Preclinical Pharmacokinetics and Acute Toxicity in Rats of 5-{[(2E)-3-Bromo-3-carboxyprop-2-enoyl]amino}-2-hydroxybenzoic Acid: A Novel 5-Aminosalicylic Acid Derivative with Potent Anti-Inflammatory Activity.

Compound 5-{[(2E)-3-bromo-3-carboxyprop-2-enoyl]amino}-2-hydroxybenzoic acid (C1), a new 5-aminosalicylic acid (5-ASA) derivative, has proven to be an antioxidant in vitro and an anti-inflammatory agent in mice. The in vivo inhibition of myeloperoxidase was comparable to that of indomethacin. The aim of this study was to take another step in the preclinical evaluation of C1 by examining acute toxicity with the up-and-down OECD method and pharmacokinetic profiles by administration of the compound to Wistar rats through intravenous (i.v.), oral (p.o.), and intraperitoneal (i.p.) routes. According to the Globally Harmonized System, C1 belongs to categories 4 and 5 for the i.p. and p.o. routes, respectively. An RP-HPLC method for C1 quantification in plasma was successfully validated. Regarding the pharmacokinetic profile, the elimination half-life was approximately 0.9 h with a clearance of 24 mL/min after i.v. administration of C1 (50 mg/kg). After p.o. administration (50 mg/kg), the maximum plasma concentration was reached at 33 min, the oral bioavailability was about 77%, and the compound was amply distributed to all tissues evaluated. Therefore, C1 administered p.o. in rats is suitable for reaching the colon where it can exert its effect, suggesting an important advantage over 5-ASA and indomethacin in treating ulcerative colitis and Crohn's disease.

In vitro release kinetics of graft matrices from Lannea coromandelica (Houtt) gum for treatment of colonic diseases by 5-ASA.

Polyacrylamide graft Jhingan gum (Jh-g-PAMs) was synthesized adopting microwave assisted graft co-polymerization technique. The synthesized graft copolymer was characterized by various analytical techniques such as Elemental analysis, FTIR, TGA, XRD and NMR. Following standard protocol, drug matrix tablets using 5-Aminosalicylic Acid (5-ASA) were prepared and swelling and erosion studies were carried out in different pH dissolution media. The result revealed that maximum swelling and erosion took place in pH 7.4 while the lowest was recorded in pH 1.2. The 'in vitro' drug release studies revealed that grades with higher grafting % exhibited more sustained release. The highest sustained release was observed in Jh-g-PAM 3 (%G 1231) in pH 1.2 while the least was observed in native gum in pH 7.4. Furthermore, the kinetic studies revealed that 'n' values in all dissolution media lies within 0.5-1.0 which suggested non-Fickian diffusion mode of release. From the above results, it can be said that controlled release of 5-ASA using graft material was successful and hence it can be explored for treatment of colon related diseases.

Mechanistic studies on genotoxicity and carcinogenicity of salicylazosulfapyridine an anti-inflammatory medicine.

Salicylazosulfapyridine (SASP), which has been in clinical use for over 50 years, was reported by the National Toxicology Program to increase rat (F344 strain) urinary bladder and mouse (B6C3F1 hybrid) liver tumours under ad libitum (AL) feeding conditions, while under a feed restriction (FR) regimen, these tumours were not increased. The present investigations were undertaken to assess the implications of these results for the safety of SASP in humans. SASP and its 2 major metabolites, 5-aminosalicylic acid (ASA) and sulfapyridine (SP) were tested for in vivo induction of micronuclei in mouse bone marrow cells with or without prefolic treatment and for in vivo formation of DNA adducts in rat and mouse liver and urinary bladder. None exhibited mutagenicity or DNA reactivity. SASP and SP have induced sister chromatid exchanges and micronuclei (MN) in cultured human lymphocytes in the absence of liver activation enzymes and in B6C3F1 mice (but not in rats) MN in bone marrow and peripheral RBC. Treatment with folate reduces the frequency of MN. Perhaps the short (28 days) RBC lifespan in mouse underlies the sensitivity of this species. Thus, SASP without folate supplementation is an aneuploidogen. In a 2-year study in AL fed SASP-treated (high dose 337.5 mg/kg) rats, urinary pH was increased and urinary specific gravity was reduced at 60 weeks. At the end, this SASP group showed urothelial hyperplasia and papillomas in the urinary bladders of male rats primarily. In the FR high dose SASP group, the hyperplasia was reduced from 82% to 14%. At the end of 2 years, the incidence of multi-organ leukemia was reduced in both AL and FR high dose SASP groups. Thus, SASP caused intraluminal bladder changes in the rat (especially males) consisting of chronic urothelial stimulation, concretions, hyperplasia which resulted in neoplasia. In the mouse, because of species differences in liver ratios (mouse > rat) and, increasing (3 times higher) liver perfusion in the mouse, compared to the rat, there was hepatocellular toxicity and resulting preneoplasia and neoplasia within 2 years. These findings occurred in all AL SASP groups (flat curve without dose response); but were reduced under FR conditions. In this species, the multiorgan lymphoma incidence was reduced in both AL and FR high dose SASP groups. Thus, SASP and its major metabolites are not genotoxic. Folate deficiency associated with SASP administration is probably responsible for aneuploidy in lymphocytes and erythrocytes. SASP does not induce neoplasia directly in either livers, urinary bladders or other organs. Accordingly, SASP is judged to pose no carcinogenic risk to humans.

