Plethysmography and impulse oscillometry assessment of tiotropium and ipratropium bromide; a randomized, double-blind, placebo-controlled, cross-over study in healthy subjects.

AIMS: Spirometry, plethysmography and impulse oscillometry (IOS) measure different aspects of lung function. These methods have not been compared for their ability to assess long- and short-acting anticholinergic agents. We therefore performed a double-blind, placebo-controlled, four-way cross-over study in 30 healthy subjects. METHODS: Single doses of tiotropium bromide (Tio) 54 and 18 mcg, ipratropium bromide (IB) 40 mcg and placebo were administered. Specific conductance (sGaw), total lung capacity (TLC), inspiratory capacity (IC) and residual volume (RV) were measured using plethysmography, while IOS measured resistance (R5-25) and reactance (RF and X5). Pulmonary function was measured for 26 h post dose. RESULTS: Tio caused significant improvements in sGaw, forced expiratory voume in 1 s (FEV(1)), maximum mid-expiratory flow (MMEF) and R5-R25 at time points up to 26 h, with no clear differences between doses. IB improved the same parameters, but only up to 8 h. The weighted mean change (0-24 h) caused by Tio 54 mcg compared with placebo for FEV(1) was 240 ml (95% confidence interval 180, 300), while for sGaw the ratio of geometric means (Tio compared with placebo) was 1.35 (1.28, 1.41). Neither drug caused consistent statistically significant changes in RF, forced vital capacity, TLC or IC over 26 h. RV was significantly improved from 8 to 24 h by Tio 54 mcg only. CONCLUSIONS: In addition to spirometry, IOS resistance measurements and sGaw can distinguish between the effects of long- and shortacting anticholinergic effects in healthy subjects.

Pranlukast, a cysteinyl leukotriene receptor antagonist, attenuates allergen-induced early- and late-phase bronchoconstriction and airway hyperresponsiveness in asthmatic subjects.

BACKGROUND: The cysteinyl leukotrienes (cysLTs) have been implicated in the pathogenesis of allergen-induced airway responses. The purpose of this study was to evaluate the effects of pretreatment with the cysLT receptor antagonist pranlukast on allergen-induced early asthmatic responses (EARs) and late asthmatic responses (LARs) and on allergen-induced airway hyperresponsiveness (AHR). METHODS: Ten atopic, nonsmoking patients with mild asthma and previously demonstrated early- and late-phase allergen-induced asthmatic responses participated in a double-blind, placebo-controlled, cross-over study, comparing treatment with either 450 mg pranlukast given twice daily or placebo for 5.5 days. A methacholine challenge was performed before administration of medication, and the result was expressed as the PC20. An allergen inhalation challenge was performed on the morning of the fifth day of treatment 2 hours after administration of medication. Methacholine challenges were also performed 2 hours after medication on days 4 and 6 (24 hours before and 24 hours after allergen administration) to examine allergen-induced AHR. RESULTS: Pranlukast attenuated allergen-induced early responses, late responses, and AHR. The mean (SEM) maximal percent fall in FEV1 from baseline during the early response was 30.0% (5.1%) during placebo treatment and 15.5% (3.5%) during pranlukast treatment (mean difference, 14.5%; 95% confidence interval [CI], 5.3 to 23.7; P = .007), with a mean protection afforded by pranlukast of 48.3%. The mean maximal percent fall in FEV1 during the late response was 34.7% (5.3%) during placebo treatment and 24.0% (4.4%) during pranlukast treatment (mean difference, 10.7%; 95% CI, 4.1 to 17.3; P = .006), with a mean protection afforded by pranlukast of 30.8%. The mean allergen-induced shift in PC20 was -1.76 (0.32) doubling doses during placebo treatment and -0.38 (0.31) doubling doses during pranlukast treatment (mean difference, -1.38 doubling doses; 95% CI, 0.44 to 2.32; P = .012), with a mean protection afforded by pranlukast of 78.4%. CONCLUSION: These results demonstrate that pranlukast can attenuate allergen-induced early and late airways responses and AHR and adds further support for an important role for the cysLTs in mediating allergen-induced asthmatic responses.

Results of the first U.S. double-blind, placebo-controlled, multicenter clinical study in asthma with pranlukast, a novel leukotriene receptor antagonist.

