International Journal of Pharmaceutics, 75 (1991) 97-115
`0 1991
`ADONIS 037851739100307L
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`Drug Delivery Research Group, The School of Pharmacy, The Queen’s University of BeIfast, 97 Lisburn Road, Berfast BT9 7BL (U.K.)
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`97 IJP 02512 Invited Reviews Peptide and protein drugs: I. Therapeutic applications, absorption and parenteral administration X.H. Zhou and A. Li Wan PO
`(Received 20 February 1991) (Modified version received 4 May 1991) (Accepted 10 May 1991) Key words: Peptide delivery; Protein delivery; Stability; Bioavailability; Absorption barrier; Proteolytic activity; Absorption enhancer; Proteinase inhibitor; Liposome Summary In this first part of a two-part review of peptide and protein drugs, the pertinent terminology is introduced and the therapeutic applications of those drugs summarised. Their absorption and the methodology commonly used for study on it are discussed. Approaches to optimising delivery of the peptide and protein drugs are highlighted.
`A. Li Wan PO, Drug Delivery Research Group, The School of Pharmacy, The Queen’s University of Belfast, 97 Lisburn Road, Belfast BT9 7BL, U.K. physical properties, including molecular size, sus- ceptibility to proteolytic breakdown, rapid plasma clearance, immunogenicity and denaturation, which make them unsuitable for delivery using the normal absorption routes and in particular, the oral route. In part one of this review protein and peptide drugs are considered with particular emphasis on their pharmacological profiles, po- tential routes of delivery and their associated problems. Recent major reviews on the subject include the general article by Gardner (1984) on the intestinal absorption of intact peptides and pro- teins and that by Humphrey and Ringrose (1986) on the absorption, metabolism and excretion of peptide and related drugs. In a further review, Lee (1988) discussed enzymic barriers to peptide and protein absorption. Banga and Chien (1988)
`With the recent advances in recombinant DNA technology, the commercial production of pro- teins and peptides for pharmaceutical purpose is now routine. The list of available therapeutic agents produced by this technology is expanding rapidly to include interferon, macrophage activa- tion factors, tissue plasminogen activator, neu- ropeptides and experimental agents that may have potential in cardiovascular disease, inflammation, contraception and so on. Unfortunately, protein and peptide drugs possess some chemical and
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`Introduction
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`Correspondence:
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`Elsevier Science Publishers B.V. Ah rights reserved 0378-5173/91/$03.50
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`National Formuhy,
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`1989).
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`98 broadened the scope and considered systemic delivery of those agents in general. Terminology Peptide or protein drugs are derived from amino acids by peptide bond linkages. Proteins are large peptides. Peptides containing less than eight amino acid residues are called small pep- tides. Peptide drugs in this group include enalapril, lisinopril and thyroid releasing hor- mone analogues. The term polypeptide drugs refers to peptide drugs with eight or more amino acid residues and includes cyclosporin, leuproline and luliberin. Polypeptide drugs containing from about 50 to as many as 2500 amino acid residues are named protein drugs. These include insulin, growth hormone and interferons. Some protein drugs, such as insulin or IgG containing two or more polypeptide chains, are called oligomeric proteins and their component chains are termed subunits or protomers. Therapeutic Uses of Peptide and Protein Drugs Peptide and protein drugs can be conveniently classified according to their activity profiles as follows: TABLE 1
`Some exogenous enzymes have been used as enzyme replacement therapy in the treatment of enzyme deficiency diseases such as lysosomal storage and mannosidosis (Table 1). Because en- zyme deficiency in humans is usually genetic in origin, enzyme replacement is often the only available therapy. Some exogenous enzymes have also been utilized in the treatment of diseases other than inborn enzyme deficiency. Good ex- amples include t-PA (tissue plasminogen activa- tors), urokinase and streptokinase. These en- zymes activate circulating plasminogen and fibrin clot-associated plasminogen equally well and, be- cause of this, they have been marketed in the U.K. and U.S.A. (Robinson and Sobel, 1986; British
`Hormones represent the largest class of pro- tein or peptide drugs used in medical therapy. All hormones have ‘target cells’ on which they act and these may be located in a specific organ or be more widely distributed in the body. Some hor- Enzymes Therapeutic application Adenosine deaminase Dextranase P-Fructofuranosidase cY-Mannosidase t_-Asparaginase &Glucosidase Tissue plasminogen activators Urokinase Streptokinase Thromhin-like enzymes of snake venoms Enzyme deficiency Lysosomal storage Storage disease Mannosidosis Cancer Adult Gaucher’s disease Thrombosis Thrombosis Thrombosis Thrombosis Reference Hershfield et al. (1987) Colley and Ryman (1974) Gregoriadis and Ryman (1972b) Pate1 and Ryman (1974) Fishman and Citri (1975) Abuchowski et al. (1984) Braidman and Gregoriadis (1976) Robinson and Sobel(1986) Robinson and Sobel(1986) Robinson and Sobel(1986) Kornalik (1985)
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`Thrombin-like enzymes of snake venoms have also been devel- oped for dissolving blood clots through enhanced release of fibrinopeptides from fibrinogen (Kornalik, 1985).
