Vol. 91, No. 4, 1979 December
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
BIOCHEMICAL
Pages 1295-1301
28, 1979
--IN VITRO
AND --IN VIVO STUDIES WITH ADRIAMYCIN Eric A. Forssen and Zolthn A. TSk&*
LIPOSOMES
University of Southern California, School of Pharmacy and School of Medicine, Department of Biochemistry Comprehensive Cancer Center, Los Angeles, California Received
October
26,1979
Summary: Liposome entrapped adriamycin retains its full cytotoxic potential when tested under in vitro conditions against murine leukemia L-1210 cells. --In vivo drug dizrmon studies indicate that, relative to the free drug, a lower proportion of adriamycin administered in the liposome form is delivered to the heart and kidneys at one and four hours after injection. When administered to normal mice in high doses, anionic adriamycin liposomes appear less harmful than equal doses of the free drug as judged by alterations in animal weight gain. In these studies, a nova1 double packing procedure has been used for the entrapment of adriamycin in phospholipid vesicles. INTRODUCTION A number mycin-D
of chemotherapeutic
have been encapsulated
for overcoming toxicities. primarily
to entrap
of the vesicles. phospholipid
membranes
us to report
resulting
developed
MATERIALS choline
reviewed
compounds
in our laboratory activity
(1,2,3).
and actino-
have been useful and reducing
Liposomes
drug
have been used
into the aqueous compartment
materials
minimizes
(Adr) can destabilize
(4,5).
this disruptive
of liposome-Adr
--in vivo distribution
Ara-C
such as adriamycin
in loss of entrapped
on the --in vitro
reported
These preparations,
drugs by incorporation
ampholiphilic
methotrexate,
--in vivo tissue distribution
has been extensively
cells as well as their altered have been previously
altering
hydrophilic
However
The approach enabling
in liposomes.
drug resistances,
This subject
agents including
complexes
and toxicity.
A portion
effect against
thus L-1210
of these findings
(6).
AND METHODS
Adriamycin (provided by Adria Laboratories, Columbus, Oh.) is bound to phosphatidyl(PC) (Sigma Chem. Co, St. Louis, MO.) by adding a solution of the drug in normal
*Address: USC Comprehensive Cancer Center, Cancer Research Labs. Rm. 208, 1303 N. Mission Road, Los Angeles, CA 90033. Abbreviations used: Adriamycin: Adr; Cholesterol: Ch; Phosphatidyl Choline: PC; Arabinosylcytosine: Ara-C. 0006-291X/79/241295-07$01.00/0 12%
Copyright All rights
@ 1979 by Academic Press, Inc. ofreproduction in anyform reserved.
Vol. 91, No. 4, ~1979
BIOCHEMICAL
AND BIOPHYSICAL
RESEARCH COMMUNlCATlONS
saline to a two-fold molor excess of the dried lipid. This material is then sonicated for 5 min per ml under a nitrogen atmosphere using a needle probe type sonicator (Braun Sonic-1410) set at 100 watts. The resulting drug-lipid mixture is then entrapped within anionic liposomes consisting of a 7:3:1 molar ratio of PC, cholesterol (Ch) and phosphatidyl serine (Sigma Chem. Co.) or cationic liposomes containing PC, Ch and stearylamine (Eastman Org. Chem. Rochester, N.Y.) 7:3:0.5. This liposome suspension is sonicated as above at 100 watts for 5 min per ml. Unentrapped Adr is separated from liposomes by gel filtration on Sephadex G-50. Using this procedure, between five and ten percent of the starting Adr can be entrapped. The stability of Adr-liposomes was determined by incubating the liposomes in plasma at 37’C with continuous agitation. Liposomes (0.2 ml) containing [ H] - Adr (Morave Biochemicals, City of Industry, CA. Specific activity: 600mCi/mmole) were added to a dialysis bag (m.w. cutoff = 3500), (Spectrum-Medical Ind., Los Angeles, Ca.) containing 2 ml human plasma. The dialysis bag was placed into a cylinder containing 50 ml of the same plasma. Samples of the dialysate were removed and counted by liquid scintillation. Leakage was expressed as a percentage of the original