Synthesis of polyfluoro ketones for selective inhibition of human phospholipase A2 enzymes

The development of selective inhibitors for individual PLA2 enzymes is necessary in order to target PLA2-specific signaling pathways, but it is challenging due to the observed promiscuity of known PLA2 inhibitors. In the current work, we present the development and application of a variety of synthetic routes to produce pentafluoro, tetrafluoro, and trifluoro derivatives of activated carbonyl groups in order to screen for selective inhibitors and characterize the chemical properties that can lead to selective inhibition. Our results demonstrate that the pentafluoroethyl ketone functionality favors selective inhibition of the GVIA iPLA2, a very important enzyme for which specific, potent, reversible inhibitors are needed. We find that 1,1,1,2,2-pentafluoro-7-phenyl-heptan-3-one (FKGK11) is a selective inhibitor of GVIA iPLA2 (XI(50) = 0.0073). Furthermore, we conclude that the introduction of an additional fluorine atom at the α′ position of a trifluoromethyl ketone constitutes an important strategy for the development of new potent GVIA iPLA2 inhibitors. © 2008 American Chemical Society

Investigating the metabolic fingerprint of term infants with normal and increased fetal growth

An NMR metabolomic approach was employed to highlight the metabolic changes underlying prenatal disorders and determine metabolites that could serve as potential markers in relation to large for gestational age (LGA) newborns. In this holistic study, multivariate data analysis elicited information from the NMR spectra and probed to metabolic signatures of macrosomic fetuses. Moreover, metabolic trends that characterize LGA from mothers diagnosed with gestational diabetes mellitus (LGA-GDM), as well as LGA from mothers not diagnosed with GDM (LGA-NGDM) were framed. Results obtained from maternal and umbilical cord (UC) samples indicated that LGA fetuses present alterations especially in the aminoacid metabolism as compared to Appropriate for Gestational Age (AGA) cases. Clear discrimination of LGA-NGDM from LGA-GDM was achieved both in maternal and in UC samples' blood. The role of glutamine and alanine together with four essential (valine, leucine, isoleucine, threonine) aminoacids, as well as the role of glycerol and glucose is emphasized for the case of maternal LGA samples' differentiation. Glycine and histidine only contributed to the differentiation of UC samples, the former characterized the AGA cases, while the latter was ascribed to both LGA-GDM and LGA-NGDM cases. Interestingly, both UC and maternal LGA-GDM samples were characterized by increased levels of N-acetylglutamic and acetoacetic acids. The OPLS-DA models were validated with permutation testing and ROC curves. In conclusion, this study indicates that NMR metabolomics may enable the detection of metabolic changes associated with LGA prenatal disorders. © 2016 The Royal Society of Chemistry

Steroidal Cardiac Na+/K (+) ATPase Inhibitors Exhibit Strong Anti-Cancer Potential in vitro and in Prostate and Lung Cancer Xenografts in vivo

Sodium potassium pump (Na+/K+ ATPase) is a validated pharmacological target for the treatment of congestive heart failure. Recent data with inotropic drugs such as digoxin & digitoxin (digitalis) suggest a potent anti-cancer action of these drugs and promote Na+/K+ ATPase as a novel therapeutic target in cancer. However, digitalis have narrow therapeutic indices, are pro-arrhythmic and are considered non-developable drugs by the pharmaceutical industry. On the contrary, a series of recently-developed steroidal inhibitors showed better pharmacological properties and clinical activities in cardiac patients. Their anti-cancer activity however, remained unknown. In this study, we synthesized seventeen steroidal cardiac inhibitors and explored for the first time their anti-cancer activity in vitro and in vivo. Our results indicate potent anti-cancer actions of steroidal cardiac inhibitors in multiple cell lines from different tumor panels including multi-drug resistant cells. Furthermore, the most potent compound identified in our studies, the 3-[(R)-3-pyrrolidinyl] oxime derivative 3, showed outstanding potencies (as measured by GI(50), TGI and LC50 values) in most cells in vitro, was selectively cytotoxic in cancer versus normal cells showing a therapeutic index of 31.7 and exhibited significant tumor growth inhibition in prostate and lung xenografts in vivo. Collectively, our results suggest that previously described cardiac Na+/K+ ATPase inhibitors have potent anti-cancer actions and may thus constitute strong re-purposing candidates for further cancer drug development

