Dependence Potential of the Synthetic Cannabinoids JWH-073, JWH-081, and JWH-210: In Vivo and In Vitro Approaches

Synthetic cannabinoids (CBs) such as the JWH series have caused social problems concerning their abuse liability. Because the JWH series produces euphoric and hallucinogenic effects, they have been distributed illegally under street names such as "Spice" and "Smoke". Many countries including Korea have started to schedule some of the JWH series compounds as controlled substances, but there are a number of JWH series chemicals that remain uncontrolled by law. In this study, three synthetic CBs with different binding affinities to the CB1 receptor (JWH-073, 081, and 210) and Delta9-tetrahydrocannabinol (Delta9-THC) were evaluated for their potential for psychological dependence. The conditioned place preference test (unbiased method) and self-administration test (fixed ratio of 1) using rodents were conducted. Ki values of the three synthetic cannabinoids were calculated as supplementary data using a receptor binding assay and overexpressed CB1 protein membranes to compare dependence potential with CB1 receptor binding affinity. All mice administered JWH-073, 081, or 210 showed significantly increased time spent at unpreferred space in a dose-dependence manner in the conditioned place preference test. In contrast, all tested substances except Delta9-THC showed aversion phenomenon at high doses in the conditioned place preference test. The order of affinity to the CB1 receptor in the receptor binding assay was JWH-210 > JWH-081 >> JWH-073, which was in agreement with the results from the conditioned place preference test. However, no change in self-administration was observed. These findings suggest the possibility to predict dependence potential of synthetic CBs through a receptor binding assay at the screening level.

Dependence Potential of Quetiapine: Behavioral Pharmacology in Rodents

Quetiapine is an atypical or second-generation antipsychotic agent and has been a subject of a series of case report and suggested to have the potential for misuse or abuse. However, it is not a controlled substance and is not generally considered addictive. In this study, we examined quetiapine's dependence potential and abuse liability through animal behavioral tests using rodents to study the mechanism of quetiapine. Molecular biology techniques were also used to find out the action mechanisms of the drug. In the animal behavioral tests, quetiapine did not show any positive effect on the experimental animals in the climbing, jumping, and conditioned place preference tests. However, in the head twitch and self-administration tests, the experimental animals showed significant positive responses. In addition, the action mechanism of quetiapine was found being related to dopamine and serotonin release. These results demonstrate that quetiapine affects the neurological systems related to abuse liability and has the potential to lead psychological dependence, as well.

Gene Expression Analysis for Statin-induced Cytotoxicity from Rat Primary Hepatocytes

Statins are competitive inhibitors of hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase and used most frequently to reduce plasma cholesterol levels and to decrease cardiovascular events. However, statins also have been reported to have undesirable side effects such as myotoxicity and hepatotoxicity associated with their intrinsic efficacy mechanisms. Clinical studies recurrently reported that statin therapy elevated the level of liver enzymes such as ALT and AST in patients suggesting possible liver toxicity due to statins. This observation has been drawn great attention since statins are the most prescribed drugs and statin-therapy was extended to a larger number of high-risk patients. Here we employed rat primary hepatocytes and microarray technique to understand underlying mechanism responsible for statin-induced liver toxicity on cell level. We isolated genes whose expressions were commonly modulated by statin treatments and examined their biological functions. It is of interest that those genes have function related to response to stress in particular immunity and defense in cells. Our study provided the basic information on cellular mechanism of statin-induced cytotoxicity and may serve for finding indicator genes of statin-induced toxicity in rat primary hepatocytes.

