A General Approach to Optically Pure, α-Methyl Non-Natural Amino Acids: Enabling Unique Peptides as Drug Candidates.

An α-methyl, non-natural amino acid (NNAA) building block equipped with an alkyl halide tail that could be readily transformed into an organozinc was prepared. This single organometallic was cross-coupled to an array of heterocyclic electrophiles using the Pd-PEPPSI-IHeptCl catalyst to produce a wide selection of optically pure α-methyl NNAAs. With these in hand, non-natural peptides are being produced for evaluation in a variety of therapeutic areas in drug discovery.

The History of Flow Chemistry at Eli Lilly and Company.

Flow chemistry was initially used for speed to early phase material delivery in the development laboratories, scaling up chemical transformations that we would not or could not scale up batch for safety reasons. Some early examples included a Newman Kwart Rearrangement, Claisen rearrangement, hydroformylation, and thermal imidazole cyclization. Next, flow chemistry was used to enable safe scale up of hazardous chemistries to manufacturing plants. Examples included high pressure hydrogenation, aerobic oxidation, and Grignard formation reactions. More recently, flow chemistry was used in Small Volume Continuous (SVC) processes, where highly potent oncolytic molecules were produced by fully continuous processes at about 10 kg/day including reaction, extraction, distillation, and crystallization, using disposable equipment contained in fume hoods.

Contributions of nitric oxide synthase isozymes to exhaled nitric oxide and hypoxic pulmonary vasoconstriction in rabbit lungs.

We investigated the source(s) for exhaled nitric oxide (NO) in isolated, perfused rabbits lungs by using isozyme-specific nitric oxide synthase (NOS) inhibitors and antibodies. Each inhibitor was studied under normoxia and hypoxia. Only nitro-L-arginine methyl ester (L-NAME, a nonselective NOS inhibitor) reduced exhaled NO and increased hypoxic pulmonary vasoconstriction (HPV), in contrast to 1400W, an inhibitor of inducible NOS (iNOS), and 7-nitroindazole, an inhibitor of neuronal NOS (nNOS). Acetylcholine-mediated stimulation of vascular endothelial NOS (eNOS) increased exhaled NO and could only be inhibited by L-NAME. Selective inhibition of airway and alveolar epithelial NO production by nebulized L-NAME decreased exhaled NO and increased hypoxic pulmonary artery pressure. Immunohistochemistry demonstrated extensive staining for eNOS in the epithelia, vasculature, and lymphatic tissue. There was no staining for iNOS but moderate staining for nNOS in the ciliated cells of the epithelia, lymphoid tissue, and cartilage cells. Our findings show virtually all exhaled NO in the rabbit lung is produced by eNOS, which is present throughout the airways, alveoli, and vessels. Both vascular and epithelial-derived NO modulate HPV.

Effects of S-nitrosation and cross-linking of hemoglobin on hypoxic pulmonary vasoconstriction in isolated rat lungs.

Free hemoglobin (Hb) and red blood cells augment hypoxic pulmonary vasoconstriction (HPV) by scavenging nitric oxide (NO). S-nitrosation of Hb (SNO-Hb) may confer vasodilatory properties by allowing release of NO during deoxygenation and/or by interaction with small-molecular weight thiols. Likewise, cross-linking of free Hb may limit its vasoconstrictive effect by preventing abluminal movement of the molecule. We compared the effects of free SNO-Hb and Hb intramolecularly cross-linked at the beta-cysteine 93 residue [Bis(maleidophenyl)-polyethylene glycol2000HbA (Bis-Mal-PEGHb)] to those of free oxyHb on pulmonary artery pressure (PAP), HPV, and exhaled NO (eNO) in isolated, perfused rat lungs. Ventilation of lungs with anoxic gas for 5 minutes reduced perfusate PO2 to 11+/-1.0 Torr. Addition of SNO-Hb or Bis-Mal-PEGHb (100 micromol/L) to buffer perfusate increased normoxic PAP and augmented HPV in similar magnitude as free oxyHb, but had no effect on eNO. Addition of the allosteric modulator inositol hexaphosphate to increase Hb P50 and the thiol glutathione (GSH) to allow removal of NO from Hb via transnitrosation to the perfusate did not reduce augmentation of HPV by SNO-Hb or increase eNO. GSH resulted in an approximately 50% reduction in perfusate [S-nitrosothiol], in association with an increase in perfusate [metHb]. Free SNO-Hb is a net NO scavenger and pulmonary vasoconstrictor in this model, although thiol-mediated release of NO from SNO-Hb does occur. However, release of NO from SNO-Hb was not influenced by deoxygenation-mediated allosteric changes in Hb across a broad range of oxyHb saturation. Cross-linking of Hb does not limit its pulmonary vasoconstrictor effects.

The effect of a bioresorbable matrix barrier in endodontic surgery on the rate of periapical healing: an in vivo study.

The purpose of this prospective clinical study was to assess the rate of healing of periapical bony defects created at the time of endodontic periapical surgery by measuring the densitometric ratio change in periapical osseous tissues, after periapical endodontic surgery was performed, by using or not using Guidor bioresorbable membrane material. Periapical surgeries were performed on 25 patients where nonsurgical root canal therapy had failed and a periapical lesion was present. For inclusion in the study, the osseous defect to be analyzed had to be confined to the apical area, with bone covering the entire root surface coronally, and an intact lingual cortical plate had to be present. A series of radiographs at the 3-, 6-, and 12-month recalls were compared with the radiograph taken immediately after surgery by using digital imaging and a densitometric ratio that gave a numerical estimation of osseous healing. Using repeated-measure ANOVA, it was shown that there was no statistical difference between the rate of healing in those cases where a Guidor resorbable membrane was used and those cases where no membrane was used. The results suggest that placement of a guided tissue membrane over the bony opening created during an endodontic periapical surgical procedure has no beneficial effect on the rate of healing and the added expense to the patient would not be warranted in these cases.