Tumor Inhibition Assay Research Articles

Peptide-Guided System with Programmable Subcellular Translocation for Targeted Therapy and Bypassing Multidrug Resistance.

Nonselectivity and drug resistance are two major obstacles for cancer treatment. Although great advances have been made toward cell targeting or discovering novel delivery pathways, it is still desirable to simultaneously overcome the two hurdles for successful cancer theranostics. Herein, a peptide-guided system was tailored by modular integration of a cancer biomarker-specific peptide, a mitochondria-targeting motif and a cell toxin. Cell imaging analysis revealed that the dual-targeting peptide-drug conjugate (PDC) features in cancer cell-specific uptake, strong drug retention, and programmable intracellular translocation. Facilitated by in situ bond cleavage, PDC successfully diverted the toxic effect of nucleus-localized drug to mitochondria. Mechanism investigation demonstrated that the cell damage pathway of the drug was also transformed, which is beneficial to reverse drug resistance in cancer cells. The effectiveness of PDC for cancer therapy was further demonstrated by in vivo imaging and tumor inhibition assay. With intravenous injection, targeted accumulation in the tumor site, and tumor suppressing efficacy without side effects exhibit its perspective for cancer treatment. The dual-targeting peptide-drug conjugate featuring tailored transportation route highlights a promising and generally applicable way to enhance the overall therapeutic index of conventional anticancer drugs.

A novel anti-microtubule agent with carbazole and benzohydrazide structures suppresses tumor cell growth in vivo

BackgroundThe mitotic spindles are among the most successful targets of anti-cancer chemotherapy, and they still hold promise as targets for novel drugs. The anti-mitotic drugs in current clinical use, including taxanes, epothilones, vinca alkaloids, and halichondrins, are all microtubule-targeting agents. Although these drugs are effective for cancer chemotherapy, they have some critical problems; e.g., neurotoxicity caused by damage to neuronal microtubules, as well as innate or acquired drug resistance. To overcome these problems, a great deal of effort has been expended on development of novel anti-mitotics. MethodsWe identified novel microtubule-targeting agents with carbazole and benzohydrazide structures: N′-[(9-ethyl-9H-carbazol-3-yl)methylene]-2-methylbenzohydrazide (code number HND-007) and its related compounds. We investigated their activities against cancer cells using various methods including cell growth assay, immunofluorescence analysis, cell cycle analysis, tubulin polymerization assay, and tumor inhibition assay in nude mice. ResultsHND-007 inhibits tubulin polymerization in vitro and blocks microtubule formation and centrosome separation in cancer cells. Consequently, it suppresses the growth of various cancer cell lines, with IC50 values in the range 1.3–4.6μM. In addition, HND-007 can inhibit the growth of taxane-resistant cancer cells that overexpress P-glycoprotein. Finally, HND-007 can inhibit HeLa cell tumor growth in nude mice. Conclusions and general significanceTaken together, these findings suggest that HND-007 is a promising lead compound for development of novel anti-mitotic, anti-microtubule chemotherapeutic agents.

Simultaneous delivery of therapeutic antagomirs with paclitaxel for the management of metastatic tumors by a pH-responsive anti-microbial peptide-mediated liposomal delivery system

The roles of microRNAs (miRNAs) in the regulation of metastasis have been widely recognized in the recent years. Mir-10b antagomir (antagomir-10b) was shown to impede metastasis through the down-regulation of mir-10b; however, it could not stunt the growth of primary tumors. In this study we showed that the co-delivery of antagomir-10b with paclitaxel (PTX) by a novel liposomal delivery system modified with an anti-microbial peptide [D]-H6L9 (D-Lip) could significantly both hinder the migration of 4T1 cells and induce evident cellular apoptosis and cell death in the meantime. The histidines in the sequence of [D]-H6L9 allowed the peptide to get protonated under pH5.0 (mimicking the lysosome/endosome environment), and strong membrane lytic effect could thus be activated, leading to the escape of liposomes from the lysosomes and the decrease of of mir-10b expression. The in vivo and ex vivo fluorescence imaging showed that D-Lip could reach 4T1 tumors efficaciously. Incorporation of PTX did not influence the antagomir-10b delivery effect of D-Lip; for the in vivo tumor inhibition assay, compared with all the other groups, the combination of antagomir-10b and PTX delivered by D-Lip could prominently delay the growth of 4T1 tumors and reduce the lung metastases at the same time, and the expression of Hoxd10 in tumors was also significantly up-regulated. Taken together, these results demonstrated that D-Lip could act as a sufficient tool in co-delivering antagomir-10b and PTX.