Distribution and concentrations of 5-aminosalicylic acid in rectosigmoid biopsy specimens after rectal administration.

PURPOSE: Using the autofluorescent properties of 5-aminosalicylic acid (5-ASA), we studied the penetration and distribution of this molecule in human colonic biopsies at different time intervals after administration of 5-ASA enemas. METHODS: Fluorescence scores of rectosigmoidal biopsy specimens were compared with 5-ASA and acetyl-5-aminosalicylic (Ac-5-ASA) concentrations, determined by high-performance liquid chromatography, in adjacent biopsies and in serum samples. RESULTS: 5-ASA penetrates into the rectal mucosa and into the epithelial cells after local application by means of an enema. We found a characteristic 5-ASA staining of two intramucosal structures that need further identification: intraepithelial triangular configurations and "5-ASA scavengers" in the lamina propria. Fluorescence scores correlate well with 5-ASA concentrations in adjacent biopsies (r = 0.67; P < 0.005) and correlate even better with serum concentrations of 5-ASA (r = 0.84; P < 0.005) and Ac-5-ASA (r = 0.80; P < 0.005), hence reflecting the amount of systemically absorbed and metabolized 5-ASA. CONCLUSION: 5-ASA penetrates into the rectal mucosa after local application. Local availability, assessed by means of fluorescence microscopy, correlates well with serum concentrations.

Distribution of mesalazine enemas in active and quiescent ulcerative colitis.

BACKGROUND: The efficacy of mesalazine enemas depends on intraluminal concentration of the drug and is therefore limited by the enema distribution in the colon. Active ulcerative colitis changes colon motility and this leads to uncertainty about enema spread. AIM: To assess the influence of disease activity on enema distribution, we conducted a physician-blinded, longitudinal study of the retrograde spread of three mesalazine enemas. METHODS: Thirty-one patients with mild to moderate ulcerative colitis were subdivided into three groups, and treated with 2 g mesalazine in 30 mL (group I, n = 10), 4 g mesalazine in 60 mL (group II, n = 12) or 1 g mesalazine in 100 mL (group III, n = 9). All patients received oral mesalazine 500 mg t.d.s. Enemas were labelled by adding 10 MBq (99mTc)technetium-sulphur colloid. Anterior scintigraphic images were taken at the start of the study and after 12 weeks of therapy; retrograde spread was assessed by calculating the percentage of the enema in each colonic segment. RESULTS: The activity score of ulcerative colitis diminished significantly after 12 weeks of treatment, but five patients dropped out of the study. At the start of treatment enema activity in group I was mainly concentrated in the sigmoid (99%); in group II activity was found in the rectum (9%), the sigmoid (61%) and the descending colon (15%); in group III activity was distributed between the sigmoid (66%) and descending colon (25%). The colonic distribution of mesalazine enemas was not influenced by disease activity. CONCLUSION: Volume, but not disease activity, is the important determinant of retrograde colonic spread of mesalazine enemas in ulcerative colitis.

Colonic spreading of a non-chlorofluorocarbon mesalazine rectal foam enema in patients with quiescent ulcerative colitis.

BACKGROUND: Rectal foam enemas provide for drug delivery to the distal colon for treatment of left sided ulcerative colitis. However, currently available formulations contain chlorofluorocarbons which are due to be phased out in the near future. The objective of this study was therefore to determine the degree of dispersion of a newly developed non-chlorofluorocarbon rectal foam preparation in ulcerative colitis patients. METHODS: This was an open label non-controlled study of a single administration of a mesalazine foam enema (two actuations containing 2 g of mesalazine in approximately 120 mL foam) in 10 patients with quiescent ulcerative colitis. Spreading of the 99mTc-labelled foam enema was assessed over a 4-h period by the non-invasive technique of gamma scintigraphy. RESULTS: All patients retained the enema for the full 4-h imaging period. In nine out of the 10 patients, the enema was observed to spread as far as the descending colon and on average 23% of the dose was present in the descending colon at 4 h post-dose. CONCLUSIONS: The extent of spreading observed in the study supports the use of the formulation in the treatment of left sided ulcerative colitis.

