Alenka Boban Blagaic

Pentadecapeptide BPC 157 and its effects on a NSAID toxicity model : Diclofenac-induced gastrointestinal, liver, and encephalopathy lesions

AIMS We attempted to fully antagonize the extensive toxicity caused by NSAIDs (using diclofenac as a prototype). MAIN METHODS Herein, we used the stable gastric pentadecapeptide BPC 157 (GEPPPGKPADDAGLV, MW 1419), an anti-ulcer peptide shown to be efficient in inflammatory bowel disease clinical trials (PL 14736) and various wound treatments with no toxicity reported. This peptide was given to antagonize combined gastrointestinal, liver, and brain toxicity induced by diclofenac (12.5mg/kg intraperitoneally, once daily for 3 days) in rats. KEY FINDINGS Already considered a drug that can reverse the toxic side effects of NSAIDs, BPC 157 (10 μg/kg, 10 ng/kg) was strongly effective throughout the entire experiment when given (i) intraperitoneally immediately after diclofenac or (ii) per-orally in drinking water (0.16 μg/mL, 0.16 ng/mL). Without BPC 157 treatment, at 3h following the last diclofenac challenge, we encountered a complex deleterious circuit of diclofenac toxicity characterized by severe gastric, intestinal and liver lesions, increased bilirubin, aspartate transaminase (AST), alanine transaminase (ALT) serum values, increased liver weight, prolonged sedation/unconsciousness (after any diclofenac challenge) and finally hepatic encephalopathy (brain edema particularly located in the cerebral cortex and cerebellum, more in white than in gray matter, damaged red neurons, particularly in the cerebral cortex and cerebellar nuclei, Purkinje cells and less commonly in the hippocampal neurons). SIGNIFICANCE The very extensive antagonization of diclofenac toxicity achieved with BPC 157 (μg-/ng-regimen, intraperitoneally, per-orally) may encourage its further use as a therapy to counteract diclofenac- and other NSAID-induced toxicity.

Stable gastric pentadecapeptide BPC 157 in trials for inflammatory bowel disease (PL-10, PLD-116, PL14736, Pliva, Croatia) heals ileoileal anastomosis in the rat.

PurposeGastric pentadecapeptide BPC 157 (BPC 157), which has been shown to be safe in clinical trials for inflammatory bowel disease (PL-10, PLD-116, PL14736, Pliva, Croatia), may be able to cure intestinal anastomosis dehiscence. This antiulcer peptide shows no toxicity, is limit test negative, and a lethal dose is not achieved. It is stable in human gastric juice. In comparison with other standard treatments it is more effective for ulcers and various wounds, and can be used without a carrier needed for other peptides, both locally and systemically (i.e., perorally, parenterally). We studied the effectiveness of BPC 157 for ileoileal anastomosis healing in rats.MethodsWe assessed ileoileal anastomosis dehiscence macroscopically, histologically, and biomechanically (volume [ml] infused through a syringe-perfusion pump system (1 ml/10 s), and pressure [mmHg] to leak induction [catheter connected to a chamber and a monitor, at 10 cm proximal to anastomosis]), at 1, 2, 3, 4, 5, 6, 7, and 14 days. BPC 157 (10 µg, 10 ng, 10 pg/kg i.p. (or saline [5 ml/kg]) was first administered after surgery, while it was last given 24 h before either assessment or sacrifice.ResultsThroughout the experiment, both higher doses of BPC 157 were shown to improve all parameters of anastomotic wound healing. The formation of adhesions remained slight, the blood vessels were filled with blood, and a mild intestinal passage obstruction was only temporarily observed. Anastomosis without leakage induces markedly higher volume and pressure values, with a continuous increase toward healthy values. From day 1, edema was markedly attenuated and the number of granulocytes decreased, while from days 4 or 5 necrosis decreased and granulation tissue, reticulin, and collagen formation substantially increased, thus resulting in increased epithelization.ConclusionThis study showed BPC 157 to have a beneficial effect on ileoileal anastomosis healing in the rat.

Traumatic brain injury in mice and pentadecapeptide BPC 157 effect.

