stimulation, but completely recycled to the cell surface; however, B2R destruction can be achieved following limited proteolysis (extracellular trypsin, neutrophil proteases), a plausible down-regulation mechanism in pathology. (2) The inducible B1Rs, stimulated by des-Arg9-kinins, are not phosphorylated nor internalized upon agonist stimulation, but rather undergo a reversible redistribution to caveolae-related rafts. B2Rs are also subjected to this translocation, but only transiently (before endocytosis). (3) Based on the analysis of rabbit aortic smooth muscle cells, B1R induction by cytokines is dependent on nuclear factor KB in rabbit vascular tissue, but exogenous kinins acting on either receptor type do not induce B1R expression.

Substituted phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamides as antimitotics. Antiproliferative, antiangiogenic and antitumoral activity, and quantitative structure-activity relationships.

The importance of the bridge linking the two phenyl moieties of substituted phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonates (PIB-SOs) was assessed using a sulfonamide group, which is a bioisostere of sulfonate and ethenyl groups. Forty one phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamide (PIB-SA) derivatives were prepared and biologically evaluated. PIB-SAs exhibit antiproliferative activities at the nanomolar level against sixteen cancer cell lines, block the cell cycle progression in G(2)/M phase, leading to cytoskeleton disruption and anoikis. These results were subjected to CoMFA and CoMSIA analyses to establish quantitative structure-activity relationships. These results evidence that the sulfonate and sulfonamide moieties are reciprocal bioisosteres and that phenylimidazolidin-2-one could mimic the trimethoxyphenyl moiety found in the structure of numerous potent antimicrotubule agents. Finally, compounds 16 and 17 exhibited potent antitumor and antiangiogenic activities on HT-1080 fibrosarcoma cells grafted onto chick chorioallantoic membrane similar to CA-4 without significant toxicity for the chick embryos, making this class of compounds a promising class of anticancer agents.

Ultra-small gadolinium oxide nanoparticles to image brain cancer cells in vivo with MRI.

The majority of contrast agents used in magnetic resonance imaging (MRI) is based on the rare-earth element gadolinium. Gadolinium-based nanoparticles could find promising applications in pre-clinical diagnostic procedures of certain types of cancer, such as glioblastoma multiforme. This is one of the most malignant, lethal and poorly accessible forms of cancer. Recent advances in colloidal nanocrystal synthesis have led to the development of ultra-small crystals of gadolinium oxide (US-Gd(2)O(3), 2-3 nm diameter). As of today, this is the smallest and the densest of all Gd-containing nanoparticles. Cancer cells labeled with a sufficient quantity of this compound appear bright in T(1)-weighted MRI images. Here we demonstrate that US-Gd(2)O(3) can be used to label GL-261 glioblastoma multiforme cells, followed by localization and visualization in vivo using MRI. Very high amounts of Gd are efficiently internalized and retained in cells, as confirmed with TEM and ICP-MS. Labeled cells were visualized in vivo at 1.5 T using the chicken embryo model. This is one more step toward the development of "positively contrasted" cell tracking procedures with MRI.

Design, synthesis, biological evaluation, and structure-activity relationships of substituted phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonates as new tubulin inhibitors mimicking combretastatin A-4.

Sixty-one phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonates (PIB-SOs) and 13 of their tetrahydro-2-oxopyrimidin-1(2H)-yl analogues (PPB-SOs) were prepared and biologically evaluated. The antiproliferative activities of PIB-SOs on 16 cancer cell lines are in the nanomolar range and unaffected in cancer cells resistant to colchicine, paclitaxel, and vinblastine or overexpressing the P-glycoprotein. None of the PPB-SOs exhibit significant antiproliferative activity. PIB-SOs block the cell cycle progression in the G(2)/M phase and bind to the colchicine-binding site on ß-tubulin leading to cytoskeleton disruption and cell death. Chick chorioallantoic membrane tumor assays show that compounds 36, 44, and 45 efficiently block angiogenesis and tumor growth at least at similar levels as combretastatin A-4 (CA-4) and exhibit low to very low toxicity on the chick embryos. PIB-SOs were subjected to CoMFA and CoMSIA analyses to establish quantitative structure-activity relationships.