Intracellular phospholipase A2 group IVA and group VIA play important roles in Wallerian degeneration and axon regeneration after peripheral nerve injury

We provide evidence that two members of the intracellular phospholipase A2 family, namely calcium-dependent group IVA (cPLA2 GIVA) and calcium-independent group VIA (iPLA2 GVIA) may play important roles in Wallerian degeneration in the mouse sciatic nerve. We assessed the roles of these PLA2s in cPLA2 GIVA-/- mice, and mice treated with a selective inhibitor of iPLA2 GVIA (FKGK11). Additionally, the effects of both these PLA2s were assessed by treating cPLA2 GIVA-/- mice with the iPLA2 inhibitor. Our data suggest that iPLA2 GVIA may play more of a role in the early stages of myelin breakdown, while cPLA2 GIVA may play a greater role in myelin clearance by macrophages. Our results also show that the delayed myelin clearance and Wallerian degeneration after sciatic nerve crush injury in mice lacking cPLA2 and iPLA2 activities is accompanied by a delay in axon regeneration, target re-innervation and functional recovery. These results indicate that the intracellular PLA 2s (cPLA2 GIVA and iPLA2 GVIA) contribute significantly to various aspects of Wallerian degeneration in injured peripheral nerves, which is then essential for successful axon regeneration. This work has implications for injury responses and recovery after peripheral nerve injuries in humans, as well as for understanding the slow clearance of myelin after CNS injury and its potential consequences for axon regeneration. © The Author (2008). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved

Global metabolic stress of isoeffort continuous and high intensity interval aerobic exercise: a comparative 1H NMR metabonomic study

The overall metabolic/energetic stress occurring during an exercise-bout is proposed as the main driving force for long-term training adaptations. Continuous and high-intensity interval exercises (HIIE) are currently prescribed to acquire the muscular and metabolic benefits of aerobic training. We applied 1H NMR-based metabonomics to compare the overall metabolic perturbation and the activation of individual bio-energetic pathways of three popular aerobic exercises matched for effort/strain. Nine men performed continuous, long-interval (3min-bouts), and short-interval (30s-bouts) exercise-bouts under isoeffort conditions. Blood was collected before and after exercise. The multivariate PCA and OPLS-DA models showed a distinct separation of pre- and post-exercise samples in three protocols. The two models did not discriminate the post-exercise overall metabolic profiles of three exercise types. Analysis focused on muscle bio-energetic pathways revealed an extensive up-regulation of carbohydrate-lipid metabolism and TCA-cycle in all three protocols; there were only few differentiations among protocols in post-exercise abundance of molecules when long-interval bouts were performed. In conclusion, continuous and HIIE exercise protocols, when performed with similar effort/strain, induce comparable global metabolic stress despite their marked differences in work-bout intensities. This study highlights the importance of NMR-metabonomics in comprehensive monitoring of metabolic consequences of exercise training in blood of athletes and exercising individuals

Differing roles for members of the phospholipase A2 superfamily in experimental autoimmune encephalomyelitis