Spread and distribution of 5-ASA colonic foam and 5-ASA enema in patients with ulcerative colitis.

Rectal treatment with enemas, foams, and suppositories is the most efficient method of delivering an adequate quantity of locally active drugs to the distal colon. In a pilot study carried out by colonoscopy in four patients, it was observed that 4 g 5-ASA in 20 ml foam spread up or beyond the splenic flexure and more extensively than 2 g 5-ASA in 10 ml foam. Therefore we have undertaken a study in order to compare by scintigraphy the colonic distribution of 4 g 5-ASA foam versus 4 g 5-ASA in 100 ml liquid enemas in 10 patients with ulcerative colitis using a crossover randomized design. Both preparations were labeled with 100 MBq [99mTc]sulfur colloid before administration. Anterior scans were taken at intervals for 4 hr. Activity, expressed as a percentage of total radioactivity, was measured in the rectum, sigmoid, descending, transverse, and ascending colon. Six patients had the same extent of spread with the two formulations; in three patients with foam and in one patient with enema a greater spread was observed. The foam reached the upper limit of disease in all cases, while enema failed in two cases. The maximum spread with foam was observed within 30 min in nine of 10 patients compared with seven of 10 after enema. Compared to enema, foam distributes more uniformly and seems to persist longer in the descending and sigmoid colon. The 5-ASA colonic foam shows some more favorable characteristics than enema for the local treatment of left-sided ulcerative colitis.

Distribution of mesalamine enemas in patients with active distal ulcerative colitis.

Mesalamine (5-aminosalicylic acid), a topically administered anti-inflammatory agent, is effective treatment by enema for distal ulcerative colitis; it lacks many of the side effects of orally administered sulfasalazine. In this study, we determined the colonic distribution of a 60-ml mesalamine enema in eight patients (five women and three men, 18 to 48 years old) with active distal ulcerative colitis that ranged from 12 to 40 cm proximal to the anal verge. On 3 consecutive days, each patient self-administered a 4-g (60-ml) 5-aminosalicylic acid enema that contained 3.7 MBq of [99mTc]technetium-sulfur colloid. Anterior and posterior images were obtained at 0, 30, 60, 120, and 240 minutes. During the 4-hour study period, all patients retained the enemas. The enemas spread to the sigmoid region in 24 of 24 studies, to the splenic flexure region in 22 of 24, and to the transverse colon in 1 of 24. Most of the enema was retained in the sigmoid colon. Therefore, we conclude that a 60-ml enema, when administered as recommended clinically, routinely flows retrograde as far as the splenic flexure but rarely spreads beyond this point. These results support the use of intrarectally administered 5-aminosalicylic acid for segmental colitis of the descending colon.

Retrograde distribution of a new 5-aminosalicylic acid enema in patients with ulcerative colitis.

Retrograde colonic distribution of a new 4 g 5-aminosalicylic acid enema (mesalazine) was investigated in seven patients with ulcerative colitis, five of whom were in remission. The enema was labeled with 99m-technetium and imaged by a gamma camera. A median of 86 percent (range 57-90) of the enema had spread beyond the rectum during the first two hours after administration. In four of the five examined patients with left-sided ulcerative colitis, the diseased mucosa was covered within the first 4 hours. In the fifth patient, with involvement of the descending colon, the enema did not spread beyond the very tortuous sigmoid colon 4 or even 8 hours after enema administration.

Availability of mesalazine (5-aminosalicylic acid) from enemas and suppositories during steady-state conditions.

The local and systemic bioavailability of a mesalazine enema (Pentasa, Ferring A/S, Denmark) and a mesalazine suppository (Pentasa, Ferring) was assessed during steady-state conditions. Eleven healthy subjects took 1 g of the enema or the suppository twice daily for 1 week, with a drug-free period of at least 1 week in between. At the end of each treatment period the urine and faeces were collected for 48 h, and the concentrations of mesalazine and the metabolite acetyl-mesalazine were measured. Plasma concentrations of drug and metabolite were measured hourly during a 12-h dose interval. The faecal water concentration of mesalazine was significantly higher after suppository treatment (55.7 mmol/l) compared with enema treatment (31.7 mmol/l) (p less than 0.01). The systemic absorption was low; 15% of daily mesalazine dose was recovered in urine after enema treatment and 10% after suppositores (p less than 0.01). Plasma concentrations were low, and no accumulation of either mesalazine or acetyl-mesalazine occurred. In conclusion, the enema and the suppository can be continuously administered as 1 g of mesalazine twice daily, respectively, giving high faecal water concentrations of mesalazine and a low systemic absorption.

