The mammalian target of rapamycin, mTOR, is a central regulator of cell growth. Its activity is regulated by Rheb, a Ras-like small guanosine triphosphatase (GTPase), in response to growth factor stimulation and nutrient availability. We show that Rheb regulates mTOR through FKBP38, a member of the FK506-binding protein (FKBP) family that is structurally related to FKBP12. FKBP38 binds to mTOR and inhibits its activity in a manner similar to that of the FKBP12-rapamycin complex. Rheb interacts directly with FKBP38 and prevents its association with mTOR in a guanosine 5'-triphosphate (GTP)-dependent manner. Our findings suggest that FKBP38 is an endogenous inhibitor of mTOR, whose inhibitory activity is antagonized by Rheb in response to growth factor stimulation and nutrient availability.

• Recurrent somatic mutations in MAP2K1 were identified in 33% of LCH lesions with wild-type BRAF. The mutant MAPK kinase 1 proteins activate ERK.• The ability of MAPK pathway inhibitors to suppress MAPK kinase and ERK phosphorylation in vitro was dependent on the specific LCH mutation.Langerhans cell histiocytosis (LCH) is a myeloproliferative disorder characterized by lesions composed of pathological CD207 1 dendritic cells with an inflammatory infiltrate.BRAFV600E remains the only recurrent mutation reported in LCH. In order to evaluate the spectrum of somatic mutations in LCH, whole exome sequencing was performed on matched LCH and normal tissue samples obtained from 41 patients. Lesions from other histiocytic disorders, juvenile xanthogranuloma, Erdheim-Chester disease, and RosaiDorfman disease were also evaluated. All of the lesions from histiocytic disorders were characterized by an extremely low overall rate of somatic mutations. Notably, 33% (7/21) of LCH cases with wild-type BRAF and none (0/20) with BRAFV600E harbored somatic mutations in MAP2K1 (6 in-frame deletions and 1 missense mutation) that induced extracellular signal-regulated kinase (ERK) phosphorylation in vitro. Single cases of somatic mutations of the mitogen-activated protein kinase (MAPK) pathway genes ARAF and ERBB3 were also detected. The ability of MAPK pathway inhibitors to suppress MAPK kinase and ERK phosphorylation in cell culture and primary tumor models was dependent on the specific LCH mutation. The findings of this study support a model in which ERK activation is a universal end point in LCH arising from pathological activation of upstream signaling proteins. (Blood. 2014;124(19):3007-3015)

The Hippo pathway regulates organ size and tumorigenesis in Drosophila and mammals and is altered in a variety of human cancers, yet it remains unclear if the Hippo pathway is of prognostic significance to cancer patients. Here we show that the key targets of Hippo signaling, transcriptional coactivators Yki and Yap, play a conserved role in promoting ovarian cancer in flies and humans, respectively. Whereas studies linking Yap to cancer in other tissues have focused on overall Yap levels, we show for the first time that subcellular levels of Yap show an exceptionally strong association with poor patient survival. Specifically, high levels of nuclear Yap (nYap), or low levels of cytoplasmic phosphorylated Yap (cpYap), associated with poor survival from ovarian cancer. Patients with both high nYap and low cpYap had ∼50% lower 5-year survival, and this combination is an independent prognostic marker for survival, with an exceptionally high hazard ratio of 7.8. We find that Yap2 is the predominantly expressed Yap isoform in both the ovarian surface epithelium (OSE) and epithelial ovarian cancers. Overexpression of Yap2 and phosphorylation-defective Yap2-5SA in immortalized OSE cells resulted in increased cell proliferation, resistance to cisplatin-induced apoptosis, faster cell migration, and anchorage-independent growth, whereas Yap knockdown resulted in increased sensitivity to cisplatin-induced apoptosis. Findings argue that the Hippo signaling pathway defines an important pathway in progression of ovarian cancer. Cancer Res; 70(21); 8517-25. ©2010 AACR.

In Saccharomyces cerevisiae, the Tap42-phosphatase complexes are major targets of the Tor kinases in the rapamycinsensitive signaling pathway. The immunosuppressive agent, rapamycin, induces a prompt activation of the Tap42-associated phosphatases, which is vitally important in Tor-mediated transcriptional regulation. However, the mechanism for the rapid phosphatase activation is poorly understood. In this study, we show that the Tap42-phosphatase complexes exist mainly on membrane structures through their association with Tor complex 1 (TORC1). Rapamycin abrogates this association and releases the Tap42-phosphatase complexes into the cytosol. Disassembly of the Tap42-phosphatase complexes occurs subsequently, following the release but at a much slower rate, presumably caused by Tap42 dephosphorylation. Release of the Tap42-phosphatase complexes from membrane structures also occurs when cells are deprived of nutrient. These findings suggest that the association of the Tap42-phosphatase complexes with TORC1 represents an important mechanism by which nutrient controls Tor signaling activity. In addition, our data support a model in which rapamycin acts not by inhibiting the kinase activity of Tor but by disrupting its interaction with downstream targets.

