Drug delivery systems,second edition o Folate X /PEG3350 图wpg Entrapping Folate Recepto Released araC ■ Figure 1.8 Possible mechanism of intracellular araC delivery by FR-targeted,cationic, lipid-based,pH-sensitive liposomes.At first,the folate-derivatized liposomes are taken into the cell by binding to the FRs on the plasma membrane and FR-mediated endocytosis.This is followed by acidification of the endosome,which results in protonation of the anionic lipid component and generation of a net positive surface charge on the liposomes.Finally,the electrostatic interactions between the liposomal and endosomal membranes result in bilayer fusion and the cytosolic delivery of the encapsulated araC.(With permission,Elsevier,J.Control.Rel.,80,309-319,2002) mechanical properties of the liposomal membranes,their thermodynamics and colloidal properties of the system.Often,stability tests stress a system to limits beyond those to which the product will ever be subjected. High-temperature testing(greater than 25C;see Figure 1.8)is frequently used for heterogeneous products.Phase-transition temperatures for a lipo somal system are critical,but the changes,should this cu,are reversible On the other hand,under frozen conditions,ice crystals are f ormed.Ceran polymers are known to retard or suppress ice crystal growth.Aging studies involving determinations of zeta potential and dielectric constants are usu- ally performed.These analyses determine the status of structural alterations in liposomal vesicles.In summary,in liposomal preparations,each test con- dition indicating stability of the vesicles should e press conditions for micro scopic observation (e.floc culation),particle-size profiles,rheological pro- files,extent of leakage,and chemical and physical stability. IV.Liposomes as carriers of therapeutic agents Application Since 1972,when Gregoriadis proposed the use of liposomes as carriers of enzymes in the treatment of lysosomal storage diseases,the application of
12 Drug delivery systems, second edition mechanical properties of the liposomal membranes, their thermodynamics, and colloidal properties of the system. Often, stability tests stress a system to limits beyond those to which the product will ever be subjected. High-temperature testing (greater than 25∞C; see Figure 1.8) is frequently used for heterogeneous products. Phase-transition temperatures for a liposomal system are critical, but the changes, should this occur, are reversible. On the other hand, under frozen conditions, ice crystals are formed. Certain polymers are known to retard or suppress ice crystal growth. Aging studies involving determinations of zeta potential and dielectric constants are usually performed. These analyses determine the status of structural alterations in liposomal vesicles. In summary, in liposomal preparations, each test condition indicating stability of the vesicles should express conditions for microscopic observation (e.g., flocculation), particle-size profiles, rheological pro- files, extent of leakage, and chemical and physical stability. IV. Liposomes as carriers of therapeutic agents A. Application Since 1972, when Gregoriadis proposed the use of liposomes as carriers of enzymes in the treatment of lysosomal storage diseases, the application of Figure 1.8 Possible mechanism of intracellular araC delivery by FR-targeted, cationic, lipid-based, pH-sensitive liposomes. At first, the folate-derivatized liposomes are taken into the cell by binding to the FRs on the plasma membrane and FR-mediated endocytosis. This is followed by acidification of the endosome, which results in protonation of the anionic lipid component and generation of a net positive surface charge on the liposomes. Finally, the electrostatic interactions between the liposomal and endosomal membranes result in bilayer fusion and the cytosolic delivery of the encapsulated araC. (With permission, Elsevier, J. Control. Rel., 80, 309–319, 2002.) + + + + + Folate PEG3350 Liposomes Entrapping araC Folate Receptor Released araC H + H
