Human Merkel cell polyomavirus infection II. MCV is a common human infection that can be detected by conformational capsid epitope immunoassays

Merkel cell polyomavirus (MCV) is a newly‐discovered human tumor virus found in ～80% of Merkel cell carcinoma (MCC). The rate of MCV infection among persons without MCC is unknown. We developed a MCV virus‐like particle (VLP) enzyme‐linked immunoassay (EIA) that does not cross‐react with human BK or murine polyomaviruses. Peptide mapping of the MCV VP1 gene and immunoblotting with denatured MCV VLP are less sensitive than the MCV EIA in detecting MCV antibodies suggesting antibody reactivity in this assay primarily targets conformational but not linear epitopes. Among MCC patients, all 21 (100%) patients tested with MCV‐positive tumors had high serum MCV IgG but not high MCV IgM levels. Only 3 of 6 (50%) MCC patients with MCV‐negative tumors were positive for MCV antibodies. Sera from most adults, including 107 of 166 (64%) blood donors, 63 of 100 (63%) commercial donors and 37 of 50 (74%) systemic lupus erythematosus patients, show evidence for prior MCV exposure. Age‐specific MCV prevalence was determined by examining a cross‐sectional distribution of 150 Langerhans cell histiocytosis (an unrelated neoplasm) patient sera. MCV prevalence increases from 50% among children age 15 years or younger to 80% among persons older than 50 years. We did not find evidence for vertical transmission among infants. Although past exposure to MCV is common among all adult groups, MCC patients have a markedly elevated MCV IgG response compared with control patients. Our study demonstrates that MCV is a widespread but previously unrecognized human infection. © 2009 UICC     

Mocetinostat (MGCD0103): a review of an isotype-specific histone deacetylase inhibitor

INTRODUCTION: HDAC inhibitors (HDACIs) have the potential to restore gene expression and display antitumor effects in vitro. As single agents, HDACIs have clinical activity in lymphoma. In myeloid leukemias, combinations of DNA methylation inhibitors and HDACIs are promising. Other combinations are being studied in solid tumors. AREAS COVERED: This article covers basic information and an update on preclinical and clinical experience with the oral isotype-selective HDACI MGCD0103 (mocetinostat) in hematological malignancies and solid tumors. It also examines data concerning MGCD0103 from recent conferences and articles through to November 2010, including new data regarding responses in lymphoma and toxicities. EXPERT OPINION: MGCD0103 is well-tolerated and exhibits favorable pharmacokinetic and pharmacodynamic profiles, demonstrating target inhibition and clinical responses. It induces cell death and autophagy, synergizes with proteasomal inhibitors and affects non-histone targets, such as microtubules. In 2008, new patient enrollment in trials was temporarily suspended due to potential cardiac complications. This restriction was lifted in 2009 as no correlation between MGCD0103 exposure and pericardial effusions was found. New patient enrollment in MGCD0103 clinical trials requires the exclusion of patients diagnosed with significant cardiac abnormalities prior to enrollment. Clinical and pharmacodynamic data support a three-times-weekly administration at a 90 mg fixed dose. MGCD0103 displays promising antitumor activity in several hematological diseases.     

Drug sensitivity prediction by CpG island methylation profile in the NCI-60 cancer cell line panel

Aberrant promoter hypermethylation and associated gene silencing are epigenetic hallmarks of tumorigenesis. It has been suggested that aberrant DNA methylation can affect the sensitivity of cancers to antineoplastic agents by altering expression of genes critical to drug response. To study this issue, we used bisulfite PCR to assess DNA methylation of 32 promoter-associated CpG islands in human cancer cell lines from the National Cancer Institute (NCI) drug-screening panel (NCI-60 panel). The frequency of aberrant hypermethylation of these islands ranged from 2% to 81% in NCI-60 cancer cells, and provided a database that can be analyzed for the sensitivity to approximately 30,000 drugs tested in this panel. By correlating drug activity with DNA methylation, we identified a list of methylation markers that predict sensitivity to chemotherapeutic drugs. Among them, hypermethylation of the p53 homologue p73 and associated gene silencing was strongly correlated with sensitivity to alkylating agents. We used small interfering RNA to down-regulate p73 expression in multiple cell lines, including the resistant cell lines TK10 (renal cancer) and SKMEL28 (melanoma). Down-regulating p73 substantially increased sensitivity to commonly used alkylating agents, including cisplatin, indicating that epigenetic silencing of p73 directly modulates drug sensitivity. Our results confirm that epigenetic profiles are useful in identifying molecular mediators for cancer drug sensitivity (pharmaco-epigenomics).     

