Note: Descriptions are shown in the official language in which they were submitted.
WO90t1407~PCT/US90/02685
~'METHOD OF TREATMENT OF HEPATITIS 2 0 ~ ~ ~ 3
FIELD OF THE INVENTION
The present invention relates to a method for
treating hepatitis B.
5BACKGROUND OF THE INVENTION
Chronic infection which the hepatitis B virus
(HBV) affects approximately 5~ of the world's population.
Chronic carriers o~ hepatitis B are at a increased risk of
morbidity and mortality due to chronic liver disease, and
a proportion of these will ultimately develop cirrhosis
and/or hepatocellular carcinoma. At present, there is no
therapy of proven benefit for chronic hepatitis B.
Although ~-interferon has shown yreat promise in a subset
of patients treated for prolonged perlod of time, the
response rates overall have, unfortunately, been disap-
po~ntlngly low.
rrhe human hepatiki~ B virus ia a member o~ a
~amily o~ viru~ known a~ hepadnavlru~e~. O~h~r virus~s
ln this ~amily ar~ thc woodchuck hepat~ti~ vlru~, the
ground squirrel h~patitl~ v.lru~, ~nd the duc}; h~pat1ti~
virus. 'rhese animal viruses hav@ been invaluable models
~or characterization of hepadnaviruses and delineation o~
their unu~ual replicative cycle. These viruses replic~te
asymmetrically through an RNA template which re~uires
reverse transcriptase activity, cf. Summers, Cell 29:403-
415, 1982.
The 2', 3'-dideoxynucleosides are nucleosides
which recently have been shown to have potent antiviral ~- -
activity against the reverse transcriptase activity of the
30 human immunodeficiency virus, HIV, as described by Mitsuya, -
et al. in Proc. Natl. Acad. Sci. USA 1986; 83:1911-1915.
The most potent of these analogues is 2', 3'-dideoxycy-
tidine, or DDC, which inhibits HIV in cell culture in
concentrations as low as 10 nM.
SUMMARY OF THE INVENTION
It is an object o~ t~e present invention -to
overcome the above-mentioned deficiencies ln the prlor ~rt.
WO90~14n79 PCT/US90/02685
~5~3~ 2 - ~
It is another object of the present invention to
provide methods for treating hepatitis B.
It i5 further object of the present invention to
provide compositions for tre ting hepatitis B.
According to the present invention, hepatitis B
can be treated by administering 2', 3'-dideoxycytidine to
a patient infected with hepatitis B. The 2', 3'-
dideoxycytidine, following anabolic phosphorylation,
inhib~ts the reverse transcriptase of the hepatitis B
virus.
While the exact mechanisms of the antiviral
activity of the compositions according to the present
invention are unknown, it is believed that the mechanism of
action of DDC is inhibition oP viral polymerases, in
particular, reverse transcriptases. DDC is a nucleoside
analogue, and i~ appears to prev~nt the ~ormation oE normal
phosphodiester linkages once ~t become~ .incorpora-ted .LntQ
A growing DN~ chain 'rhl~ proco.3 leads to "ch~ tenmin~
tion." DDC ha~ a high aP~lnity ~or rQverse transcriptase,
and, there~ore, may inhibit replication of hepatitis B
virus by preventing reverse transcription from the pregeno-
mic RNA template. This inter~erence in replication would
lead to a decrease in serum levels of virus and a gradual
fall in the amounts of hepatitis B virus DNA in the liver.
DDC is particularly attractive as antiviral agent
because it is absorbed orally and has comparatively minimal
side effects under the conditions used.
BRIEF DESCRIPTION OF THE DRAWINGS
Figure l shows the ln vitro effects of 2', 3'-
dideoxycytidine triphosphate on the DNA polymerase reaction
of the human and duck hepatitis B viruses.
Figure 2 shows changes in mean serum DNA
polymerase activity among Peking ducks chronically in~ected
with duck hepatitis B virus who received ~DC or no
treatment.
Figure 3 shows changes in mean serum duck
hepatitis B virus DNA levels among Peking ducks chronically
- , : . , : , : : .~ - :: . : , , . : , . .
,.: :................ . . : . : ' ' .
Wogo/14079 PCT/US90/0268;
~ 2~a~33
infected with duck hepatitis B virus who received either
DDC or no treatment.
Figure 4 shows liver duck hepatitis B virus DNA
levels in ducks before and after treatment with DDC.
Figure 5 shows autoradiograms of duck hepatitis
B virus DNA analyses done on liver tissue taken before and
on the sixth day of treatment with DDC.
