AIMTo prepare 8-chloro-adenosine (8-Cl-A) long circulation liposomes with high entrapped efficiency and prolonged action-time of 8-Cl-A in vivo.

METHODSTo prepare 8-Cl-A long circulation liposomes of nanometer size by improved multiple emulsion. The entrapped efficiency, size and size distribution of 8-Cl-A long circulation liposomes were determined, and its pharmacokinetics in rats was evaluated.

RESULTSThe entrapped efficiency of 8-Cl-A long circulation liposomes was 62.70% and mean diameter of the liposomes was 79.9 nm. The pharmacokinetics studies indicated that 8-Cl-A long circulation liposomes showed higher drug concentration and larger AUC values than that of 8-Cl-A after iv to rats.

CONCLUSION8-Cl-A long circulation liposomes could prolong the action-time of 8-Cl-A in vivo.

2-Chloroadenosine ; administration & dosage ; analogs & derivatives ; pharmacokinetics ; Animals ; Antineoplastic Agents ; administration & dosage ; pharmacokinetics ; Area Under Curve ; Delayed-Action Preparations ; Liposomes ; Male ; Nanostructures ; Particle Size ; Rats ; Rats, Wistar

Previous reports raised question as to whether 8-chloro-cyclic adenosine 3,5-monophosphate (8-Cl-cAMP) is a prodrug for its metabolite, 8-Cl-adenosine which exerts growth inhibition in a broad spectrum of cancer cells. The present study was carried out to clarify overall cellular affects of 8-Cl-cAMP and 8-Cl-adenosine on SK-N-DZ human neuroblastoma cells by ystematically characterizing gene expression using radioactive human cDNA microarray. Microarray was prepared with PCR-amplified cDNA of 2,304 known genes spotted on nylon membranes, employing (1)P-labeled cDNAs of SK-N-DZ cells as a probe. the expression levels of approximately 100 cDNAs, representing about 8% of the total DNA elements on the array, were altered in 8-Cl-adenosine- or 8-Cl-cAMP-treated cells, respectively. The genome-wide expression of the two samples exhibited partial overlaps; different sets of up-regulated genes but the same set of down-regulated genes. 8-Cl-adenosine treatment up- egulated genes involved in differentiation and development (LIM protein, connexin 26, neogenin, neurofilament triplet L protein and p21( WAF1/CIP1)) and immune response such as natural killer cells protein 4, and down-regulated ones involved in proliferation and transformation (transforming growth factor-beta, DYRK2, urokinase-type plasminogen activator and proteins involved in transcription and translation) which were in close parallel with those by 8-Cl-cAMP. Our results indicated that the two drugs shared common genomic pathways for the down-regulation of certain genes, but used distinct pathways for the up-regulation of different gene clusters. Based on the findings, we suggest that the anti-cancer activity of 8-Cl-cAMP results at least in part through 8-Cl-adenosine. Thus, the systematic use of DNA arrays can provide insight into the dynamic cellular pathways involved in anticancer activities of chemotherapeutics.
2-Chloroadenosine/*analogs & derivatives/chemistry/*pharmacology ; 8-Bromo Cyclic Adenosine Monophosphate/*analogs & derivatives/chemistry/*pharmacology ; Antineoplastic Agents/chemistry/*pharmacology ; Blotting, Western ; *Gene Expression Profiling ; Gene Expression Regulation, Neoplastic/*drug effects ; Genome, Human ; Human ; Neuroblastoma/*genetics ; Oligonucleotide Array Sequence Analysis ; Reproducibility of Results ; Tumor Cells, Cultured ; Up-Regulation/drug effects

OBJECTIVETo study the effect of 8-chloro-adenosine (8-Cl-Ado) on the sensitivity of human hepatoma and breast cancer cell lines to TRAIL-induced apoptosis in vitro and its mechanisms.

METHODSRecombinant soluble TRAIL (rsTRAIL) or 8-Cl-Ado was used to treat hepatoma cell line BEL-7402 and breast cancer cell line MCF-7 in vitro. MTT assay was used to evaluate cell viability. The effect of cotreatment with rsTRAIL and 8-Cl-Ado was analyzed. NF-kappaB activity reporter plasmid was designed to measure the activity of transcription factor NF-kappaB. After transient transfection with the reporter plasmid, which contains NF-kappaB-responsive elements, into the cell lines, cells were treated with rsTRAIL and/or 8-Cl-Ado, then the activity of the reporter gene luciferase was determined. Different kinds of caspase inhibitors were used to measure the effect of caspases in the rsTRAIL and/or 8-Cl-Ado induced apoptosis.

RESULTS8-Cl-Ado could greatly enhance sensitivity of BEL-7402 and MCF-7 cells to reTRAIL. Treatment with 8-Cl-Ado and rsTRAIL inactivated transcription factor NF-kappaB and induced apoptosis in BEL-7402, but not in MCF-7. Caspase family inhibitor could not prevent apoptosis induced by 8-Cl-Ado and rsTRAIL in BEL-7402 cells, however, it could block apoptosis in MCF-7 cells, indicating that two different apoptosis pathways in MCF-7 and BEL-7402 might exist, one was caspase dependent and the other caspase independent. Moreover, all of the inhibitors of caspse-3, -8 and -9 could not block apoptosis induced by the co-treatment.

CONCLUSION8-chloro-adenosine can enhance the sensitivity of human hepatoma cell line BEL-7402 and breast cancer cell line MCF-7 to rsTRAIL, even though MCF-7 is TRAIL-resistant. 8-Cl-Ado combined with rsTRAIL can trigger different signal pathways in MCF-7 and BEL-7402, which are caspase dependent and independent, respectively.

2-Chloroadenosine ; analogs & derivatives ; pharmacology ; Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; pharmacology ; Breast Neoplasms ; pathology ; Carcinoma, Hepatocellular ; pathology ; Cell Line, Tumor ; Humans ; Liver Neoplasms ; pathology ; Membrane Glycoproteins ; pharmacology ; NF-kappa B ; metabolism ; TNF-Related Apoptosis-Inducing Ligand ; Transfection ; Tumor Necrosis Factor-alpha ; pharmacology