Synthesis, physicochemical studies and potential applications of high-molecular-weight ferrocene-based poly(azomethine)ester and its soluble terpolymers

A new ferrocene-based poly(azomethine)ester (P) was synthesized via solution polycondensation of 1,1′-di(chlorocarboxyl) ferrocene with a novel preformed 4-((4-(4-(4-hydroxybenzylideneamino)phenoxy)phenylimino)methyl)phenol (SB). The condensation copolymers with good solubility were obtained using commercially available aliphatic (1,3-propan-diol, 1,6-hexan-diol and poly(dimethylsiloxane), hydroxyl-terminated (n = 550)) and aromatic ((1,1,1,3,3,3-hexaflouro)bisphenol propane and bisphenol A) along with the synthesized monomer (SB) and 1,1′-di(chlorocarboxyl) ferrocene. The obtained compounds were characterized by elemental analysis, FTIR and 1H NMR spectrophotometric techniques. The structure–property relation was studied in term of solubility measurements, WAXRD and UV–visible spectroscopic techniques. Average molecular weights, Mw obtained by laser light scattering method were found to be of the order 105 g/mol. Thermogravimetric curves along with polarized optical micrographs were also recorded. SEM–EDX and AFM techniques were employed to study the surface morphology of the synthesized material. Whereas the redox properties were explored using cyclic voltammetric investigations. The biological activity of the synthesized material was studied by utilizing various biological assays (antibacterical, brine shrimp cytotoxicity, antitumor potato disc, DPPH free radical scavenging and free radical induced oxidative DNA damage analysis). In case of brine shrimp (Artemia salina) lethality assay PSi and PF showed LD50 < 1 ppm indicating highly toxic behavior. PF resulted IC50 = 37.78 in tumor inhibition assay. The Schiff base (SB) was highly antioxidant with IC50 = 2.05 ppm and fully protected the plasmid DNA from damage. All the compounds studied were found potent antioxidants, highly cytotoxic and more importantly DNA protecting. The influence of aromatic and aliphatic segments on the properties of organometallic poly(azomethine)ester is discussed.

Identification of a New Cytotoxic T-Cell Epitope p675 of Human Telomerase Reverse Transcriptase

The identification of new tumor-associated antigens and epitopes is the prerequisite for cancer immunotherapy. The human telomerase catalytic subunit human telomerase reverse transcriptase (hTERT) is an attractive target for tumor therapy. The hTERT:540-548 peptide (p540, ILAKFLHWL) was previously identified as an HLA-A*0201-restricted T-cell epitope. However, other MHC-restricted epitopes in this antigen are urgently needed for an effective T-cell-based cancer treatment. The goal of this study was to identify other immunogenic peptides of hTERT restricted by HLA-A*0201 molecule, which accounts for about 15% of the Chinese population. Computer algorithms were used for epitope prediction, followed by analyzation on the plate of Discovery Studio 2.5. The candidate peptides were verified by T2-cell-binding assay, ELISPOT analysis, flow cytometry examination, and in vitro and in vivo tumor inhibition assay. Six peptides were predicted by computer algorithms. Of these peptides, p540 and p675 (LLGASVLGL) were identified with lower binding free energies than the other four peptides, binding to HLA-A*02 more strongly, and elicited stronger IFN-γ secretion of patients' peripheral blood mononuclear cells. The CTL clone against p675 elicited a significant cancer cell inhibition effect HLA-A*0201 restrictly. We identified p675 as a new HLA-A*0201-restricted T-cell epitope, and the CTL clone against it could suppress tumor growth effectively.

