Professional Documents
Culture Documents
Abbreviations
CFTR-qstic fibrosis trammembrane receptor; DWE-dioleoylphosphatidyl ethanolamine; ECF+pidermal growth factor;
FDA-Food and Drug Admini~ration; HSV-herpes simpex virus; It_--interleukin; WV-large unilarnellar vesicle;
MDR-multidrug resistance; MI.V-multilarnellar vesicle; MTP-PE-muramyl tripeptide phosphatidyl ethanolamine;
PEG-P&polyethylene glycol modified phosphatidyl ethanolamine;
POPC--I-palmitoyl-2-oleoylphosphatidylcholine; SUV-small unilarnellar vesicle.
Conventional drugs
ABLC (Abelcet) Amphotericin B The Liposome Company, Marketed in UK and Luxembourg. Awaiting
Princeton, USA approval for treatment of aspergilfosis
AmBisome Amphotericin B NeXstar Pharmaceuticals Inc, Marketed in certain countries in Europe
Boulder, USA
Amphocil Amphotericin B Sequus Pharmaceuticals I Awaiting FDA approval
Doxil (DOX-SL) Doxorubicin Menlo Park, USA FDA-accelerated approval for treatment
of Kaposi’s sarcoma
TLC D-99 Doxorubicin The Liposome Company Phase I I I
Princeton, USA
TLC C-S3 Prostaglandin Et The Liposome Company Phase II
Princeton, USA
DaunoXome Daunorubicin NeXstar Pharmaceuticals Inc. Approval for treatment of Kaposi’s sarcoma;
Boulder, USA in Phase II trials for breast cancer, small
cell lung cancer, leukemia and lymphoma
AR-121 Nystatin Argus Pharmaceuticals Inc, Phase II
Tretinoin (AR-623) All-trans retinoic acid The Woodlands, Texas, USA Phase II (leukemia) and phase I
(Kaposi’s sarcoma)
Annamycin Argus Pharmaceuticals Inc, Phase I
The Woodlands, Texas, USA
Vincristine INEX Pharmaceuticals Corporation, Phase I
Vancouver, Canada
Proteins
OncoLipin IL-2 Oncotherapeutics, Phase II (kidney cancer)
OncoVax IL-3 and cancer tumor antigen New Jersey, USA Phase I
modified phosphatidyl ethanolamine (PEG-PE), that cytotoxic drugs, such as doxorubicin, accumulate within
engender long circulation lifetimes when incorporated these sites preferentially compared with conventional
into liposomes [5-7] Alternatively, the presence of liposomes [9,11,14].
entrapped cytotoxic drug can also lead to extended
circulation times [8]. It has been demonstrated that
increased circulation lifetimes enhance the opportunity
for liposomes, administered systemically, to leave the Targeted delivery
vascular compartment and enter certain extravascular It is envisioned that the next generation of liposomal
regions [9-1] Tumors, for example, exhibit leaky blood pharmaceuticals will consist of drug-loaded liposomes
vessels that have a reduced ability to retain circulating with surface-associated targeting information (Pig. I).
macromolecules [12, 13]. Liposomes can extravasate in Site-directing targeting ligands, such as monoclonal
these regions, thus leading to preferential accumulation antibodies, can be attached to liposomes by either
within tumors. Studies have now clearly demonstrated covalent or non-covalent methods [15-17]. The advent
that long-circulating liposomes containing PEG-PE or of novel PEG-PE lipids that allow targeting ligands to
60 /
!
700 Pharmaceutical biotechnology
be conjugated at the distal ends of the PEG spacer leading to enhanced fusogenic tendencies in low pH
has afforded both effective target binding in vitro and compartments such as endosomes [30]. The inclusion
prolonged circulation times.[le,lP-211. of lipids that are able to form non-bilayer phases,
such as dioleoylphosphatidyl ethanolamine (DOPE), can
To date, only two studies have demonstrated the
promote destabilization of the bilayer, inducing fusion
improved therapeutic activity of liposomal drugs i n
events. DOPE has been particularly useful for cationic
vivo achieved through the use of antibody-mediated
liposomes complexed with plasmid DNA for gene
targeting [22,23], with both studies employing a mon-
delivery [31*,32].
oclonal antibody against lung endothelial thrombo-
modulin (mAb 34A) and intravenously injected tumor
cells. The use of immunoliposomes may be limited
because of their potential immunogenicity [24].