The phospholipase A2 (PLA2) superfamily hydrolyzes phospholipids to release free fatty acids and lysophospholipids, some of which can mediate inflammation and demyelination, hallmarks of the CNS autoimmune disease multiple sclerosis. The expression of two of the intracellular PLA2s (cPLA2 GIVA and iPLA2 GVIA) and two of the secreted PLA2s (sPLA2 GIIA and sPLA2 GV) are increased in different stages of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. We show using small molecule inhibitors, that cPLA2 GIVA plays a role in the onset, and iPLA2 GVIA in the onset and progression of EAE. We also show a potential role for sPLA2 in the later remission phase. These studies demonstrate that selective inhibition of iPLA2 can ameliorate disease progression when treatment is started before or after the onset of symptoms. The effects of these inhibitors on lesion burden, chemokine and cytokine expression as well as on the lipid profile provide insights into their potential modes of action. iPLA2 is also expressed by macrophages and other immune cells in multiple sclerosis lesions. Our results therefore suggest that iPLA2 might be an excellent target to block for the treatment of CNS autoimmune diseases, such as multiple sclerosis. © 2009 Oxford University Press on behalf of the Guarantors of Brain. All rights reserved

Phospholipase A2 superfamily members play divergent roles after spinal cord injury

Spinal cord injury (SCI) results in permanent loss of motor functions. A significant aspect of the tissue damage and functional loss may be preventable as it occurs, secondary to the trauma. We show that the phospholipase A 2 (PLA2) superfamily plays important roles in SCI. PLA2 enzymes hydrolyze membrane glycerophospholipids to yield a free fatty acid and lysophospholipid. Some free fatty acids (arachidonic acid) give rise to eicosanoids that promote inflammation, while some lysophospholipids (lysophosphatidylcholine) cause demyelination. We show in a mouse model of SCI that two cytosolic forms [calcium-dependent PLA2 group IVA (cPLA 2 GIVA) and calcium-independent PLA2 group VIA (iPLA 2 GVIA)], and a secreted form [secreted PLA2group IIA (sPLA2 GIIA)] are up-regulated. Using selective inhibitors and null mice, we show that these PLA2s play differing roles. cPLA2 GIVA mediates protection, whereas sPLA2 GIIA and, to a lesser extent, iPLA2 GVIA are detrimental. Furthermore, completely blocking all three PLA2s worsens outcome, while the most beneficial effects are seen by partial inhibition of all three. The partial inhibitor enhances expression of cPLA2 and mediates its beneficial effects via the prostaglandin EP1 receptor. These findings indicate that drugs that inhibit detrimental forms of PLA2 (sPLA2 and iPLA2) and upregulate the protective form (cPLA2) may be useful for the treatment of SCI. © FASEB

Chemistry of GABAB receptor ligands: focus on agonists and antagonists

Since the discovery of GABAB receptor by Norman G. Bowery and coworkers in 1980, a striking endeavour was made by industrial and academic researchers to develop GABAB receptor ligands for therapeutic application in a variety of diseases associated with dysfunctions of the gabaergic system. Although baclofen (Lioresal) is still the only approved GABAB receptor agonist, this sustained research effort has produced many new compounds which are able to exert GABAB agonist, partial agonist or antagonist activity. This chapter presents an overview of the outcomes in this field, with a special focus on the chemistry, structure–activity relationship and mechanism of action of several GABA and baclofen analogues, derivatives and bioisosteres. © Springer International Publishing Switzerland 2016

Phospholipase A 2

Phospholipases represent one of the earliest enzyme activities to be identified and studied, and the phospholipase A2 superfamily traces its roots to the identification of lytic actions of snake venom at the end of the 19th century. Both electrostatic and hydrophobic interactions contribute to the interfacial binding of sPLA2 to anionic phospholipid membranes. The interaction between basic residues on the binding surface with anionic vesicles plays an important role in interfacial binding. The major functions will be summarized below and include the ability to kill Gram-positive and Gram-negative bacteria, thereby affecting host defense against bacterial infections. sPLA2 may be involved in the pathogensis of inflammatory bowel disease including Crohn's disease and ulcerative colitis. GIIA sPLA2 protein and mRNA were detected in Paneth cells of the small intestinal mucosa in the intestine in Crohn's disease patients