Extent of methionine limitation in peak-, early-, and mid-lactation dairy cows.

Five multiparous, ruminally and duodenally cannulated Holstein cows were assigned to 5 x 5 Latin squares at wk 2 (experiment 1), wk 11 to 13 (experiment 2), and wk 17 to 19 postpartum (experiment 3) to determine extent of Met limitation. Treatments were duodenally infused and consisted of 10 g/d of l-Lys plus 0, 3.5, 7.0, 10.5, or 16.0 g/d of dl-Met in experiments 1 and 2 and 8 g/d of l-Lys plus 0, 5, 10, 15, or 20 g/d of dl-Met in experiment 3. Calculated Lys contributions to total AA (TAA) in duodenal digesta for control treatments were 8.6, 7.5, and 9.0% for experiments 1, 2, and 3, respectively. Methionine contributions to TAA for the 5 infusion treatments were 1.9, 2.1, 2.2, 2.4, and 2.7% for experiment 1; 2.1, 2.3, 2.4, 2.5, and 2.7% for experiment 2; and 1.8, 2.0, 2.2, 2.4, and 2.5% for experiment 3, respectively. Milk protein yield increased linearly in experiments 1 and 2, indicating that Met contribution to TAA in duodenal digesta for maximal milk protein synthesis exceeded 2.7 for early-lactation cows. In experiment 2, a quadratic relationship was found between level of infused Met and milk protein content, with the response reaching a plateau when 12.2 g of Met was infused, corresponding with a Met contribution to TAA in duodenal digesta of 2.4%. In experiment 3, milk protein content increased quadratically, but milk yield declined linearly with increasing levels of infused Met; hence, milk protein yield was unaffected by treatment. The calculated plateau point of the milk protein content response curve was determined to be 12.4 g of infused Met, which corresponds to a Met contribution to TAA in duodenal digesta of 2.3%. Experiment 3 results indicate that the required level of Met in duodenal digesta for maximizing milk protein yield is lower than that required for maximizing milk protein content.

Influence of tannic acid application on alfalfa hay: in vitro rumen fermentation, serum metabolites and nitrogen balance in sheep.

Alfalfa protein is poorly utilised by ruminants due to its rapid degradation in rumen. The objective of the study was to assess the influence of spraying tannic acid (TA) on chopped alfalfa hay on in vitro rumen fermentation and nitrogen (N) retention by sheep. Alfalfa hay with and without TA was fed to sheep to determine nutrient digestibility and N balance. TA was sprayed on chopped alfalfa at three concentrations to determine its effect on in vitro fermentation of dry matter (DM) and N balance in sheep. Final TA concentrations were 0, 30, 60 and 90 g TA per kg DM. The control was sprayed with the same amount of water but without TA. In vitro DM degradation and the production of gas, ammonium-N (NH4-N) and short-chain fatty acid (SCFA) were measured. TA-sprayed alfalfa and the control were fed to sheep to determine nutrient digestibility and N retention. Addition of TA had no influence on the extent and rate of gas production but significantly decreased NH4-N concentration at 30 (P < 0.05), 60 and 90 (P < 0.0001) g/kg DM. Addition of polyethylene glycol (PEG) to TA-sprayed alfalfa increased NH4-N to a level comparable to non-TA-sprayed alfalfa. Spraying of alfalfa with TA significantly decreased (P < 0.05) isovalerate but did not affect the total and individual SCFA acid production. Tannic acid significantly (P < 0.05) reduced in vitro true degradability of DM (IVTD) after 24 h incubation at levels of 60 and 90 g TA per kg DM. Neutral-detergent fibre digestibility (dNDF) after 24 h (P < 0.01), 60 and 90 (P < 0.0001) g TA per kg DM. The effect of TA on either IVTD or dNDF was not significant (P > 0.05) after 48 h of incubation. There was a strong linear relationship between percentage increase in gas production due to PEG and protein precipitation capacity (R2 = 0.94). N digestibility was significantly reduced with all three levels of TA additions. However, the proportion of urine-N to total N output was reduced by adding 60 g (P < 0.05) and 90 g (P < 0.01) TA per kg DM. Serum metabolites and liver enzymes were not affected by TA (P > 0.05). Higher faecal N as the TA level increased indicates incomplete dissociation of tannin-protein complexes post ruminally. Factors affecting dissociation of tannin-protein complexes need further study.