Gastric pentadecapeptide BPC 157 (GEPPPGKPADDAGLV, an anti-ulcer peptide, efficient in inflammatory bowel disease trials (PL 14736), no toxicity reported, improved muscle crush injury. After an induced traumatic brain injury (TBI) in mice by a falling weight, BPC 157 regimens (10.0microg, 10.0ng/kgi.p.) demonstrated a marked attenuation of damage with an improved early outcome and a minimal postponed mortality throughout a 24h post-injury period. Ultimately, the traumatic lesions (subarachnoidal and intraventricular haemorrhage, brain laceration, haemorrhagic laceration) were less intense and consecutive brain edema had considerably improved. Given prophylactically (30 min before TBI) the improved conscious/unconscious/death ratio in TBI-mice was after force impulses of 0.068 Ns, 0.093 Ns, 0.113 Ns, 0.130 Ns, 0.145 Ns, and 0.159 Ns. Counteraction (with a reduction of unconsciousness, lower mortality) with both microg- and ng-regimens included the force impulses of 0.068-0.145 Ns. A higher regimen presented effectiveness also against the maximal force impulse (0.159 Ns). Furthermore, BPC 157 application immediately prior to injury was beneficial in mice subjected to force impulses of 0.093 Ns-TBI. For a more severe force impulse (0.130 Ns, 0.145 Ns, or 0159 Ns), the time-relation to improve the conscious/unconscious/death ratio was: 5 min (0.130 Ns-TBI), 20 min (0.145 Ns-TBI) or 30 min (0.159 Ns-TBI).

Ibuprofen hepatic encephalopathy, hepatomegaly, gastric lesion and gastric pentadecapeptide BPC 157 in rats.

Chronic ibuprofen (0.4 g/kg intraperitoneally, once daily for 4 weeks) evidenced a series of pathologies, not previously reported in ibuprofen-dosed rats, namely hepatic encephalopathy, gastric lesions, hepatomegaly, increased AST and ALT serum values with prolonged sedation/unconsciousness, and weight loss. In particular, ibuprofen toxicity was brain edema, particularly in the cerebellum, with the white matter being more affected than in gray matter. In addition, damaged and red neurons, in the absence of anti-inflammatory reaction was observed, particularly in the cerebral cortex and cerebellar nuclei, but was also present although to a lesser extent in the hippocampus, dentate nucleus and Purkinje cells. An anti-ulcer peptide shown to have no toxicity, the stable gastric pentadecapeptide BPC 157 (GEPPPGKPADDAGLV, MW 1419, 10 μg, 10 ng/kg) inhibited the pathology seen with ibuprofen (i) when given intraperitoneally, immediately after ibuprofen daily or (ii) when given in drinking water (0.16 μg, 0.16 ng/ml). Counteracted were all adverse effects, such as hepatic encephalopathy, the gastric lesions, hepatomegaly, increased liver serum values. In addition, BPC 157 treated rats showed no behavioral disturbances and maintained normal weight gain. Thus, apart from efficacy in inflammatory bowel disease and various wound treatments, BPC 157 was also effective when given after ibuprofen.

Pentadecapeptide BPC 157 and anaphylactoid reaction in rats and mice after intravenous dextran and white egg administration.

Anesthetized mice or rats received intravenously 6%, 10%, 20%, 40%, 60%, 80%, and 90% dextran and/or white egg (1ml/rat or 0.15ml/mouse) into their tails. Medication (/kg b.w., 5ml/kg) was given intraperitoneally (BPC 157 10µg, 1µg, 10ng, and 10pg/kg, chloropyramine 20mg/kg, and cimetidine 10mg/kg intraperitoneally, alone or in combination while controls received an equivolume of saline), immediately after challenge or, alternatively, at 5min after or 24 or 48h before challenge. The effect was assessed at 5, 10, 20 and 30min after dextran and/or white egg challenge. We commonly noted prominent edema involving the face, upper and lower lip, snout, paws and scrotum (presented with extreme cyanosis), poor respiration and the number of fatalities after dextran and/or white egg application. Contrary, BPC 157 regimens (10µg, 1µg, 10ng, and 10pg/kg) effectively, may both prevent anaphylactoid reactions that may arise from dextran and/or white egg application and furthermore, rescue already advanced reactions when given after the challenge. Chloropyramine and cimetidine given alone were only moderately effective. When given together with BPC 157, the observed effect correlates with the strong effect of BPC 157 given alone.

Pentadecapeptide BPC 157 Counteracts Failure of Lower Esophageal and Pyloric Sphincter Induced by NSAIDs in Rats