radioactivity. Anionic liposomes containing [ HI-Adr were utilized for --in vivo distribution studies. Swiss mice (Simonson Labs; Gilroy, Ca.) weighing about 20 grams each, received [ HIAdr in either free or entrapped form by tail vein injections. Mice were then sacrificed at one and four hours following drug administration. The brain, heart, kidneys, liver, lungs and spleen were removed and prepared for liquid scintillation counting using tissue solubilizer (Protosol, New England Nuclear; Boston, Ma.). The distribution of [ HI-Adr to each organ was expressed as a percentage of the total radioactivity measured for all organs examined. Murine leukemia L-1210 Moore cells, adapted to cell culture, were grown in suspension at 37’C in RPMI-1640 media supplemented with 10% fetal calf serum (Grand Island Biological, Co., Grand Island, N.Y.), cells were treated with Adr in either the free or entrapped form at concentrations ranging from 10 to 10 molar. Liposome treated cells received Adr in anionic vesicles, prepared as described above. For controls free Adr was added to cells alone or with empty anionic liposomes containing the same amount Other controls received either empty anionic liposomes of phospholipid and cholesterol. without Adr or no treatment. The Adr concentration required for 50% inhibition of cell growth (Ic ) was determined for each group of L-1210 cells tested. All samples were done in duplicate. Swiss mice, weighing 20 grams each, were administered two intravenous Adr doses (lOmg/kg) with a one week interval in a high dose toxicity study. One group was injected with the free drug while a second was given Adr entrapped in anionic liposomes. Controls received normal saline. Animal body-weights were determined daily for the first two weeks and twice weekly thereafter. RESULTS The stability --in vitro.
of both cationic
While almost
hours of incubation, 1% in cationic
liposomes
in plasma
30% of free Adr escaped into the dialysate
less than 10% of entraped
liposomes
these values increased
and anionic
Adr in anionic
was found in the dialysate,
Figure
to about 20% and 5% for anionic
was demonstrated
during
liposomes
the first two and less than
1. At the end of ten hours,
and cationic
liposomes
respectively.
The 1~5~ for Ll210 cells treated with free Adr, either with or without empty liposomes was 1.9x10 -8 molar. Anionic liposome-Adr produced only a slightly lower level
1296
Vol. 91, No. 4, 1979
BIOCHEMICAL
AND BIOPHYSICAL
RESEARCH COMMUNICATIONS
50
FREE
0
1
2
Figure 1: The stability at 37OC.
those cells treated
with
anionic
The relative after
injection
anionic
liposomes
liposomes
display
organ distribution
liposomes
deliver
a lower
Much of the liposomal
as evidenced
by the enhanced
decrease
free or liposome between
an average
receiving
weight
Adr alone.
group during
no inherent
proportion
form,
was seen for
that over the concentration
toxicity. Adr at one and four hours
these studies demonstrate
of the administered
uptake.
is shown on Figure Over a period
that
dose to brain, heart
At four hours following
of Adr to cardiac,
gain of 56
Also notable
by dialysis
Adr is taken up by the reticuloendothelial
the two groups is revealed.
showed
a
7
of cell growth
alone, indicating
system
injection
an even
renal and brain tissue can be noted.
in the changes of net body-weight
entrapped
LIPOSOMES
No inhibition
2. Most notably,
hepatic
in the distribution
A comparison
CATIONIC 6
of free and liposomal
in Figure
and kidneys.
greater
LIPOSOMES
of liposomes in human plasma as determined
anionic
is illustrated
ANIONIC
4 5 OF INCUBATION
with an IcSO of 5x10 -8 molar.
of cytotoxicity
range tested,
3 HOURS
ADRIAMYCIN
for mice receiving 3. A significant
Adr in either
difference
of ten weeks the liposome-Adr
10% compared
is the less pronounced
to only 18 weight
(p
0.001) group
14% seen in the group loss for the liposome
the first two weeks of the study.