Pranlukast (SB 205312; ONO-1078), a potent, orally active selective cysteinyl-leukotriene receptor antagonist (LTRA), was developed in Japan for the treatment of asthma. This article reports results of the initial U.S. clinical evaluation of pranlukast. The primary objective of this multicenter study was to evaluate the safety and tolerability of pranlukast administered at doses of 337.5 mg b.i.d. and 450 mg b.i.d. in 65 patients with mild to moderate asthma. Pranlukast, a novel LTRA, is safe and well tolerated at doses of 337.5 mg b.i.d. and 450 mg b.i.d. Pranlukast has demonstrated clinical activity in patients with asthma.

A placebo-controlled multicenter study of auranofin in the treatment of patients with corticosteroid-dependent asthma. Auranofin Multicenter Drug Trial.

BACKGROUND: Previous clinical studies have demonstrated that injectable gold salts and the oral gold compound, auranofin, possess significant steroid-sparing effects in the treatment of asthma. OBJECTIVES: The objectives of this investigation were to determine whether auranofin could reduce oral corticosteroid requirements and to evaluate the safety of auranofin in the treatment of chronic corticosteroid-dependent asthma. METHODS: Patients with asthma were eligible if they required at least 10 mg of prednisone per day for control and prevention of asthma exacerbations. Two hundred seventy-nine patients with chronic corticosteroid-dependent asthma (requiring > or = 10 mg/day) were randomized to receive auranofin, 3 mg twice daily, or placebo during an 8-month clinical trial, which was divided into three phases including: a 4-week baseline period (phase I), a 6-month double-blind treatment and steroid reduction period (phase II), and a 4-week posttreatment observation period during which steroid and auranofin doses or placebo doses were maintained at levels achieved by the end of phase II (phase III). The primary efficacy variable was "therapeutic success" or reduction of daily corticosteroid use by 50% or more. RESULTS: The proportion of patients in the auranofin group achieving therapeutic success (41%) was significantly higher than that in the placebo group (27%) (p = 0.01). This effect was greatest in patients requiring 10 to 19 mg of oral prednisone per day at baseline (p < 0.001). In all treated patients, including those who did and did not complete the trial, significant reduction (> or = 50% of baseline) in oral corticosteroid dosage was achieved in the auranofin group (60%) compared with the placebo group (32%) (p < 0.001). There were no significant differences between treatment groups in symptoms, concomitant medication use, or lung function. Mean serum total IgE levels decreased significantly from baseline in the auranofin group (-44.63 IU/ml) compared with the placebo group (p = 0.001). Gastrointestinal and cutaneous adverse events were greater in the auranofin group. CONCLUSIONS: Auranofin demonstrated a steroid-sparing effect without concomitant worsening of symptoms or lung function and appeared to be more effective in patients dependent on 10 to 19 mg of prednisone per day. Therefore this study has demonstrated that auranofin is useful as a steroid-sparing agent in the treatment of chronic corticosteroid-dependent asthma.

Reaction of antipolynucleotide antibody from systemic lupus erythematosus patient serum with double-stranded DNA complexed to protein.

Antipolynucleotide antibody from systemic lupus erythematosus patient serum was found to interact with common determinants of double-stranded DNA (dsDNA) complexed to protein within nucleosomes, and with protein-free denatured DNA. In contrast, purified dsDNA isolated from nucleosomes manifested decreased reactivity with antipolynucleotide antibody. These data suggest that immunoassays using denatured DNA antigen substrate are useful for identifying a broad spectrum of antipolynucleotide antibodies that are reactive with dsDNA complexed to protein.

Inhibition of leukocyte adherence by potassium chloride extract of allogeneic squamous cell carcinoma of the human uterine cervix.

Leukocyte adherence inhibition (LAI) assay was used to evaluate cell-mediated immunity in patients with invasive squamous cell carcinoma of the cervix. The reactivity of the peripheral blood leukocytes of these patients was evaluated after incubation with pooled extracts of allogeneic squamous cell carcinoma of the cervix. One hundred sixty-seven sets of LAI assays were performed on 54 individuals, including 23 patients with Stage I squamous cell carcinoma of the cervix, 9 patients with other stages of this tumor, 9 patients with unrelated tumors and 13 normal healthy volunteers. A protein concentration of one milligram per milliliter in the tumor extract and 10% fetal bovine serum in the feeding media gave the best results. Eighty-seven percent (28/32) of patients with squamous cell carcinoma of the cervix showed marked specific reactivity. No difference was found in the LAI indices of different stages of the disease.

Sequential in vitro reactivity of lymphocytes from patients with cervical squamous malignancy in a cytotoxicity assay.

