Antitumor agents. 256. Conjugation of paclitaxel with other antitumor agents: evaluation of novel conjugates as cytotoxic agents

Sixteen different taxoid conjugates were prepared by linking various anticancer compounds, including camptothecin (CPT), epipodophyllotoxin (EP), colchicine (COL), and glycyrrhetinic acid (GA), at the 2'- or 7-position on paclitaxel (TXL, 1) through an ester, imine, amine, or amide bond. Newly synthesized conjugates were evaluated for cytotoxic activity against replication of several human tumor cell lines. Among them, TXL-CPT conjugates, 8-10, were more potent than TXL itself against the human prostate carcinoma cell line PC-3 (ED(50)=14.8, 3.1, 19.4nM compared with 55.5nM), and conjugate 10 was also 8-fold more active than TXL against the LN-CAP prostate cancer cell line. These compounds also possessed anti-angiogenesis ability as well as lower inhibitory effects against a normal cell line (MRC-5). Thus, conjugates 8-10 are possible antitumor drug candidates, particularly for prostate cancer.     

Conjugated platinum(IV)-peptide complexes for targeting angiogenic tumor vasculature

The integrins alpha vbeta3 and alpha vbeta5 and the membrane-spanning surface protein aminopeptidase N (APN) are highly expressed in tumor-induced angiogenesis, making them attractive targets for therapeutic intervention. Both integrins and APN recognize a broad range of peptides containing RGD (Arg-Gly-Asp) and NGR (Asn-Gly-Arg) motifs, respectively. Here, we describe the design, synthesis, and characterization of a series of mono- and difunctionalized platinum(IV) complexes in which a conjugated peptide motif, containing RGD, (CRGDC)c, (RGDfK)c, or NGR, is appended as a "tumor-homing device" to target tumor endothelial cells selectively over healthy cells. Platinum(IV)-peptide complexes with nonspecific amino acids or peptide moieties were prepared as controls. Concentration-response curves of these compounds were evaluated against primary proliferating endothelial cells and tumor cell lines and compared to those of cisplatin, a well-described platinum-based chemotherapeutic agent. The Pt(IV)-RGD conjugates were highly and specifically cytotoxic to cell lines containing alpha vbeta3 and alpha vbeta5, approaching the activity of cisplatin. The Pt(IV)-NGR complexes were less active than Pt(IV)-RGD-containing compounds but more active than nonspecific Pt-peptide controls. Integrin alpha vbeta3 mediated, at least in part, the anti-proliferative effect of a Pt(IV)-RGD conjugate, as demonstrated by a decreased inhibitory response when endothelial cells were either (1) incubated with an excess of alpha vbeta3/alpha vbeta5-specific RGD pentapeptides or (2) transfected with RNAi for beta 3, but not beta 1, integrins. These results suggest a rational approach to improved chemotherapy with Pt(IV)-peptide conjugates by selective drug delivery to the tumor compartment.     

Antitumor agents. 258. Syntheses and evaluation of dietary antioxidant--taxoid conjugates as novel cytotoxic agents

Various dietary antioxidants, including vitamins, flavonoids, curcumin, and a coumarin, were conjugated with paclitaxel (1) through an ester linkage. The newly synthesized compounds were evaluated for cytotoxic activity against several human tumor cell lines as well as the corresponding normal cell lines. Interestingly, most tested conjugates selectively inhibited the growth of 1A9 (ovarian) and KB (nasopharyngeal) tumor cells without activity against other cell lines. Particularly, conjugates 16 and 20 were highly active against 1A9 (ED(50) value of 0.005 microg/mL) as well as KB (ED(50) values of 0.005 and 0.14 microg/mL, respectively) cells. Compound 22b, the glycinate ester salt of vitamin E conjugated with 1, appears to be a promising lead for further development as a clinical trial candidate as it exhibited strong inhibitory activity against Panc-1 (pancreatic cancer) with less effect on the related E6E7 (normal) cell line.     