The risks of aspirin/NSAIDs to induce esophageal and other GI tract lesions have been documented, whilst their effect on lower esophagea sphincter (LES) and pyloric sphincter (PS) pressure is far less studied. Thereby, we tested in rats various NSAIDs and a safe stable gastric pentadecapeptide BPC 157(GEPPPGKPADDAGLV, MW 1419), LD1 not achieved, since successful in inflammatory bowel disease trials, and counteracts esophagitis, sphincters failure, gastrointestinal ulcer and skin ulcer, gastro- or colo-cutaneous fistulas in rats, and particularly counteracts NSAIDs-lesions(1-6). Diclofenac (DI) (12.5 or 40 mg/kg ip, once daily, for 1, 2 or 3 days), ibuprofen (IB) (400 mg/day/kg for 4 weeks), paracetamol (PA) (150 mg/kg ip, 250 mg/kg ig, 5 g/kg ip once), aspirin (AS) (400 mg/kg i.p. or i.g.), BPC 157 immediately after NSAIDs (10 ug/kg i.p. or i.g.), LES and PS pressure (cm H2O), 10 rats at least per group, assessed as described before (3). NSAIDs (control). Regularly, fall of pressure occurred rapidly and persisted (i.e., DI, 12.5 mg: 30 mins: 45.0±2.5 PS, 55.0±2.5 LES - 3h: 43.8±2.5 PS, 56.0±2.5 LES -41.0±3.0 PS, 48.6±3.5 LES (3 days) ; PA 250 mg ig: 60.0±1.5 PS, 63.0±1.0 LES (15 mins) ; PA 5g ip: 35.0±5.5 PS, 40.0±3.5 LES (3h) ; IB, 4 weeks: 33.0±3.5 PS, 43.0±3.0 LES ; AS 400mg ip, 3h: 60.0±1.5 PS, 62.0±0.5 LES (3h) ; AS 400mg ig, 3h: 58.0±1.5 PS, 61.0±1.0 LES ; BPC 157. Regularly, initial fall of pressure was minimized and pressure values restored to normal values (i.e., DI, 12.5 mg: 30 mins: 66.0±2.5 PS, 70.0±2.5 LES - 3h: 62.8±2.5 PS, 69.0±3.5 LES – 62.2±3.0 PS, 72.6±5.5 LES (3 days) ; PA 250 mg ig: 70.0±1.5 PS, 73.0±1.0 LES (15 mins) ; PA 5g ip: 68.0±5.5 PS, 70.0±2.5 LES (3h) ; IB, 4 weeks: 70.0±5.5 PS, 73.0±6.0 LES ; AS 400mg ip, 3h: 65.0±0.5 PS, 70.0±0.8 LES (3h) ; AS 400mg ig, 3h: 68.0±1.5 PS, 71.0±0.9 LES ; ; (vs. control p<0.05). Finally, these BPC 157 effects correlate with attenuated injuries of liver (PA+BPC157 ; IB+BPC157), stomach (AS+BPC 157, IB+BPC 157, DI+BPC157), and small intestine (DI+BPC157). All tested NSAIDs decrease pressure in LES and PS, whilst BPC 157 counteracts their effects and restored LES and PS function. Literature. 1.Curr Pharm Des. 2010 ; 16:1224-34, 2. Dig Dis Sci. 2009 ; 54:2070-83, 3. J Pharmacol Sci. 2008 ; 108:7-17, 4. Dig Dis Sci. 2009 ; 54:46-56, 5. J Pharmacol Sci. 2007 ; 104:7-18. 6. J Physiol Pharmacol. 2010 ; 61:241-50.

The Healing of Colocutaneous Fistulas and Bile Duct Ligation. The Role of Pentadecapeptide BPC 157

The bile duct obstruction is well described as the possible complication of IBD. Since, the pentadecapeptide BPC 157 has finished the second phase of clinical trials for IBD therapy, we assess its effect onto the IBD complications. The previous studies have shown pentadecapeptide BPC 157 effective in healing colocutaneous fistulas (J Pharmacol Sci, 2008), gastrocutaneous fistulas (Dig Dis Sci, 2009). Therefore we suggest it as possible agent in therapy of colocutaneous fistulas at condition of bile duct ligation. 40 male Wistar Albino rats under deep anesthesis underwent surgical procedure. A colocutaneous fistula was created (J Pharm Sci, 2008). At the same act the bile duct ligation was performed at 20 rats in order to cause bile duct obstruction. The animals were randomly assigned into two groups ; half of them were in control group that was treated with saline (5mL/kg, i.p.), while the other received pentadecapeptide BPC 157 (10µg/kg). The treatment was once daily, the first dose was applied immediately after the operation and the animals received the last dose 24h before sacrifice. At the end of each experimental period (2 days, 6 days ) the animals were sacrificed and the biomechanical, functional, macroscopic and microscopic assessment were performed. In animals that underwent the bile duct ligation the healing of anastomosis was impaired in comparison to other animals without bile duct ligature. On the second postoperative day the majority of animals treated with the pentadecaptide BPC 157 started to defecate through anus, while the control animals defecate through fistula. At the sixth postoperative day the control were able to sustain only a small volume of water without leakage through fistula, while the animals treated with BPC 157 sustained larger volume. At the animals treated with pentadecapeptide BPC 157 the diameters of fistula were diminished , at both external and internal sides of fistula. The controls developed jaundice and the liver enzymes were raised, which was not found in animals treated with pentadecapeptide BPC 157. The bile duct ligation has impaired the healing of colocutaneous fistulas. The pentadecapeptide BPC 157 has provided sufficient healing of colocutaneous fistulas and reduced the jaundice the same as liver enzyme raised levels caused by bile duct ligation.

