Gastrointestinal mucoadhesive patch system (GI-MAPS) for oral administration of G-CSF, a model protein

Abstract

A new gastrointestinal mucoadhesive patch system (GI-MAPS) has been designed for the oral delivery of protein drugs. The system consists of four layered films, 3.0×3.0 mm², contained in an enteric capsule. The 40 μm backing layer is made of a water-insoluble polymer, ethyl cellulose (EC). The surface layer is made of an enteric pH-sensitive polymer such as hydroxypropylmethylcellulose phthalate (HP-55^®), Eudragit^® L100 or S100 and was coated with an adhesive layer. The middle layer, drug-containing layer, made of cellulose membrane is attached to the EC backing layer by a heating press method. Both drug and pharmaceutical additives including an organic acid, citric acid, and a non-ionic surfactant, polyoxyethylated castor oil derivative (HCO-60^®), were formulated in the middle layer. The surface layer was attached to the middle layer by an adhesive layer made of carboxyvinyl polymer (Hiviswako^® 103). Fluorescein (FL), 30 mg, was first used as a model drug for oral administration of GI-MAPS having different surface layers in beagle dogs. The plasma FL concentration vs. time profiles demonstrated that the targeting of the systems was obtained, because the T_max, the time when plasma FL concentrations reaches to its maximum lelev, was 2.33±0.82 h for HP-55 system, 3.33±0.41 h for Eudragit L100 system and 5.00±0.00 h for Eudragit S100 system. The same three kinds of GI-MAPSs containing 125 μg of recombinant human granulocyte colony-stimulating factor (G-CSF) were prepared and orally administered to dogs and the increase in total white blood cell (WBC) counts were measured as the pharmacological index for G-CSF. Comparison with the total increase of WBCs after iv injection of the same amount of G-CSF (125 μg) indicated the pharmacological availabilities (PA) of G-CSF were 23%, 5.5% and 6.0% for Eudragit L100, HP-55 and Eudragit S100 systems. By decreasing the amount of HCO-60 and citric acid, the PA of G-CSF decreased. These results suggest the usefulness of GI-MAPS for the oral administration of proteins.

Introduction

With the development of gene-technology, many useful proteins such as recombinant human granulocyte colony-stimulating factor (G-CSF) and erythropoietin are capable of being used clinically with great effectiveness [1], [2], [3], [4]. However, their administration route is limited to iv or subcutaneous injection [5]. The oral route has several attractive advantages for the administration of proteins, including the avoidance of pain and discomfort associated with injections, the elimination of possible infections caused by the use of needles, and increased QOL of the patients [6]. However, there are several barriers for the oral delivery of proteins including degradation from the acidic pH of the stomach, hydrolytic degradation by proteolytic enzymes, metabolism by luminal, brush border and cytosolic enzymes, and poor membrane permeability across the intestinal epithelium [7]. To overcome the problem of proteolytic degradation, several new pharmaceutical technologies have been proposed [8], [9], [10], [11], [12], [13], [14]. Among these, lipid systems such as “Macrulin^®” and “Macritonin^®” which contain insulin and calcitonin are now under clinical phase II-III trials [15], [16]. However, even in those systems, attack by proteolytic enzymes in the GI tract cannot be prevented.

We have been studying oral protein delivery systems using G-CSF as a model protein. Our previous studies showed that hydrolytic degradation in the gastrointestinal (GI) tract played an important role on the pharmacological availability (PA) of G-CSF after oral administration to rats [17], [18], [19], [20]. Since hydrolytic enzyme activity is lower in the colon than in the small intestine, we developed two types of colon delivery systems, i.e., intestinal luminal pressure-controlled colon delivery capsules (PCDCs) and time-controlled colon delivery capsules (TCDCs) [21], [22]. Using PCDCs, we orally administered G-CSF at a dose of 250 μg/dog. As a pharmacological index of G-CSF, total white blood cell (WBC) counts were measured. Comparison of the WBC increase occurring after oral administration of G-CSF in PCDCs to that obtained after iv injection in dogs, the comparative PA of oral G-CSF was approximately 7.0% [21]. These results suggested that proteolytic enzymatic activity also exists in the colon. Therefore, we concluded that the hydrolytic degradation of G-CSF in the GI tract must be prevented in order to increase the PA of G-CSF.

Recent studies showed that hydrolytic activity occurring in the brush border membrane and cytosol is still high [23], [24], [25]. However, the GI luminal hydrolytic enzyme activity is by far higher than the hydrolytic enzymatic activity of the brush border membrane and cytosolic enzymes. To protect orally administered G-CSF from attack by the intestinal luminal hydrolytic enzymes, a new technology is needed.

Patch preparations have been increasingly used clinically, i.e., TTSs of nitroglycerin and nicotine, and they contribute to the therapy of many diseases [26]. Drugs contained in this system are protected from removal by the backing layer. We assumed that a mucoadhesive patch system could protect proteins against degradation by the attack of the proteolytic enzymes localized in the GI lumen. Therefore, we designed a novel GI mucoadhesive patch system (GI-MAPS), which delivers protein drugs to the targeted intestinal site, is adhesive to the small intestinal wall and prevents the attack by luminal proteolytic enzymes via a backing layer made of water-insoluble polymer.

In this report, an explorative study of this novel system was first performed with fluorescein (FL) as a model drug. The usefulness of GI-MAPS for oral protein delivery was evaluated in dogs after oral administration of G-CSF formulated in GI-MAPS by measuring the total WBC count as a pharmacological index of G-CSF.