Immunotargeting of daunomycin to localized and metastatic human colon adenocarcinoma in athymic mice

Summary A monoclonal antibody (designated SF25), which recognizes a protein antigen expressed on a large number of human colon carcinomas, was used for drug targeting. Daunomycin-antibody conjugates were prepared by two previously described procedures. In one, the drug was bound to the antibody through a spacer of small molecular mass (cis-aconitic acid), while in the other a dextran bridge served as the link between drug and antibody. High substitution rates of drug to antibody were obtained using the latter binding procedure. Both conjugates were tested in vitro against two human colon carcinoma cell lines, LS180 and KM-12. The efficacy of a daunomycin-dextran-SF25 antibody conjugate was tested against colon carcinoma LS180 tumors transplanted at different sites into athymic mice. The specific conjugate was significantly more inhibitory to a subcutaneous tumor growth than its components or their mixture. SF25 antibody alone showed antitumoral effects against all three forms of transplanted tumor tested, namely, local, metastatic or intrahepatic, whereas daunomycin, on its own, was effective only against the subcutaneous tumor. Binding of daunomycin to dextran partially improved its inhibitory activity against the metastatic tumor. The conjugate, daunomycin-dextran-SF25 antibody reduced the number of metastatic foci, increased the survival rate and delayed death. Yet against lymph node metastases it was not significantly better than a mixture of both constituents. However, results obtained with an intrahepatic tumor, a model that mimics the natural progression of the disease, resembled those described with the subcutaneous tumor. Daunomycin-dextran-SF25 antibody was significantly more effective than all components separately and than a mixture of drug and antibody, provided a highly drug-substituted conjugate was used.     

Synthesis and human carbonic anhydrase I,II, VA,and XII inhibition with novel amino acid–sulphonamide conjugates

Abstract

A series of amino acid–sulphonamide conjugates was prepared through benzotriazole mediated coupling reactions and characterised by ¹H-NMR, ¹³C-NMR, MS, and FTIR spectroscopic techniques as well as elemental analysis. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activity of the new compounds was determined against four human (h) isoforms, hCA I, hCA II, hCA VA, and hCA XII. Most of the synthesised compounds showed effective in vitro CA inhibitory properties. The new amino acid–sulphonamide conjugates showed potent inhibitory activity against hCA II, some of them at subnanomolar levels, exhibiting more effective inhibitory activity compared to the standard drug acetazolamide. Some of these sulphonamides were also found to be effective inhibitors of hCA I, hCA VA, and hCA XII, with activity from the low to high nanomolar range.     

Design, synthesis and antitumor evaluation of phenyl N-mustard-quinazoline conjugates

A series of N-mustard-quinazoline conjugates was synthesized and subjected to antitumor studies. The N-mustard pharmacophore was attached at the C-6 of the 4-anilinoquinazolines via a urea linker. To study the structure-activity relationships of these conjugates, various substituents were introduced to the C-4 anilino moiety. The preliminary antitumor studies revealed that these agents exhibited significant antitumor activity in inhibiting various human tumor cell growths in vitro. Compounds 21b, 21g, and 21h were selected for further antitumor activity evaluation against human breast carcinoma MX-1 and prostate PC-3 xenograft in animal model. These agents showed 54-75% tumor suppression with low toxicity (5-7% body-weight changes). We also demonstrate that the newly synthesized compounds are able to induce DNA cross-linking through alkaline agarose gel shift assay and inhibited cell cycle arrest at G2/M phase.     

