Novel P2X7 Antagonist Ameliorates the Early Phase of ALS Disease and Decreases Inflammation and Autophagy in SOD1-G93A Mouse Model.

Amyotrophic lateral sclerosis (ALS) is a disease with a resilient neuroinflammatory component caused by activated microglia and infiltrated immune cells. How to successfully balance neuroprotective versus neurotoxic actions through the use of anti-inflammatory agents is still under debate. There has been a boost of awareness regarding the role of extracellular ATP and purinergic receptors in modulating the physiological and pathological mechanisms in the nervous system. Particularly in ALS, it is known that the purinergic ionotropic P2X7 receptor plays a dual role in disease progression by acting at different cellular and molecular levels. In this context, we previously demonstrated that the P2X7 receptor antagonist, brilliant blue G, reduces neuroinflammation and ameliorates some of the pathological features of ALS in the SOD1-G93A mouse model. Here, we test the novel, noncommercially available, and centrally permeant Axxam proprietary P2X7 antagonist, AXX71, in SOD1-G93A mice, by assessing some behavioral and molecular parameters, among which are disease progression, survival, gliosis, and motor neuron wealth. We demonstrate that AXX71 affects the early symptomatic phase of the disease by reducing microglia-related proinflammatory markers and autophagy without affecting the anti-inflammatory markers or motor neuron survival. Our results suggest that P2X7 modulation can be further investigated as a therapeutic strategy in preclinical studies, and exploited in ALS clinical trials.

P2X7 antagonists for CNS indications: recent patent disclosures.

P2X7, a ligand-gated purinergic ion channel, has been at the center of intense efforts in the pharmaceutical industry in the last 15 years due to the growing appreciation of its role in inflammation. Since 2008-2009, increased focus on CNS available compounds has led to the publication of various patents on behalf of several pharmaceutical companies. This patent review aims at analyzing the recent patent literature (2008-2016) with a particular emphasis on those patents that are thought to deal with CNS penetrant compounds on the basis of their physicochemical features, the assays described in the patents and the uses these compounds are claimed for.

Ca(2+) release-activated Ca(2+) channel inhibitors.

Ca(2+) release-activated Ca(2+) (CRAC) channels are becoming important targets for therapeutic intervention in several areas of disease, including immunology, allergy and cancer. In parallel to the progression towards reliable methods for measuring CRAC currents and their inhibition, patents have been generated by several companies. In this Patent Review, an analysis of the patents in the CRAC channel inhibition filed is presented. A discussion of the biological methods used in the patents is included. The general interest in this area is growing fast with almost 80% of the patents issued after 2010.

Microarray analysis of 1,25(OH)2D3 regulated gene expression in human primary osteoblasts.

Though extensive studies have been conducted, questions regarding the molecular effectors and pathways underlying the regulatory role of 1,25(OH)(2)D(3) in human osteoblasts other than cell differentiation and matrix protein production remain unanswered. This study aims to identify genes and pathways that are modulated by 1,25(OH)(2)D(3) treatment in human osteoblasts. Primary osteoblast cultures obtained from human bone tissue samples were treated with 1,25(OH)(2)D(3) (10(-7) M) for 24 h and their transcritptomes were profiled by microarray analysis using the Affymetrix GeneChip. Statistical analysis was conducted to identify genes whose expression is significantly modulated following 1,25(OH)(2)D(3) treatment. One hundred and fifty-eight genes were found to be differentially expressed. Of these, 136 were upregulated, indicating clear transcriptional activation by 1,25(OH)(2)D(3). Biostatistical evaluation of microarray data by Ingenuity Pathways Analysis (IPA) revealed a relevant modulation of genes involved in vitamin D metabolism (CYP24), immune functions (CD14), neurotransmitter transporters (SLC1A1, SLC22A3), and coagulation [thrombomodulin (THBD), tissue plasminogen activator (PLAT), endothelial protein C receptor (PROCR), thrombin receptor (F2R)]. We identified a restricted number of highly regulated genes and confirmed their differential expression by real-time quantitative PCR (RT qPCR). The present genome-wide microarray analysis on 1,25(OH)(2)D(3) -treated human osteoblasts reveals an interplay of critical regulatory and metabolic pathways and supports the hypothesis that 1,25(OH)(2)D(3) can modulate the coagulation process through osteoblasts, activates osteoclastogenesis through inflammation signaling, modulates the effects of monoamines by affecting their reuptake.

The GeneTrawler: mapping potential drug targets in human and rat tissues.

Expression data are an important element of target identification and validation. The authors have established an automated high-throughput method based on real time quantitative polymerase chain reaction, called the GeneTrawler, for the characterization of pharmaceutical targets on an annotated collection of human tissues. The authors have conducted a variability analysis of the system, which demonstrates that the majority of the variability between expression levels determined is due to biologic variation between samples, rather than technical variation due to imprecision of the method. Gene expression maps, generated with this carefully controlled system provide a large, reliable, consistent data set. The authors have used this system to characterize the expression of > 100 genes, and here they show the expression profile of SUR1 in order to illustrate its use. The authors were able to confirm SUR1 expression in the lung, which was suggested on the basis of pharmacologic experiments but has not previously been confirmed by mRNA detection. The data also show SUR1 expression in tissues that have been associated with some of the side effects seen with SUR1 modulators. This and other examples demonstrate that the GeneTrawler is useful to gauge the suitability of a prospective therapeutic target, to fully exploit a known drug target, or to identify and help validate new hypothetical druggable targets to fuel drug discovery pipelines.