Chapter one:Site-specific drug delivery using liposomes as carriers 13 liposomes has been extended to a variety of drugs,such as antineoplastic 9-52 aont utiizing lpoomess drug sarries in the tratment of ntrace lular infections,53 liposomes have also been used as carriers of amphotericin B in the treatment of mycotic infections,such as histoplasmosis,s4 cryptococ- cosis,and candidiasis.s4 Lopez-Berestein et al.s5 reported that liposomal phBis effective in the treatment of candida and aspergillus infec. emia patients who have not responded to the nonencapsulated drug.The increase in amphotericin's efficacy by encapsulation in liposomes is associated with reduced toxicitv.> Incorporation of lipophilic amphotericin within liposomes might result in a facilitated transfer of the drug to fungal cells.In turn,this selective nsfer of a form the kinetics or tissue distribution,may also play an important role.s Antibacterial activity with liposome encapsulation has been reported by Sunamoto and co-workers (in experimental Legionnaires'disease).58 They showed that uptake of IV-injected liposomes by circulating monocytes and rophages ting the vesicles itoyl derivative of amylopectin.After IV inject liposomes were found to preferentially distribute in the lungs. Macrophages have an affinity for liposomes,and this property has been utilized in the use of these vesicles as carriers of immunomodulators to create macrophages cytotoxic to metastatic tumor cells.As a result,macrophage e as an i ant b arrier against the prolife of tumor cells Activation of macrophages to induce tumor cytotoxicity occurs as a result of exposure to a variety of immunomodulating substances such as lymphokines,s9 y-interferon,and muramyl dipeptide(MDP).60-63 Liposomes are known to increase the adjuvant activity of MDP.Adju- vants Fare nonspe cific immune stimulants that boost immuno MDP m sponses to weak an olecule for exam are high y poten adjuvants in tests for vaccination against bovine viral diarrhea.Although i is unknown how this process occurs,activated macrophages can selectively kill tumor cells.Activated macrophages have been considered in the man- biological hete rogeneity of tun Although preliminary results with liposome-encapsulated immunomod. ulators are encouraging,successful application in the treatment of patients with liver metastasis may be hampered by unfavorable macrophage- to-tumor cell ratios in many metastatic tumors.s4 Therefore,it would appe that therapeutic reg en sdesigned to stimulate mac crophag have to be used -mediated other treatment modalities. Successful targeting of liposomes,at least to solid tumors located outside the main circulatory system,faces numerous challenges.As described by
Chapter one: Site-specific drug delivery using liposomes as carriers 13 liposomes has been extended to a variety of drugs, such as antineoplastic agents,16,46,47 antimicrobial compounds,42,48 and immunomodulators.49–52 In addition to utilizing liposomes as drug carriers in the treatment of intracellular infections,53 liposomes have also been used as carriers of amphotericin B in the treatment of mycotic infections, such as histoplasmosis,54 cryptococcosis, and candidiasis.54 Lopez-Berestein et al.55 reported that liposomal amphotericin B is effective in the treatment of candida and aspergillus infections in leukemia patients who have not responded to the nonencapsulated drug. The increase in amphotericin’s efficacy by encapsulation in liposomes is associated with reduced toxicity.56 Incorporation of lipophilic amphotericin within liposomes might result in a facilitated transfer of the drug to fungal cells. In turn, this selective transfer of amphotericin from liposomes to fungal cells may form the molecular basis of the reduced toxicity. Other factors, such as altered kinetics or tissue distribution, may also play an important role.57 Antibacterial activity with liposome encapsulation has been reported by Sunamoto and co-workers (in experimental Legionnaires’ disease).58 They showed that uptake of IV-injected liposomes by circulating monocytes and alveolar macrophages can be increased by coating the vesicles with a palmitoyl derivative of amylopectin. After IV injection, the amylopectin-modified liposomes were found to preferentially distribute in the lungs. Macrophages have an affinity for liposomes, and this property has been utilized in the use of these vesicles as carriers of immunomodulators to create macrophages cytotoxic to metastatic tumor cells. As a result, macrophages serve as an important barrier against the proliferation and metastatic spread of tumor cells. Activation of macrophages to induce tumor cytotoxicity occurs as a result of exposure to a variety of immunomodulating substances, such as lymphokines,59 g-interferon, and muramyl dipeptide (MDP).60–63 Liposomes are known to increase the adjuvant activity of MDP. Adjuvants are nonspecific immune stimulants that boost immunoresponses to weak antigenic molecules. MDP micelles, for example, are highly potent adjuvants in tests for