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`Hormones
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`Therapeutic application of some enzymes
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`Immunomodulating peptides and proteins
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`Endogenous
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`immunomodulating agents
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`Enzyme inhibitors
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`immunomodulating agents
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`dium inflatum
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`Tolypocla-
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`99 mones like luliberin (luteinizing hormone releas- ing hormone, LHRH) function solely to bring about the release of other hormones from differ- ent endocrine glands. It is also well known that many hormones act by means of a second mes- senger and quite often this is cyclic AMP (CAMP) which is formed from ATP. On reaching its re- ceptor in the cell membrane, the hormone causes the release of CAMP, which is the actual regula- tor of the metabolic process. In this way, the physiological effect of one molecule of the hor- mone is amplified many times (Wills, 1985). Be- cause hormones are very specific and a tiny amount can produce large pharmacological ef- fects, they are ideal for biotechnological develop- ment which is more suitable for relatively small outputs. Perhaps the best known hormone drug is insulin which has been used as an endocrinother- apeutic agent since the 1920’s (Banting and Best, 1922).
`These agents are now produced by molecular genetic approaches. Well-known examples are the interferons (IFNs) which are families of inducible secretory proteins produced by eukaryotic cells in response to viral and other stimuli. Interferons are not directly antiviral but they act prophylacti- cally by inducing antiviral proteins. These protect cells from viral infection by inhibiting virus-di- rected translation and transcription (Moore and Dawson, 1989). Another example is interleukin-2 (IL-2) which exerts its biological effect through cell surface receptors on activated T and B cells and on NK cells (natural killer cell). Interleukin-2 has been administered clinically in attempts to restore immunocompetence in patients suffering from the acquired immunodeficiency syndrome (AIDS), and to improve the immunocompetence of cancer patients (Dawson and Moore, 1989). Exogenous
`Enzyme inhibitors have been used as drugs for a long time. These include proteins such as apro- tinin, and peptide drugs such as enalapril and lisinopril. Captopril is an inhibitor of angiotensin converting enzyme (ACE), which catalyses in vivo generation of angiotensin II from the decapep- tide, angiotensin I, to constrict arterioles and increase cardiac output, leading to hypertension in man. Captopril is now a widely used antihyper- tensive agent (Romankiewicz et al., 1983). Enalapril and lisinopril are subsequent develop- ments which are also becoming widely adopted for the treatment of hypertension and congestive heart failure (Todd and Heel, 1986; Lancaster and Todd, 1988).
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`Some exogenous immunomodulating agents are also used to promote immunocompetence in man. For example, cyclosporin (CS-41, a cyclic undecapeptide which is isolated from
`Gams, is widely used as an im- munosuppressive (Calne et al., 1978; Cantarovich et al., 1987; Mehta et al., 1988; Borel, 19891, whereas muramyl dipeptide has been used as an immunological adjuvant (Kreuger et al., 1984; Bomford, 1989).
