Inhibition of Protein Phosphatase 2A (PP2A) Prevents Mcl-1 Protein Dephosphorylation at the Thr-163/Ser-159 Phosphodegron,Dramatically Reducing Expression in Mcl-1-amplified Lymphoma Cells

Abundant, sustained expression of prosurvival Mcl-1 is an important determinant of viability and drug resistance in cancer cells. The Mcl-1 protein contains PEST sequences (enriched in proline, glutamic acid, serine, and threonine) and is normally subject to rapid turnover via multiple different pathways. One of these pathways involves a phosphodegron in the PEST region, where Thr-163 phosphorylation primes for Ser-159 phosphorylation by glycogen synthase kinase-3. Turnover via this phosphodegron-targeted pathway is reduced in Mcl-1-overexpressing BL41-3 Burkitt lymphoma and other cancer cells; turnover is further slowed in the presence of phorbol ester-induced ERK activation, resulting in Mcl-1 stabilization and an exacerbation of chemoresistance. The present studies focused on Mcl-1 dephosphorylation, which was also found to profoundly influence turnover. Exposure of BL41-3 cells to an inhibitor of protein phosphatase 2A (PP2A), okadaic acid, resulted in a rapid increase in phosphorylation at Thr-163 and Ser-159, along with a precipitous decrease in Mcl-1 expression. The decline in Mcl-1 expression preceded the appearance of cell death markers and was not slowed in the presence of phorbol ester. Upon exposure to calyculin A, which also potently inhibits PP2A, versus tautomycin, which does not, only the former increased Thr-163/Ser-159 phosphorylation and decreased Mcl-1 expression. Mcl-1 co-immunoprecipitated with PP2A upon transfection into CHO cells, and PP2A/Aα knockdown recapitulated the increase in Mcl-1 phosphorylation and decrease in expression. In sum, inhibition of PP2A prevents Mcl-1 dephosphorylation and results in rapid loss of this prosurvival protein in chemoresistant cancer cells.     

Bone microenvironment and androgen status modulate subcellular localization of ErbB3 in prostate cancer cells

ErbB-3, an ErbB receptor tyrosine kinase, has been implicated in the pathogenesis of several malignancies, including prostate cancer. We found that ErbB-3 expression was up-regulated in prostate cancer cells within lymph node and bone metastases. Despite being a plasma membrane protein, ErbB-3 was also detected in the nuclei of the prostate cancer cells in the metastatic specimens. Because most metastatic specimens were from men who had undergone androgen ablation, we examined the primary tumors from patients who have undergone hormone deprivation therapy and found that a significant fraction of these specimens showed nuclear localization of ErbB3. We thus assessed the effect of androgens and the bone microenvironment on the nuclear translocation of ErbB-3 by using xenograft tumor models generated from bone-derived prostate cancer cell lines, MDA PCa 2b, and PC-3. In subcutaneous tumors, ErbB-3 was predominantly in the membrane/cytoplasm; however, it was present in the nuclei of the tumor cells in the femur. Castration of mice bearing subcutaneous MDA PCa 2b tumors induced a transient nuclear translocation of ErbB-3, with relocalization to the membrane/cytoplasm upon tumor recurrence. These findings suggest that the bone microenvironment and androgen status influence the subcellular localization of ErbB-3 in prostate cancer cells. We speculate that nuclear localization of ErbB-3 may aid prostate cancer cell survival during androgen ablation and progression of prostate cancer in bone.     

Effect of different thermal conditions on the pre-imaginal biology of Culex apicinus (Philippi, 1865) (Diptera: Culicidae).

The time of development and survival of the pre-imaginal period and the adult body size of Culex apicinus were analyzed in individuals reared from the 1st instar larva to adult emergence under laboratory and field conditions. In the laboratory, insects were exposed to three constant temperatures (15, 20, and 25 degrees C) and a photoperiod of 14:10 (L:D). In the field, temperature and photoperiod were not manipulated; during the study period water temperature ranged between 15.5 and 24.2 degrees C, and photoperiod changed from 13:11 to 14:10 (L:D). Survival to the imaginal stage at 15 degrees C (85.4%) and in the field (88.8%) was higher than that at 20 degrees C (45.8%) and 25 degrees C (8.3%). Based on femur and wing length measurements, the mosquitoes reared under field conditions (mean water temperature = 20.5 degrees C) were smaller than those laboratory-reared, even when the time of development in the field (29.5 - 36 days) was longer than that observed at 15 degrees C (24 days), 20 degrees C (15 days), and 25 degrees C (11 days). Male emergence was earlier than that of females (protandry) only in the field. Although laboratory conditions included a photoperiod and temperature range comparable with the observed values in the field, the disagreements between field and laboratory results suggest that the characteristics examined in this work could be affected by the joint variation of several factors not controlled in field conditions, such as photoperiod, temperature regime, and/or food quality of larval habitat.     

Phosphorus Availability Determines the Response of Tundra Ecosystem Carbon Stocks to Nitrogen Enrichment

Northern permafrost soils contain important carbon stocks. Here we report the long-term response of carbon stocks in high Arctic dwarf shrub tundra to short-term, low-level nutrient enrichment. Twenty years after experimental nitrogen addition, carbon stocks in vegetation and organic soil had almost halved. In contrast, where phosphorus was added with nitrogen, carbon storage increased by more than 50%. These responses were explained by changes in the depths of the moss and organic soil layers. Nitrogen apparently stimulated decomposition, reducing carbon stocks, whilst phosphorus and nitrogen co-stimulated moss productivity, increasing organic matter accumulation. The altered structure of moss and soil layers changed soil thermal regimes, which may further influence decomposition of soil carbon. If climate warming increases phosphorus availability, any increases in nitrogen enrichment from soil warming or expanding human activity in the Arctic may result in increased carbon sequestration. Where phosphorus is limiting in tundra areas, however, nitrogen enrichment may result in carbon loss.     