2', 3' dideoxycytidine can be used for treating
hepatitis B in patients 90 infected. The DDC i5 well
absorbed orally, and is generally well tolerated. In
humans, the dose-limiting toxicity has been a peripheral
neuropathy which can be significantly reduced by lowering
the dose.
~n ~ DDC triphosphate had little e~ect on
DNA polymerase activit~ o~ clther duck hapatlti~ B vlru~ or
human hepatlti~ ~ viru~. ~rev.lous ras~arch~r~ hav~ u~d
~he ~n ~i~ a~say ~to ~a~s antlvlral ~atlvlt~ in
hepatitis B, c~. Nord@n~elt, et al., ~ ~h- ~l~iQ}~
Scand. Sect. _ 87:75-76, 1979; and He~s, et al., Antimic.
Aaents Chemo. lg~ 50, 19~ lowever, it has now been
di~covered that khis assessment may be unreliable as ~
means of screening antiviral agents. The ~NA polymerase
activity measured in serum from humans and ducks infected
with hepadnaviruses may represent only one of the viral
enzymes necessary for replication, and this activity may be
relatively resistant to inhibition.
In c~ntrast, in ducks chronically infected with
duck hepatitis B virus, DDC exhibited potent antiviral
activity when given for six days in doses similar to those
used in human antiviral trials, cf. Yarchoan, et al.,
Lancet 1:76-81, 1988. The de~rees of inhibition of both
DNA polymerase activity and duck hepatitis B DNA were
similar (67~ and 69~, respectively and were comparab~e to
the degrees o~ inhib.ition of ~he~e ~ar~ers reparted in
s~udies of other an~iviral agents use~ in trea~ment of
chronic hepatitis B. The antiviral effect was only part-
ial, however, in that no duck became completely negative
~090/l4079 PCT/VS90/02685
?~5~13-~ 4 _ ~
for duck hepatitis B DNA or DNA polymerase activity, and
levels of these viral markers began to rise soon after the
DDC therapy was stopped. These findings are si~ilar to
those reported with other antiviral agents used in chronic
hepatitis B. A promising finding following DDC administra-
tion, however, was that some inhibition of DNA polymerase
activity and duck hepatitis B DNA was still observed for as
long as twelve days after therapy was stopped. This
observation is contrary to findings with adenine arabino-
side and acyclovir, wherein ~ollowing withdrawal of theseagents, serum levels o~ duck hepatitis B virus often
rebound to above pretreatment levels (Hirota, et al.,
Hepatology 7:24-28, }987).
The effect of 2', 3'-dideoxycytidine was assessed
in 16 Pekiny ducks chronically in~ected with the duck
hepatitis ~ virus ~DHBV). Nine ducks were given DDC at the
rate o~ 11 mg/m2 intravqnou~ly evary ~ix hours, and ~v~n
ducks received no tre~tmcnt. Scrum D~IBV ~NA ~nd DN~
polymaras~ act.Lvity d~crea~d ln Qv~ry duek tr~at~d wl~h
DDC. The m~an lnhibition oE DNA polym~r~8Q ~nd D~IBV DNA on
the third day o~ treatment me~sured 6~ (p~0.01) and 73~
(p~0.01), respec~ively. The inhibition of DNA polymerase
persisted a~ter treatment was stopped, and four ducks
continued to show greater than 50~ inhibition twelve days
after stopping treatment. DHBV DNA, which was measured in
total cellular DNA extracted from liver biopsies obtained
before and on the last day of treatment with DDC, showed
an average inhibition of 96~ in three ducks treated with
DDC, but showed no decrease in the remaining five ducks.
DUCKS AND TREATMENT SCHEDULE
An initial group of twenty ducks chronically
infected with DHBV were supplemented later with twenty
ducklings who were obtained within a day of hatching and
inoculated intraperitoneally with 100 ~1 of serum pooled
~rom DHBV carrier ducks. The inoculum contain~d approx.i-
mately 1.3 x 10~ vlrions per ml. serum as estimated by slot
blot hybridization.
WO90/14079 2 ~ 5 a ~ 3 3
.
The ducks were maintained and screened at monthly
intervals for persistence of DHBV DNA and DNA polymerase
activity in serum. At approximately four months of age,
eighteen ducks with high levels of DNA polymerase activity
were selected for study.