Synthesis and biological evaluations of some Schiff-base esters of ferrocenyl aniline and simple aniline

Abstract The synthesis and biological studies of some long chain esters containing Schiff bases and their ferrocenyl analogues were carried out. The 4-amino ferrocene was prepared by the reported method. Long chain esters were synthesized by the condensation of different aliphatic acids with the corresponding aldehyde. The esters were then reacted with aniline as well as with 4-aminophenyl ferrocene to give corresponding Schiff bases. All the synthesized compounds were analyzed by elemental, FTIR and proton NMR studies, were also investigated for a range of biological activities. Determined by crown gall tumor inhibition assay. Antioxidant and DNA protective activities were determined by DPPH free radical scavenging assay and OH radical induced oxidative DNA damage assay, respectively. Among all test compounds, o-hydroxy-p-n-octadecanoyloxy-benzylidine-p-ferrocenyl aniline (FA2.1: a ferrocene containing Schiff base) showed highest antitumor, DPPH free radical scavenging and DNA protective activities.

Measuring Melanoma-Specific Cytotoxic T Lymphocytes Elicited by Dendritic Cell Vaccines with a Tumor Inhibition Assay In Vitro

Improving cancer vaccines depends on assays measuring elicited tumor-specific T-cell immunity. Cytotoxic effector cells are essential for tumor clearance and are commonly evaluated using 51Cr release from labeled target cells after a short (4 hours) incubation with T cells. The authors used a tumor inhibition assay (TIA) that assesses the capacity of cytotoxic T lymphocytes (CTLs) to control the survival/growth of EGFP-labeled tumor cell lines. TIA was validated using CD8+ T cells primed in vitro against melanoma and breast cancer cells. TIA was then used to assess the CTL function of cultured CD8+ T cells isolated from patients with metastatic melanoma who underwent vaccination with peptide-pulsed CD34+ HPCs-derived DCs. After the DC vaccination, T cells from six of eight patients yielded CTLs that could inhibit the survival/growth of melanoma cells. The results of TIA correlated with killing of tumor cells in a standard 4-hour 51Cr release assay, yet TIA allowed detection of CTL activities that appeared marginal in the 51Cr release assay. Thus, TIA might prove valuable for measuring spontaneous and induced antigen-specific cytotoxic T cells.

Screening of Panamanian Medicinal Plants for Brine Shrimp Toxicity, Crown Gall Tumor Inhibition, Cytotoxicity and DNA Intercalation

ude extracts from 20 Panamanian plants used in traditional medicine have been screened for brine shrimp toxicity, tumor inhibition, DNA-intercalation and cytotoxicity tests in a clonogenic assay in V79 suspension culture gassed with air or nitrogen. Of the 20 plant extracts, 8 (40%) proved active in DNA intercalation. The most active were the methanol extracts of Bidens pilosa (root), Jatropha curcas (leaf), Pereskia bleo (whole plant), Isertia hypoleuca (leaf), Cyperus luzulae (root), Vernonia patens (leaf and stem), Alibertia edulis (stem) and Justicia graciliflora (aerial parts). Sixteen (80%) of the 20 methanol plant extracts were active in potato disc tumor inhibition assay; Cyperus luzulae (root), Justicia graciliflora (aerial parts), Pereskia bleo (whole plant) and Vernonia patens (leaf and stem) being the most active. Cytotoxicity in the clonogenic assay was observed only in three plant extracts: Cyperus luzulae, Piper auritum and Psychotria correae, whereas high brine shrimp toxicity (LC 50 &lt;100 p.p.m.) was observed in 13(65%) of the 20 plant extracts tested. These results appear to show a good correlation between the three bioassays: brine shrimp toxicity, tumor inhibition and DNA intercalation. The latter bioassay is rapid, economical, and can be used in activity guided fractionation of plant extracts.