DISCUSSION Liposomes actinomycin-D
have been used to increase resistance
(7). Alternatively,
tumor cell drug uptake liposomes
1297
and to overcome
may also function
as protective
Vol.
91,
No,
4, 1979
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
90 60 70
One
60 __
50
-
T
25
Hour
After
Injection
m
Free
A&
m
Liposome
Adr
20
15
BRAIN
B Four
0
BRAIN
Figure
vehicles potentially therapeutic
2.
by altering toxic agents
KIDNEY
HEART
Relative organ distribution Adr at one and four hours normal
substances is Adr
patterns into which
distribution tissues
has pronounced
1298
(8). cardiac
After
Injection
LUNG
of free and aniomic after injections.
of drug sensitive
LIVER
Hours
and One and
liposome
entrapped
decreasing of the
commonly
renal
toxicities.
the
uptake
of
used
chemo-
Therefore
BIOCHEMICAL
Vol. 91, No. 4, 1979
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
160
150
80
Figure
IL
2
3 WEEKS
3: Body-weight charts for trapped Adr, 10 mg/kg.
it is of interest
decreases
1
to study
in toxicity.
the disruptive
the
4 FOLLOWING
which
liposome
between
Adr and liposomes
permiability
causing
a loss of entrapped
peutic packing
vesicles
and toxicological procedure
containing
studies.
for liposome
7 INJECTION
8
9
10
either free or liposome enstandard deviations from five
mode of Adr administration
entrapped
been difficult
it exerts on phospholipid
interactions
tion of phospholipid
6 FIRST
mice receiving Bars represent
It has, however,
action
5
to entrap membranes
for potential
Adr in liposomes
(5). This has made difficult
the high levels of Adr needed
These difficulties production
which
membrane
the produc-
for --in vivo thera-
have been overcome allows
due to
(9). Studies on the direct
have shown that the drug can increase solutes
mice.
by our double
5 to 10% of the starting
Adr
to be entrapped. Due to the strong membranes,
it is difficult
the slower
rate of leakage
somes does indicate loss of entrapped
that,
affinity
of Adr to serum proteins
to determine
actual
into the dialysing under physiological
efflux
and to cellulose
rate from liposomes.
plasma from both anionic conditions,
drug do not occur.
1299
of dialysis However,
and cationic
rapid lysis of Adr vesicles
lipoand
Vol. 91, No. 4, 1979
BIOCHEMICAL
AND BIOPHYSICAL
The IcSO value of 5x10m8M for anionic ished level of cell killing However
when compared
these --in vitro
or its entrapment
liposomes
The --in vivo distribution the proportion
of injected
to the liver
of phospholipid
vesicles
toxic
with Adr therapy
free drug.
and a greater support
immediately
Mice in the liposome-Adr final
weight-gain
heart
of this drug.
in anionic
liposomes,
and kidneys
is significantly
since renal and cardic
The increased
during
proportion
form reflects
following
that liposomes
The successful
entrapment
of Adr de-
the active
form produces
each injection
when compared
group also show a more rapid recovery
exert
a protective
action
removal
system (11).
at the end of the ten week experiment.
the concept
toxicities
the course of Adr treatment
of Adr in the liposome
choline
action
cells of reticuloendothelial
recorded
The administration
severe loss of body weight
the cytotoxic
in the liposome
weight
dimin-
of Adr to phosphatidyl
when entrapped
noting (10).
by the phagocytic
in animal
effects.
that,
This is worth
when administered
The changes overall
do not prevent
Adr taken up by the brain,
have both been associated livered
that the binding
data indicate
less than that for the free drug.
does show a slightly
to that for the free drug (1~5~ of 1.9x10e8M).
tests demonstrate
in anionic
Adr liposomes
RESEARCH COMMUNICATIONS
against
reflect a less to the
of body weight
These observations the toxic
effects
of Adr.
presented useful their
of Adr in liposomes,
here suggest that these complexes
for reducing potential
Adr cardiotoxicity.
cytotoxicity
against
the --in vitro
should be further
These complexes Adr-resistant
tumor
and --in vivo studies
investigated
may also be useful
as potentially to evaluate
cell lines (12).