`Vaccines derived from the infective microor- ganisms are introduced into the mammalian body to induce antibody formation against the path- ogens. Well-known examples include measles vac- cine and polio vaccine. It is anticipated that an increasing number of such vaccines will be biotechnologically produced, to give more specific and pronounced antigenic responses.
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`Vaccines
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`Antimicrobial agents
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`A number of antimicrobial agents are peptide drugs, for example, the penicillins, cephalo- sporins, polymyxin B sulphate, actinomycin and bleomycin. Structurally, these drugs are small peptides, mostly containing a non-peptide moiety. All of these antimicrobial drugs are microbial metabolites.
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`Absorption of Peptide and Protein Drugs
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`Analytical problems
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`Several methods have been employed for studying the absorption of peptide and protein
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`Instability of protein and peptide drugs
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`100 drugs. However, high molecular weight proteins and polypeptides present some unique difficul- ties. Techniques such as gel filtration and ion-ex- change HPLC usually have to be used. Even so, it is still very difficult to assay them in the presence of body fluids such as blood and urine. In such cases, radioassays or radioimmunoassays are of- ten the most appropriate and hence, these tech- niques have been widely used in the measure- ment of the bioavailability of peptide or protein drugs. However, radioassays may be non-specific, and many chemical assay procedures may by themselves influence the conformation of protein drugs, thereby causing the loss of their biological activities. The entity being chemically assayed may not be the biologically active moiety and in such cases, in vitro or in vivo bioassays are often used during absorption studies. For protein/ peptide hormones, the measurement of pharma- cological responses may be the assay method of choice. For enzymes or enzyme inhibitors, spe- cific enzyme reactions may be the best analytical method. The bioavailability of immunomodulat- ing and antimicrobial agents may be evaluated using some specific animal models and indicator microorganisms. For example, the prophylactic TABLE 2
`Effect factor Physical instability Aggregation Precipitation Protein or peptide drugs Interferon-y Bovine growth hormone Insulin Reference Hsu and Arakawa (1985) Arakawa et al. (1987) Brems et al. (1986) Brems et al. (1988) Brennan et al. (1985) Lougheed et al. (1980) Chemical instability p Elimination Deamidation Lysozyme Phosvitin Bovine growth hormone Human growth hormone Insulin r-Immunoglobulin Epidermal growth factor Prolactin Gastrin releasing peptide ACTH Nashef et al. (1977) Sen et al. (1977) Lewis and Cheever (196.5) Lewis et al. (1970) Becker et al. (1988) Berson and Yalow (1966) Fisher and Porter (1981) Minta and Painter (1972) Diaugustine et al. (1987) Graf et al. (1970) McDonald et al. (1983) Graf et al. (1971) Bhatt et al. (1990) Disulphide exchange Racemization Lysozyme Ribonuclease A ACTH Volkin and Khbanov (1987) Zale and Klibanov (1986) Geiger and Clarke (1987) Meinwald et al. (1986) Oxidation Corticotropin Dedman et al. (1961) Q-, P-Melanotropins Dixon (1956) Parathyroid hormone Tashjian et al. (1964) Gastrin Morley et al. (1965) Calcitonin Riniker et al. (1968) Corticotropin releasing factor Vale et al. (1981)
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`Liposomes as peptide and protein carrier
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`Liposome Peptide or composition protein Route Animal model Reference Phosphatidyl- choline : cholesterol 712 Phosphatidyl- choline : cholesterol 717 semipurified glucocerebroside P-glucosidase highly purified glucocerebroside fi-glucosidase i.v. i.v. man man Belchetz et al. (1977) Gregoriadis et al. (1982) Phosphatidyl- choline : cholesterol : phosphatidic acid 7:2:1 Dimyristoyl phosphatidyl- choline : choles- terol : dicetyl phosphate 1:0.75:0.11 bacterial amyloglucosidase cholera toxin human malaria sporozoite antigen i.v. i.v. man rabbit Tyre11 et al. (1976) Alving et al. (1986) Phosphatidyli- nositol Phosphatidyl- choline : choles- terol : dicetyl phosphate 10:2: 1 Phosphatidyl- choline : choles- terol : dicetyl phosphate 3:9:1 Phosphatidyl- choline : phos- phatidylserine 713 Phosphatidyl- choline : choles- terol : dicetyl phosphate 7:1:2 Phosphatidyl- choline : choles- terol : phospha- tidic acid 20: 1..5:0.2 insulin insulin insulin i.v. oral oral muramyl peptide i.v. hwme adenovirus type 5 hexon protein i.v. mouse rat rat Dapergolas and Gregoriadis (1976) Pate1 and Ryman (1976) rat Tanaka et al. (1975) mouse guinea-pig Fidler et al. (1985) Sessa and Weissmann (1970) mouse Six et al. (1988) Sessa and Weissmann (1970) Phosphatidyl- choline : choles- terol : phospha- tidic acid 7:1:2
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`TABLE 3
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`102 TABLE 3
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`Liposme
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`Ccontinuedf
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`Peptide or composition profein Microcapsules catalase Route is. Animal model
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`ItlOUSe
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`i.V.