Nine ducks with the highest levels of DNA polyme-
rase activity were administered DDC in a dose of 11 mg/m2
intravenously every six hours for 5iX days. Two ducks were
given adenine arabinoside monopho~phate ~Ara-AMP; vidara-
bine monophosphate: Parke, Davis, Detroit, Michigan) at arate o~ 400 and 1000 my/mZ intramuscularly twice daily for
six days. Seven ducks received no treatment. Blood was
drawn from a wing vein be~ore treatment, twice during
treatment (days 3 and 6), and twice thereafter (days 10 and
18). Liver biopsies were per~ormed under general anesthe-
sia before and on the last day o~ treatment. Tis~ue was
processed for light microscopy. Sections ~or D~lBV DNA
determination w~re ~rozen .immcdiat~ly and ~torcd at -70C
until re~u:ired.
~ c ~ v~
Serum DNA polymerase activity was determined by
measuring 3~- thymidine incorporation into puri~ied Dane
particles by the method of Feinberg, et al., ~nalYt~
Biochem. 132:6-13, 1983. The ln ~ e~ects o~ DDC as a
nucleotide analogue on DHBV and HBV were assessed using the
DNA polymerase reaction. A range of concentrations of DDC
triphosphate were incubated with purified Dane particles
for one hour at 37C, and the DNA polymerase reaction was
then performed~
DHBV DNA was analyzed by molecular hybridization
~sing a 3.0 kb, full-length DHBV DNA clone in cACYC184.
The DHBV DNA insert was freed from plasmid A49 by digestion
with EcoRl and agarose gel electrophoresis. The DHBV DNA
was radiolabelled with 32p using the random primer method of
Feinberg, et al., ~kl~-, to a speci~ic activity of 3 x la~
to 1 x 10 cpm/~g.
DHBV DNA was detected in serum ~nd liver tissue
by slot blot analysis. For analysis of DHBV DNA in serum,
W~0/14079 PCT/US90/02fi8;
2 ~ 5 3 ~ - 6 - ~
10~1 of serum was denatured with 1 ~l of 1 M NaOH for five
minutes. The mixture was then neutralized by adding 90 ~1
of 1 M ammonium acetate. For analysis o~ DHBV DNA in liver
biopsy specimens, approximately 100 mg of minced liver was
homogenized in 10 ml of ice cold 50 mm Tris, pH 8.5, 10 mM
EDTA and 1% SDS. The crude liver homogenate was digested
with proteinase K (200 ~g/mk) for two hours at 50~C. Total
cellular DNA was prepared by two extractions with a mixture
of phenol and chloroform (1:1) in Tris pH 7.5. DNA was
precipitated with ab~olute ethanol and diluted to a con-
centration of ~pproximately 2 mg o~ DNA/ml in TE buffer.
One hundred microliters of the DNA sample
prepared from serum or liver was spotted onto a nitrocellu-
lose filter premoistened with 1 M ammonium acetate using a
slot blot apparatus and vacuum manifold. The membr~ne was
air dried and baked in a v~cuum oven at 80'C ~or two hours
and hybr~dlzed at ~0C with the D~IBV ~NA prob~. ~h~
hybridizcd m~mbran~ w~re @xpo~ed to X-ray ~llm ~or ~
and 72 hour~, and the r~ultln~ a~ltoracllo~relm~ w~r~ ~ann~d
using Zcnith 9c~nnincJ ~en~l~om~t~r. Th@ amount Or ~ V
DNA was quanti~ied by camparing the autoradiographic
signals for each sample with those o~ known amounts of
cloned DHBV DNA dotted on the same filter diluted in normal
serum or normal duck liver homogenate.
Liver tissue DHBV DNA was also analyzed by
Southern hybridiization. ~en micrograms of total cellular
DNA was subjected to horizontal slab gel electrophoresis in
1% agarose and transferred to nitrocellulose paper by the
method of Southern, J. Mol. Biol. 98:503-517, 1975; as
modified by Wahl, et al., Proc. Natl. Acad. Sci. USA 76:
3683-3687, 1979. Hybridization and autoradiography were
carried out as described above.
PHARMACOKINE~TIC STUDY OF DDC LEVELS
Serial serum levels of DDC were monitored in one
duck after the initial dose o~ the drug was administered.
Blood was drawn before an IV bolus o~ ~C an~ ~0 minute~,
1, 2, 3, and 6 hours therea~ter. DDC was measured in sera
by high performance liquid chromatography.
.
