Pantothenate Kinase Activation Restores Brain Coenzyme A in a Mouse Model of Pantothenate Kinase-Associated Neurodegeneration.

Pantothenate kinase-associated neurodegeneration (PKAN) is characterized by a motor disorder with combinations of dystonia, parkinsonism, and spasticity, leading to premature death. PKAN is caused by mutations in the PANK2 gene that result in loss or reduction of PANK2 protein function. PANK2 is one of three kinases that initiate and regulate coenzyme A biosynthesis from vitamin B5, and the ability of BBP-671, an allosteric activator of pantothenate kinases, to enter the brain and elevate coenzyme A was investigated. The metabolic stability, protein binding, and membrane permeability of BBP-671 all suggest that it has the physical properties required to cross the blood-brain barrier. BBP-671 was detected in plasma, liver, cerebrospinal fluid, and brain following oral administration in rodents, demonstrating the ability of BBP-671 to penetrate the brain. The pharmacokinetic and pharmacodynamic properties of orally administered BBP-671 evaluated in cannulated rats showed that coenzyme A (CoA) concentrations were elevated in blood, liver, and brain. BBP-671 elevation of whole-blood acetyl-CoA served as a peripheral pharmacodynamic marker and provided a suitable method to assess target engagement. BBP-671 treatment elevated brain coenzyme A concentrations and improved movement and body weight in a PKAN mouse model. Thus, BBP-671 crosses the blood-brain barrier to correct the brain CoA deficiency in a PKAN mouse model, resulting in improved locomotion and survival and providing a preclinical foundation for the development of BBP-671 as a potential treatment of PKAN. SIGNIFICANCE STATEMENT: The blood-brain barrier represents a major hurdle for drugs targeting brain metabolism. This work describes the pharmacokinetic/pharmacodynamic properties of BBP-671, a pantothenate kinase activator. BBP-671 crosses the blood-brain barrier to correct the neuron-specific coenzyme A (CoA) deficiency and improve motor function in a mouse model of pantothenate kinase-associated neurodegeneration. The central role of CoA and acetyl-CoA in intermediary metabolism suggests that pantothenate kinase activators may be useful in modifying neurological metabolic disorders.

Wolff/Cope Approach to the AB Ring of the Sesterterpenoid Variecolin.

A stereoselective synthesis of the AB ring of the complex sesterterpenoid variecolin is presented. Our strategy features the development of a tandem Wolff/Cope rearrangement of α-diazo cyclobutyl ketones for the construction of fused, 8-membered carbocycles. Preliminary studies revealed a facile Wolff rearrangement but a difficult vinyl ketene cyclobutane Cope rearrangement. We have leveraged an efficient microwave-promoted tandem rearrangement to prepare the desired functionalized cyclooctadienones that we envision as potential key intermediates in the convergent synthesis of variecolin.

Hydroxyproline-derived pseudoenantiomeric [2.2.1] bicyclic phosphines: asymmetric synthesis of (+)- and (-)-pyrrolines.

We have prepared two new diastereoisomeric 2-aza-5-phosphabicyclo[2.2.1]heptanes from naturally occurring trans-4-hydroxy-L-proline in six chemical operations. These syntheses are concise and highly efficient, with straightforward purification. When we used these chiral phosphines as catalysts for reactions of γ-substituted allenoates with imines, we obtained enantiomerically enriched pyrrolines in good yields with excellent enantioselectivities. These two diastereoisomeric phosphines functioned as pseudoenantiomers, providing their chiral pyrrolines with opposite absolute configurations.

Phosphine-catalyzed synthesis of highly functionalized coumarins.

2-Styrenyl allenoates are converted into cyclopentene-fused dihydrocoumarins through phosphine-catalyzed regio- and diastereoselective [3 + 2] cycloadditions. Remarkably, changing the solvent from THF to benzene promotes the conversion of the 2-(2-nitrostyrenyl) allenoate into a tricyclic nitronate through a previously undocumented mode of phosphine catalysis. This nitronate was subjected to efficient face-, regio-, and exo-selective 1,3-dipolar cycloadditions to provide tetracyclic coumarin derivatives.

Phosphine-catalyzed synthesis of 1,3-dioxan-4-ylidenes.

[reaction: see text] A phosphine-catalyzed reaction of an allenoate with aldehydes furnished (2,6-diaryl-[1,3]dioxan-4-ylidene)-acetates 4 in excellent to moderate yields with complete diastereoselectivity and high E/Z-selectivities. Upon removal of the acetal functionality in this domino reaction product 4, delta-hydroxy-beta-ketoester 11 was obtained. The reported vinylphosphonium-based approach provides a new way to achieve a synthesis of delta-hydroxy-beta-ketoesters that differs from the classical dianion-based approach.

Requirement of Cys399 for processing of the human ecto-ATPase (NTPDase2) and its implications for determination of the activities of splice variants of the enzyme.

Ecto-ATPase (CD39L1) corresponds to the type 2 enzyme of the ecto-nucleoside triphosphate diphosphohydrolase family (E-NTPDase). We have isolated from human ECV304 cells three cDNAs with high homology with members of the E-NTPDase family that encode predicted proteins of 495, 472, and 450 amino acids. Sequencing of a genomic DNA clone confirmed that these three sequences correspond to splice variants of the human ecto-ATPase (NTPDase2 alpha,-2 beta, and -2 gamma). Although all three enzyme forms were expressed heterologously to similar levels in Chinese hamster ovary cells clone K-1 (CHO-K1) cells, only the 495-amino acid protein (NTPDase2 alpha exhibited ecto-ATPase activity. Immunolocalization studies demonstrated that NTPDase2 alpha is fully processed and trafficked to the plasma membrane, whereas the NTPDase2 beta and -2 gamma splice variants were retained in not fully glycosylated forms in the endoplasmic reticulum. The potential roles of two highly conserved residues, Cys399 and Asn443, in the activity and cellular trafficking of the ecto-ATPase were examined. Mutation of Cys399, which is absent in NTPDase2 beta and -2 gamma, produced a protein completely devoid of nucleotidase activity, while mutation of Asn443 to Asp resulted in substantial loss of activity. Neither the Cys399 nor Asn443 mutants were fully glycosylated, and both were retained in the endoplasmic reticulum. These results indicate that the lack of ecto-nucleotidase activity exhibited by NTPDase2 beta and -2 gamma and the C399S mutant, as well as the large reduction of activity in the N443D mutant are due to alterations in the folding/maturation of these proteins.