The in vitro reactivity of peripheral blood lymphocytes from patients with cervical squamous malignancy was prospectively followed over a relatively long period of time. In 12 of 14 patients with preinvasive cervical lesions, reactivity was present at the time of initial diagnosis. Three months after treatment, reactivity was still present in only one of 12 (8%). Six months after the treatment, no significantly reactivity could be detected in any of them. In the group with invasive squamous cell carcinoma, four patients developed recurrence in the course of follow-up. Reactivity was present in all at the time of initial diagnosis. In three of them, cytotoxic reactivity was retained up to the time of death, in spite of clinical deterioration. One lost the reactivity after the third course of chemotherapy. Fourteen patients with invasive squamous cell carcinoma were followed up to 24 months without any evidence of recurrence. In all of this group, the cell-mediated cytotoxicity was nonreactive at 9 months. We conclude that: (1) patients with squamous cell carcinoma of the cervix demonstrated cell-mediated immune responses which disappeared 3 to 9 months after effective treatment (apparent cure) and (2) with persistent disease, in spite of marked clinical deterioration and inanition, cell-mediated cytotoxicity was demonstrable until the time of death.

Partial characterization of serum suppressor factors in patients with cervical squamous carcinoma.

A cell-mediated cytotoxicity system was used to characterize the suppressor factors in sera from patients with invasive squamous cell carcinoma of the uterine cervix. The immunoglobulin-M fraction of the sera of 17 patients with different stages of invasive squamous cell carcinoma of the cervix, separated by Sephadex gel filtration, were tested. All showed marked cytotoxic suppressor activity, including two cases in which sera showed only mild activity. The immunoglobulin-G fraction of sera from 13 patients with invasive squamous cell carcinoma of the cervix was separated by ion exchange chromatography. The cytotoxic suppressor activity of the immunoglobulin-G fraction proved to be comparable in effect to that of the whole sera. By using increasing dilutions of IgG and IgM fractions, it could be demonstrated that IgM fraction retained its activity at 0.01 +/- 0.005 mg/ml whereas IgG fraction was devoid of activity at this concentration, suggesting that these two immunoglobulin fractions act independently. It is suggested that the cytotoxic suppressor activity in sera from patients with invasive squamous cell carcinoma of the cervix resides in the gamma globulin fraction and that both immunoglobulin-M and immunoglobulin-G of their sera manifest the cytotoxic suppressor activity independently. (Am J Reprod Immunol. 2:199-203.)

Pathology of metastasizing tumors in nitrosomethyl urea-induced rat mammary carcinoma.

The metastatic capacity of rat mammary tumors induced with N-nitrosomethyl urea was tested in BUF/N inbred female rats by successive transplantation. After the first and second passage, tumor cells appeared diffusely distributed throughout the bone marrow and spleen, confirming results reported by others; no other metastases were observed. After six successive transplantations, large, well-defined tumor nodules were observed in the liver, spleen, lung, and the peritoneal surface of the intestines in 40% of the injected animals. The morphology of the primary and metastatic tumors was compared by light microscopy. The tumors appeared to be adenocarcinomas with differing degrees of differentiation. No morphological differences could be observed between the primary and the metastatic tumors.

Cytotoxic blocking activity in invasive squamous cell carcinoma of the human uterine cervix.

A cell-mediated cytotoxicity assay was used to evaluate the specific reactivity of patients with squamous cell carcinoma to an established cell line derived from squamous cell carcinoma of the cervix. Heat inactivated sera from patients with squamous cell carcinoma could very effectively nullify this specific cytotoxicity (P less than 0.01). There was no significant difference between the cytotoxic blocking activity of the sera from patients with different stages of the cancer in this experimental setup.

Cell-mediated cytotoxicity in preinvasive and invasive squamous cell carcinoma of the cervix.

Cell-mediated immunity was tested by coincubation of target cell line (2043) of human squamous cell carcinoma and peripheral blood leukocytes (PBL) of 22 patients with squamous cell neoplasia of the uterine cervix, 9 patients with other tumors, and 9 normal females. The percentage of cell reduction in mild dysplasia was 90.1 +/- 6.8% (.001 less than P less than .005), in moderate dysplasia was 91.1 +/- 6.4% (.001 less than P less than .005), in severe dysplasia was 91.6 +/- 15.6% (P less than .001), in carcinoma in situ was 85.0 +/ 2.6% (P less than .001), and in invasive squamous cell carcinoma of the cervix was 85.0 +/- 6.9% (P less than .001). Peripheral blood leukocytes of patients with squamous neoplasia did not show any significant cytocoxicity against cell line SKOV-3, developed from an ovarian adenocarcinoma, nor did the PBL of patients with "other tumors" show any significant cytotoxicity against cell line 2043. This study shows that even in early stages of preinvasive squamous cell carcinoma of the cervix, the PBL are sensitized against the neoplastic process, confirming that different stages of cervical intraepithelial neoplasia and invasive squamous cell carcinoma are different intensities of the same biologic process.