Effects of rumen-protected choline and monensin on milk production and metabolism of periparturient dairy cows.

Choline and monensin may be supplemented during the transition period with the objectives of aiding in fat metabolism and improving energy balance, respectively. The objectives of this study were to determine the effects of supplementing rumen-protected choline (RPC) and monensin in a controlled-release capsule (CRC) on metabolism, dry matter intake, milk production, and liver function in transition dairy cattle. Three weeks before expected calving, 182 Holsteins were randomly assigned to receive one of the following: a monensin CRC, 56 g/d of RPC until 28 d in milk, CRC + RPC, or neither supplement (control). Blood samples were collected at enrollment, 1 wk before calving, and in the first and second weeks after calving. Liver biopsies were obtained from multiparous cows randomly selected from each treatment group within 24 h and again 3 wk postpartum. Daily milk production was recorded through 60 d in milk. There were no interactions of the effects of RPC and CRC on any of the outcomes measured. Overall, cows that received RPC produced 1.2 kg/d more milk in the first 60 d of lactation, but this effect was attributable to an increase in milk production of 4.4 kg/d among cows with a body condition score > or =4 at 3 wk before calving; fat cows that received RPC ate 1.1 kg of DM/d more from wk 3 before calving through wk 4 after calving. Monensin supplementation significantly increased serum concentrations of glucose and urea, lowered concentrations of beta-hydroxybutyric acid and aspartate aminotransferase in the peripartum period, and increased liver glycogen content at 3 wk into lactation. The metabolic effects of CRC are consistent with previous studies, and the effects on liver are novel. The mechanism by which RPC increased milk production was not revealed in this study and merits further research.

Response to conjugated linoleic acid in dairy cows differing in energy and protein status.

The trans-10, cis-12 conjugated linoleic acid (CLA) isomer inhibits milk fat synthesis, whereas milk yield and synthesis of other milk components generally remain unchanged in established lactation. However, in some CLA studies increases in milk yield, milk protein yield, or both have been observed in cows limited in energy, either in early lactation or when grazing pasture. Our objective was to evaluate the performance and monitor peripheral tissue responses to homeostatic signals regulating lipolysis and glucose uptake with CLA supplementation when cows were limited in metabolizable energy in combination with moderate or excess metabolizable protein supply. Holstein cows (n = 48; 112 +/- 5 d in milk; mean +/- SE) were provided ad libitum access to a diet that met energy and protein requirements for a 16-d standardization interval. Based on performance during this interval, the Cornell Net Carbohydrate and Protein System was used to design energy-limiting rations that provided 80% of metabolizable energy requirements, and these were fed throughout the treatment periods. Cows were randomly allocated to 4 treatments, in a 2-period crossover design. Treatments were 1) moderate metabolizable protein (MP) supply, 2) moderate MP supply + CLA, 3) excess MP supply, and 4) excess MP supply + CLA. Moderate and excess MP supply were at 88 and 117%, respectively, of the MP requirement established during the standardization period, as estimated by the Cornell Net Carbohydrate and Protein System. Each experimental period comprised 16 d, with crossover of CLA within each protein level. The lipid-encapsulated CLA supplement provided 12 g/d of trans-10, cis-12 CLA. Conjugated linoleic acid treatment reduced milk fat yield by 21% but increased milk yield and milk protein yield by 2.6 and 2.8%, respectively. Milk yield and content and yield of both milk protein and fat were unaltered by either protein treatment alone or in combination with CLA. Basal concentrations of glucose, insulin, and nonesterified fatty acids were unaffected by CLA supplementation. The fractional rate of glucose clearance in response to an insulin challenge and the nonesterified fatty acid response to an epinephrine challenge were also not altered by either CLA treatment or MP supply. Overall, the results demonstrate that CLA supplementation when cows are energy-limited allows for repartitioning of nutrients, resulting in increased yields of milk and milk protein, and this can occur without changes in whole-body glucose homeostasis and adipose tissue response to lipolytic stimuli.

RNA-interference effectors and their delivery.