ACKNOWLEDGEMENT We appreciate the skillful technical help of Ms. Mila Suva. This work was supported by Am. Heart Association, Grant No. 613 and by NIH, Grant No. CA-21271.
REFERENCES 1. 2. 3. . 4.
Kimelberg, H.K., and Mayhew, E.G. (1978) CRC Crit. Rev. Tox, , 25-79. Papahadjopoulos, D. (ed.) (1978) Ann, N.Y. Acad. Sci., 308. Tyrrell, D.A., Heath, T.D., Colley, CM. and Ryman, B.E.-(1976) Biochim. Biophys. Acta., -457, 259-302. Tritton, T.R., Murphree, S.A., an d Sartorelli, AC. (1978) Biochem, Biophys. Res. Comm., 84, 802-808.
1300
Vol. 91, No. 4, 1979
5. 6. 7. 8. 9. 10. 11. 12.
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Tritton,
T.R., Murphree, S.A., and Ssrtorelli, A.C. (1977) Biochem. Pharm., -26, 2319-2323. Forssen, E.A., Kharasch, N., and T&b, Z.A., (1979) Amer. Assoc. Cancer Res. (Proceedings), 20, 188. Papahadjopoulos, D., %ste, G., Vail, W.J., and Biedler, J.L. (1976) Cancer Res., 36,2988-2994. RahmayY.E., Hanson, W.R., Barucha, J., Ainsworth, E.J., and Jaroslow, EN., (1978) Ann. New York Acad. Sci., 308, 325-342. Schioppocassi, G. and Schwartz, H.S., (197TRes. Comm. Chem. Path. and Pharm., 18, 519-531. Burke,xF., Laucius, J.F., Brodovsky, H.S. and Soriano, R.Z., (1977) Arch. Intern. Med., 137, 385-388. Ryman, B.E. et al (1978) Ann. New York Acad. Sci., 308, 281-307. Dana, K. (1976) Acta Path. et Micro. Scan., 256A, 6-80.
1301
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
BIOCHEMICAL
Pages 1295-1301
28, 1979
--IN VITRO
AND --IN VIVO STUDIES WITH ADRIAMYCIN Eric A. Forssen and Zolthn A. TSk&*
LIPOSOMES
University of Southern California, School of Pharmacy and School of Medicine, Department of Biochemistry Comprehensive Cancer Center, Los Angeles, California Received
October
26,1979
Summary: Liposome entrapped adriamycin retains its full cytotoxic potential when tested under in vitro conditions against murine leukemia L-1210 cells. --In vivo drug dizrmon studies indicate that, relative to the free drug, a lower proportion of adriamycin administered in the liposome form is delivered to the heart and kidneys at one and four hours after injection. When administered to normal mice in high doses, anionic adriamycin liposomes appear less harmful than equal doses of the free drug as judged by alterations in animal weight gain. In these studies, a nova1 double packing procedure has been used for the entrapment of adriamycin in phospholipid vesicles. INTRODUCTION A number mycin-D
of chemotherapeutic
have been encapsulated
for overcoming toxicities. primarily
to entrap
of the vesicles. phospholipid
membranes
us to report
resulting
developed
MATERIALS choline
reviewed
compounds
in our laboratory activity
(1,2,3).
and actino-
have been useful and reducing
Liposomes
drug
have been used
into the aqueous compartment
materials
minimizes
(Adr) can destabilize
(4,5).
this disruptive
of liposome-Adr
--in vivo distribution
Ara-C
such as adriamycin
in loss of entrapped
on the --in vitro
reported
These preparations,
drugs by incorporation
ampholiphilic
methotrexate,
--in vivo tissue distribution
has been extensively
cells as well as their altered have been previously
altering
hydrophilic
However
The approach enabling
in liposomes.
drug resistances,
This subject
agents including
complexes
and toxicity.
A portion
effect against
thus L-1210
of these findings
(6).