In addition to antibodies, glycolipids (e.g. galactose [25]
Conventional drugs
and mannose [26]), proteins (e.g. transferrin [27] and
asialofetuin [28*]), and vitamins (e.g. folic acid [18’,29])
have been used to target specific cells via cell surface A vast literature describes the feasibility of formulating
receptors. a wide range of conventional drugs in liposomes, often
resulting in enhanced therapeutic activity and/or re-
duced toxicity compared with the free drug. In general,
intracellular delivery altered pharmacokinetics for liposomal drugs can lead
Liposomes can facilitate the intracellular delivery of to enhanced drug bioavailability to specific target cells
drugs by fusing with the target cell. Alterations in the that reside in the circulation, or more importantly,
lipid composition can render liposomes pH sensitive, to extravascular disease sites such as tumors. Recent
(a)
. Drug
“t Fusogenic protein
n Fusogenic lipid
(e.g. DOPE)
Fig. 1. Types of liposomal delivery. (a) Liposomes prepared from natural or synthetic phospholipids containing an encapsulated drug. This type
of drug delivery reduces toxicity, maintains or enhances activity and facilitates accumulation in the disease site. (b) Conventional liposornes
that incorporate lipids enhancing circulation lifetimes. Delivery in these molecules improves access to the disease site and reduces interaction
with phagocytic cells of the reticulo-endothelial system. (c) Conventional liposomes with lipids that enhance circulation lifetimes and have
surface-associated targeting information. Drug delivery using this type of liposome results in an improved therapeutic index and target cell
specific delivery. (d) Fusogenic liposomes with DOPE or fusogenic proteins. This method allows intracellular drug delivery.
Advances in liposomal drugdelivery systems Chonn and Cullis 701
advances include liposomal formulations of all-trans of rat liver macrophages by Iiposomal doxorubicin can be
retinoic acid [33, 34] and daunorubicin [35-38], which substantial [63]. Whether this finding applies to humans
has recently received Food and Drug Administration remains to be seen. To date, severe hepatic toxicities have
(FDA) approval as a first-line treatment of AIDS-related not been reported in any clinical trial.
advanced Kaposi’s sarcoma. Notable examples are given
The increase in therapeutic index of liposomal dox-
below.
orubicin most likely results from the ‘passive* targeting
to tumor sites of liposomes, which because of the
leaky vasculature, exhibit increased extravasation. This is
Amphotericin B particularly relevant for liposomes with long circulation
Liposomal amphotericin B drugs are presently approved lifetimes. At the tumor site, liposomes appear to act
for sale in certain European countries and are nearing as a depot for slow release of drug. This model is
regulatory approval in North America. Acute toxicities supported by the findings of Suzuki et al. [64] indicating
associated with amphotericin B are markedly reduced that liposomal doxorubicin remaining on the cell
with liposomal formulations, without losing broad-spec- surface is more cytotoxic than endocytosed liposomal
trum antifungal activity. Early studies on a variety of doxorubicin. Furthermore, several reports indicate that
formulations of liposomal amphotericin B demonstrated hyperthermia induces the release of doxorubicin from
the successful treatment of fungal infections in mice long-circulating liposomes and enhances their antitumor
[39,40]. Recent studies have focused on understanding efficacy [59-61].
the possible mechanisms for reduced toxicities, which
include altered pharmacokinetics [41, 42] and increased Natural or acquired resistance to doxorubicin may limit
association with high-density lipoproteins [43,44]. the clinical use of liposomal doxorubicin. Different
ways of overcoming multidrug resistance, including
Although most applications involve am intravenous route the use of modulators that can inhibit drug ef-
of administration to treat systemic fungal infections, flux mediated by P-glycoprotein [65], have been
liposomal amphotericin B can also lbe administered in explored and have proved effective in in vitro sys-
an aerosolized form, resulting in treatment of systemic tems. Several successful attempts have also been de-
Candida albicans or Cryptococcus neoformans infections in scribed, at least in vitro, to overcome multidrug
mice [45,46]. resistance by employing structurally different analogs
of anthracyclines entrapped in liposomes [66,67*].
For instance, the non-cross-resistant anthracycline an-
Doxorubicin tibiotic, annamycin, formulated in dimyristoylphos-
Phase III clinical trials on liposomal doxorubicin are phatidylcholine/dimyristoylphosphatidylglycerol SUVs
ongoing. As demonstrated in several pre-clinical and or MLVs, is more effective than doxorubicin against
clinical reports, the administration of liposomal doxoru- several tumor models, and multidrug resistance shows
bicin significantly reduces drug-associated cardiotoxi- only partial cmssresistance to annamycin, both in vitro
city beause cardiac uptake of liposome-encapsulated and in vivo [67*].
doxorubicin is substantially reduced compared with
the free drug. A variety of lipo:some-doxorubicin
formulations have been described. These include PEG- Vincristine
stabilized liposomes [14], as well as conventional egg The benefits of prolonged drug bioavailability as a result
phosphatidylcholine/cholesterol LUVs [8]. R e c e n t l y of administering the drug in a liposomal form is perhaps
described formulations that extend the circulation haIf- best exemplified by liposomal vincristine, an important
life of doxorubicin i n c l u d e dipalmitoylphosphatidyl- anticancer drug effective against a wide variety of neo-
choline/cholesterol (1 :l) liposomes containing 10 mol% plasms. Vincristine is a cell cycle specific drug that arrests
paImityl-D-glucuronide, a uronic acid derivative [47], cell growth exclusively during metaphase by attaching
and fluorinated liposomes [48]. Targeted liposomal dox- to the growing ends of microtubules and inhibiting
orubicin systems have recently been described [29,4Y]; their assembly. As such, prolonged exposure of neoplastic
however, their efficacy, compared with non-targeted cells to vincristine should greatly enhance its therapeutic
systems in vivo, has yet to be determined. index. Indeed, increased drug retention and increased
Of the above types of formulation, reports of PEG- circulation longevity, as achieved by encapsulating
coated liposomal doxorubicin dominate the recent vincristine in distearoylphosphatidylcholine/cholesterol
literature [5@,51-61]. Much attention has focused on LUVs with an internal pH of 2.0, act synergistically
the use of liposomal doxorubicin in the treatment to significantly enhance the circulation lifetime of
of AIDS-related Kaposi’s sarcoma [55-58,62]. In the encapsulated vincristine, the extent and duration of
prolonged use of liposomal doxorubicin for AIDS- tumor exposure to vincristine, and ultimately, the
related Kaposi’s sarcoma, hand-foot syndrome may be therapeutic activity of vincristine [68’,69].