vaccination against bovine viral diarrhea. Although it is unknown how this process occurs, activated macrophages can selectively kill tumor cells. Activated macrophages have been considered in the management of metastatic cancer, which is often seriously hampered by the biological heterogeneity of tumor cells with respect to growth rate and sensitivity to various cytotoxic drugs. Although preliminary results with liposome-encapsulated immunomodulators are encouraging, successful application in the treatment of patients with liver metastasis may be hampered by unfavorable macrophageto-tumor cell ratios in many metastatic tumors.64 Therefore, it would appear that therapeutic regimens designed to stimulate macrophage-mediated tumor cytotoxicity will have to be used in combination with other treatment modalities.65–67 Successful targeting of liposomes, at least to solid tumors located outside the main circulatory system, faces numerous challenges. As described by
14 Drug delivery systems,second edition Roerdink et al.,4 selective introduction of antineoplastic drugs into tumo cells in vivo by means of liposomes is currently a difficult task.However application of liposomes as a drug delivery system for antitumor drugs may be of great benefit in diminishing toxicity of encapsulated compounds by altering their pharmacokinetics or tissue distribution.In addition,liposomes can serve as sustained-or ntrolled-release syster for cytost such as cytosine arabinoside.The therapeutic effect of this cell-cycle-specific drug is enhanced by liposomal encapsulation,possibly by maintaining ther- apeutically favorable drug levels for a prolonged period of time following leakage from the liposomes,or,alternatively,from macrophages that have phago cytosed the drug-loaded liposomes P isi g examp le of a lipo omal delivery system for an antit drug has been the use of doxorubicin in liposome-encapsulated form.4 Dox- orubicin,an anthracycline antibiotic,is useful in the treatment of a variety of solid neoplasms,lymphomas,and leukemias.Its clinical use,however,is limited by its cardiotoxicity.Several investigators have shown that entrap ment of doxorubicin within liposomes greatly reduces itscardiotoxicity with- out loss of ant umor activity. n The me nism respons sible for doxoru cin's increased therapeutic index is not fully understood,but may involve low uptake of the liposomal drug by the myocardium'4or prolonged release of the drug from macrophage depots.72 While in the bloodstream,liposomes may be susceptible to destabilizing effects of serum proteins,resultin ng in the e ape of er sulated water-sol- uble compounds In additio high-den found to penetrate liposomal bilayers.This process is accompanied by loss of phosphatidylcholine from the liposomes to the HDL.?3.74 High susceptibility to phosphapidylcholine loss was found at the gel-to-liquid phase-transition temperatures of the lipos somal lipids,while oth ar mperatures the osomes wer erelatively et loss of phospholipid can be prevented by incorporation of cho lesterol into the liposomal membranes,thereby causing obstruction to pen- etration of serum lipoproteins.This may result in an increased stability of liposomes.76 Manufacturers Table 1.1 pre ents a list of liposon techno ogy-based research and devel- opment fi ns,the ir products,and r th Ir us ge In addition to those presented in the table,the following industria establishments are also involved in liposome delivery research and devel- opment:American Bioproducts,American Lecithin Co.,Applied Genetics, Argus Pharmaceuticals,Becton Dickinson&Co.Bristol-My rs Squibb,Bro des ph a ESCA A s Corp. Fou tain Ph ticals enzyme Inc.,Nichiyu Liposome Co.,Pharmos Ltd.,Ribilmmunochem Research,Inc.,Schering AG,Schering-Plough Corp.,Somatogen,Inc.,Struc- ture Probe,and Vical,Inc.According to FIND/SVP's report on the total
14 Drug delivery systems, second edition Roerdink et al.,14 selective introduction of antineoplastic drugs into tumor cells in vivo by means of liposomes is currently a difficult task. However, application of liposomes as a drug delivery system for antitumor drugs may be of great benefit in diminishing toxicity of encapsulated compounds by altering their pharmacokinetics or tissue distribution. In addition, liposomes can serve as a sustained- or controlled-release system for cytostatic drugs, such as cytosine arabinoside. The therapeutic effect of this cell-cycle-specific drug is enhanced by liposomal encapsulation, possibly by maintaining therapeutically favorable drug levels for a prolonged