Woso/14079 PCT/US90/0268S
- 7 -
20~3~
STATISTICAL ANALYSES
Data were compared using Student's test, the
Shapiro-Wilk test for normal distribution, and Spearman's
rank correlation coefficient. Mean and standard deviations
of serum DNA polymerase levels were calculated after
logarithmic transformation of the data. Changes in serum
and liver levels of these viral makers were expressed as
percent inhibition of the pretreatment levels.
IN VITRO EFFECTS OF DDC T~IP}IOSPHA~E ON DNA POLYMER~SE
~ t~o, DDC triphosphate had little e~fect on
the DNA dependent DNA polymerase activity of either HBV or
DHBV, as shown in Figure 1. There was no inhibition of
either viral DNA polymerase activity at concentrates below
10 ~M and less than 20% inhibition at 100 ~M DDC. At this
concentration, cellular DN~ polymerase activity is also
inhiblted by DDC.
Pl~sma lev~ls o~ DDC a~tor an XV bolu~ o-~ 2.5 my
(11 mg/m2) were ~6 ~M ~t tcn mi~utos ~nd loss that 1 ~M at
six hours (data not shown). The estimated peak level of
DDC was 60 mM and the estimate half~ e was approximately
thirty minutes.
IN VIVO EFFECTS OF DDC ON DUCKS CH~ONICALLY ~NFECTED
Antiviral therapy was tolerated well, and all
ducks survived therapy and liver biopsy. No duck showed
obvious evidence of drug toxicity.
Serum levels of DHBV DNA polymerase decreased in
all nine ducks given DDC, but in none of the controls, as
shown in Figure 2. The mean inhibition of DNA polymerase
activity measured on the third day of treatment was 64~.
The difference between the pretreatment and day 3 value was
statistically significant (p< 0.01). The inhibition of DNA
polymerase persisted after treatment was stopped, and four
o~ nine ducks treated with DDC continued to show greater
than 50~ inhibi~ion ~welv~ day~ a~ter stopping ~reatment
(mean inhibition on day 18 - 55~).
WO~0/l4079 PCT/US90/0268~
20~5~33 8 - ~ ~
serum levels of DHBV DNA also decreased in all
nine ducks during therapy, but in none of the controls, as
shown in the Table. The mean percentage inhibition of DHBV
DNA levels was 73~ of d~y 3 of treatment (p<0.01), as shown
in Figure 3. The inhibition of serum levels of this viral
marker persisted for at least twelve days after stopping
DDC therapy.
Tre~tes~ with 2 ' 3'-D~deoKy~tidine tDDC)_or
Adenine Arabinoside MonoPhosphate (Ara-AMPl
Duck Hepatitis B Virus DNA (pa/10 ul~
Time DDC Ara-AMP Control
(9) (2) ~7)
Pre 13.1 t 1.3~.3 and g.7 3.0 ~ .L.5
Day 3 .6 ~ 1.6*0.6 ~nd 8.6 3.9 ~ ~.6
Ds~y 6 ~ l.fi and ~.~ 3.0
Day 10 3.1 L 1.33.9 ~d 1.7 ~.~ t 1.~
Day 1~ 8.3 ~ 1.71.0 and 5.~ 2.9 ~ 1.5
Data expressed as geometric mean (~ relative standard
error).
* p<.01 compared to pre values by Student's paired t test.
Treatment of two ducks wlth Ara-AMP yielded
results similar to those reported by others, cf. Hirota, et
al., op. cit. DNA polymerase and DHBV DNA levels decreased
by 71~ and 100~ during therapy, as shown in the Table, but
levels of these viral markers rapidly rose to greater than
pretreatment values within four days of stopping the
intramuscular injections.
Pretreatment DNA polymerase levels correlated
with DHBV DNA levels in ducks treated with DDC ~p ~ 0.01).
In addition, successive chan~es in DNA polymeras~s levels
correslated with succesaive ahange~ in DN~ polymeras~ l~v~15
on days 3, 6, and 10 o~ treatmeSnt.
Results o~ measuring DHBV DNA by slot blot
analysis o~ total cellular DN~ extracted ~rom liver bio-
WO90/14079 PCT/US90/02685
~ 9 ~ 205~3
psies before and on the last day of treatment with DDC areshown in Figure 4. Three ducks showed a marked inhibition
of DHBV DNA after treatment with DDC taverage inhibition,
96~), three showed mild inhibition (mean, 7.7%), and two
ducks demonstrated a 60% increase. Southern blot analysis
of liver DHBV from before and on the last day of treatment
showed that the decrease in total DHBV DNA was attributable
to a global decrease in DHBV DNA replicative intermediates,
as shown in Figure 5.
