Blockade of the dopamine (DA) transporter (DAT) is central to the effects of cocaine, and the DAT has been a focus of discovery research on cocaine-abuse medications. JHW007, an analogue of benztropine, binds to the DAT but is less effective than cocaine in producing psychomotor stimulant and subjective effects, and has been shown to antagonize these effects of cocaine in mice (Desai et al., J. Neurosci. 2005). We tested the hypothesis that the antagonism of cocaine by JHW 007 is mediated by a reduction in DA transmission. We compared the interactions of cocaine with JHW 007 or WIN 35,428 (a cocaine-like DAT inhibitor) on extracellular DA levels, measured by brain microdialysis in the shell of the nucleus accumbens of Swiss-Webster mice. All drugs dose dependently increased DA levels, with JHW 007 having a slower onset and lower effectiveness than cocaine or WIN 35428. A 4-hr pretreatment with JHW 007 (3–10 mg/kg i.p.) attenuated the elevation of DA levels produced by cocaine (10, 20, and 40 mg/kg). WIN 35428 (0.3, 1.0 mg/kg) administered with cocaine produced effects on DA levels that were not different from the addition of the effects of each drug alone. The present data suggest that antagonism of the behavioral effects of cocaine by JHW 007 results from a reduced cocaine-stimulated DA transmission. Results with WIN 35428 suggest that not all DAT blockers can reduce the neurochemical effects of cocaine, and further support consideration of JHW007 as a lead compound in the search for medications to treat cocaine abuse.
Gene expression profiling has the potential to produce new insights into complex biologic systems. To test the value of complement DNA arrays in identifying pathways involved in organ transplant rejection, we examined the gene expression profiles of rat heart allografts from recipients treated with or without immunosuppression to prevent acute allograft rejection.Heterotopic heart transplantation was performed using ACI or Lewis donors and Lewis recipients. Recipients were treated with tacrolimus (Tac) or cyclosporine (CsA) at the equivalent effective doses, and graft hearts were harvested on days 3, 5, and 7. A commercial microarray was used to measure gene expression levels of 588 genes in day 5 grafts. Selected genes were analyzed by reverse transcriptase-polymerase chain reaction.The expression levels of 118 genes were perturbed in the untreated allograft in comparison with the isograft control, of which 77 genes were categorized as candidate genes for Tac- or CsA-mediated immunosuppression or both, and 41 as genes associated with other pathways. Among the 77 candidate genes, 55 genes shared the same response to suppression by both drugs, including inducible nitric oxide synthase, interferon-gamma, and interferon regulatory factor 1. Drug-specific effects were observed in 22 genes: Fourteen genes were exclusively reversed by Tac and eight by CsA.Gene expression profiling reveals a large variety of genes affected during acute rejection, indicating that multiple metabolic pathways, including immune and nonimmune responses, are involved in the local graft rejection events. The differences and similarities of the gene expression profiles relative to the two immunosuppressants may provide more detailed therapeutic approaches for optimal immunosuppression.
K-Ras-induced lung cancer is a very common disease, for which there are currently no effective therapies. Because therapy directly targeting the activity of oncogenic Ras has been unsuccessful, a different approach for novel therapy design is to identify critical Ras downstream oncogenic targets. Given that oncogenic Ras proteins activate the transcription factor NF-kappaB, and the importance of NF-kappaB in oncogenesis, we hypothesized that NF-kappaB would be an important K-Ras target in lung cancer. To address this hypothesis, we generated a NF-kappaB-EGFP reporter mouse model of K-Ras-induced lung cancer and determined that K-Ras activates NF-kappaB in lung tumors in situ. Furthermore, a mouse model was generated where activation of oncogenic K-Ras in lung cells was coupled with inactivation of the NF-kappaB subunit p65/RelA. In this model, deletion of p65/RelA reduces the number of K-Ras-induced lung tumors both in the presence and in the absence of the tumor suppressor p53. Lung tumors with loss of p65/RelA have higher numbers of apoptotic cells, reduced spread, and lower grade. Using lung cell lines expressing oncogenic K-Ras, we show that NF-kappaB is activated in these cells in a K-Ras-dependent manner and that NF-kappaB activation by K-Ras requires inhibitor of kappaB kinase beta (IKKbeta) kinase activity. Taken together, these results show the importance of the NF-kappaB subunit p65/RelA in K-Ras-induced lung transformation and identify IKKbeta as a potential therapeutic target for K-Ras-induced lung cancer.
Despite proven therapy options for estrogen receptor (ER)-positive breast tumors, a substantial number of ER+ cancer patients exhibit relapse with associated metastasis. Loss of expression of RasGAPs leads to poor outcomes in several cancers, including breast cancer. Mining the TCGA breast cancer RNA-sequencing dataset revealed that low expression of the RasGAP DAB2IP was associated with a significant decrease in relapse-free survival in Luminal A breast cancer patients. Immunostaining demonstrated that DAB2IP loss occurred in grade 2 tumors and higher. Consistent with this, genes upregulated in DAB2IP-low Luminal A tumors were shared with more aggressive tumor subtypes and were associated with proliferation, metastasis, and altered ER signaling. Low DAB2IP expression in ER+ breast cancer cells was associated with increased proliferation, enhanced stemness phenotypes, and activation of IKK, the upstream regulator of the transcription factor NF-kB. Integrating cell-based ChIP-sequencing with motif analysis and TCGA RNA-seq data, we identified a set of candidate NF-kB target genes upregulated with loss of DAB2IP linked with several oncogenic phenotypes, including altered RNA processing. This study provides insight into mechanisms associated with aggressiveness and recurrence within a subset of the typically less aggressive Luminal A breast cancer intrinsic subtype.