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`i.V,
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`terol :
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`Reference ~. Chang and Poznansky (1968) Phosphatidyi- choline : choles- terol : dicetyl phosphate 7:2:1 amyloglucosidase i.v. rat Gregoriadis and Ryman (1972a) Phos~hatidyl- choline : choies- terol : phospba- tidic acid ?:2:1 yeast invertase Phosphatidyl- choke : choks- feral : phospha- tidic acid 7:2:1 neuraminidase Phosphat~I- choline : choies- terol : phospha- tidic acid 7:2:1 dexfranase Phosphatidyl- choline : choles- terol : phospha- tidic acid 7:2:I ru-mannosidase Dipalmitoyl- phosphatidyl- choline a-amylase PhosphatidyI- choline : choles-
`i.v. rat rat rat rat amoeba rat mouse mouse rat man Gregoriadis and Ryman (197Zb) Gregoriadis et al. (1974a)
`iv. i.v. i.v. i.v. i.v.
`and Ryman j19f4) Pate1 and Ryman (1974) Batzri and Korn (1975) Wisse and Gregoriadis (1975) Neerunjun and Gregoriadis (1976) Dapargolas et al. (1976) Grqoriadis and Neerunjun (1974) Gregoriadis et al. 11974bf
`phospha- tidic acid 7:2:1 horseradish peroxidase PhosphafidyI- choline : choles- terol : Phospha- tidic acid ?:2:1 asparaginase Phosphatidyli- n&to1 glucose oxidase Phosphatidyl- choline : choles- terol 7:2 albumin Phosphatidyl- choline : chobs- terol : phospha- tidic acid 7:2:1 albumin
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`Colley
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`d)
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`103 TABLE 3
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`(continued)
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`Liposome composition Peptide or protein Route Animal model Reference Phosphatidyl- choline : choles- terol : dicetyl phosphate 61612 albumin i.v. mouse Heath et al. (1976) Phosphatidyl- choline : choles- terol : dicetyl phosphate 7:2:1 diphtheria toxoid Phosphatidyl- choline : choles- terol : phospha- tidic acid 7:2:1 fetuin Phosphatidyl- choline : choles- terol : phospha- tidic acid 4:2:1 antia- glucosidase Phosphatidyl- choline : choles- terol : phosphati- dylethanolamine 1O:lO:l monoclonal anti-Thy1 IgGl Distearoylphos- phatidylcholine: (2-puridyldithio)- propionol-dipal- mitoylphospha- tidylcholine: cholesterol 0.99 : 0.01 : 1 monoclonal anti-Thy1 IgGl a Liposomes TRH Phosphatidyl- choline : phos- phatidylserine 713 superoxide dismutase Phosphatidylcholine: phosphatidic acid 15.3 : 0.1 factor VIII i.v. iv. i.v. i.v. iv. rat rat mouse mouse i.v. intratracheal injection cat rat oral i.p. man mouse Gregoriadis and Alison (1974) Gregoriadis and Neerunjun (1975) De Barsy et al. (1975) Debs et al. (1987) Wolff and Gregoriadis (1984) Kumashiro et al. (1986) Padmanabhan et al. (1985) Sakuragawa et al. (1985) Hashimoto et al. (1986) Dipalmitoyl- subunits of phosphatidyl- monoclonal IgM choline : cholesterol : m-ma- leimidobenzoyl- (dipalmitoyl- phosphatidyl)- ethanolamine 25 : 17.5 : 2.5 a The composition of liposomes was not mentioned in the paper.