RNA-interference, the mechanism by which the expression of a specific protein can be reduced or eliminated, has emerged as a potential way to treat disease. RNA-interference effectors, such as small interfering RNA and small hairpin RNA, are double-stranded nucleic acid sequences expressly designed to have homology to sections of the target protein's mRNA, and when present in the cytosol trigger hydrolysis of the mRNA through the RNA-interference pathway. Because RNA-interference effectors are macromolecular and typically polyanionic, their efficacy is poor when not accompanied by a delivery vehicle. This review outlines the RNA-interference mechanism and discusses the delivery of RNA-interference effectors, with an emphasis on synthetic polymer-based delivery systems.

Dietary forage and nonfiber carbohydrate contents influence B-vitamin intake, duodenal flow, and apparent ruminal synthesis in lactating dairy cows.

The objective of this experiment was to quantify intakes, duodenal flows, and ruminal apparent synthesis (AS) of B-vitamins in lactating dairy cows fed diets varying in forage and nonfiber carbohydrate (NFC) contents. Eight (4 primiparous and 4 multiparous) ruminally and duodenally cannulated Holstein cows were assigned to 4 dietary treatments in a replicated 21-d period, 4 x 4 Latin square design with a 2 x 2 factorial treatment arrangement. Diets, fed as TMR, contained (DM basis) 2 levels of forage (35 and 60%) and 2 levels of NFC (30 and 40%). The forage portion of the diets contained 50% corn silage, 33% alfalfa hay, and 17% grass hay. Soybean hulls and beet pulp (2:1) and corn meal and ground barley (2:1) were included to achieve desired NFC concentrations. No supplemental B-vitamins were fed. B-vitamin AS was calculated as the amount of a specific B-vitamin flowing to the duodenum minus its daily orts-corrected intake. Dry matter and organic matter intakes were higher for cows fed the 35% forage diets and the 40% NFC diets. Increasing dietary forage content decreased ruminal AS of pyridoxine, folic acid, and B12. Increasing dietary NFC content increased ruminal AS of nicotinic acid, nicotinamide, niacin, pyridoxal, B6, and folic acid but decreased AS of B12. Across diets, amounts of B-vitamins synthesized were highest for niacin, followed by riboflavin, B12, thiamin, B6, and folic acid. Biotin AS values were negative for all diets, suggesting either no ruminal synthesis or that destruction by ruminal microflora was greater than synthesis. B-vitamin intake, duodenal flow, and ruminal synthesis are influenced by dietary forage and NFC contents.

Improving intestinal amino acid supply of pre- and postpartum dairy cows with rumen-protected methionine and lysine.

Eighty-four Holstein cows were assigned to a randomized block experiment to determine effects of supplementing pre- and postpartum diets containing highLys protein supplements with rumen-protected Met and Lys. Before parturition, cows received a basal diet with 1) no rumen-protected amino acids (AA), 2) 10.5 g/d of Met from rumen-protected Met, or 3) 10.2 g/d of Met and 16.0 g/d of Lys from rumen-protected Met plus Lys. After parturition, cows continued to receive AA treatments but switched to diets balanced for 16.0 or 18.5% crude protein (CP). Diets were corn-based; supplemental protein was provided by soybean products and blood meal. Cows received treatments through d 105 of lactation. Compared with basal and Met-supplemented diets, Met + Lys supplementation increased yield of energy-corrected milk, fat, and protein, and tended to increase production of 3.5% fat-corrected milk. Significant CP x AA interactions were observed only for milk protein and fat content. Supplementation of the 16% CP diet with Met and Met + Lys had no effect on milk true protein and fat content. However, Met and Met + Lys supplementation of the 18.5% CP diet increased milk protein content by 0.21 and 0.14 percentage units, respectively, and Met supplementation increased fat content by 0.26 percentage units. Results of this study indicate that early-lactation cows fed corn-based diets are responsive to increased intestinal supplies of Lys and Met and that the responses depend on dietary CP concentration, supply of metabolizable protein, and intestinal digestibility of the rumen-undegradable fraction of supplemental proteins.

Pain-free results with sumatriptan taken at the first sign of migraine pain: 2 randomized, double-blind, placebo-controlled studies.