Steady-state pharmacokinetics of a new 4-gram 5-aminosalicylic acid retention enema in patients with ulcerative colitis in remission.

The pharmacokinetic profile of a new 4-g 5-aminosalicyclic acid (5-ASA) retention enema, Mesasal, was investigated. Nine patients with ulcerative colitis in remission and one patient with mild disease activity received one enema for seven consecutive nights. They were admitted to hospital for administration of the eighth enema. Plasma concentration and urinary excretion of 5-ASA and acetyl-5-aminosalicyclic acid (Ac-5-ASA) were studied for 45 h and faecal excretion for 24 h after administration of the last enema. The median peak plasma concentration of 5-ASA was 0.92 (range, 0.59-1.87) micrograms/ml at a median of 11 h after administration, and of Ac-5-ASA 1.62 (range, 1.03-4.36) micrograms/ml at a median of 12 h after administration. On average, the plasma concentration of Ac-5-ASA was almost twice that of 5-ASA at each sampling period. At 24 h after administration the median plasma concentration for 5-ASA was 0.12 (range, 0-0.77) micrograms/ml and for Ac-5-ASA 0.36 (range, 0.01-1.6) micrograms/ml. At 45 h after administration low levels of both 5-ASA (less than 0.2 micrograms/ml) and Ac-5-ASA (less than 0.3 microgram/ml) were noted in two patients, low levels of only Ac-5-ASA (less than 0.1 microgram/ml) in two patients, and neither 5-ASA nor Ac-5-ASA in the other six patients. All patients had detectable urinary levels of both 5-ASA and Ac-5-ASA during the first 4 h after administration. Median urinary recovery during 45 h was 12.6% (range, 5.6-22.2%), indicating a low absorption at steady-state conditions.(ABSTRACT TRUNCATED AT 250 WORDS)

Mesalamine in ulcerative colitis.

Sulfasalazine has been used for many years in the management of ulcerative colitis. As many as 20 percent of patients treated with it experience intolerable adverse effects usually attributed to its sulfapyridine component. The other active component is 5-aminosalicylic acid (5-ASA); the only 5-ASA enema preparation currently available in the U.S. is mesalamine (Rowasa, Reid-Rowell) containing 5-ASA 4 g in 60 mL. In clinical trials, mesalamine has proved efficacious in treating refractory cases of distal ulcerative colitis, proctitis, and proctosigmoiditis. Because of its high cost compared with more conventional treatment modalities, it should be reserved for cases that are either refractory or intolerant to conventional treatment.

Topical treatment of ulcerative colitis.

It is clear that the therapeutic options available for the treatment of ulcerative proctosigmoiditis have increased over the last few years and in the future additional therapies will be available. Therapy will have to be individualized. Studies to date have generally failed to confirm the superiority of one form of treatment over another, although there is some evidence that high-dose 5-ASA enemas are superior to hydrocortisone enemas. Patients who fail to respond to one form of therapy may respond to another therapeutic modality. Continued evaluation of topical therapies provides not only an opportunity to improve the treatment prospects for patients but also allows for examination of potential mechanisms of action of therapeutically active compounds. Future research directions should also include assessment of combination therapy, the more widespread use of suppositories, and strategies to encourage patient compliance. Larger multicenter trials are needed to assess the effectiveness of some of the newer compounds. The role of topical therapy for patients with Crohn's disease confined to the left colon requires evaluation.

Mesalazine: an overview of key preclinical studies.

Studies were undertaken to evaluate the clinical relevance of toxicologic assessments of mesalazine and to establish mesalazine pharamcokinetics. In the toxicology studies groups of rats and dogs received various doses of mesalazine, ranging from 40 to 320 mg/kg of body weight, for 6 or 12 months. Evidence of renal papillary necrosis was found at 60 or 100 mg/kg in dogs and 320 mg/kg in rats. Data are presented to show that drug absorption after the clinical doses of mesalazine given to human patients is much lower than after the potentially nephrotoxic doses given to experimental animals. In the pharmacokinetic studies plasma from rats and dogs given various doses of mesalazine was analysed for mesalazine and its major metabolites. The results demonstrated that mesalazine in pharmaceutic preparations is not likely to present a nephrotoxic risk to patients, given the low systemic exposure achieved. After 12 months of mesalazine administration to dogs, many showed an ocular condition diagnosed as keratoconjunctivitis sicca. Experimental and epidemiologic evidence demonstrates that this condition is peculiar to dogs, and there are no reports of untoward ocular effects in patients receiving long-term therapy with mesalazine.