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`104
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`usefulness of intranasal IFN-P against rhinovirus infection was determined using healthy volun- teers or animals (Higgins et al., 1986). Assay methods available for monitoring the absorption of small peptide drugs are freely avail- able and routine methods include reverse-phase HPLC, TLC, and fluorescence techniques.
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`Stability
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`Irrespective of which dosage form is used, pep- tide or protein decomposition may be a problem. Drug breakdown can take place both in the for- mulation and when present in tissue fluids. First pass metabolism and enzymic breakdown are dis- cussed in greater detail further on. Non-enzymic breakdown may be of two types: chemical and physical changes. Physical changes include aggre- gation and precipitation and are usually induced by high concentrations of co-solvents which may be used in some formulations or by injudicious choice of ionic strengths. Loss of conformation not only leads to poor absorption but also to loss of activity. Chemical changes include p-elimina- tion, deamidation, disulphide exchange, racem- ization and oxidation. Examples of peptides and proteins which have been reported to be unstable are shown in Table 2 along with some of the reported mechanisms of breakdown.
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`For systemic delivery of peptide and protein drugs, parenteral administration is currently al- most universally required in order to achieve consistent therapeutic activities. This is because of the drugs’ susceptibility to breakdown by gas- tric acid and the proteolytic enzymes in the gas- trointestinal tract. In addition, peptides and pro- teins are high-molecular-weight substances and thus do not easily cross the intestinal mucosa. Therefore, the oral bioavailabilities of most intact peptides and proteins are very low. Of the parenteral routes, only intravenous (i.v.> administration is usually efficient in delivering protein and peptide drugs to the systemic circula- tion. For example, optimal blood levels of protein or peptide drugs, such as y-globulin (Buckley, 1982), can be achieved by the intravenous route. Generally, intramuscular or subcutaneous injec- tions are less efficient due to the absorption and diffusion barriers presented by the muscle mass and connective tissues under the skin. However, insulin can be efficiently administered by subcu- taneous injections (Nora et al., 1964; Koivisto and Felig, 1978) although hydrolysis is still significant (Berger et al., 1979). While most peptide/protein drugs can be effi- ciently delivered to the systemic circulation by parenteral injections, poor disposition profiles lead to sub-optimal therapeutic benefits without high dosing frequencies. Such frequent injections, besides being unpleasant to the patients, also lead to usual complications such as throm- bophlebitis and tissue necrosis. In attempts to improve the disposition profile and the efficiency of delivery of parenterally ad- ministered peptides and protein drugs, many in- vestigators have reported on liposomal systems. Examples of enzymes and monoclonal antibodies which have been formulated as liposomal systems for intravenous administration are shown in Table 3. Also included are some liposomal systems in- tended for oral administration. Despite the ex- tensive evaluation of such systems and experi- mental results (Gregoriadis, 1976; Goosen, 1987) indicating that insulin absorption is greatly en- hanced in animals by liposomal encapsulation of the hormone, no insulin liposomal system is cur- rently in commercial use. One biodegradable implant in current use in humans in goserelin acetate formulated in a biodegradable matrix of lactide-glycolide co-poly- mer. Systems which are designed with an enzymi- tally controlled feed-back mechanism have also been described. Fischel-Ghodsian et al. (1988), for example, reported on an insulin system con- sisting of insulin and glucose oxidase dispersed in an ethylene/vinyl acetate polymer matrix. In the presence of glucose oxidase, glucose is converted into gluconic acid. This acid lowers the pH and increases the solubility of entrapped insulin which is then released faster. Consequently, some feed- back control between glucose and insulin is thereby established.