Homeostatic regulation of h‐conductance controls intrinsic excitability and stabilizes the threshold for synaptic modification in CA1 neurons

Key points

• We determined the contribution of the hyperpolarization‐activated cationic (h) current (I _h) to the homeostatic regulation of CA1 pyramidal cells in vitro using chronic treatments (48 h) that either increase (picrotoxin) or decrease (kynurenate) neuronal activity.
• The h‐conductance was found to be up‐ or down‐regulated following chronic activity enhancement or activity deprivation, respectively. This bidirectional plasticity of I _h was found to subsequently alter both apparent input resistance and intrinsic neuronal excitability.
• Bidirectional homeostatic plasticity of I _h also determined EPSP waveform and EPSP summation tested at 5–30 Hz.
• Long‐term synaptic modification induced by repetitive stimulation of the Schaffer collaterals was found to be constant across treatments in the presence of I _h but not when I _h was blocked pharmacologically. Thus, bidirectional homeostatic regulation of I _h stabilizes induction of long‐term synaptic modification in CA1 pyramidal neurons that depends on EPSP summation.

Abstract

The hyperpolarization‐activated cationic (h) current is a voltage‐shock absorber, highly expressed in the dendrites of CA1 pyramidal neurons. Up‐regulation of I _h has been reported following episodes of intense network activity but the effect of activity deprivation on I _h and the functional consequence of homeostatic regulation of I _h remain unclear. We determined here the contribution of I _h to the homeostatic regulation of CA1 pyramidal cell excitability. Intrinsic neuronal excitability was decreased in neurons treated for 2–3 days with the GABA_A channel blocker picrotoxin (PiTx) but increased in neurons treated (2–3 days) with the glutamate receptor antagonist kynurenate (Kyn). Membrane capacitance remained unchanged after treatment but the apparent input resistance was reduced for PiTx‐treated neurons and enhanced for Kyn‐treated neurons. Maximal I _h conductance was up‐regulated after chronic hyperactivity but down‐regulated following chronic hypoactivity. Up‐regulation of I _h in PiTx‐treated cultures was found to accelerate EPSP kinetics and reduce temporal summation of EPSPs whereas opposite effects were observed in Kyn‐treated cultures, indicating that homeostatic regulation of I _h may control the induction of synaptic modification depending on EPSP summation. In fact, stimulation of the Schaffer collaterals at 3–10 Hz induced differential levels of plasticity in PiTx‐treated and Kyn‐treated neurons when I _h was blocked pharmacologically but not in control conditions. These data indicate that homeostatic regulation of I _h normalizes the threshold for long‐term synaptic modification that depends on EPSP summation. In conclusion, bidirectional homeostatic regulation of I _h not only controls spiking activity but also stabilizes the threshold for long‐term potentiation induced in CA1 pyramidal neurons by repetitive stimulation.

Abbreviations

BCM

Bienenstock–Cooper–Monroe rule

fEPSP

field EPSP

I_h

hyperpolarization‐activated cationic current

Kyn

kynurenate

LTD

long‐term depression

LTP

long‐term potentiation; PiTx, picrotoxin

     

Use of an innovative, affordable, and open-source short message service-based tool to monitor malaria in remote areas of Uganda