Current medications for drug abuse have had only limited success. Anti-addiction vaccines to elicit antibodies that block the pharmacological effects of drugs have great potential for treating drug abuse. We review the status for two vaccines that are undergoing clinical trials (cocaine and nicotine) and two that are still in pre-clinical development (methamphetamine and heroin). We also outline the challenges and ethical concerns for anti-addiction vaccine development and their use as future therapeutics.

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BACKGROUND Vaccines have treatment potential for methamphetamine (MA) addiction. We tested whether a conjugate vaccine against MA (succinyl-methamphetamine–keyhole limpet hemocyanin carrier protein; SMA-KLH) would generate MA antibodies and alter MA-induced behaviors. METHODS Mice were injected with SMA-KLH and received booster administrations 3-and 20-weeks later. Serum antibody titers reached peak levels by 4–6 weeks, remained at a modest level through 18-weeks, peaked again at 22-wks after the second boost, and were still elevated at 35-weeks. At 7 weeks, groups of vaccinated and non-vaccinated mice were administered one of three MA doses (1, 2, or 3 mg/kg) to assess locomotor activity. RESULTS Non-vaccinated mice showed dose-dependent effects of MA with hypolocomotion at the lowest dose and elevated activity levels at the highest dose. Both dose effects were reduced in SMA-KLH groups, particularly low dose-induced hypolocomotion at later times post MA administration. Separate groups of vaccinated and non-vaccinated mice were trained in MA place conditioning at 30-weeks with either 0 (vehicle) or 0.5 mg/kg MA. Although times spent in the MA-paired side did not differ between groups on Test vs. Baseline sessions, SMA-KLH mice conditioned with MA showed reduced conditioned approach behaviors and decreased conditioned activity levels compared to control groups. CONCLUSION These data suggest SMA-KLH attenuates the ability of MA to support place conditioning and reduces or delays its locomotor effects. Overall, results support SMA-KLH as a candidate MA vaccine.

The trichorhinophalangeal syndrome 1 (TRPS-1) gene is a novel GATA transcription factor family member. Previously, using a gene expression profiling and immunohistochemistry (IHC) screen, we identified TRPS-1 as a highly prevalent gene in breast cancer (BC), expressed in >90% of estrogen receptor alpha (ERα)(+) and ERα(-) BC subtypes. TRPS-1 was also shown to be expressed in prostate cancer where it was shown to play a proapoptotic function during androgen withdrawal possibly through regulating antioxidant metabolism. The role of TRPS-1 and its prognostic significance in hormone-dependent and hormone-independent BC however is not known. In this study, we developed a new quantitative IHC (qIHC) method to further study TRPS-1 as a marker and possible prognostic indicator in BC. By using this method, a quantitative parameter for TRPS-1 expression called a quick score (QS) was derived from the measured labeling index and mean optical density after IHC and applied to a set of 152 stage II/III BC patients from 1993 to 2006 who did not receive preoperative chemotherapy. Associations between QS and tumor characteristics were evaluated using the Kruskal-Wallis test. A wide range of TRPS-1 QS was found among the sample set with higher TRPS-1 QS significantly associated with tumor ERα (p = 0.023 for QS and p = 0.028 for Allred score), progesterone receptor (p = 0.009), and GATA-3 (p < 0.0001). TRPS-1 QS was also positively associated with HER2 status (p = 0.026). Further analysis of different ductal structures in ten BC cases revealed that TRPS-1 expression was expressed at low levels in the remaining normal ducts and in areas of usual ductal hyperplasia but showed marked increase in expression in ductal carcinoma in situ and invasive carcinoma lesions in the tissue. An analysis of TRPS-1 expression in association with overall survival in the 152 stage II/III sample set also revealed that TRPS-1 QS (≥4.0) was significantly associated with improved survival (p = 0.0165). Patients with TRPS-1 QS <4 had a hazard ratio of 2 (p = 0.019) after univariate Cox proportional hazards analysis. In summary, this new qIHC approach was found to reveal critical differences in TRPS-1 expression in primary BC samples and found that it is a promising prognostic marker that should be further evaluated as a possible tumor suppressor gene facilitating improved survival in different subtypes of BC.