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`Parenteral routes of delivery
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`General Approaches to Optimizing Absorption and Disposition To optimize the absorption of high-molecular- weight protein and peptide drugs across absorp- tion barriers, several approaches are available: (i) inhibition of their enzymic degradation; (ii) in- creasing their permeability across the relevant membrane; and (iii) improving their resistance to breakdown by structural modification. Inhibitors of proteolytic enzymes Protease inhibitors have been known for sev- eral years to increase the absorption of protein drugs (Laskowski et al., 1958). Table 4 lists the different protease inhibitors which have been used in investigations of the delivery of peptide and protein drugs. Aprotinin, a bovine pancreatic kallikrein in- hibitor, consists of a single-chain polypeptide containing 58 amino acid residues with a molecu- lar weight of 6500 (Kassell et al., 1965). It has been used to inhibit plasmin, trypsin, chy- motrypsin and various intracellular proteases (Trautschold et al., 1967). It was demonstrated, in an early study, that when insulin and aprotinin TABLE 4 105 were injected together into a loop of the jejunum, a significant drop in blood glucose was observed. In contrast, no significant drop in blood glucose was found when the insulin was injected alone (Laskowski et al., 1958). Similar results have also been reported by several other workers (Berger et al., 1980; Fredenberg et al., 1981; William et al., 1983; Dandona et al., 1985; Linde and Gun- narson, 1985; Owens et al., 19881. However, some recent studies provided conflicting results, at least with respect to insulin and calcitonin absorption by nasal administration (Hanson et al., 1986; Aungst and Rogers, 1988). When the effects of laureth-9, sodium salicylate, Na,EDTA and aprotinin on insulin absorption via the rectal, nasal and buccal tissues were examined by Aungst and Rogers (19881, aprotinin was found to be ineffective, either alone or in combination with laureth-9. Hanson and his co-workers (1986) ex- amined the effects of several protease inhibitors, including bile salt, fatty acid derivative, aprotinin, kallikrein inhibitor, RG-1, bestatin, fusidic acid, chemostatin, benzamidine, chymotrypsin in- hibitor, trypsin inhibitor III-O and leupeptin on intranasal delivery of calcitonin, and found that aprotinin in vitro did not inhibit proteolytic activ- ity of nasal extracts. In vivo the inhibitor did not
`Compound Route Peptide Animal Reference studied model Aprotinin intestinal insulin rat Ziv and Kidron (19871, Laskowski et al. (1958) RNase rat Ziv and Kidron (1987) S.C. a insulin man Owens et al. (19881, Wnde and Gunnarsson (1985), Berger et al. (1980) Soybean intestinal insulin rat Ziv and Kidron (1987) ttypsin RNase rat Ziv and Kidron (1987) inhibitor FK-448 b intestinal insulin rat Yokoo et al. (1988) (chymotrypsin inhibitor) Boroleucine ’ nasal Leu-enkephalin rat Hussain et al. (1989) Borovaline ’ nasal Leu-enkephalin rat Hussain et al. (1989) a Subcutaneous delivery. b 4-(4-Isopropylpiperazinocarbonyl)phenyl-1,2,3,4-tetrahydro-l-naphthoate methanesulphonate. ’ cr-Aminoboronic acid derivatives.
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`Inhibitors of proteolytic enzymes used in investigation of the delivery of peptide and protein drugs
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`106
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`Absorption enhancers
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`enhance the serum calcium drop observed. These results are supported by the study of Deurloo et al. (1989). The co-administration of sodium tau- rodihydrofusidate with aprotinin also failed to increase significantly insulin bioavailability in rab- bits via the nasal route. Clearly, further studies are required to define better the effects of apro- tinin on the absorption of peptide and protein drugs. More recently, cr-aminoboronic acid deriva- tives, such as boroleucine, which are potent and reversible inhibitors of aminopeptidase, have been used to stabilize peptide drugs during their in- tranasal absorption (Hussain et al., 1989). When these inhibitors were compared with other known peptidase inhibitors, bestatin [an inhibitor of leucine aminopeptidase, aminopeptidase B, and aminopeptidase N (Suda et al., 1976)] and puromycin [an inhibitor of aminopeptidase B and N but not leucine aminopeptidase (McDonald et al., 1964)], using leucine enkephalin as substrate in rat nasal perfusate, it was found that bestatin and puromycin were less effective than boro- leucine, even at concentrations lOO- and lOOO- times higher, respectively.