OBJECTIVE: To evaluate the efficacy and tolerability of sumatriptan, 50-mg and 100-mg tablets, compared with placebo for treatment of migraine at the first sign of pain. PATIENTS AND METHODS: Two identical multicenter randomized, double-blind, placebo-controlled, single-attack studies were conducted from May through November 2000 in adults (aged 18-65 years). Patients treated migraine at the first sign of pain, while pain was mild, but not more than 2 hours after onset with oral sumatriptan, 50 mg or 100 mg, or matching placebo. The primary end point was pain-free relief at 2 hours after treatment with 50 mg of sumatriptan compared with placebo. RESULTS: There were 354 patients in study 1 and 337 patients in study 2. Significantly more patients treated with sumatriptan, 50 mg and 100 mg, were completely free from pain 2 and 4 hours after treatment vs patients treated with placebo (at 2 hours, 50% and 57% vs 29%; at 4 hours, 61% and 68% vs 30%; for both, P < .001). Also, significantly more patients treated with sumatriptan, 50 mg and 100 mg, were migraine-free (no pain or associated symptoms) vs those treated with placebo at 2 and 4 hours after treatment (at 2 hours, 43% and 49% vs 24%; at 4 hours, 54% and 63% vs 28%; for both, P < .001). The incidence of overall adverse events was low with the 50- and 100-mg dose of sumatriptan (placebo, 7%; sumatriptan at 50 mg, 14%; sumatriptan at 100 mg, 16%). CONCLUSIONS: Treatment of migraine at the first sign of pain with sumatriptan, 50-mg and 100-mg tablets, provides superior pain-free relief at 2 and 4 hours after treatment compared with placebo. Results of these studies suggest that sumatriptan at 100 mg may be more efficacious than at 50 mg when used in the early treatment paradigm. Because these studies were not powered to detect statistical differences between active doses, studies to investigate this finding are warranted.

Demographic and migraine characteristics of adolescents with migraine: Glaxo Wellcome clinical trials' database.

OBJECTIVE: To describe the demographics and migraine characteristics of patients in the Glaxo Wellcome adolescent clinical trials' database. METHODS: Data from 8 sumatriptan (tablet and nasal spray) and naratriptan (tablet) trials (6 placebo controlled and 2 open label) were reviewed. Adolescents aged 12 to 17 years who had participated in migraine clinical trials and used at least 1 dose of study medication were summarized using descriptive statistics. Patient demographic (gender, age, race, height, and weight) and migraine (diagnosis, pain location and intensity, time and day of migraine onset and treatment, and associated symptoms) characteristics were examined. RESULTS: One thousand nine hundred thirty-two adolescents with migraine were identified; mean age was 14.1 years (standard deviation, 1.64; range, 11 to 18) and 54% of patients were female. More males were represented in the 12- to 14-year-old group (646 [73%] of 885) than in the 15- to 17-year-old group (234 [26%] of 885). Most patients reported migraine without aura (67%, 1121 of 1672), unilateral migraine pain (58%, 458 of 787), and pulsating pain (74%, 582 of 790). Migraine was aggravated by physical activity in most of the adolescents (88%, 526 of 598). Most migraine attacks (73%, 1363 of 1858) began between 6 am and 6 pm, and proportionately more attacks occurred Monday through Wednesday. Pretreatment vomiting was experienced by 5% (97 of 1830) of patients, nausea by 53% (983 of 1849), and photophobia or phonophobia (or both) by 88% (1628 of 1858) of patients. The incidence of associated symptoms was directly related to pretreatment headache severity. CONCLUSIONS: In this large clinical trials' database, adolescents had migraine without aura characterized by unilateral and pulsating pain and aggravated by activity. The incidence of associated symptoms was directly related to pretreatment pain intensity. More migraines occurred Monday through Wednesday during typical school hours. These data may facilitate clinicians' efforts to tailor migraine therapy to the needs of this patient population.

Monocharged inhibitors of mast cell tryptase derived from potent and selective dibasic inhibitors.

Truncation of potent and selective dibasic inhibitors afforded monocharged inhibitors of human mast-cell tryptase. Using two classes of analogues as lead structures, several monocharged derivatives were identified with K(i) values ranging from 0.084 to 0.21 microM against the enzyme.