Quality health management requires timely and accurate data, and paper-based reporting does not fill this role adequately. The introduction of malaria rapid diagnostic tests and the availability of wireless communications present an opportunity to open direct data transmission and feedback between peripheral health workers and central managers. In November 2009, the Uganda Ministry of Health deployed a short message service-based reporting system in two districts. At a set-up cost of $100/health facility, local technician support of $ 400 per month, and a cost of $0.53/week/clinic, the SMS reporting system was started at more than 140 clinics. Positivity rates for rapid diagnostic tests and artemisinin combination therapy stock outs were 48% and 54% in Kabale and 71% and 54% in Gulu, among other reports, at more than 85% health facilities reporting weekly and without monetary incentives or additional supervision. The SMS-based reporting systems have potential to improve timeliness in reporting of specific, time-sensitive metrics at modest cost, while by-passing current bottlenecks in the flow of data. With the development of specific capacity to manage stock data at district level, the availability of timely data offers potential to address commodity distribution problems and reduce stock-outs.     

Harnessing the p53-PUMA Axis to Overcome DNA Damage Resistance in Renal Cell Carcinoma

Resistance to DNA damage–induced apoptosis is a hallmark of cancer and a major cause of treatment failure and lethal disease outcome. A tumor entity that is largely resistant to DNA-damaging therapies including chemo- or radiotherapy is renal cell carcinoma (RCC). This study was designed to explore the underlying molecular mechanisms of DNA damage resistance in RCC to develop strategies to resensitize tumor cells to DNA damage–induced apoptosis. Here, we show that apoptosis-resistant RCC cells have a disconnect between activation of p53 and upregulation of the downstream proapoptotic protein p53 upregulated modulator of apoptosis (PUMA). We demonstrate that this disconnect is not caused by gene-specific repression through CCCTC-binding factor (CTCF) but instead by aberrant chromatin compaction. Treatment with an HDAC inhibitor was found to effectively reactivate PUMA expression on the mRNA and protein level and to revert resistance to DNA damage–induced cell death. Ectopic expression of PUMA was found to resensitize a panel of RCC cell lines to four different DNA-damaging agents tested. Remarkably, all RCC cell lines analyzed were wild-type for p53, and a knockdown was likewise able to sensitize RCC cells to acute genotoxic stress. Taken together, our results indicate that DNA damage resistance in RCC is reversible, involves the p53-PUMA axis, and is potentially targetable to improve the oncological outcomes of RCC patients.     

Endocannabinoids Tune Intrinsic Excitability in O-LM Interneurons by Direct Modulation of Postsynaptic Kv7 Channels

KCNQ-Kv7 channels are found at the axon initial segment of pyramidal neurons, where they control cell firing and membrane potential. In oriens lacunosum moleculare (O-LM) interneurons, these channels are mainly expressed in the dendrites, suggesting a peculiar function of Kv7 channels in these neurons. Here, we show that Kv7 channel activity is upregulated following induction of presynaptic long-term synaptic depression (LTD) in O-LM interneurons from rats of both sex, thus resulting in a synergistic long-term depression of intrinsic excitability (LTD-IE). Both LTD and LTD-IE involve endocannabinoid (eCB) biosynthesis for induction. However, although LTD is dependent on cannabinoid type 1 receptors, LTD-IE is not. Molecular modeling shows a strong interaction of eCBs with Kv7.2/3 channel, suggesting a persistent action of these lipids on Kv7 channel activity. Our data thus unveil a major role for eCB synthesis in triggering both synaptic and intrinsic depression in O-LM interneurons.SIGNIFICANCE STATEMENT In principal cells, Kv7 channels are essentially located at the axon initial segment. In contrast, in O-LM interneurons, Kv7 channels are highly expressed in the dendrites, suggesting a singular role of these channels in O-LM cell function. Here, we show that LTD of excitatory inputs in O-LM interneurons is associated with an upregulation of Kv7 channels, thus resulting in a synergistic LTD of LTD-IE. Both forms of plasticity are mediated by the biosynthesis of eCBs. Stimulation of CB1 receptors induces LTD, whereas the direct interaction of eCBs with Kv7 channels induces LTD-IE. Our results thus provide a previously unexpected involvement of eCBs in long-lasting plasticity of intrinsic excitability in GABAergic interneurons.