a limiting toxicity [55]. With regard to liposomal The development of a liposomal formulation of vin-
doxorubicin-induced toxicities, a recent report indicates cristine, e m p l o y i n g sphingomyelin/cholesterol LUVs
that the depletion and impairment of phagocytic activity with an internal pH of 4.0 or 2.0 has recently been de-
62
702 Pharmaceutical biotechnology
scribed ]7(r*]. This formulation displays significantly en- resulting from vaccination with liposome-formulated
hanced stability and antitumor properties compared with antigen were higher than those with alum-associated
distearoylphosphatidylcholine/cholesterol LW systems antigen, and these antibody responses were enhanced
[70”]. Substantially increased vincristine accumulation, by concurrent administration of IL-7 liposomes. In
compared with the free drug, is observed in both addition, immunogenicity of alum-associated herpes
peritoneal ascitic murine P388 tumors and subcutaneous simplex virus (HSV) gD antigen can b e enhanced by
solid A431 h u m a n xenograft tumors. In addition, a a recombinant IL-7 liposomal formulation, resulting in
recent report of a liposomal vincristine formulation a significantly reduced severity and course of primary
employing PEG-PE shows an enhanced therapeutic HSV-2 infection. The sustained release of IL-7, over
index for vincristine entrapped in liposomes against sub- a period of >6 days, contributes to the observed effects
cutaneously or intraperitoneally implanted P388 tumor ]83’1.
cells [71]. Recent reports also indicate that unencapsulated cy-
As is the case for several toxic conventional drugs, tokines, at relatively low doses, augment the ther-
liposomal vincristine exhibits reduced toxicity compared apeutic effects of liposomal reagents [84*,85]. For
with the free drug [72]. Particularly notable is the greatly instance, unencapsulated recombinant IL-2 adminis-
enhanced efficacy that can be achieved for liposomal tered intraperitoneally (10 000 U day-r). in combination
vincristine compared with equivalent doses of the bee with intravenously administered phosphatidylcholine/
drug. Liposomal vincristine is currently in clinical trials. phosphatidylserine (1: 1) MLVs containing a synthetic
peptide derived from C-reactive protein (RS-83277),
significantly inhibited tumor metastases and prolonged
survival of C57B1/6 mice bearing established pulmonary
Proteins and peptides metastases of fibrosarcoma T241. The combination
therapy was accompanied by an increase in the number
of Thy1.2 cells in the lungs of RS-83277 MLV/IL-
The majority of current hposomal protein formulations 2 treated animals compared with those receiving
are still in various preclinical research stages (recently RS-83277 MLVs alone.
reviewed in [73]), with one Iiposomal interleukin (IL)-2
drug entering a phase II clinical trial for kidney cancer.
For the production of artificial blood substitutes, the Liposomal muramyl tripeptide
use of liposomes to encapsulate hemoglobin is actively Muramyl tripeptide phosphatidyl ethanolamine (MTP-
being investigated for the in vivo delivery of hemoglobin PE) is a synthetic lipophilic analog of muramyl dipep-
without many of the inherent toxicities associated with tide, the smallest component of a mycobacterium
the delivery of the free molecule (recently reviewed capable of stimulating the immune system. MTP-PE
in [74-76]). Another area of intense research is the is a potent monocyte/macrophage activator and is
application of liposomes exhibiting improved adjuvancy currently under clinical investigation against metastatic
for vaccine development. melanoma and osteosarcoma (reviewed in [86]). Of
major interest to the development of liposomal MTP-PE
immunomodulators is the recent finding that repeated
Immunomodulaton: interleukins per os (oral) administration of l-palmitoyl-2-oleoylphos-
The feasibility of formulating cytokines in MLVs phatidylcholine (POPC) MLVs containing a synthetic
[77-791 and in sterically stabilized SUVs [80,81*] muramyl tripeptide, CGP 19835A (Ciba-Geigy, Basel,
has recently been demonstrated. These Iiposomal cy- Switzerland), increased the tumoricidal activity of
tokines show great promise as immunoadjuvants for alveolar and peritoneal macrophages against renal cell
vaccine development. IL-2 enapsulated in sterically carcinoma [87**]. Liposomes were rapidly absorbed
stabilized SUVs (65 nm in diameter) is significantly in the intestine and reached the systemic circulation
more effective than free IL-2 both in increasing within 4 h, as determined by the biodistribution of
leukocyte number in the blood and spleen and in trig- radioactively labeled, or N-4-nitrobenzo-2-oxa-1,3-
gering spleen lymphokine-activated killer-cell activity diazolephosphatidyl ethanolamine fluorescently labeled
[81-J. Co-injection of phosphatidylcholine/cholesterol POPC-CGP 19835A liposomes. The mechanism of
(1:l) MLVs containing IL-6 (50 000 U IL-6 mouse-l) liposomal muramyl tripeptide antitumor activity is linked
or 65 kDa heat-shock protein antigen (0.03 pg mouse-r to its activation of monocyte/macrophage tumoricidal
or 0.3 pg mouse-r) significantly enhanced secondary function, as shown by several recent reports [88-90].