Poly(lactic acid)-poly(ethylene glycol) nanoparticles as new carriers for the delivery of plasmid DNA.

The purpose of the present work was to produce and characterize poly(lactic acid)-poly(ethylene glycol) (PLA-PEG) nanoparticles (size lower than 300 nm) containing a high loading of plasmid DNA in a free form or co-encapsulated with either poly(vinyl alcohol) (PVA) or poly(vinylpyrrolidone) (PVP). The plasmid alone or with PVA or PVP was encapsulated by two different techniques: an optimized w/o/w emulsion-solvent evaporation technique as well as by a new w/o emulsion-solvent diffusion technique. Particle size, zeta potential, plasmid DNA loading and in vitro release were determined for the three plasmid-loaded formulations. The influence of the initial plasmid loadings (5, 10, 20 microg plasmid DNA/mg PLA-PEG) on those parameters was also investigated. The plasmid loaded into the nanoparticles and released in vitro was quantified by fluorimetry and the different molecular forms were identified by gel electrophoresis. PLA-PEG nanoparticles containing plasmid DNA in a free form or co-encapsulated with PVA or PVP were obtained in the range size of 150-300 nm and with a negative zeta potential, both parameters being affected by the preparation technique. Encapsulation efficiencies were high irrespective of the presence of PVA or PVP (60-90%) and were slightly affected by the preparation technique and by the initial loading. The final plasmid DNA loading in the nanoparticles was up to 10-12 microg plasmid DNA/mg polymer. Plasmid DNA release kinetics varied depending on the plasmid incorporation technique: nanoparticles prepared by the w/o diffusion technique released their content rapidly whereas those obtained by the w/o/w showed an initial burst followed by a slow release for at least 28 days. No significant influence of the plasmid DNA loading and of the co-encapsulation of PVP or PVA on the in vitro release rate was observed. In all cases the conversion of the supercoiled form to the open circular and linear forms was detected. In conclusion, plasmid DNA can be very efficiently encapsulated, either in a free form or in combination with PVP and PVA, into PLA-PEG nanoparticles. Additionally, depending on the processing conditions, these nanoparticles release plasmid DNA either very rapidly or in a controlled manner.

Dibasic inhibitors of human mast cell tryptase. Part 3: identification of a series of potent and selective inhibitors containing the benzamidine functionality.

A survey of charged groups and linkers for a series of symmetrical and unsymmetrical dibasic inhibitors is described, leading to several classes of potent and selective inhibitors. In particular, the benzamidine functionality was identified as the most potent charged group investigated.

Naratriptan as short-term prophylaxis of menstrually associated migraine: a randomized, double-blind, placebo-controlled study.

OBJECTIVE: To determine the efficacy of naratriptan 1-mg and 2.5-mg tablets twice daily compared with placebo as short-term prophylaxis of menstrually associated migraine. BACKGROUND: Approximately 60% of women with migraine report headaches associated with their menstrual cycles. Results from an open-label study suggest that short-term administration of sumatriptan is useful in the prophylaxis of menstrually associated migraine. METHODS: A randomized, double-blind, three-arm, parallel-group, placebo-controlled study was conducted in women aged 18 years or older with a history of migraine with or without aura, as defined by the International Headache Society, of at least 6 months. Two dose strengths of naratriptan (1 mg, 2.5 mg) or identical-appearing placebo tablets (1:1:1) were administered twice daily for 5 days starting 2 days prior to the expected onset of menses across four perimenstrual periods. End points included the number of menstrually associated migraines, total migraine days, peak headache severity, lost work/activity time, migraine-related quality of life, and incidence of adverse events. RESULTS: Overall, the intent-to-treat population comprised 206 women (naratriptan 1 mg, n = 70; naratriptan 2.5 mg, n = 70, and placebo, n = 66); 171 women treated four perimenstrual periods. Significantly more perimenstrual periods per subject treated with naratriptan, 1 mg, were headache-free compared with placebo (50% versus 25%, P =.003). Naratriptan, 1 mg, significantly reduced the number of menstrually associated migraines (2.0 versus 4.0, P <.05) and menstrually associated migraine days (4.2 versus 7.0, P <.01) compared with placebo. More patients treated with naratriptan, 1 mg, were headache-free across all treated perimenstrual periods compared with placebo (23% versus 8%). No difference in headache severity was observed in breakthrough headaches. The incidence and severity of adverse events was similar across treatment groups. Naratriptan, 2.5 mg, was not statistically superior to placebo for any measure. CONCLUSIONS: Naratriptan, 1 mg, with tolerability similar to placebo, is an effective, short-term, prophylactic treatment for menstrually associated migraine.

