The Rho signaling pathway regulates the cytoskeleton and motility and plays an important role in neuronal growth inhibition. Here we demonstrate that inactivation of Rho or its downstream target Rho-associated kinase (ROK) stimulated neurite growth in primary cells of cortical neurons plated on myelin or chondroitin sulfate proteoglycan substrates. Furthermore, treatment either with C3 transferase (C3) to inactivate Rho or with Y27632 to inhibit ROK was sufficient to stimulate axon regeneration and recovery of hindlimb function after spinal cord injury (SCI) in adult mice. Injured mice were treated with a single injection of Rho or Rho-associated kinase inhibitors delivered in a protein adhesive at the lesion site. Treated animals showed long-distance regeneration of anterogradely labeled corticospinal axons and increased levels of GAP-43 mRNA in the motor cortex. Behaviorally, inactivation of Rho pathway induced rapid recovery of locomotion and progressive recuperation of forelimb-hindlimb coordination. These findings provide evidence that the Rho signaling pathway is a potential target for therapeutic interventions after spinal cord injury.

The influence of a bulky 5-position substituent on the amide isomer equilibrium N-terminal to proline has been explored via the synthesis and analysis of N-(acetyl)proline N′-methylamide (1) and its respective cis-and trans-5-tert-butylproline amide diastereomers 2 and 3. The relative populations of the amide cis-and trans-isomers as well as the energy barriers for amide isomerization of 1-3 in D 2 O were ascertained using NMR with coalescence and magnetization transfer experiments. The relative populations of free C-terminal amide and hydrogen-bonded amide in the γ-turn conformation were also estimated by integrating the N-H stretch absorbances in the FT-IR spectra of 1-3 in CHCl 3 and CCl 4 . In the prolyl peptides, the 5-tert-butyl substituent was found to exhibit profound effects on the amide isomer equilibrium, on the energy barrier for amide isomerization, and on the stability of the γ-turn conformation. Steric interactions between the 5-position substituent and the N-acetyl group disfavor the amide trans-and augment the cis-isomer population: 25% in 1, 48% in 2, and 66% in 3. In the case of cis-5-tert-butylproline 2, the energy barrier for amide isomerization is observed to be 3.9 kcal/mol lower than that of 1. On the other hand, the amide isomerization barrier for trans-5-tert-butylproline 3 is similar to that for 1. Only a single amide N-H stretch band is observed at 3454 cm -1 in the FT-IR spectrum of 3 in CHCl 3 and indicates that the NH group is free of intramolecular hydrogen bonding. Hence, trans-5-tert-butylproline amide 3 does not adopt a seven-membered γ-turn conformation, which is a favored conformer for 1 and 2 in CHCl 3 . Maps, in which the ψ-and ω-dihedral angles are plotted at 30°intervals against the calculated energy of the local minimum conformation, predict qualitatively and display clearly all of the observed effects of the 5-tert-butyl substituent on the amide isomer in the N-(acetyl)-proline N′-methylamides. The results of this study suggest the use of 5-tert-butylprolines to prepare both X-Pro cis-amide isomers and twisted amide surrogates for examining prolyl residue conformations in bioactive peptides.

A three-step synthesis of JV-(9-(9-phenylfluorenyl))-L-alaninal (5) from alanine is described. Exposure to silica or nonnucleophilic base causes no detectable racemization of this -amino aldehyde. The 9-(9-phenylfluorenyl) N-protecting group also maintains the configurational integrity of -amino aldehyde 5 during C-C bond-forming reactions, providing enantiomerically pure products from Wittig reaction, aldol condensation, and addition of Grignard reagent.The synthetic applications of optically active N-protected examino aldehydes have increased dramatically with the rising interest in the synthesis of polyfunctional amino acids,1 peptide analogues,1

Progressive cystic kidney degeneration underlies diverse renal diseases, including the most common cause of kidney failure, autosomal dominant Polycystic Kidney Disease (PKD). Genetic analyses of patients and animal models have identified several key drivers of this disease. The precise molecular and cellular changes underlying cystogenesis remain, however, elusive. Drosophila mutants lacking the translational regulator Bicaudal C (BicC, the fly ortholog of vertebrate BICC1 implicated in renal cystogenesis) exhibited progressive cystic degeneration of the renal tubules (so called “Malpighian” tubules) and reduced renal function. The BicC protein was shown to bind to Drosophila (d-) myc mRNA in tubules. Elevation of d-Myc protein levels was a cause of tubular degeneration in BicC mutants. Activation of the Target of Rapamycin (TOR) kinase pathway, another common feature of PKD, was found in BicC mutant flies. Rapamycin administration substantially reduced the cystic phenotype in flies. We present new mechanistic insight on BicC function and propose that Drosophila may serve as a genetically tractable model for dissecting the evolutionarily-conserved molecular mechanisms of renal cystogenesis.

Mimicry of bioactive conformations is critical for peptide-based medicinal chemistry because such peptidomimetics may augment stability, enhance affinity, and increase specificity. Azapeptides are peptidomimetics in which the α-carbon(s) of one or more amino acid residues are substituted by nitrogen. The resulting semicarbazide analogues have been shown to reinforce β-turn conformation through the combination of lone pair-lone pair repulsion of the adjacent hydrazine nitrogen and urea planarity. Substitution of a semicarbazide for an amino amide residue in a peptide may retain biological activity and add benefits such as improved metabolic stability. The applications of azapeptides include receptor ligands, enzyme inhibitors, prodrugs, probes, and imaging agents. Moreover, azapeptides have proven therapeutic utility. For example, the aza-glycinamide analogue of the luteinizing hormone-releasing hormone analogue Zoladex is a potent long-acting agonist currently used in the clinic for the treatment of prostate and breast cancer. However, the use of azapeptides was hampered by tedious solution-phase synthetic routes for selective hydrazine functionalization. A remarkable stride to overcome this bottleneck was made in 2009 through the introduction of the submonomer procedure for azapeptide synthesis, which enabled addition of diverse side chains onto a common semicarbazone intermediate, providing a means to construct azapeptide libraries by solution- and solid-phase chemistry. In brief, aza residues are introduced into the peptide chain using the submonomer strategy by semicarbazone incorporation, deprotonation, N-alkylation, and orthogonal deprotection. Amino acylation of the resulting semicarbazide and elongation gives the desired azapeptide. Since the initial report, a number of chemical transformations have taken advantage of the orthogonal chemistry of semicarbazone residues (e.g., Michael additions and N-arylations). In addition, libraries have been synthesized from libraries by diversification of aza-propargylglycine (e.g., A coupling reactions, [1,3]-dipolar cycloadditions, and 5-exo-dig cyclizations) and aza-chloroalkylglycine residues. In addition, oxidation of aza-glycine residues has afforded azopeptides that react in pericyclic reactions (e.g., Diels-Alder and Alder-ene chemistry). The bulk of these transformations of aza-glycine residues have been developed by the Lubell laboratory, which has applied such chemistry in the synthesis of ligands with promising biological activity for treating diseases such as cancer and age-related macular degeneration. Azapeptide analogues of growth hormone-releasing peptide-6 (His-d-Trp-Ala-Trp-d-Phe-Lys-NH, GHRP-6) have for example been pursued as ligands of the cluster of differentiation 36 receptor (CD36) and show promising activity for the development of treatments for angiogenesis-related diseases, such as age-related macular degeneration, as well as for atherosclerosis. Azapeptides have also been employed to make a series of conformationally constrained second mito...