AND METHODS
Adriamycin (provided by Adria Laboratories, Columbus, Oh.) is bound to phosphatidyl(PC) (Sigma Chem. Co, St. Louis, MO.) by adding a solution of the drug in normal
*Address: USC Comprehensive Cancer Center, Cancer Research Labs. Rm. 208, 1303 N. Mission Road, Los Angeles, CA 90033. Abbreviations used: Adriamycin: Adr; Cholesterol: Ch; Phosphatidyl Choline: PC; Arabinosylcytosine: Ara-C. 0006-291X/79/241295-07$01.00/0 12%
Copyright All rights
@ 1979 by Academic Press, Inc. ofreproduction in anyform reserved.
Vol. 91, No. 4, ~1979
BIOCHEMICAL
AND BIOPHYSICAL
RESEARCH COMMUNlCATlONS
saline to a two-fold molor excess of the dried lipid. This material is then sonicated for 5 min per ml under a nitrogen atmosphere using a needle probe type sonicator (Braun Sonic-1410) set at 100 watts. The resulting drug-lipid mixture is then entrapped within anionic liposomes consisting of a 7:3:1 molar ratio of PC, cholesterol (Ch) and phosphatidyl serine (Sigma Chem. Co.) or cationic liposomes containing PC, Ch and stearylamine (Eastman Org. Chem. Rochester, N.Y.) 7:3:0.5. This liposome suspension is sonicated as above at 100 watts for 5 min per ml. Unentrapped Adr is separated from liposomes by gel filtration on Sephadex G-50. Using this procedure, between five and ten percent of the starting Adr can be entrapped. The stability of Adr-liposomes was determined by incubating the liposomes in plasma at 37’C with continuous agitation. Liposomes (0.2 ml) containing [ H] - Adr (Morave Biochemicals, City of Industry, CA. Specific activity: 600mCi/mmole) were added to a dialysis bag (m.w. cutoff = 3500), (Spectrum-Medical Ind., Los Angeles, Ca.) containing 2 ml human plasma. The dialysis bag was placed into a cylinder containing 50 ml of the same plasma. Samples of the dialysate were removed and counted by liquid scintillation. Leakage was expressed as a percentage of the original radioactivity. Anionic liposomes containing [ HI-Adr were utilized for --in vivo distribution studies. Swiss mice (Simonson Labs; Gilroy, Ca.) weighing about 20 grams each, received [ HIAdr in either free or entrapped form by tail vein injections. Mice were then sacrificed at one and four hours following drug administration. The brain, heart, kidneys, liver, lungs and spleen were removed and prepared for liquid scintillation counting using tissue solubilizer (Protosol, New England Nuclear; Boston, Ma.). The distribution of [ HI-Adr to each organ was expressed as a percentage of the total radioactivity measured for all organs examined. Murine leukemia L-1210 Moore cells, adapted to cell culture, were grown in suspension at 37’C in RPMI-1640 media supplemented with 10% fetal calf serum (Grand Island Biological, Co., Grand Island, N.Y.), cells were treated with Adr in either the free or entrapped form at concentrations ranging from 10 to 10 molar. Liposome treated cells received Adr in anionic vesicles, prepared as described above. For controls free Adr was added to cells alone or with empty anionic liposomes containing the same amount Other controls received either empty anionic liposomes of phospholipid and cholesterol. without Adr or no treatment. The Adr concentration required for 50% inhibition of cell growth (Ic ) was determined for each group of L-1210 cells tested. All samples were done in duplicate. Swiss mice, weighing 20 grams each, were administered two intravenous Adr doses (lOmg/kg) with a one week interval in a high dose toxicity study. One group was injected with the free drug while a second was given Adr entrapped in anionic liposomes. Controls received normal saline. Animal body-weights were determined daily for the first two weeks and twice weekly thereafter. RESULTS The stability --in vitro.
of both cationic
While almost
hours of incubation, 1% in cationic
liposomes
in plasma
30% of free Adr escaped into the dialysate
less than 10% of entraped
liposomes
these values increased
and anionic
Adr in anionic
was found in the dialysate,
Figure
to about 20% and 5% for anionic
was demonstrated
during
liposomes
the first two and less than
1. At the end of ten hours,
and cationic
liposomes
respectively.