Effects of Mercuric Chloride on Gene Expression in NRK-52E Cells

Mercuric chloride, a model nephrotoxicant was used to elucidate time- and dose-dependent global gene expression changes associated with proximal tubular toxicity. Rat kidney cell lines NRK-52E cells were exposed for 2, 6 and 12 hours and with 3 different doses of mercuric chloride. Cell viability assay showed that mercuric chloride had toxic effects on NRK-52E cells causing 20% cell death (IC20) at 40micrometer concentration. We set this IC20 as high dose concentration and 1/5 and 1/25 concentration of LC20 were used as mid and low concentration, respectively. Analyses of microarray data revealed that 738 genes were differentially expressed (more than two-fold change and p<0.05) by low concentration of mercuric chloride at least one time point in NRK-52E cells. 317 and 2,499 genes were differentially expressed at mid and high concentration of mercuric chloride, respectively. These deregulated genes showed a primary involvement with protein trafficking (CAV2, CANX, CORO1B), detoxification (GSTs) and immunity and defense (HMOX1, NQO1). Several of these genes were previously reported to be up-regulated in proximal tubule cells treated with nephrotoxicants and might be aid in promoting the predictive biomarkers for nephrotoxicity.

Analysis of Gene Expression in Mouse Spinal Cord-derived Neural Precursor Cells During Neuronal Differentiation

The differentiation of neural precursor cells (NPCs) into neurons and astrocytes is a process that is tightly controlled by complicated and ill-defined gene networks. To extend our knowledge to gene networks, we performed a temporal analysis of gene expression during the differentiation (2, 4, and 8 days) of spinal cord-derived NPCs using oligonucleotide microarray technology. Out of 32,996 genes analyzed, 1878 exhibited significant changes in expression level (fold change>2, p<0.05) at least once throughout the differentiation process. These 1878 genes were classified into 12 groups by k-means clustering, based on their expression patterns. K-means clustering analysis revealed that the genes involved in astrogenesis were categorized into the clusters containing constantly upregulated genes, whereas the genes involved in neurogenesis were grouped to the cluster showing a sudden decrease in gene expression on Day 8. Functional analysis of the differentially expressedgenes indicated the enrichment of genes for Pax6- NeuroD signaling-TGFb-SMAD and BMP-SMAD-which suggest the implication of these genes in the differentiation of NPCs and, in particular, key roles for Nova1 and TGFBR1 in the neurogenesis/astrogenesis of mouse spinal cord.

High-concentration Epigallocatechin Gallate Treatment Causes Endoplasmic Reticulum Stress-mediated Cell Death in HepG2 Cells

Epigallocatechin gallate (EGCG), a well-known antioxidant molecule, has been reported to cause hepatotoxicity when used in excess. However, the mechanism underlying EGCG-induced hepatotoxicity is still unclear. To better understand the mode of action of EGCG-induced hepatotoxicity, we examined the effect of EGCG on human hepatic gene expression in HepG2 cells using microarrays. Analyses of microarray data revealed more than 1300 differentially expressed genes with a variety of biological processes. Upregulated genes showed a primary involvement with protein-related biological processes, such as protein synthesis, protein modification, and protein trafficking, while downregulated genes demonstrated a strong association with lipid transport. Genes involved in cellular stress responses were highly upregulated by EGCG treatment, in particular genes involved in endoplasmic reticulum (ER) stress, such as GADD153, GADD34, and ATF3. In addition, changes in genes responsible for cholesterol synthesis and lipid transport were also observed, which explains the high accumulation of EGCG-induced lipids. We also identified other regulatory genes that might aid in clarifying the molecular mechanism underlying EGCG-induced hepatotoxicity

Gene Expression Profiling in C57BL/6 Mice Treated with the Anorectic Drugs Sibutramine and Phendimetrazine and Their Mechanistic Implications

