IFN-stimulatory gene factor 15 (ISG15) is a ubiquitin-like protein, which is conjugated to many cellular proteins. However, its role in protein degradation is unclear. Here, we show that ISG15 is highly elevated and extensively conjugated to cellular proteins in many tumors and tumor cell lines. The increased levels of ISG15 in tumor cells were found to be associated with decreased levels of polyubiquitinated proteins. Specific knockdown of ISG15 expression using ISG15-specific small interfering RNA (siRNA) was shown to increase the levels of polyubiquitinated proteins, suggesting an antagonistic role of ISG15 in regulating ubiquitin-mediated protein turnover. Moreover, siRNA-mediated down-regulation of the major E2 for ISG15 (UbcH8), which blocked the formation of ISG15 protein conjugates, also increased the levels of polyubiquitinated proteins. Together, our results suggest that the ISG15 pathway, which is deregulated during tumorigenesis, negatively regulates the ubiquitin/proteasome pathway by interfering with protein polyubiquitination/ degradation. (Cancer Res 2006; 66(2): 921-8)

Low doses of anticancer drugs have been shown to enhance antitumor immune response and increase the efficacy of immunotherapy. The molecular basis for such effects remains elusive, although selective depletion of T regulatory cells has been demonstrated. In the current studies, we demonstrate that topotecan (TPT), a topoisomerase I-targeting drug with a well-defined mechanism of action, stimulates major histocompatibility complex class I (MHC I) expression in breast cancer cells through elevated expression/secretion of interferon-β (IFN-β) and activation of type I IFN signaling. First, we show that TPT treatment elevates the expression of both total and cell-surface MHC I in breast cancer cells. Second, conditioned media from TPT-treated breast cancer ZR-75-1 cells induce elevated expression of cell-surface MHC I in drug-naïve recipient cells, suggesting the involvement of cytokines and/or other secreted molecules. Consistently, TPT-treated cells exhibit elevated expression of multiple cytokines such as IFN-β, TNF-α, IL-6 and IL-8. Third, either knocking down the type I interferon receptor subunit 1 (IFNAR1) or addition of neutralizing antibody against IFN-β results in reduced MHC I expression in TPT-treated cells. Together, these results suggest that TPT induces increased IFN-β autocrine/paracrine signaling through type I IFN receptor, resulting in the elevated MHC I expression in tumor cells. Studies have also demonstrated that other chemotherapeutic agents (e.g. etoposide, cisplatin, paclitaxel and vinblastine) similarly induce increased IFN-β secretion and elevated MHC I expression. In addition, conditioned media from γ-irradiated donor cells are shown to induce IFN-β-dependent MHC I expression in unirradiated recipient cells. In the aggregate, our results suggest that many cancer therapeutics induce elevated tumor antigen presentation through MHC I, which could represent a common mechanism for enhanced antitumor immune response through T cell cytotoxicity during metronomic chemotherapy, as well as increased efficacy of combined chemo- (or radio-)/immuno-therapy.

BackgroundAntipsychotics have been linked to prolongation of the QT interval. However, little is known about the risk of ventricular arrhythmia (VA) and/or sudden cardiac death (SCD) associated with individual antipsychotic drug use. This study was designed to investigate the association between specific antipsychotic drugs and the risk of VA and/or SCD.Methods and ResultsWe conducted a case‐crossover study using a nation‐wide population‐based sample obtained from Taiwan's National Health Insurance Research Database. A total of 17 718 patients with incident VA and/or SCD were enrolled. Conditional logistic regression models were applied to examine the effects of antipsychotic drug use on the risk of VA/SCD during various case and control time windows of 7, 14, and 28 days. The effect of the potency of a human ether‐à‐go‐go‐related gene (hERG) potassium channel blockade was also assessed. Antipsychotic drug use was associated with a 1.53‐fold increased risk of VA and/or SCD. Antipsychotic drugs with increased risk included clothiapine, haloperidol, prochlorperazine, thioridazine, olanzapine, quetiapine, risperidone, and sulpiride. The association was significantly higher among those with short‐term use. Antipsychotics with a high potency of the hERG potassium channel blockade had the highest risk of VA and/or SCD.ConclusionUse of antipsychotic drugs is associated with an increased risk of VA and/or SCD. Careful evaluations of the risks and benefits of antipsychotic treatment are highly recommended.

Tumor cells are known to exhibit highly varied sensitivity to camptothecins (CPT; e.g., irinotecan and topotecan). However, the factors that determine CPT sensitivity/ resistance are largely unknown. Recent studies have shown that the ubiquitin-like protein, IFN-stimulated gene 15 (ISG15), which is highly elevated in many human cancers and tumor cell lines, antagonizes the ubiquitin/ proteasome pathway. In the present study, we show that ISG15 is a determinant for CPT sensitivity/resistance possibly through its effect on proteasome-mediated repair of topoisomerase I (TOP1)-DNA covalent complexes. First, short hairpin RNA-mediated knockdown of either ISG15 or UbcH8 (major E2 for ISG15) in breast cancer ZR-75-1 cells decreased CPT sensitivity, suggesting that ISG15 overexpression in tumors could be a factor affecting intrinsic CPT sensitivity in tumor cells. Second, the level of ISG15 was found to be significantly reduced in several tumor cells selected for resistance to CPT, suggesting that altered ISG15 regulation could be a significant determinant for acquired CPT resistance. Parallel to reduced CPT sensitivity, short hairpin RNAmediated knockdown of either ISG15 or UbcH8 in ZR-75-1 cells resulted in increased proteasomal degradation of CPT-induced TOP1-DNA covalent complexes. Taken together, these results suggest that ISG15, which interferes with proteasome-mediated repair of TOP1-DNA covalent complexes, is a potential tumor biomarker for CPT sensitivity.

SUMMARY Innate immunity controls pathogen replication and spread. Yet, certain pathogens, such as Hepatitis C Virus (HCV), escape immune elimination and establish persistent infections that promote chronic inflammation and related diseases. Whereas HCV regulatory proteins that attenuate anti-viral responses are known, those that promote inflammation and liver injury remain to be identified. Here we describe that transient expression of HCV RNA-dependent RNA polymerase (RdRp), NS5B, in mouse liver and human hepatocytes results in production of small RNA species that activate innate immune signaling via TBK1-IRF3 and NF-κB and induce cytokine production including type I interferons (IFN) and IL-6. NS5B-expression also results in liver damage.