antibody responses at antigen dosages where other
adjuvants (e.g. Ribi or dimethyldioctadecylammonium-
bromide) exhibit no adjuvant activity [79]. Liposomal
formulations of IL-7 have been shown to enhance Antisense oligonucleotides, ribozymes and genes
the immune responses of mice vaccinated with either
alum-associated or liposome-formulated recombinant Antisense molecules and ribozymes present interesting
HIV envelope protein env-2-3SF2 [82]. Antibody titers challenges for delivery systems. The efficacy of these
63
Advances in liposomal drug-delivery systems Chonn and Cullis 703
drugs is dependent on their ability to gain entry into promote fusion with target cells [94*]. Wang et al. [ 102**]
cells in an intact form; however, they are particularly describe the use of phosphatidylcholine/cholesterol(3:2)
susceptible to degradation by nucleases in the bio- containing 0.5mol% folate conjugated to PEG-dis-
logical milieu and usually cannot cross the target cell tearoylphosphatidyl ethanolamine to deliver antisense
membrane. For example, in general, phosphodiester oligonucleotides against human epidermal growth factor
antisense oligonucleotides have been reported to have (EGF) (up to 2.0x 107 molecules cell-l) in a folate-spe-
little or no inhibitory effect in culture because they are cific manner, as free folic acid competes with EGF
rapidly degraded in the culture medium. In addition, uptake.
these molecules are highly charged and can activate
An interesting approach to increase the association
the complement system, resulting in the generation
of antisense oligonucleotides with liposomes involves
of anaphylatoxins and other immunomodulators. The
coupling antisense oligonucleotides to cholesterol via
potential of liposomes to encapsulate antisense oligonu-
a reversible disulfide bond [103*,104-1071. Using this
cleotides or DNA, protecting them from nucleases and
method, the association of oligonucleotides with im-
complement, represents a great advantage over other
munoliposomes is improved by a factor of -10. The
drug carriers, such as polymers or immunoconjugates.
capacity of modified oligonucleotides directed against
The further potential for fusogenic liposomes to promote
the tat gene of HIV-1 to inhibit HIV-l proliferation in
intracellular delivery of these compounds is also of major
acutely infected cells has been found to be the same as
importance. The application of liposomes to deliver
the unmodified oligonucleotide on an equimolar basis
antisense oligonucleotides, ribozymes and genes is an
(IC50 =0.1 pM) [103*].
area of intense research.
To date, only a few papers have reported the use
of cationic liposomes to deliver ribozymes, a class
Antisense oligonucleotides and ribozymes of RNA molecule that possesses enzymatic cleavage
Several reports demonstrate the feasibility of employing activity [108’,109,1 l@,l 11.1. Ribozymes, being RNA
liposomal systems to deliver antisense oligonucleotides, molecules, are highly susceptible to nuclease digestion.
with the accompanying significant enhancement of effi- Their stability is markedly increased in vitro in the
cacy in vitro and in vitro [91,92,93*,9,4*]. Cellular uptake presence of cationic liposomes, with >30% remaining
of fluorescently labeled oligonucleotides is significantly intact after a 60min incubation in medium containing
enhanced by cationic liposomes, as assessed by confocal 10% fetal bovine serum. The feasibility of using a variety
laser scanning microscopy, flow cytofluorometry and of cationic liposomes. to deliver ribozyrnes into cultured
laser-scanning microscopy. Intact oligonucleotides are cells in vitro has recently been described for ribozymes
found in the cytoplasm and nucleus only when they are directed against leukocyte-type 12-lipoxygenase mRNA
delivered by cationic liposomes. [lOS*], bcr-abl mRNA [l lO$ or multiple drug resistance
The overwhelming conclusion from studies to date is (MDR)-1 mRNA [ill’]. Liposome-mediated transfer
that liposomes are able to resolve the problems of ex- of ribozymes against MDR-1 mRNA was shown to
tracellular degradation by nucleases and poor membrane reverse the MDR phenotype of adriamycin-resistant
permeability that are inherent for oligonucleotide drugs. and vindesine-resistant human pleural mesothelioma
This has been achieved using a variety of liposomal cell lines and restored sensitivity to chemotherapeutic
compositions, with the majority employing cationic drugs [Ill’]. As with antisense oligonucleotides, it is
lipids and DOPE [93*,95-971. A recent report describes likely that liposomal systems will provide significant
an extensive physicochemical study of the aggregation advantages to the delivery of ribozyme molecules in
and fusion reactions that occur during the formation vivo. The development of such liposomal formulations
of oligonucleotide and cationic liposomal complexes in is advancing rapidly
solution [98]. Furthermore, several approaches to encap-
sulate antisense oligonucleotides have been described.