Recently, obesity has become a worldwide public health concern and the use of anorectic drugs has drastically increased. In this study, sibutramine and phendimetrazine, representative marketed anorectics, were repeatedly administered per os on a daily basis into C57BL/6 mice and the effects of these drugs on food intakes, body weight changes and gene expression profiles were monitored for up to following 7 days. Methamphetamine, which has a potent anorectic effect, was used as a positive control. Anorectic effects were sustained only for two days by phendimetrazine or methamphetamine, but for six days by sibutramine. The modulations of gene expressions in the hypothalamus and the striatum were investigated using microarrays on day 2 and day 7 post-administration, which corresponded to the anorectic period and a return of appetite respectively, for all three drugs tested. Differences in overall gene expression profiles in the stratum on day 2 for sibutramine and phendimetrazine seems to reflect difference between the two in terms of the onsets of drug tolerance. According to microarray findings, the Ankrd26 gene appears to have an important anorectic role, whereas the up-regulation of the olfaction system appeared to be involved in the drug tolerance of anorectics. The microarray data presented in this study demonstrates the usefulness of gene expression analysis for gathering information on the efficacy and safety of anorectic drugs.

cDNA Microarray Analysis of Differential Gene Expression in Gastric Cancer Cells Sensitive and Resistant to 5-Fluorouracil and Cisplatin / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Gastric cancer is one of the most prevalent cancers worldwide. 5-fluorouracil (5-FU) and cisplatin are the most commonly used drugs for the treatment of gastric cancer. However, a significant number of tumors often fail to respond to chemotherapy. MATERIALS AND METHODS: To better understand the molecular mechanisms underlying drug resistance in gastric cancer the gene expression in gastric cancer cells, which were either sensitive or resistant to 5-FU and cisplatin, were examined using cDNA microarray analysis. To confirm the differential gene expression, as determined using the microarray, semiquantitative RT-PCR was performed on a subset of differentially expressed cDNAs. RESULTS: 69 and 45 genes, which were either up-regulated (9 and 22 genes) or down-regulated (60 and 25 genes), were identified in 5-FU- and cisplatin-resistant cells, respectively. Several genes, such as adaptor-related protein complex 1 and baculoviral IAP repeat-containing 3, were up-regulated in both drug-resistant cell types. Several genes, such as the ras homolog gene family, tropomyosin, tumor rejection antigen, protein disulfide isomerase-related protein, melanocortin 1 receptor, defensin, cyclophilin B, dual specificity phosphatase 8 and hepatocyte nuclear factor 3, were down-regulated in both drug-resistant cell types. CONCLUSION: These findings show that cDNA microarray analysis can be used to obtain gene expression profiles that reflect the effect of anticancer drugs on gastric cancer cells. Such data may lead to the assigning of signature expression profiles of drug-resistant tumors, which may help predict responses to drugs and assist in the design of tailored therapeutic regimens to overcome drug resistance.

The Differential Gene Expression Profiles between Sensitive and Resistant Breast Cancer Cells to Adriamycin by cDNA Microarray / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Adriamycin(R) is one of the most commonly used drugs in the treatment of breast cancer. This study was performed to understand the molecular mechanisms of drug resistance in breast cancer cells. MATERIALS AND METHODS: We have analyzed the MCF-7 breast cell line and its adriamycin-resistant variants, MCF-7/ADR using human 10 K element cDNA microarrays. RESULTS: We defined 68 genes that were up-regulated (14 genes) or down-regulated (54 genes) in adriamycin resistant breast cancer cells. Several genes, such as G protein-coupled receptor kinase 5, phospholipase A2, guanylate cyclase 1, vimentin, matrix metalloproteinase 1 are up-regulated in drug resistant cells. Several genes, such as interferon, alpha-inducible protein 27, forkhead box M1, mitogen-activated protein kinase 6, regulator of mitotic spindle assembly 1 and tumor necrosis factor superfamily are down-regulated in adriamycin resistant cells. The altered expression of genes observed in microarray was verified by RT-PCR. CONCLUSION: These findings show that cDNA microarray analysis can be used to obtain gene expression profiles reflecting the effect of anticancer drugs on breast cancer cells. Such data may lead to the assigning of signature expression profiles of drug-resistant tumors which may help predict responses to drugs and assist in the design of tailored therapeutic regimens to overcome drug resistance.