BPC 157 and Standard Angiogenic Growth Factors. Gastrointestinal Tract Healing, Lessons from Tendon, Ligament, Muscle and Bone Healing

Commonly, the angiogenic growth factors signify healing. However, gastrointestinal ulceration is still poorly understood particularly with respect to a general pharmacological/pathophysiological role of various angiogenic growth factors implemented in growth factors wound healing concept. Thereby, we focused on the stable gastric pentadecapeptide BPC 157, a peptide given always alone vs. standard peptidergic angiogenic growth factors (EGF, FGF, VEGF), and numerous carriers. Further, we reviewed how the gastrointestinal tract healing could be generally perceived (i) in terms of angiogenic growth factors, and/or (ii) through the healing of extragastrointestinal tissues healing, such as tendon, ligament, muscle and bone, and vice versa. Respected were the beneficial effects obtained with free peptides or peptides with different carriers; EGF, FGF, VEGF, and BPC 157, their presentation along with injuries, and a healing commonality, providing their implementation in both gastrointestinal ulcer healing and tendon, ligament, muscle and bone healing. Only BPC 157 was consistently effective in all of the models of acute/chronic injury of esophagus, stomach, duodenum and lower gastrointestinal tract, intraperitoneally, per-orally or locally. Unlike bFGF-, EGF-, VEGF-gastrointestinal tract studies demonstrating improved healing, most of the studies on tendon, muscle and bone injuries provide evidence of their (increased) presentation along with the various procedures used to produce beneficial effects, compared to fewer studies in vitro, while in vivo healing has a limited number of studies, commonly limited to local application, diverse healing evidence with diverse carriers and delivery systems. Contrary to this, BPC 157 - using same regimens like in gastrointestinal healing studies - improves tendon, ligament and bone healing, accurately implementing its own angiogenic effect in the healing. Thus, we claim that just BPC 157 represents in practice a pharmacological and pathophysiological role of various peptidergic growth factors.

M1275 Ibuprofen, Hepatic Encephalopathy, Hepatomegaly, Gastric Lesion and Gastric Pentadecapeptide BPC 157 in Rats

Diclofenac Encephalopathy,Liver and Gastrointestinal Lesions in Rat and Stable Gastric Pentadecapeptide BPC 157 Spomenko Ilic, Domagoj Drmic, Danijela Kolenc, Marijana Coric, Luka Brcic, Robert Klicek, Bozo Radic, Marko Sever, Viktor Duzel, Marinko Filipovic, Mihovil Ivica, Alenka Boban Blagaic, Tomislav Ani, Ivan Zoricic, Miroslav Gjurasin, Zeljko Romic, Senka Dzidic, Sven Seiwerth, Predrag Sikiric

M1274 Diclofenac Encephalopathy, Liver and Gastrointestinal Lesions in Rat and Stable Gastric Pentadecapeptide BPC 157

Combined diclofenac encephalopathy, liver and gastrointestinal lesions have not yet been established in rats. The stable gastric pentadecapeptide, BPC 157 (GEPPPGKPADDAGLV, MW 1419, efficient in inflammatory bowel disease trials (PL 14736) and various wound treatment, no toxicity reported) is an anti-ulcer peptide with hepatoprotective effects that may also affectmany central disturbances. Diclofenac (12.5mg/kg)was given intraperitoneally once daily for 3 subsequent days. BPC 157 (10μg/kg, 10ng/kg) was given either (i) intraperi-toneally immediately after diclofenac or (ii) per-orally in drinking water (0.16 μg/ml, 0.16 ng/ml) up until the end of the experiment. At 3 h following the last diclofenac challenge, we evidenced severe gastric, intestinal and liver lesions, increased bilirubin, AST, ALT serum values, liver weight, prolonged sedation/unconsciousness (after any diclofenac challenge) and finally (hepatic) encephalopathy (Fig.1, C (control) B (BPC 157)). Brain edema was particularly present in the cerebral cortex and cerebellum, more in white than in gray matter, damaged (balloonized) red neurons were particularly expressed in the cerebral cortex and cerebellar nuclei, Purkinje cells and less expressed in hippocampal neurons. This was consistently counteracted in diclofenac-rats that received BPC 157 (μg- or ng-regimen, intraperitoneally or per-orally). In conclusion, the successful counteraction of combined diclofenac encephalopathy, liver and gastrointestinal lesions by BPC 157 regimens means that besides inflammatory bowel disease, diclofenac toxicity may be a new domain for possible BPC 157 therapy.