A probe sonication method employing phosphatidyl- Genes
choline/cholesterol/dipalmitoylphosphatidylethanol- Several reviews on the use of liposomes to deliver
amine covalently coupled to L-polylysine (5.5:3.0:1.5) genes have appeared recently (see [112-l 14]; this issue,
has recently been shown to result in liposomes with Cunliffe, Thatcher and Craig, pp 709-713). Although
a diameter of 1 10-140nm and encapsulation efficien- the utility of cationic liposomes in the delivery of
cies ranging from 55% to 100% depending on the reporter genes was noted in the early 1980s, we are
oligonucleotide [99]. The use of immunoliposomes has only now beginning to characterize these systems and
also been described [100]. Aigner and Caroni [lOl*] to understand the cellular processes -that are required.
report the use of liposomes composed of phosphatidyl- For instance, the role of DOPE in mediating cytosolic
choline/phosphatidylserine (10:1) and myelin proteins delivery of plasmid DNA has now been elucidated
derived from adult rat spinal cord or sciatic nerve to [32,115]. It has been shown that the principal route
deliver antisense oligonucleotides to dorsal root ganglion of cationic liposome-mediated gene transfer occurs after
neurons. In addition, liposomes containing viral fusion endocytosis [116*]. Recent electron microscopy studies
proteins, derived from Sendai virus, have been used to have attempted to reveal the structural features of plasmid
64
704 Pharmaceutical biotechnology
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m j Bfof Chem 1994, 269:2S5&2561. human serum. B&him Siophys Acta 1994, 1194~61-68.
A comprehensive stdy of structure/function relationships of cationic 49. Park JW, Hong K, Carter P, Asgari H, Guo LY, Keller GA,
lipids and neutral lipids for optimization of cationic liposomemediatd . W&h C, Shalaby R, Kotts C, Wood WI et al.: Devtbpment of
gene transfer. anti*Wfa imnnmolii for ancer therapy. Proc Nat/
32. Farbood .H, Setbina .N, Huang L: like rok of diileoyl . Acad. Sci USA_ 1995, 92:1327-1331.
.. - _-_
p in ationic r i meditd gene Describes the coupling to liposomes of anti-p18SntR2 antibodies,
tmwfu. Biochim Siophvs Acia 1995, 1235:289-295. which retain their antiproliferative activity. lmmunofiposomes are rapidly
internalized via receptor-mediated endocytosis. Specific delivery of
33. Parthasarathy R, Sacks PC, Hanis 0, Brock H, Mehta K: doxorubicin-loaded anti-p18SHEa immunofiposomes to tumor cells in
tntenction of ritd all-tr~inoic acid with vivo was assessed 24 h after a single intraperitoneal injection, yielding
quamous urcbwmu c&s. cancer chemh Pham?aco/ 1994, a tumor:muxle ratio of 2.69 for doxorubicin levels on the basis of
X527-534. percentage injected dose per gramme of tissue.
66 i
706 Pharmaceutical biotechnology
50. ‘Cabizon A, Catane R, Uziely 8, Kaufman 6, Safra T, Cohen o f a d r i i m y c i n enmted in immunoliposomes. Biochim
. R, Manin F, Huang A, Barer&& Y: Prolonged circulation Siophys Acta 1994, 1224~445-453.
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of doxorubicin encapsulated in polyethyienC_glycol coated 65. Sela S, Husein SR, Pearson JW, Long0 DL, Rahman A: Reversal
liposomes. Cancer Res 1994, 54:987-992. of multidrug resistance in human coton cancer cells expressing
In ,this -study, the pharrnacokinetics in cancer patients of Doxil the human MDRI gene by liposomes in combination with
(doxorubicin encapsulated in PEG-containing liposomes) at two doses, monoclinal antibody or verapamil. j Narl Cancer fnu 1995,
2Smgm-2 and SOrngm-2, was compared with free doxorubicin. With 871123-l 28.
Doxil, -30% of the injaed dose was cleared from plasma, with an initial 66. Perez-Soler R, Ling YH, Zou Y, Priebe W: C e l l u l a r
distribution half-life of l-3 h. The rest of the Doxil was cleared slowly pharmacology of the partially nonsrorwaistant anthracycline
(second half-life of 45 h). Concomitant with the extended circulation annamycin entrapped in liposomes in KB and RR-VI cells.
times was an accumulation in malignant effusions, oeakina between 3 Cancer Chemother Pharmacol 1994, 34:109-l 18.
days and 7 days. For free doxorubici; the first half-Iif; “al&were 0.07 h
(25 mgm-*t and 0.06 h (SOmg m-2) and the second half-life values were 67. Zou Y, Ling H, Van NT, Priebe W, Perez-S&r R: Antitumor
8.7 h (2Smgm-2) and 10.4h (SOmgm-2). l activity of free and liposome-cntrapped annamycin, a iipophilic
anthtacycline antibiotic with non-cross-resistance properties.