The 1~5~ for Ll210 cells treated with free Adr, either with or without empty liposomes was 1.9x10 -8 molar. Anionic liposome-Adr produced only a slightly lower level
1296
Vol. 91, No. 4, 1979
BIOCHEMICAL
AND BIOPHYSICAL
RESEARCH COMMUNICATIONS
50
FREE
0
1
2
Figure 1: The stability at 37OC.
those cells treated
with
anionic
The relative after
injection
anionic
liposomes
liposomes
display
organ distribution
liposomes
deliver
a lower
Much of the liposomal
as evidenced
by the enhanced
decrease
free or liposome between
an average
receiving
weight
Adr alone.
group during
no inherent
proportion
form,
was seen for
that over the concentration
toxicity. Adr at one and four hours
these studies demonstrate
of the administered
uptake.
is shown on Figure Over a period
that
dose to brain, heart
At four hours following
of Adr to cardiac,
gain of 56
Also notable
by dialysis
Adr is taken up by the reticuloendothelial
the two groups is revealed.
showed
a
7
of cell growth
alone, indicating
system
injection
an even
renal and brain tissue can be noted.
in the changes of net body-weight
entrapped
LIPOSOMES
No inhibition
2. Most notably,
hepatic
in the distribution
A comparison
CATIONIC 6
of free and liposomal
in Figure
and kidneys.
greater
LIPOSOMES
of liposomes in human plasma as determined
anionic
is illustrated
ANIONIC
4 5 OF INCUBATION
with an IcSO of 5x10 -8 molar.
of cytotoxicity
range tested,
3 HOURS
ADRIAMYCIN
for mice receiving 3. A significant
Adr in either
difference
of ten weeks the liposome-Adr
10% compared
is the less pronounced
to only 18 weight
(p
0.001) group
14% seen in the group loss for the liposome
the first two weeks of the study.
DISCUSSION Liposomes actinomycin-D
have been used to increase resistance
(7). Alternatively,
tumor cell drug uptake liposomes
1297
and to overcome
may also function
as protective
Vol.
91,
No,
4, 1979
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
90 60 70
One
60 __
50
-
T
25
Hour
After
Injection
m
Free
A&
m
Liposome
Adr
20
15
BRAIN
B Four
0
BRAIN
Figure
vehicles potentially therapeutic
2.
by altering toxic agents
KIDNEY
HEART
Relative organ distribution Adr at one and four hours normal
substances is Adr
patterns into which
distribution tissues
has pronounced
1298
(8). cardiac
After
Injection
LUNG
of free and aniomic after injections.
of drug sensitive
LIVER
Hours
and One and
liposome
entrapped
decreasing of the
commonly
renal
toxicities.
the
uptake
of
used
chemo-
Therefore
BIOCHEMICAL
Vol. 91, No. 4, 1979
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
160
150
80
Figure
IL
2
3 WEEKS
3: Body-weight charts for trapped Adr, 10 mg/kg.
it is of interest
decreases
1
to study
in toxicity.
the disruptive
the
4 FOLLOWING
which
liposome
between
Adr and liposomes
permiability
causing
a loss of entrapped
peutic packing
vesicles
and toxicological procedure
containing
studies.
for liposome
7 INJECTION
8
9
10
either free or liposome enstandard deviations from five
mode of Adr administration
entrapped
been difficult
it exerts on phospholipid
interactions
tion of phospholipid
6 FIRST
mice receiving Bars represent
It has, however,
action
5
to entrap membranes
for potential
Adr in liposomes
(5). This has made difficult
the high levels of Adr needed
These difficulties production
which
membrane
the produc-
for --in vivo thera-
have been overcome allows
due to
(9). Studies on the direct
have shown that the drug can increase solutes
mice.
by our double
5 to 10% of the starting
Adr
to be entrapped. Due to the strong membranes,
it is difficult
the slower
rate of leakage
somes does indicate loss of entrapped
that,
affinity
of Adr to serum proteins
to determine
actual
into the dialysing under physiological
efflux
and to cellulose
rate from liposomes.
plasma from both anionic conditions,
drug do not occur.