The influence of carbohydrate ligands on the cytotoxicity of liposomes bearing a methotrexate-diglyceride conjugate in human acute leukemia cell cultures

The efficiency of the chemotherapeutic agent methotrexate (MTX) in cancer treatment is limited by the frequent development of the drug resistance of tumor cells. We had previously shown in vitro using human acute leukemia cells with various sensitivity to MTX (T-lymphoblastic CCRF-CEM line and resistant CEM/MTX subline) that MTX incorporation into liposomes in the form of a lipophilic prodrug, diglyceride conjugate (MTX-DG), allows for the overcoming of cell resistance due to the impaired active transmembrane transport. In this work, we have studied the profile of binding with carbohydrates of the cell lines mentioned using carbohydrate fluorescent probes (poly(acryl amide) conjugates). Lipophilic conjugates of tetrasaccharide SiaLe^X, 6′-HSO₃LacNAc, and also inactive pentaol for incorporation into liposomes, have been synthesized. The cytotoxicity of MTX-DG liposomes equipped with the SiaLe_X ligand toward the sensitive CCRF-CEM cell culture was demonstrated to be 3.5 times higher than that of MTX-DG liposomes bearing the control inactive pentaol. The activity of MTX liposomes bearing 6′-HSO₃LacNAc toward resistant CEM/MTX was 1.6-fold increased. The use of carbohydrate ligands as molecular addresses for drug-carrying liposomes as a potential method of treating heterogeneous tumor tissue is discussed.     

Synthesis, biological evaluation and molecular docking studies of 3-(1,3-diphenyl-1H-pyrazol-4-yl)-N-phenylacrylamide derivatives as inhibitors of HDAC activity

In present study, a series of 3-(1,3-diphenyl-1H-pyrazol-4-yl)-N-phenylacrylamide derivatives (5a-8d) were designed, synthesized, and evaluated for HDAC inhibition and tumor cell antiproliferation. All of these compounds are reported for the first time, the chemical structures of these compounds were confirmed by means of (1)H NMR, ESI-MS and elemental analyzes. Among the compounds, compound 8c showed the most potent biological activity against HCT116 cancer cell line (IC(50) of 0.42 ± 0.02 μM for HDAC-1 and IC(50)=0.62 ± 0.02 for HCT116). Docking simulation was performed to position compound 8c into the HDAC active site to determine the probable binding model. The results of antiproliferative assay and western-blot demonstrated that compound 8c with potent inhibitory activity in tumor growth inhibition may be a potential anticancer agent against HCT116 cancer cell.     

Preparation,characterization, and in vitro efficacy of O-carboxymethyl chitosan conjugate of melphalan

A series of melphalan-O-carboxymethyl chitosan (Mel-OCM-chitosan) conjugates with different spacers were prepared and structurally characterized. All conjugates showed satisfactory water-solubility (160-217 times of Mel solubility). In vitro drug release behaviors by both chemical and enzymatic hydrolysis were investigated. The prodrugs released Mel rapidly within papain and lysosomal enzymes of about 40–75%, while released only about 4–5% in buffer and plasma, which suggested that the conjugates have good plasma stability and the hydrolysis in both papain and lysosomes occurs mostly via enzymolysis. It was found that the spacers have important effect on the drug content, water solubility, drug release properties and cytotoxicity of Mel-OCM-chitosan conjugates. Cytotoxicity studies by MTT assay demonstrated that these conjugates had 52–70% of cytotoxicity against RPMI8226 cells in vitro as compared with free Mel, indicating the conjugates did not lose anti-cancer activity of Mel. Overall these studies indicated Mel-OCM-chitosan conjugates as potential prodrugs for cancer treatment.     

Quinazolino linked 4β-amidopodophyllotoxin conjugates regulate angiogenic pathway and control breast cancer cell proliferation