51. Cabizon A, lsacson R. Libson E. Kaufman 6. Uziely 6, Catane, Cancer Res 1994, 54~1479-1484.
R, Ben-Dor CC, Rabello E, Cass Y, Peretz T ef al.: Clinical These authors assess the ability of annamycin to overcome multidrug
studies of liposome+n capsulated doxorubicin. Acta Oncol resistance in vifro and in viva. They repon the following findings:
1994, 33:779-786. first, MDR cell lines (KB-VI, 8226/Dox, P388/Dox and CEM/Vbl) display
partial lack of cross-resistance to annamycin in vitro; second, liposomal
52. Vaage J, Barbera-Guillem E, Abra R, Huang A, Working P:
annamycin is more effective than doxorubicin in the treatment of
Tissue distribution and therapeutic effect of intravenous free
or encapsulated liposomal doxorubicin on human prostate lung tumors, subcutaneous tumors, and tumors expressing the MDR
phenotype in viva; and third, MLVs are more effective than SUVs.
carcinoma xenografts. Cancer 1994, 73:1478-l 484.
68. B o m a n N L , Masin 0, M a y e r LO, Cullis PR, Bally M B :
53. Vaage J, Donovan 0, Loftus T, Abra R, Working P,
. liposomal vincristine which exhibits increased drug retention
Huang A: Chemoprevention and therapy of mouse mammary
and increased circulation longevity cures mice bearing P388
carcinomas with doxorubicin encapsulated in sterically
tumors. Cancer Res 1994, 54:2830-2833.
stabiliied liposomes. Cancer 1994, 73:2366-2371.
In this report, use of a low internal pH and incorporation of ganglioside
54. Vaage J, Donovan D, Loflus T, Uster P, Working P: Prophylaxis C&I into the liposome acts synergistically to significantly enhance the
and therapy of mouse manunar), carcinomas with doxotubicin circulation lifetime of encapsulated vincristine. Therapeutic activity of
and vincristii encapsulated in sterically stabiiii lies. the resulting liposomal vincristine preparation is dramatically improved,
Eur ) Cancer 1995, 31A:367-372. resulting in cures in >SO% of mice inoculated with P388 lymphocytic
leukemia.
5s. G o r d o n KB, T a j u d d i n A , Guitan J, Kutei T M , E r a m o
LR, VonRoenn J: Hand-foot syndrome associated with 69. M a y e r LO, Masin 0, N a y a r R , B o m a n N L , Bally M E :
liilated doxorubicin tbenpy. Cancer 1995, Pharmacology of liposomal vincristine in mice bearing 11210
75:2169-2173. ascitic a n d B16/616 solid tumors. Br I C a n c e r 1 9 9 5 ,
71:482-488.
56. Bogner jR, Kronawitter U, Rolinski B, Truebenbach K, Goebel
FD: lii doxon&zin in the t r e a t m e n t o f a d v a n c e d 70. Webb MS, Harasym TO, Masin 0, Bally MB, Mayer LO:
AIDS-related Uaposi sarcoma. 1 Acquired Immune Deficiency . . SphinRomydin-cholesterol loosomes sign*Xcultly enhance the
syndromes 1994, 7z463-468. p~ha&c&inctic a n d ther+utic prGpefties bf vi&i
in murine and human tumour models. Br J Cancer 1995,
57. Wagner D, Kern WV, Kern P: L@osomal doxorubicin in AIDS- 72:89&904.
related Kaposi% sarcomaz long-term expeGences. CIin Invest Significantly enhanced stability and therapeutic activity is observed with
1994, 72:417-423. a liposornal formulation that is composed of sphingomyeli~cholesterol,
with an internal pH of 4.0 or 2.0. The im@roved circulation lifetime
58. Harrison M, Tomlinson 0, Stewart 5: Liimai-entrapped of vincristine in sphingomyelin/cholestetol liposomes correlates with
doxorubii an active agent in AIDS-related Kaposi’s sarcoma. increased vincristine accumulation in peritoneal ascitic murine P388
/ C/in Oncol 1995, 13:914-920. tumors and in subcutaneous solid A431 human xenograft tumors.
Increased vincristine delivery to tumors is accompanied by increased
SY. Huang SK, Stauffer PR, Hong K, Guo JW, Phillips TL,
antiturnor efficacy with >SO% cures in mice bearing ascitic P388 tumors.
Huang A, Papahadjopoulos D: Liposoma and hypertbermia
Furthem\ore, the time required for a 100% increase in the solid mass of
in mice: increased tumor uptake and therapeutic efficacy
human A431 xenograft tumors was delayed by >40 days compared with
of doxorubicin en sterically stabilized liposomes. Cancer Res
a delay of 7 days for free vincristine.