1299
of dialysis However,
and cationic
rapid lysis of Adr vesicles
lipoand
Vol. 91, No. 4, 1979
BIOCHEMICAL
AND BIOPHYSICAL
The IcSO value of 5x10m8M for anionic ished level of cell killing However
when compared
these --in vitro
or its entrapment
liposomes
The --in vivo distribution the proportion
of injected
to the liver
of phospholipid
vesicles
toxic
with Adr therapy
free drug.
and a greater support
immediately
Mice in the liposome-Adr final
weight-gain
heart
of this drug.
in anionic
liposomes,
and kidneys
is significantly
since renal and cardic
The increased
during
proportion
form reflects
following
that liposomes
The successful
entrapment
of Adr de-
the active
form produces
each injection
when compared
group also show a more rapid recovery
exert
a protective
action
removal
system (11).
at the end of the ten week experiment.
the concept
toxicities
the course of Adr treatment
of Adr in the liposome
choline
action
cells of reticuloendothelial
recorded
The administration
severe loss of body weight
the cytotoxic
in the liposome
weight
dimin-
of Adr to phosphatidyl
when entrapped
noting (10).
by the phagocytic
in animal
effects.
that,
This is worth
when administered
The changes overall
do not prevent
Adr taken up by the brain,
have both been associated livered
that the binding
data indicate
less than that for the free drug.
does show a slightly
to that for the free drug (1~5~ of 1.9x10e8M).
tests demonstrate
in anionic
Adr liposomes
RESEARCH COMMUNICATIONS
against
reflect a less to the
of body weight
These observations the toxic
effects
of Adr.
presented useful their
of Adr in liposomes,
here suggest that these complexes
for reducing potential
Adr cardiotoxicity.
cytotoxicity
against
the --in vitro
should be further
These complexes Adr-resistant
tumor
and --in vivo studies
investigated
may also be useful
as potentially to evaluate
cell lines (12).
ACKNOWLEDGEMENT We appreciate the skillful technical help of Ms. Mila Suva. This work was supported by Am. Heart Association, Grant No. 613 and by NIH, Grant No. CA-21271.
REFERENCES 1. 2. 3. . 4.
Kimelberg, H.K., and Mayhew, E.G. (1978) CRC Crit. Rev. Tox, , 25-79. Papahadjopoulos, D. (ed.) (1978) Ann, N.Y. Acad. Sci., 308. Tyrrell, D.A., Heath, T.D., Colley, CM. and Ryman, B.E.-(1976) Biochim. Biophys. Acta., -457, 259-302. Tritton, T.R., Murphree, S.A., an d Sartorelli, AC. (1978) Biochem, Biophys. Res. Comm., 84, 802-808.
1300
Vol. 91, No. 4, 1979
5. 6. 7. 8. 9. 10. 11. 12.
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Tritton,
T.R., Murphree, S.A., and Ssrtorelli, A.C. (1977) Biochem. Pharm., -26, 2319-2323. Forssen, E.A., Kharasch, N., and T&b, Z.A., (1979) Amer. Assoc. Cancer Res. (Proceedings), 20, 188. Papahadjopoulos, D., %ste, G., Vail, W.J., and Biedler, J.L. (1976) Cancer Res., 36,2988-2994. RahmayY.E., Hanson, W.R., Barucha, J., Ainsworth, E.J., and Jaroslow, EN., (1978) Ann. New York Acad. Sci., 308, 325-342. Schioppocassi, G. and Schwartz, H.S., (197TRes. Comm. Chem. Path. and Pharm., 18, 519-531. Burke,xF., Laucius, J.F., Brodovsky, H.S. and Soriano, R.Z., (1977) Arch. Intern. Med., 137, 385-388. Ryman, B.E. et al (1978) Ann. New York Acad. Sci., 308, 281-307. Dana, K. (1976) Acta Path. et Micro. Scan., 256A, 6-80.
1301