`The use of absorption enhancers has been studied extensively, particularly with respect to insulin absorption. These enhancers can be di- vided into several groups as listed in Table 5. Despite extensive use, it is very difficult to make a judgement about the relative efficacy of these bioenhancers in promoting peptide or pro- tein absorption because the results were obtained in different laboratories using different assay methods and different experimental conditions. However, it is clear that the bioavailability of most peptide and protein drugs administered by any non-parenteral route may be significantly en- hanced by some of these compounds (see Tables in part II of this review). The value of a particular enhancer depends on the route of administration used. For example, the bioavailability of ocular insulin was found to be significantly enhanced by saponin, whereas enhancement by glycocholate, which was a poten- tially good enhancer for nasal and rectal peptide and protein drug absorption, was found to be only slight (Chiou and Chuang, 1989). The mechanisms of action of the peptide ab- sorption enhancers are not clearly known, but several possibilities have been postulated. The first is increased solubility of the drugs brought about by the enhancers because proteins and peptides usually form aggregates in aqueous solu- tions. In the presence of enhancers, dissociation takes place to form monomers which are better absorbed. A second mechanism is the protection of the peptide and protein drugs from potential proteolytic hydrolysis. Both bile salts (Hirai et al., 1981b; Hanson et al., 1986; Zhou and Li Wan PO, 1991) and derivatives of fusidic acid (Deurloo et al., 1989) are known to inhibit proteolytic degra- dation of the drugs by nasal homogenates. Thirdly, binding between peptide or protein and enhancer to produce a better-absorbed entity may be a possibility. Although the effects of absorption en- hancers such as glycocholate (Hirai et al., 1981b) and sodium cholate (Zhou and Li Wan PO, 1991) on the absorption of insulin by nasal delivery are thought to be partly due to inhibition of protease, recent work suggests that compared to aminopep- tidase inhibitors such as bestatin and amastatin, cholate and its analogues are not very efficient (Hanson et al., 1986). Cholate and its analogues may also enhance the absorption of peptide and protein drugs by binding to insulin (Zhou and Li Wan PO, 1991). This would prevent the formation of enzyme-substrate complex to undergo the nec- essary conformational change which aligns the catalytic site on the protease with the susceptible bond of the substrate. Cholate and its analogues may possibly also promote the absorption of pro- teins by selectively denaturing the enzymes, al- though this is unlikely as it is difficult to identify the basis for the necessary selectivity.
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`Chemical modification is an important ap- proach for enhancing the absorption of peptides and protein drugs, especially for peptides with fewer than ten amino acid residues. Chemical modification usually results in denaturation of
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`Chemical modification
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`107 TABLE 5
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`Absorption enhancers for peptides and proteins
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`Compound Route Peptide Animal
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`model