Quantitative distribution of radiolabeled 5-aminosalicylic acid enemas in patients with left-sided ulcerative colitis.

Rectally administered suspensions of 5-aminosalicylic acid (5-ASA) are topically effective in treating left-sided ulcerative colitis. The extent to which the contents of these enemas are distributed to inflamed mucosal linings has not previously been determined. This study was undertaken to validate a technique for labeling 5-ASA with 99mTc and to quantitate the distribution of [99mTc]5-ASA in eight patients with left-sided ulcerative colitis. Eight patients underwent three colonic scintigraphic exams within five days, receiving a 60-ml radiolabeled 5-ASA enema into the unprepared rectum for each study, with sequential anterior abdominal images obtained for 4 hr. Activity within the rectum, sigmoid, descending, transverse, and ascending colon was quantitated. Over 50% of the labeled enema had advanced beyond the rectum in five of eight patients and in six of eight patients by 30 min and 60 min, respectively. The distribution of [99mTc]5-ASA was quantitatively reproducible when repeated in the same patient on different days, despite apparent visual differences. By 2 hr, the amount of the enema present within the rectum decreased significantly (P less than 0.05) compared to the initial distribution. The amount of enema present within the descending colon was increased significantly at 0.5 hr (P less than 0.05) and at 2 hr (P less than 0.01). There were no significant changes in the distribution from initial values for the sigmoid, transverse, or ascending colon at any time. In each of these cases the spread of the enema to or beyond the extent of disease was documented. In patients with left-sided ulcerative colitis, small volume [99mTc]5-ASA enemas reliably reach the area of inflammation.

Mesalazine. A review of its pharmacodynamic and pharmacokinetic properties, and therapeutic potential in chronic inflammatory bowel disease.

Mesalazine (5-aminosalicylic acid; mesalamine), the active moiety of sulphasalazine (salazosulfapyridine), is available in specially formulated oral and rectal forms for the treatment of active ulcerative colitis of mild to moderate severity and for maintenance therapy during disease remission. Tablets or capsules coated with acrylic-based resin and tablets containing microgranules coated with ethylcellulose deliver mesalazine to the distal small intestine and colon, thus avoiding the need for the carrier, sulphapyridine, which is responsible for many of the adverse effects associated with sulphasalazine. Since mesalazine is released in the small intestine from some coated preparations in contrast to sulphasalazine, these oral formulations have therapeutic potential in Crohn's disease. A limited number of therapeutic trials suggest that orally administered mesalazine 1.5 to 2.4g daily is of similar efficacy to sulphasalazine 2 to 3g daily in patients with mild to moderate ulcerative colitis. The efficacy of mesalazine enemas has been more widely investigated, a dose of 1 to 4g once daily being consistently more effective than placebo and apparently similar to enemas of prednisone 25mg or oral sulphasalazine 3g. Initial results suggest that mesalazine 4g enemas may be more effective than those containing hydrocortisone 100mg. Mesalazine and sulphasalazine in approximately equivalent oral dosages are similarly effective in maintaining remission in ulcerative colitis. Orally administered coated mesalazine is generally well tolerated by about 85% of patients allergic to or intolerant of sulphasalazine, the remainder experiencing similar reactions to both drugs. Adverse effects of mesalazine enemas are confined to local irritation and effects resulting from enema-tip insertion. Thus, orally administered coated mesalazine is a suitable alternative to sulphasalazine in the treatment of patients with mild to moderate active distal ulcerative colitis and for maintaining remission particularly in patients allergic to or intolerant of sulphasalazine. In patients who find enema therapy acceptable, mesalazine enemas are effective and well tolerated.

5-Aminosalicylic acid in patients with ulcerative colitis in remission: plasma levels after administration of a new rectal enema.

Plasma levels of total 5-aminosalicylic acid (5-ASA) were determined after a single administration of 2 g and 4 g 5-ASA enemas to 6 patients with ulcerative colitis. The mean plasma levels and AUC values confirmed that the active substance is poorly absorbed by rectal route.