Prevention of peritoneal adhesions with an in situ cross-linkable hyaluronan hydrogel delivering budesonide

https://doi.org/10.1016/j.jconrel.2007.04.016Get rights and content

Abstract

Peritoneal adhesions are tissue connections that form within the abdominopelvic cavity following surgery or other injuries. They can cause major medical complications. Barrier devices and pharmacological agents have been used to prevent adhesion formation, with mixed success. We hypothesize that an adhesion barrier which also delivers anti-adhesion drugs can address both physical and physiological causes for adhesion formation. Here, we describe an in situ cross-linking hyaluronan hydrogel (barrier device) containing the glucocorticoid receptor agonist budesonide. Budesonide was chosen because of the known role of inflammation in adhesion formation, hyaluronan because of its known biocompatibility in the peritoneum. The system, consisting of two cross-linkable precursor liquids, was applied using a double-barreled syringe, forming a flexible and durable hydrogel in less than 5 s. We applied this formulation or controls to the injured sites after the second injury in a severe repeat sidewall defect–cecum abrasion model of peritoneal adhesion formation in the rabbit. Large adhesions (median area 15.4 cm2) developed in all saline-treated animals. Adhesion formation and area were slightly mitigated in animals treated with budesonide in saline (median area 5.0 cm2) or the hydrogel without budesonide (median area 4.9 cm2). The incidence and area of adhesions were dramatically reduced in animals treated with budesonide in the hydrogel (median area 0.0 cm2). In subcutaneous injections in rats, budesonide in hydrogel reduced inflammation compared to hydrogel alone. In summary, budesonide in a hyaluronan hydrogel is easy to use and highly effective in preventing adhesions in our severe repeated injury model. It is a potentially promising system for post-surgical adhesion prevention, and suggests that the effectiveness of barrier devices can be greatly enhanced by concurrent drug delivery.

Introduction

Peritoneal adhesions are persistent tissue connections between structures in the abdomen and pelvis following surgical trauma or infection. The incidence of post-surgical adhesions is as high as 80% and often leads to severe clinical consequences such as pain, infertility, or bowel obstruction [1]. In efforts to prevent adhesions, numerous investigators have applied pharmacological agents that intervene with critical events in adhesion formation [1]. The inflammatory component of the pathogenesis of adhesion formation has been a common target for pharmacotherapy, employing a variety of steroidal anti-inflammatory drugs [2], [3], [4], [5], [6], [7]. However, the effectiveness of these agents in preventing adhesions has not been consistent in animal models [6] and clinical trials [7], especially in preventing recurrent adhesions [8]. Rapid clearance of drugs from the peritoneum could be a cause of the limited effectiveness of intraperitoneally applied drugs.

Rather than using pharmacotherapy, some investigators have devised barrier systems to keep injured abdominal contents separated during the healing process, resulting in several commercial products with differing degrees of success [9], [10], [11], [12], [13], [14], [15], [16]. These have usually been composed of hydrophilic polymers, frequently cross-linked, either ex vivo or in situ.

We have hypothesized, as have others [17], [18], [19], [20], that the limitations of drug delivery could be addressed by maintaining local concentrations of drug through controlled release methods. Conversely, the effectiveness of barrier devices could be enhanced by the co-administration of drugs. While generally encouraging, results with such approaches – including hydrogels containing anticoagulants or thrombolytics [18], [19], [20], or polymeric microspheres containing anti-inflammatory steroids [3] – have been mixed. In the latter case, the efficacy of the system may have been limited by the tendency of polymeric microspheres to encourage adhesion formation [3].

Here, we selected budesonide because it has potent glucocorticoid activity, comparable to that of dexamethasone [21], and is rapidly transformed into inactive metabolites upon systemic absorption [21]. We selected an in situ cross-linkable hyaluronan hydrogel (HAX) as a delivery vehicle for an anti-inflammatory compound. We have previously shown HAX to have excellent effectiveness in preventing peritoneal adhesions in a rabbit sidewall defect–cecum abrasion model, irrespective of the presence of nanoparticles [22], [23]. We demonstrate the anti-adhesion activity of budesonide using a rigorous repeated injury animal model. We show that the co-delivery of budesonide with HAX significantly improved anti-adhesion activity, preventing adhesions completely in the majority of tested animals.

Section snippets

Materials

Hyaluronan was purchased from Genzyme Corporation (Cambridge, MA). All other reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA) and used as received.

Preparation of in situ cross-linkable HA derivatives

In situ cross-linkable HA derivatives were synthesized and analyzed following a previously reported method [22], [24], [25], [26]. Briefly, HA–adipic dihydrazide (HA–ADH) was prepared by conjugating adipic dihydrazide to carboxylic groups in HA backbones, and HA–aldehyde (HA–CHO) was prepared by reacting HA with sodium periodate.

Preparation of budesonide–saline and budesonide–HAX (Supplemental Fig. 1)

Formation of budesonide–HAX gels

Solutions of two HA derivatives (20 mg/ml), both containing 0.13 mg/ml budesonide formed budesonide–HAX gel in 4.1 ±1.7 s upon mixing with constant stirring, which was similar to the gelation time for HAX gels without budesonide (3.9 ±1.1 sec, p = 0.72 on two-tail t-test). To observe the distribution of solid network in the budesonide–HAX gel, the gel was imaged with SEM after removing water from the hydrogel by lyophilization. Under SEM (Fig. 1), the lyophilized budesonide–HAX gel showed a porous

Discussion

Here we show that the anti-adhesion efficacy of budesonide and of the hydrogel barrier system applied separately could be significantly improved by combining the two into a drug delivery system that could maintain a high local drug concentration at the injured surface.

Budesonide is a practically water-insoluble compound, with a saturation solubility in saline of 0.027 mg/ml at 37 °C. Upon addition of the concentrated budesonide stock solution to saline, budesonide forms as a supersaturated

Acknowledgments

This research was supported by the DuPont-MIT Alliance and NIH GM073626. We thank Drs. Robert Marini and Alison Hayward for technical support for in vivo testing.

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