A series of new conjugates of quinazolino linked 4β-amidopodophyllotoxins 10aa–af and 10ba–bf were synthesized and evaluated for their anticancer activity against human pancreatic carcinoma (Panc-1) as well as breast cancer cell lines such as MCF-7 and MDA-MB-231 by employing MTT assay. Among these conjugates, some of them like 10bc, 10bd, 10be and 10bf exhibited high potency of cytotoxicity. Flow cytometric analysis showed that these conjugates arrested the cell cycle in the G2/M phase and caused the increase in expression of p53 and cyclin B1 protein with concomitant decrease in Cdk1 thereby suggesting the inhibitory action of these conjugates on mitosis. Interestingly, we observed a decrease in expression of proteins that control the tumor micro environment such as VEGF-A, STAT-3, ERK1/2, ERK-p, AKT-1 ser 473 phosphorylation in compounds treated breast cancer cells. Further, these effective conjugates have exhibited inhibitory action on integrin (αVβIII). Furthermore, the MCF-7 cells that were arrested and lost the proliferative capacity undergo mitochondrial mediated apoptosis by activation of caspases-9. Thus these conjugates have the potential to control breast cancer cell growth by effecting tumor angiogenesis and invasion.     

Design, synthesis and biological evaluation of N-phenylsulfonylnicotinamide derivatives as novel antitumor inhibitors

A series of novel N-phenylsulfonylnicotinamide derivatives (1-24) have been synthesized and evaluated as potential EGFR tyrosine kinase (TK) inhibitors. Among all the compounds, compound 10 (5-bromo-N-(4-chlorophenylsulfonyl)nicotinamide) showed the most potent growth inhibitory activity against EGFR TK and antiproliferative activity of MCF-7 cancer cell line in vitro, with IC(50) value of 0.09 and 0.07 μM. Docking simulation was performed to insert compound 10 into the EGFR TK active site to determine the probable binding model. Based on the preliminary results, compound 10 with potent inhibitory activity to tumor growth may be a potential anticancer agent.     

Selective tumor targeting by enhanced permeability and retention effect. Synthesis and antitumor activity of polyphosphazene-platinum (II) conjugates

Nanosized polyphosphazene-platinum (II) conjugates with a wide range of molecular weight from 24,000 to 115,000 were synthesized to study their tumor selectivity by enhanced permeability and retention (EPR) effect and their antitumor activity. It has been found from biodistribution study that the present polyphosphazene-Pt(II) conjugates exhibit high tumor selectivity by EPR effect with the tumor to tissue ratio (TTR) from 3.6 to 13 depending on the molecular size. These polymer conjugates have shown excellent in vivo antitumor activity against both murine and human cancer cell lines. In particular, xenograft trials of the conjugates have shown outstanding tumor inhibition effect on the stomach cancer cell line, YCC-3, which is one of the least responsive to the anticancer agents currently in clinical use, although the reason is not clearly explainable yet. The high in vivo activity seems to be attributed to the controlled-release of the antitumor active platinum (II) moiety, [GlyGluPt(dach] (dach=trans-(+/-)-1,2-diaminocyclohexane) from the phosphazene backbone by degradation in aqueous solution.     

Synthesis,biodistribution and antitumor activity of hematoporphyrin-platinum(II) conjugates

A new series of platinum(II) complexes of pegylated hematoporphyrin derivatives with controlled hydrophobic/hydrophilic balance were synthesized by introducing different kinds of poly(ethylene glycol) and amine ligands to the porphyrin ring. The antitumor activity of the porphyrin-platinum(II) conjugates was assayed in vitro and in vivo against leukemia L1210 cell line and various human tumor cell lines. The present complexes exhibited high antitumor activity and improved water solubility as well as considerable lipophilicity. In particular, complex 16 showed not only higher in vivo activity (T/C%=258) than cisplatin (T/C%=184) and carboplatin (T/C%=168), but also excellent solubility in water and organic solvent. The antitumor activity of complex 20 was superior to that of carboplatin against all human tumor cell lines tested. Moreover, some amphiphilic complexes (7 and 12) exhibited elevated tumor-localizing effect (tumor/muscle ratio>2).     