period of time following leakage from the liposomes, or, alternatively, from macrophages that have phagocytosed the drug-loaded liposomes. A promising example of a liposomal delivery system for an antitumor drug has been the use of doxorubicin in liposome-encapsulated form.14 Doxorubicin, an anthracycline antibiotic, is useful in the treatment of a variety of solid neoplasms, lymphomas, and leukemias. Its clinical use, however, is limited by its cardiotoxicity. Several investigators have shown that entrapment of doxorubicin within liposomes greatly reduces its cardiotoxicity without loss of antitumor activity.68–71 The mechanism responsible for doxorubicin’s increased therapeutic index is not fully understood, but may involve low uptake of the liposomal drug by the myocardium14 or prolonged release of the drug from macrophage depots.72 While in the bloodstream, liposomes may be susceptible to destabilizing effects of serum proteins, resulting in the escape of encapsulated water-soluble compounds. In addition, high-density lipoproteins (HDL) have been found to penetrate liposomal bilayers. This process is accompanied by loss of phosphatidylcholine from the liposomes to the HDL.73,74 High susceptibility to phosphapidylcholine loss was found at the gel-to-liquid phase-transition temperatures of the liposomal lipids, while both above and below those temperatures the liposomes were relatively stable.75 Net loss of phospholipid can be prevented by incorporation of cholesterol into the liposomal membranes, thereby causing obstruction to penetration of serum lipoproteins. This may result in an increased stability of liposomes.76 B. Manufacturers Table 1.1 presents a list of liposome technology-based research and development firms, their products, and indications for their usage.77 In addition to those presented in the table, the following industrial establishments are also involved in liposome delivery research and development: American Bioproducts, American Lecithin Co., Applied Genetics, Argus Pharmaceuticals, Becton Dickinson & Co., Bristol-Myers Squibb, Brocades Pharma ESCA Agenetics Corp., Fountain Pharmaceuticals, Genzyme Corp., IGI, Inc., Nichiyu Liposome Co., Pharmos Ltd., RibiImmunochem Research, Inc., Schering AG, Schering-Plough Corp., Somatogen, Inc., Structure Probe, and Vical, Inc. According to FIND/SVP’s report on the total
Chapter one:Site-specific drug delivery using liposomes as carriers 15 Table 1.1 Summary of activity in liposome usage Corporation Product Indications Fujisawa Vestar's liposomal Systemic fungal infections formulation of amphotericin B (AmBisome) Vestar MiKasome- Drug-resistant tuberculosis aminoglycoside antibiotic, amikacin Hemoglobin Blood substitute Cyclosporine Multidrug resistance to cancer chemotherapy Liposomes linked to Affinity for sites on specific proteins diseased cells Liposomes coated with a AIDS and other viral specific viral receptor diseases protein Boron isotope of mass 10 Cancer therapy Vescan MRI enhancer in animal tumors Teijin-Taisho Epoprostanol derivatives Mvocardial infarction Isocarbacyclin Cerebrovascular orders, chronic arterial obstruction in rats ImmunoTherapeutics Glucosamyl muramyl Delivery to the monocyte/ analog macrophage system in cancer chemotherapy Ciba-Geigy Muramyl tripeptide Cancer therapy Metastatic melanoma Genset Development of liposomes For antisense delivery Liposome Technology Amphosil (amphotericin B) aspergillosis infections Activators Plasminogen streptokinase In canines,encapsulated reversec local ischemia Amphotericin Bcholesterol Leishmaniasis sulfate-based delivery system (known as ABC D amphotericin B colloidal Advanced cancer patients doxorubicin (Lip-Dox) alth liposome (Doxil) Kaposi's sarcoma Metered d technology espiratory and systemic Liposome inhalation diseases Salbuta Beta 2 adrer formulations)
Chapter one: Site-specific drug delivery using liposomes as carriers 15 Table 1.1 Summary of activity in liposome usage Corporation Product Indications Fujisawa Vestar’s liposomal formulation of amphotericin B (AmBisome) Systemic fungal infections Vestar MiKasome — aminoglycoside antibiotic, amikacin Drug-resistant tuberculosis Hemoglobin Blood substitute Cyclosporine Multidrug resistance to cancer chemotherapy Liposomes linked to specific proteins Affinity for sites on diseased cells Liposomes coated with a specific viral receptor protein AIDS and other viral diseases Boron isotope of mass 10 Cancer therapy Vescan MRI enhancer in animal tumors Teijin-Taisho Epoprostanol derivatives Myocardial