1994, 54:2186-2191.
71. Allen TM, Newman MS, Woodle MC, Mayhew E, Uster
60. Unezaki 5, Maruyama K, Takahashi N, Koyama M, Yuda T,
PS: Phannacokimtia and anti-tumor activity of vincristine
Suginaka A, lwatsuru M: Enhanced delivery and antiturnor
encapsulated in sterically stabilized liposomes. In! / Cancer
activity of doxorubicin using long-ciradatii &rmosensitive
1995, 62:199-204.
iii containing ampbipathic poiyetbylene glycoi in
combination w i t h local $pe&emdi. Pharm Res 1 9 9 4 , 72. M a y e r C D , Baily M B , L o u g h r e y H , Masin 0, Cullis PR:
11:1180-1185. Liposomal vincristine preparations which exhibit decreased
drug toxicity and increased activity against mutine t.1210 and
61. Ning 5, Ma&cd K, Abra RM, Huang AH, Hahn GM: Hyper-
P388 tumors. Cancer Res 1990, SO:575-579.
then& induces doxontbicin release f r o m long&culating
Iii and enhances their a&-tumor efficacy. Int / Radiat 73. Weiner AL: Liposomes for protein delivery: selecting
Oruol &of Phy 1994, 29:827-834. manufacture and development processes. lmmunome~huds
1994, 4:201-209.
62. Gill PS, Rarick M, McCutchan JA, Slater L, Parker B, Muchmore
E, Bernstein-Singer M, Akil 6, Espina BM, Karilo M, Levine A: 74. Rudolph As: Encapsulated hem obin: current issues and
Systemic trutmcnt of AIDS-@&d t&p&s sarconbz results future goak. Anif Cells Blood Su
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of a mndomized trial. Am j Med 1989, 8757-61. 1994, 22:347-360.
63. Oaernen 1, Hofstede G, Ten-Kate MT, Bakker-Woudenberg 7s. Tsuchida E: Stabilized hemoglobin vesicles. Artif Cells Blood
IAJM, Scherphof CL: lipasomal doxorhicidi toxicity: Substitute lmmobil Biorechnol 1994, 221467-477.
depktii and impairment of pfqoqtii activity of liver
math. /n( / Cancer 1995, 61:7X-721. 76. Zheng S, Zheng Y, Beissinger R: Efficacy, physial properties
and pharmacokinetics of sterically-stabilized liposomc-cncap
64. Suzuki S, Watanabe 5, Uno S, Tanaka M, Mauko T, Hashimoto s u l a t e d hemo@bii. Artif C e l l s Slood Subsritu~e mmobil
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Advances in liposomal drug-delivery systems Chonn and Cullis 707
68
708 Pharmaceutical biotechnology
liposomes containing folate as a targeting ligand. Delivery (2x107 111. K i e h n t o p f M , Brach MA, Licht T, Petschauer S, Karawajew
ECF receptor moleculescell-1) was enhanced l@fold compared with . L, Kirschning C, Herrmann F: Ribozyme-mediited deavage of
non-targeted liposomes, whereas uptake of free oligonucleotides was the MDR-t transcript restores chemosensitivity in previously
barely detectable. Cell uptake was inhibited by the presence of 1 mM resistant cancer cells. EM60 j 1994, 13:4645-4652.
free folate. Phosphorothioate oligonucleotides and native phosphodiester Demonstrates in vitro the feasibility of liposome-mediated transfer of
DNA, when delivered via this method, exhibited similar growth hammerhead ribozymes directed against MDR-1 transcripts, reversing
inhibition activities. the MDR phenotype in human pleural mesothelioma cell lines that are
vindesine or doxorubicin resistant.
1 0 3 . Zelphati 0, mgner E, Lesennan L: Synthesii a n d a n t i - H I V
. activity of thiiolesterykoupkd phosphodiiter antisense 112. Ledley FD: Non-viral gene therapy. Curr Opin Biotechnoll994,
oligunuckot~kks incorporirted into immunoliposomes. Antiviral 5:626-636.
Res 1994, 25:13-25. 113. Miller N, Vile R: Targeted vedon for gene therapy. FAXB 1
Investigates the feasibility of encapsulating cholesterol-coupled antisense 1995, 9:190-199.
oligonucleotides in immunoliposomes. The authors successfully synthe-
size 1OOnm LUVs containing 80-160 oligonucleotide molecules per 114. Smith JC, Walrem RL, German JB: liposomes as agents of
liposome (with encapsulation efficiencies of ZO-30%). Oligonucleotides DNA transfer. B&him Siophys Acra 1993, 1154:327-340.
were protected from DNase I digestion. The capacity of the modified 115. Zhou X, Huang L: DNA transfection me&ted by c&ionic li-
anti-HIV antisense oligonucleotides to inhibit the replication of HIV was posomes containing lipopoiylysine: characterization and me&
the same on an equimolar basis as that of unmodified oligonucleotides, anism of action. Biochim Siophys Acta 1994, 1189: 195-203.
as demonstrated in vitro using an HIV proliferation assay in acutely in-
fected CEM cells. Furthermore, the cholesteryl-modified oligonucleotides 116. Wrobel I, Collins D: Fusion of cationic liposomes with
acquire the target specificity of the antibody on the liposome.