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`Reference Fatty acid MCFC a Caprylate Caprate Laurate LCFC b Oleate in PAGB ’ Linoleate in PAGB Linolenate in PAGB Oleic acid Bile salts Taurocholate Cholate Deoxycholate Glycocholate Chenodeo~cholate Deoxycholate (aerosol) Glycocholate Cholate Deoxycholate Cholate Glycocholate nasal nasal nasal rectal rectal rectal vaginal nasal nasal nasal nasal nasal nasal vaginal rectal rectal intestinal intestinai nasal rectal buccal sublingual Enamine derivatives of phenylglycine Ethytacetoacetate enamine of sodium p-glycine rectal Ethylacetoacetate enamine of sodium o-alanine rectal EthyIacetoacetate enamine of sodium o-leucine rectal Ethylacetoacetate enamine of sodium o-isoleucine rectal Ethylacetoacetate enamine of sodium o-phenylalanine rectal Ethylacetoacetate enamine of sodium D-phenylalanine in gelatin rectal Ethylacetoacetate enamine of sodium D-phenytglycinate rectal Ethylacetoacetate enamine of sodium oL-phenylaianine rectal insulin insulin rat rat rat insulin rat insulin rat insulin rat leuprolide rat insulin insulin insulin insulin insulin insulin leuprolide insulin insulin insulin RNase insulin insulin insulin insulin rat rat rat rat rat man rat rat rat rat rat rat rat rat rat insulin rabbit insulin rabbit insulin rabbit insulin rabbit insulin rabbit insulin dog insulin rabbit Mishima et al. (1987) Morimoto et al. (1983) Okada et al. (1982) Hirai et al. (1981a) Moses et al. (1983) Mishima et al. (1987) Moses et al. (1984) Okada et al. (1982) Ziv et al. (1981) Ziv et ai. (1987) Aungst et al. (1988) Kim et al. (1983) Kamada et al. (1981)
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`108 TABLE 5 (continued) Compound Route Peptide Animal model Reference Ethylacetoacetate enamine of sodium p-phenylglycine Ester type Glycerine-l, 3-diacetoacetate 1,2-fsopropylidene- glyceryl-3- acetoacetate Ethylaceto- acetylglycolate Polyoxyethylene lo-monolaurate Glyceryl esters of acetoacetic acid Ether type Polyoxyethylene 9-lauryl ether in PAGB Polyo~ethylene 9-lauryl ether Polyoxyethylene 24-cholesteryl ether Polyo~ethylene 5-octyf ether Polyoxyethylene lo-octyl ether Polyoxyethylene 5lauryl ether Polyoxyethylene 9-Iauryl ether Polyoxyethyiene IO-lauryl ether Polyoxyethylene 20-lauryl ether Polyoxyethylene IO-cetyl ether Polyo~ethy~ene 20-cetyl ether Polyoxyethylene lo-stearyl ether Polyoxyethylene 20-stearyl ether Poiyo~ethyiene lo-nonylphenyl ether Polyoxyethyiene lo-octylphenyl ether Polyoxyethylene 24.cholesteryl ether rectal rectal rectal rectal nasal rectal rectal rectal oral nasal nasal nasal nasal nasal nasal nasal nasal nasal nasal nasal nasal nasal lysozyme rabbit lysozyme rabbit lysozyme lysozyme insulin insulin rabbit rabbit rat rabbit calcitonin insulin ergot peptide alkaloids insulin insulin insulin insulin insulin insulin insulin insulin insulin insulin insulin insulin insulin rat Morimoto et al, (1985) dog Shichiri et al. (1978) rat rat rat rat rat rat rat rat rat rat rat rat rat rat Miyake et al. (19841 Hirai et al. (1981a) Nishihata et al. (1983) Urbancic-Smerkolj et al. (1987) Hirai et al. (1981a)
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`TABLE 5 (continued) Compound Route Peptide Animal model Reference Polyoxyethylene 20-cetyl ether (Cetomacrogol 1000) Polyoxyethylene 9-lauryl alcohol ether Polyoxyethylene lo-nonylphenyl ether Salicylates 3,5-Diiodosalicylate 5-Methoxysalicylate 5-Methoxysalicylate 5-Methoxysalicylate 5-Methoxysalicylate in PAGB (pH 5.5, M.W. 1250 000) Salicylate Organic acids Citric acid Succinic acid Tartaric acid Malonic acid Glycoside Saponin Saponin Peptide lipid Surfactin Amine Polyoxyethylene 5-oleylamine Anion Sodium lauryl sulphate Monosodium N-lauroyl L-glutamate Derivatives of fusidic acid sodium tauro- dihydrofusidate Sodium tauro- dihydrofusidate Glycyrrhetinic acid derivatives Glycyrrhizinic acid Glycyrrhetinic acid hydrogen succinate rectal rectal rectal insulin insulin insulin rectal rectal rectal rectal insulin insulin insulin gastrin pentagastrin rectal rectal calcitonin human growth hormone vaginal leuprolide vaginal insulin vaginal leuprolide vaginal leuprolide vaginal