Polymer-based gene delivery with low cytotoxicity by a unique balance of side-chain termini.

Protein expression after delivery of plasmid DNA to the cell nucleus depends on the processes of transcription and translation. Cytotoxic gene-delivery systems may compromise these processes and limit protein expression. This situation is perhaps most prevalent in current nonviral polycationic gene-delivery systems in which the polycationic nature of the delivery system can lead to cytotoxicity. To approach the problem of creating nontoxic but effective gene-delivery systems, we hypothesized that by optimizing the balance between polymer cationic density with endosomal escape moieties, effective gene transfer with low cytotoxicity could be created. As a model system, we synthesized a series of polymers whose side-chain termini varied with respect to the balance of cationic centers and endosomal escape moieties. Specifically, by polymer-analogous amidation we conjugated imidazole groups to the epsilon-amines of polylysine in varying mole ratios (73.5 mol % imidazole, 82.5 mol % imidazole, and 86.5 mol % imidazole). The primary epsilon-amine terminus of polylysine served as a model for the cationic centers, whereas the imidazole groups served as a model for the endosomal escape moieties. These polymers condensed plasmid DNA into nanostructures <150 nm and possessed little cytotoxicity in vitro. Transfection efficiency, as measured by luciferase protein expression, increased with increasing imidazole content of the polymers in a nonlinear relationship. The polymer with the highest imidazole content (86.5 mol %) mediated the highest protein expression, with levels equal to those mediated by polyethylenimine, but with little to no cytotoxicity.

Effect of early intervention with sumatriptan on migraine pain: retrospective analyses of data from three clinical trials.

OBJECTIVE: This study assessed the efficacy of sumatriptan 50- and 100-mg tablets in the treatment of migraine attacks while the pain is mild rather than moderate/severe. BACKGROUND: Results from The Spectrum Study suggested that early treatment of migraine attacks with sumatriptan 50-mg tablets while the pain is mild might enhance pain-free response and reduce headache recurrence. METHODS: Retrospective analyses of headaches treated during mild pain were performed using data from 3 studies of sumatriptan tablets (protocols S2CM09, S2BT25, and S2BT26). Our primary interest was pain-free response 2 and 4 hours after dosing; secondary interests were use of a second dose of medication, clinical disability (as measured on a 4-point disability scale), migraine-associated symptoms, meaningful pain relief (patient defined), time to meaningful relief, sustained pain-free response, and proportion of attacks in which pain had worsened 2 and 4 hours after dosing, all of which were compared in headaches treated during mild versus moderate/severe pain. RESULTS: In S2CM09, 92 patients treated 118 headaches during mild pain. Rates of pain-free response were higher 2 hours after dosing with sumatriptan 50 mg (51%) or 100 mg (67%; P < 0.05) compared with placebo (28%), and were higher with early treatment of mild pain compared with treatment of moderate/severe pain at 2 hours (sumatriptan 50 mg: mild pain, 51%; moderate/severe pain, 31%; P < 0.05; sumatriptan 100 mg: mild pain, 67%; moderate/severe pain, 36%) and 4 hours (50 mg: 75% vs 56%; 100 mg: 90% vs 61%; P < 0.05). Early intervention also resulted in less redosing than when moderate/severe pain was treated (50 mg: 21% vs 32%; 100 mg: 20% vs 29%). More attacks treated early with sumatriptan 50 or 100 mg were associated with normal function 4 hours after dosing compared with placebo (70% and 93% vs 46%, respectively). Sustained pain-free response rates 2 to 24 hours after early dosing with sumatriptan 50 or 100 mg were also higher (34% and 53%, respectively) compared with treatment of moderate/severe pain (19% and 24%, respectively). Early treatment with sumatriptan 100 mg produced significantly higher pain-free rates at 2 hours after dosing (P < 0.001) than did ergotamine plus caffeine (S2BT25: 69% vs 34%, respectively) or aspirin plus metoclopramide (S2BT26: 73% vs 25%, respectively). CONCLUSIONS: Sumatriptan 50- and 100-mg tablets are effective whether pain is mild or moderate/severe. However, treatment with sumatriptan while pain is mild provides high pain-free response rates while reducing the need for redosing, benefits not seen with ergotamine plus caffeine or aspirin plus metoclopramide.