Quantifying the hydrodynamic stress for bioprocesses

Hydrodynamic stress is an influential physical parameter for various bioprocesses, affecting the performance and viability of the living organisms. However, different approaches are in use in various computational and experimental studies to calculate this parameter (including its normal and shear subcomponents) from velocity fields without a consensus on which one is the most representative of its effect on living cells. In this letter, we investigate these different methods with clear definitions and provide our suggested approach which relies on the principal stress values providing a maximal distinction between the shear and normal components. Furthermore, a numerical comparison is presented using the computational fluid dynamics simulation of a stirred and sparged bioreactor. It is demonstrated that for this specific bioreactor, some of these methods exhibit quite similar patterns throughout the bioreactor—therefore can be considered equivalent—whereas some of them differ significantly.     

Cost-Effectiveness of Buprenorphine and Naltrexone Treatments for Heroin Dependence in Malaysia

Aims

To aid public health policymaking, we studied the cost-effectiveness of buprenorphine, naltrexone, and placebo interventions for heroin dependence in Malaysia.

Design

We estimated the cost-effectiveness ratios of three treatments for heroin dependence. We used a microcosting methodology to determine fixed, variable, and societal costs of each intervention. Cost data were collected from investigators, staff, and project records on the number and type of resources used and unit costs; societal costs for participants’ time were estimated using Malaysia’s minimum wage. Costs were estimated from a provider and societal perspective and reported in 2004 US dollars.

Setting

Muar, Malaysia.

Participants

126 patients enrolled in a randomized, double-blind, placebo-controlled clinical trial in Malaysia (2003–2005) receiving counseling and buprenorphine, naltrexone, or placebo for treatment of heroin dependence.

Measurements

Primary outcome measures included days in treatment, maximum consecutive days of heroin abstinence, days to first heroin use, and days to heroin relapse. Secondary outcome measures included treatment retention, injection drug use, illicit opiate use, AIDS Risk Inventory total score, and drug risk and sex risk subscores.

Findings

Buprenorphine was more effective and more costly than naltrexone for all primary and most secondary outcomes. Incremental cost-effectiveness ratios were below $50 for primary outcomes, mostly below $350 for secondary outcomes. Naltrexone was dominated by placebo for all secondary outcomes at almost all endpoints. Incremental treatment costs were driven mainly by medication costs, especially the price of buprenorphine.

Conclusions

Buprenorphine appears to be a cost-effective alternative to naltrexone that might enhance economic productivity and reduce drug use over a longer term.     

Molecular genetic analysis of an SNF2/brahma-related gene in Tetrahymena thermophila suggests roles in growth and nuclear development

We used a reverse genetic approach to identify three members of the SNF2 superfamily of chromatin remodeling genes in the ciliated protozoan Tetrahymena thermophila in order to investigate possible functions of ATP-dependent chromatin remodeling factors in growth and nuclear development. Comparative sequence analysis of the gene product of the Tetrahymena brahma-related gene (TtBRG1) indicates it is a member of the SNF2/BRM subgroup of the SNF2 superfamily. Northern analysis suggests that TtBRG1 has roles in growth and nuclear development in Tetrahymena. Indirect immunofluorescence analysis during nuclear development indicates that TtBrg1p localizes to both the parental and developing macronucleus of Tetrahymena during the time period corresponding to genome rearrangements. We generated germ line knockout heterokaryons for TtBRG1 and demonstrated that expression of the gene is required to complete nuclear development of Tetrahymena. In addition, the formation of distinct Pdd1p-containing structures is disturbed during the late stages of conjugation in TtBRG1 germ line knockout heterokaryons. We discuss these results in light of possible roles of SNF2-related proteins in growth and nuclear development of Tetrahymena.     

Human Papillomavirus in Head and Neck Squamous Cell Carcinoma of Unknown Primary Is a Common Event and a Strong Predictor of Survival

Background

The purpose of this study was to examine the prevalence of human papillomavirus (HPV) in patients with head and neck squamous cell carcinoma of unknown primary (CUP).

Methods

All patients diagnosed with and treated for CUP between January 1, 2000, and June 1, 2011, at two Danish medical centers were included. All patients received a thorough diagnostic work-up, including FDG-PET, before being diagnosed as CUP. We determined the HPV status in all patients using a combination of HPV DNA PCR and p16 stain. In addition, clinical information on the study patients was retrieved from clinical records.

Results

Of the identified 60 patients with CUP, 13 were shown to be positive for HPV DNA, amounting to 22% of the study population. In addition, we were able to show a clear disease-free and overall-survival benefit in the HPV-positive group, with a hazard ratio of 0.16 (95% CI: 0.038–0.67) for over-all survival. This survival benefit was also apparent when adjusted for advanced age in a multivariate Cox regression analysis.

Conclusion

A fairly large percentage of CUP cases are HPV-related, and because this is related to both the location and prognosis, we recommend HPV testing as part of the diagnostic work-up.