. mammalian cells occurs after endocytosis. Biochim Siophys
Acta 1995, 1235:29&304.
104. Shea RC, Marsters JC, Bischotberger N: Synthesis, hybridization Using a lipid mixing assay, these authors demonstrate that binding to the
properties and antiviral activity of lipid+digodeoxynud cell surface is insufficient for fusion of the cationic liposome with the
tide conjugates. Nucleic Acids Res 1990, 18:3777-3783. cell and that uptake into the endocytic pathway is required for fusion to
occur.
105. Krieg AM, Tonkinson J, Matson S, Zhao Q, Saxon M, Zhang
LM, Bhanja U, Yakubov L, Stein CA: Modiicatiat of antisense 117. Gustafsson J, Arvidson C, Karlsson C, Almgren M: Complexes
phosphodiiter oligodeoxynucleoti b y a 5’ choksteryi between cationic liposomes and DNA visualized by cryo-TEM.
moiety increases cellular association and improves efficacy. Biochim Siophys Acta 1995, 1235:3OS-312.
Proc Nat/ Acad Sci USA 1993, 90:1048-1052. 118. Gershon l-i, Ghirlando R, Cuttman SE, Minsky A: Mode
of formation and structural features of DNA-cationic
106. Boutorine AS, Gus’kova LV, lvanova EM, Kobetz ND, Zarytova
liposome complexes used for transfection. Biochemistry 1993,
VF, Ryte AS, Yurchenko LV, Vlassov VV: Synthesis of
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alkyiating oligonucleotide derivatives containing cholesterol
or phenazinium residues at their 3’-terminus and their 119. Zabner J, Fasbender Aj, Moninger T, Peollinger KA, Welsh MJ:
interadion with DNA within mammalian cells. f&3.5 Len 1989, .. Cellular and molecular barriers to gene transfer by a cationic
25&l 29-132. lipid. j Biol Ckm 1995, 270: 18997-l 9007.
Comprehensive study investigating the key cellular processes required
107. Kabannw AV, Wnogradov SV, Ovcharenko AV, Krivonos AV, for cationic lipid-mediated gene transfer.
Melik-Nubarov N S , Kiselev VI, kverin E S : A n e w dass
o f a&vi& antisense oliideotidcs c o m b i n e d with a 120. Caplen NJ, Alton EWPW, Middleton PC, Dorin JR, Stevenson
hydrophobii substii effectively inhibit influenza virus . BJ, Gao X, Durham SR, jeffery PK, Hodson ME, Coutelle C
reproduction and synthesb of virus-specific proteins in MDCK et al.: liiited CFI’R gene tvanser to the nasal
ceils. FEB.5 Left 1990, 259:327-330. epithelium of patients with cystic f&&s. Nature h&d 1995,
1:39-s.
108. Cu p, Veerapanane 0, Rossi J, Natarajan R, Thomas L, Nadler First placebo-controlled trial of liposome-mediated CFTR cDNA transfer
. J: Ribowited inhiion of expression of leukocyte-type to patients with cystic fibrosis. The safety and r&cacy of 300mg per
12-lii~ in porcine aortic vascular smooth muscle nostril of liposome-encapsulated CF7R is compared with an equivalent
&Is. Circ Res 1995, 77~14-20. dose of liposorne alone. No evidence of treatment-related toxicity, either
Describes rhe fir% chimeric hammerhead ribozyme active against an clinically or histologically, was detected in any of the treated subjects.
eicosanoid-generating mRNA Efficient &livery into porcine aortic
vascular smooth muscle cells is achiwed by transfection with cationic
liposomes.
109. Taylor NR, Kaplan BE, Swiderski P, Li H, Rossi JJ: Chime& A Chonn, INEX Pharmaceuticals Corporation. 1779 West 75th
DNA-RNA hammerhead ribozymes have enhanced in vitro Avenue, Vancouver, British Columbia V6P 6P2. Canada.
catatytll cffkimcy ad increased stability in vivo. Nucleic E-mail: achonn@inexpharm.com
Acids Res 1992, 20:4559--4565.
PR Cullis. Department of Biochemistry and Molecular Biology,
110. Leopold LH. Shore SK, Mewkirk TA, Reddy RMV, Reddy EP: The University of British Columbia, 2146 Health Sciences
. M&%nii riborymc-mcd’ied cleavage o f 6cr-a6/ mRNA i n
Mali, Vancouver, British Columbia V6T 123, Canada and
myebid kukmias. Blood 1995, 85:2162-2170.
Transfer of multi-unit ribozymes against bcr-abl mRNA into murine INEX Pharmaceuticals Corporation, 1779 West 75th Avenue,
myeloblasts transformed with the bcr-abl gene is compared using Vancouver, British Columbia V6P 6P1, Canada.
cationic liposomes and folic acid/polylysine liposomes. E-mail: pcullis@inexpharm.com
69