Design,synthesis, biological evaluation,structure-activity relationship,and toxicity of clinafloxacin-azole conjugates as novel antitubercular agents

Based on the advantages of azole molecules and fluoroquinolone drugs, we designed and synthesized 34 clinafloxacin-azole conjugates using fragment-based drug design and drug combination principles. The in vitro activities of the synthesized conjugates against Mycobacterium tuberculosis (H37Rv), Hela cell as well as Gram-positive and Gram-negative bacteria were assayed. The bioassay results revealed that most of the target molecules had anti-tuberculosis (anti-TB) activity, of which 14 compounds had very strong anti-TB activity [minimum inhibitory concentration (MIC)?<?2?μM]. In addition, the compounds with strong activity towards H37Rv had weak activity towards Gram-negative and Gram-positive bacteria, showing obvious selectivity towards H37Rv. Predicted toxicity data indicated that 27 molecules were less toxic or equivalent to that of the original drug (clinafloxacin). Especially, it is demonstrated that compound TM2l exhibited the strongest anti-TB activity (MIC?=?0.29?μM), low antibacterial activity, negligible toxicity, and good drug-likeness values, which can be considered as an ideal lead molecule for future optimization.     

Synthesis and biological evaluation of quinoxaline derivatives as specific c-Met kinase inhibitors

A series of novel quinoxaline derivatives were synthesized and evaluated for their inhibitory activity against c-Met kinase enzyme. Most of the tested compounds exhibited potent inhibitory activity. All the synthesized quinoxaline compounds were further examined against c-Met overexpressed human gastric cancer cell line (MKN-45), which showed good inhibitory activity. Among the synthesized compounds, compound 4 exhibited better tumor growth inhibition in the animal model study; we also confirmed its acceptable drug property and highly selective target activity.     

PEG-doxorubicin conjugates: influence of polymer structure on drug release, in vitro cytotoxicity, biodistribution, and antitumor activity

Polymer-drug conjugates (polymer therapeutics) are finding increasing use as novel anticancer agents. Here a series of poly(ethylene glycol) PEG-doxorubicin (Dox) conjugates were synthesized using polymers of linear or branched architecture (molecular weight 5000-20000 g/mol) and with different peptidyl linkers (GFLG, GLFG, GLG, GGRR, and RGLG). The resultant conjugates had a drug loading of 2.7-8.0 wt % Dox and contained <2.0% free drug (% total drug). All conjugates containing a GFLG linker showed approximately 30% release of Dox at 5 h irrespective of PEG molecular weight or architecture. The GLFG linker was degraded more quickly (approximately 57% Dox release at 5 h), and the other linkers more slowly (<16% release at 5 h), by lysosomal enzymes in vitro. In vitro there was no clear relationship between cytotoxicity toward B16F10 cells and the observed Dox release rate. All PEG conjugates were more than 10-fold less toxic (IC50 values > 2 microg/mL) than free Dox (IC50 value = 0.24 microg/mL). Biodistribution in mice bearing sc B16F10 tumors was assessed after administration of PEGs (5000, 10000, or 20000 g/mol) radioiodinated using the Bolton and Hunter reagent or PEG-Dox conjugates by HPLC. The 125I-labeled PEGs showed a clear relationship between Mw and blood clearance and tumor accumulation. The highest Mw PEG had the longest plasma residence time and consequently the greatest tumor targeting. The PEG-Dox conjugates showed a markedly prolonged plasma clearance and greater tumor targeting compared to free Dox, but there was no clear molecular weight-dependence on biodistribution. This was consistent with the observation that the PEG-Dox conjugates formed micelles in aqueous solution comprising 2-20 PEG-Dox molecules depending on polymer Mw and architecture. Although PEG-Dox showed greater tumor targeting than free Dox, PEG conjugation led to significantly lower anthracycline levels in heart. Preliminary experiments to assess antitumor activity against sc B16F10 in vivo showed the PEG5000linear (L)-GFLG-Dox and PEG10000branched (B)-GLFG-Dox (both 5 mg/kg Dox-equiv) to be the most active with T/C values of 146 and 143%, respectively. Free Dox did not show significant activity in this model (T/C = 121%). Dose escalation of PEG5000(L)-GFLG-Dox to 10 mg/kg Dox-equiv prolonged further animal survival (T/C = 161%). Using the Dox-sensitive model ip L1210 (where Dox displayed a T/C = 150% after single ip dose), the PEG5000(L)-GFLG-Dox displayed a maximum T/C of 141% (10 mg/kg Dox-equiv) using a once a day (x3) schedule. Further studies are warranted with PEG5000(L)-GFLG-Dox to determine its spectrum of antitumor activity and also the optimum dosing schedule before clinical testing.     