Dibasic inhibitors of human mast cell tryptase. Part 1: synthesis and optimization of a novel class of inhibitors.

The synthesis and optimization of a novel class of reversible and active-site directed dibasic inhibitors of human mast cell tryptase are described. The compounds were shown to be both remarkably potent and selective for tryptase with Ki values for optimized inhibitors in the picomolar range.

Dibasic inhibitors of human mast cell tryptase. Part 2: structure-activity relationships and requirements for potent activity.

Detailed structure activity relationships (SARs) for a series of dibasic human tryptase inhibitors are presented. The structural requirements for potent inhibitory activity are remarkably broad with a range of core template modifications being well tolerated. Optimized inhibitors demonstrate potent anti-asthmatic activity in a sheep model of allergic asthma. APC-2059, a dibasic tryptase inhibitor with subnanomolar activity, has been advanced to phase II clinical trials for the treatment of both psoriasis and ulcerative colitis.

Effectiveness of sumatriptan in reducing productivity loss due to migraine: results of a randomized, double-blind, placebo-controlled clinical trial.

OBJECTIVE: To determine the effect of sumatriptan on migraine-related workplace productivity loss. PATIENTS AND METHODS: In this randomized, double-blind, placebo-controlled, parallel-group trial, adult migraineurs self-injected 6 mg of sumatriptan or matching placebo to treat a moderate or severe migraine within the first 4 hours of a minimum of an 8-hour work shift. Outcome measures included productivity loss and number of patients returning to normal work performance 2 hours after injection and across the work shift, time to return to normal work performance, and time to headache relief. RESULTS: A total of 206 patients underwent screening, 140 (safety population) of whom returned for clinic treatment. Of these 140 patients, 119 received migraine treatment in the workplace (intent-to-treat population), 116 of whom comprised the study population. Of these 116 patients, 76 self-administered sumatriptan, and 40 self-administered placebo. Sumatriptan treatment tended to reduce median productivity loss 2 hours after injection compared with placebo (25.2 vs 29.9 minutes, respectively; P = .14). Significant reductions in productivity loss were obtained across the work shift after sumatriptan treatment compared with placebo (36.8 vs 72.6 minutes, respectively; P = .001). Significantly more sumatriptan-treated patients vs placebo-treated patients experienced shorter return to normal work performance at 2 hours (53/76 [70%] vs 12/40 [30%], respectively) and across the work shift (64/76 [84%] vs 23/40 [58%], respectively; P < .001). Significantly more sumatriptan-treated patients experienced headache relief 1 hour after injection compared with placebo-treated patients (48/76 [63%] vs 13/40 [33%], respectively; P = .004). CONCLUSION: Across an 8-hour work shift, sumatriptan was superior to placebo in reducing productivity loss due to migraine.

Design of imidazole-containing endosomolytic biopolymers for gene delivery.

The development of safe and effective gene delivery agents poses a great challenge in the quest to make human gene therapy a reality. Cationic polymers represent one important class of materials for gene delivery, but to date they have shown only moderate efficiency. Improving the efficiency will require the design of new polymers incorporating optimized gene delivery properties. For example, inefficient release of the DNA/polymer complex from endocytic vesicles into the cytoplasm is one of the primary causes of poor gene delivery. Here we report the synthesis of a biocompatible, imidazole-containing polymer designed to overcome this obstacle. DNA/polymer polyplexes incorporating this polymer were shown to have desirable physico-chemical properties for gene delivery and are essentially nontoxic. Using this system, mammalian cells in vitro were transfected in the absence of any exogenous endosomolytic agent such as chloroquine.