infarction Isocarbacyclin Cerebrovascular orders, chronic arterial obstruction in rats ImmunoTherapeutics Glucosamyl muramyl analog Delivery to the monocyte/ macrophage system in cancer chemotherapy Ciba-Geigy Muramyl tripeptide Cancer therapy Metastatic melanoma Genset Development of liposomes For antisense delivery Liposome Technology aspergillosis infections Activators Amphosil (amphotericin B) Plasminogen streptokinase reversed In canines, encapsulated local ischemia Amphotericin B cholesterol sulfate-based delivery system (known as ABCD), amphotericin B colloidal dispersion Leishmaniasis (5,12-naphthacenedione) doxorubicin (Lip-Dox) Advanced cancer patients Stealth liposomes (Doxil) Kaposi’s sarcoma Metered dose technology Liposome inhalation products Respiratory and systemic diseases Albuterol, Salbutamol (inhaled liposomal formulations) Beta 2 adrenoreceptor agonist (asthma)
6 Drug delivery systems,second edition Table 1.1 Summary of activity in liposome usage(Continued) Corporation Product Indications Technology Unlimited Development of liposomes Delivery of water and lipid-soluble material to skin,oral cav ty.lung stive tract.vagin urinary bladder,li solid tumors,and HIV-infected cells The Liposome Co. Defensins.potent ococcal infections in antifungal and antiviral AIDS patients peptides isolated from human neutrophils Mucobacterium avium (aminoglycoside intracellulare (MAD antibiotic) infections in AIDS patients TLC G-65 Amphotericin B(AB lipid Fungal infections in AlDS cor nplex)ABIC and cancer patients The Liposome Co. TLC I-16,nonionic Liver imag in CT scans iodinated contrast agent patients with advanced breast,colon,and lung cancer Univax Micro Novasome liposome Bacterial and viral vaccines Vesicular Systems technology for vaccines .g.,for pseudomonas, HIV) Liposomal adjuvant system Human influenza vaccine using TLC A-60 TLC C-53(prostaglandin E)Acute inflammatory and veso-occlusive conditions market for liposomal pharmaceuticals and diagnostics for 1995,anti-infec tives will occupy 75.2%of the market,followed by anticancers (18.8%) diagnostics(5.0%),and respiratory (1.0%).The total market for pharmaceu- tical and diagnostic liposomes reached an estimated $18 million in 1991,and this is expected to grow drama as new products gain regulator approval.The overall market for drug delivery systems is observed to reach $399 million in 1995. In Table 1.2,liposomal and conventional formulations of amphotericin B are compared in transplant recipients with systemic fungal infections.7 V.Recent advances Recent studies and examples of liposma formuations containing various entrapped ingredients are as follows:
16 Drug delivery systems, second edition market for liposomal pharmaceuticals and diagnostics for 1995, anti-infectives will occupy 75.2% of the market, followed by anticancers (18.8%), diagnostics (5.0%), and respiratory (1.0%). The total market for pharmaceutical and diagnostic liposomes reached an estimated $18 million in 1991, and this is expected to grow dramatically as new products gain regulatory approval. The overall market for drug delivery systems is observed to reach $399 million in 1995. In Table 1.2, liposomal and conventional formulations of amphotericin B are compared in transplant recipients with systemic fungal infections.78 V. Recent advances Recent studies and examples of liposomal formulations containing various entrapped ingredients are as follows:79–81 Technology Unlimited Development of liposomes Delivery of water and lipid-soluble material to skin, oral cavity, lungs, digestive tract, vagina, urinary, bladder, liver solid tumors, and HIV-infected cells The Liposome Co. Defensins, potent antifungal and antiviral peptides isolated from human neutrophils Cryptococcal infections in AIDS patients Gentamicin — (aminoglycoside antibiotic) TLC G-65 Mycobacterium avium intracellulare (MAI) infections in AIDS patients Amphotericin B (AB lipid complex) ABLC Fungal infections in AIDS and cancer patients The Liposome Co. TLC I-16, nonionic, iodinated contrast agent Liver imaging in CT scans, potential in the detection of liver metastases in patients with advanced breast, colon, and lung cancer Univax & Micro Vesicular Systems Novasome liposome technology for vaccines Bacterial and viral vaccines (e.g., for pseudomonas, HIV) Liposomal adjuvant system using TLC A-60 Human influenza vaccine TLC C-53 (prostaglandin E) Acute inflammatory and veso-occlusive conditions Table 1.1 Summary of activity in liposome usage (Continued) Corporation Product Indications