Isolation and characterization of a human liver and kidney-specific protein: the hepato-renal (H-R) antigen.

This paper reports the isolation and characterization of a soluble antigen shared by the liver and kidney of human and some other animal species. Homogenates of human liver in saline were centrifugated at 27,000 g and the supernatants were fractionated by preparative polyacrylamide gel electrophoresis. The gels were divided in sections and each was injected into rabbits; after absorption with polymerized normal human serum, the antiserum obtained by injecting one of the sections reacted only with saline extracts of human liver and kidney when tested against a variety of human tissue extracts. The absorbed antiserum, polymerized and insolubilized with glutaraldehyde, was used to purify the antigen by affinity chromatography. The purified antigen proved to be a glycoprotein containing 19 percent carbohydrate, had a molecular weight of 5.8-6.0 x 10(4) Daltons and a pI of 7.2-7.4. The antigen, relatively thermostable, was precipitated by 35-55 percent ammonium sulphate; its antigenic activity was not affected by extraction with 0.6 N perchloric acid or by incubation with ribonuclease, deoxyribonuclease or neuraminidase but was destroyed by incubation with ttypsin or chymotrypsin. Immunoperoxidase studies showed that the antigen appeared concentrated in the neclei of liver and kidney glomerular epithelial and tubular epithelial cells in humans and rats. The antigen could not be detected in human hepatomas or hypernephromas or in the rat Morris hepatoma 5123.

Isolation and characterization of a human pancreas-specific protein.

Homogenates of human pancreas in saline were centrifuged at 27 000 X g and the supernates were fractionated by preparative polyacrylamide gel electrophoresis. The gels were divided into sections and each section was injected into rabbits; after absorption with polymerized serum from apparently normal humans, the antiserum obtained by injecting one of the sections was tested against a variety of human tissue extracts but reacted only with saline extracts of human pancreas. The absorbed antiserum, polymerized and made insoluble with glutaraldehyde, was used to purify a pancreas-specific antigen by immunoaffinity batch technique. The purified antigen proved to be a protein with some carbohydrate content (180 mg/g by weight) and a molecular mass of about 2.25 X 10(5) daltons. The antigen is relatively thermostable, and precipitates in the range of 245.64-340.2 g/L saturated ammonium sulfate; its antigenic activity is not affected by incubation with ribonuclease or deoxyribonuclease, but is destroyed by incubation with trypsin or neuraminidase and by extraction with perchloric acid. Immunofluorescence studies show that the antigen is diffusely present in the cytoplasm of pancreatic acinar cells.

Leukocyte migration in ovarian carcinoma: comparison of inhibitory activity of tumor extracts.

Inhibition of migration of leukocytes from patients with serous cystadenocarcinoma of the ovary was studied by the use of several different types of ovarian carcinoma extract as antigen. KCl extract of an ovarian carconoma was found to be the most effective antigen preparation in comparison with saline, deoxycholate, and perchloric acid extracts. Low concentrations of KCl ovarian carcinoma extract significantly inhibited migration of leukocytes from 11 of 17 patients with ovarian carcinoma (migration index, less than 0.74). Leukocytes from patients with breast, colon, or endometrial carcinoma showed minimal reactivity with ovarian carcinoma KCl extract, and leukocytes from patients with ovarian carcinoma showed minimal reactivity with KCl extracts of breast, colon, and endometrial carcinoma. These results suggested that the 3 M KCl procedure is superior for the isolation of antigens active in the leukocyte migration inhibition test and that this test may be of use for the isolation of tumor-associated antigen and the immunodiagnosis of ovarian carcinoma.

Radioimmunoassay for antibodies to cytoplasmic ribosomes in human serum.

A radioimmunoassay for the detection of antibodies in human serum to tritium-labeled HeLa cell cytoplasmic ribosomes was developed with the use of Macaloid for the inhibition of endogenous ribonuclease activity. Antibodies were observed in the serum of patients with systemic lupus erythematosus in high incidence and titer. Patients with rheumatoid arthritis and chronic active hepatitis manifested a lower incidence and titer of antibodies to ribosomes, whereas serums from normal individuals and from patients with sarcoidosis, chronic glomerulonephritis, and malignant tumors showed no significant reactivity with cytoplasmic ribosomes. Maximum inhibition of the reaction was achieved with unlabeled HeLa cell ribosomes or rat liver ribosomes and partial inhibition by purified ribosomal RNA.