Antitumor agents 216. Synthesis and evaluation of paclitaxel-camptothecin conjugates as novel cytotoxic agents

Five conjugates (16-20) composed of a paclitaxel and a camptothecin derivative joined by an imine linkage were synthesized and evaluated as cytotoxic agents and as inhibitors of DNA topoisomerase I. All of the conjugates were potent inhibitors of tumor cell replication with improved activity relative to camptothecin. Significantly, compounds 16-18 were more active than paclitaxel and camptothecin against HCT-8 (colon adenocarcinoma) cell replication, and the spectrum of activity was different from a simple mixture of paclitaxel and camptothecin. All of the conjugates were significantly less potent than camptothecin as inhibitors of human topoisomerase I in vitro with 16, 18, and 19 showing only marginal activity at 50 microM. Based on activity against drug-resistant cell line replication, one could conclude that the conjugates are simply acting as 'weak taxanes', but the spectrum of activity, particularly against MCF-7 and HCT-8, strongly suggests that a novel mechanism of action has been achieved through conjugation.     

Inhibition of plasminogen activator inhibitor release in endothelial cell cultures by antisense oligodeoxyribonucleotides with a 5'-end lipophilic modification

A series of conjugates containing residues of lipophilic alcohols covalently bound to 5' end of oligodeoxyribonucleotides targeted against human plasminogen activator inhibitor (PAI-1) mRNA was synthesized via the oxathiaphospholane approach. The highest anti-PAI-1 activity in EA.hy 926 endothelial cell cultures was found for conjugates containing menthyl or heptadecanyl groups linked with an oligonucleotide complementary to a segment of human PAI-1 mRNA. The phosphodiester antisense oligonucleotides, which otherwise exhibit only limited anti-PAI-1 activity, were found to be more active than phosphorothioate oligonucleotides when conjugated to lipophilic alcohol residues. For menthyl conjugates an evidence of antisense mechanism of inhibition was found.     

Design, synthesis, single-crystal and preliminary antitumor activity of novel arenesulfonylimidazolidin-2-ones

Novel series of 1-(arenesulfonyl)imidazolidin-2-one (3a-i) and 1,3-bis(arenesulfonyl)imidazolidin-2-one (5a-i) have been synthesized and tested for their antitumor activity against 60 tumor cell lines taken from nine different organs. A significant inhibition for cancer cells was observed with series 5a-i compounds compared with the series 3a-i which showed a weak inhibition. Compounds 5a-i showed good inhibitory effect at the lung cancer HOP-92 and renal cancer CAKI-1 and UO-31 cell lines. Compound 5e showed remarkable broad-spectrum antitumor activity.     

Synthesis, biological evaluation and molecular docking studies of 1,3,4-thiadiazole derivatives containing 1,4-benzodioxan as potential antitumor agents

A series of 1,3,4-thiadiazole derivatives containing 1,4-benzodioxan (2a-2s) have been synthesized to screen for FAK inhibitory activity. Compound 2p showed the most potent biological activity against HEPG2 cancer cell line (EC(50)=10.28 μg/mL for HEPG2 and EC(50)=10.79 μM for FAK), which was comparable to the positive control. Docking simulation was performed to position compound 2p into the FAK structure active site to determine the probable binding model. The results of antiproliferative and Western-blot assay demonstrated that compound 2p possessed good antiproliferative activity against HEPG2 cancer cell line. Therefore, compound 2p with potent FAK inhibitory activity may be a potential anticancer agent against HEPG2 cancer cell.