In ~igure 5, on the lePt are slot blot analyses
of total D~BV DNA. On the right are Southern blot analyses
showing the molecular weight of cloned DHBV (approximately
3.0 kilobases [kb]) and the molecular weights of the
replicative intermediates o~ DHBV DNA found in liver before
(lanes 1 and 2) and a~ter treatment ~lanes 3 anc~ ~). DNA
in lanes 2 and ~ were digestod with EcoRl.
~ variety o~ hi~toloyLc lo~ions w~re oblervod by
light micro~copy in :Ln~ccted duck~. 'rhose ine~udQd m~cro~
vesicul~r stca~o~is and chron.Lc portal ln~llkrat~. Amony
DDC treated ducks ~h@re wer@ no Gorrela~lons noted betwQen
response to treatment and chanyes in histologic lesions.
DDC, or 2', 3,'- dideoxycytidine, comprises a
pyrimidine nuclcoside with the ribo~e moiety o~ the mole~
cule in the 2', 3'-dideoxy configuration, as illustrated
below:
~ N
OC~ ,
H~ H2 1 .~
~ ' ` '.
,
W~90/14~79 PCT/US90/02685
20~ 3~33 - lO - ~
The DDC may be in the form of carboxylic acid
esters in which the non-carbonyl moiety of the ester
grouping is selected from straight or branched chain alkyl,
alkoxyalkyl (e.g.,methoxymethyl), aralkyl (e.g., benzyl),
aryloxyalkyl ~e.g., phenoxymethyl), aryl (e.g., phenyl
optionally substituted by halogen, C1 4 alkyl or Cl,4 alkoxy);
sulfonate esters such as alkyl- or aralkylsul~onyl (e.g.,
methanesulfo~yl); and mono-, di~, and triphosphate esters.
The compounds as described ~bove also include
pharmaceutically acceptable salts thereo~. Unless other-
wise specified, any alkyl moiety present advantageously
contains ~rom 1 to 18 carbon atoms, particularly 1 to 4
carbon atoms. Any aryl moiety present in such esters
pre~erably comprises a phenyl group, including a substi-
tuted phenyl yroup.
Examples o~ pharmaceutically acceptablc ~a:lts andpharmaceutically ac~ptable derivatives o~ th~ aompound~
which can b@ u~e :In tr~atincJ hepatLtis B nocord.lnFJ to the
pre~ent invention .Lnclude b~e s~lt~ sueh a~ tho~@ d@rlved
~rom a base such as alkali metal ~soclium, l~thium, potas-
sium), alkaline earth metal (magnesium) salts, ammonium and
NXb where X is C14 alkyl. Physiologically acceptable salts
containing a hydrogen atom or any amino group include salts
of organic carboxylic acids such as acetic, lactic, tar-
taric, maleic, isothionic, lactobionic, and succinic acids;organic sulfonic acids such as methanesulfonic, ethanesul-
fonic, benzenesulfonic, and p-toluenesulfonic acid, and
inorganic acids such as hydrochloric, sulfuric, phosphoric,
and sulfamic acids. Physiologically acceptable salts of a
compound containing any hydroxy group include the anion of
said compound in combination with a suitable cation such as
Na , NHY4~, and HX4' (wherein X is C14 alkyl and X is halo-
gen).
Speci~ic examples of pharmaceutically acceptable
derivatives o~ the compound of ~ormula ~ that ~Ay be u~ed
in accordance wlth the present invention include the
monosodium salt and the following 5' esters: monophos-
phate, disodium monophosphate, diphosphate, triphosphate,
WO~o/14079 PCT/US90/02685
- 2~
acetate, 3-methylbutyratej octanoate, palmitate, 3-chloro
benzoate, 4-methylbenzoate, hydrogen succinate, pivalate,
and methylate.
Also included within the scope of this invention
5 are the pharmaceutically acceptable salts, esters, salts of
such esters, nitrile oxides, or any other covalently linked
or non-linked compound which, upon administration to the
recipient, is capable o~ providing, either directly or
indirectly, a nucleoside analogue as described above, or an
10 antivirally actlve metabolite or residue thereof. All o~
these compounds are active and relatively nontoxic at con
centrations of sufficient potency for effective inhibition
of viral infectivity and replication.
It is possible for the nucleoside of the present
15 invention to b~ administered alone in solution. ~lowever,
the ackive ingredient may be used or administer~d in a ?
pharmaceutlcal ~ormulation. These ~ormulation~ com-pri~e
the nucleos:Lde or derlvi3tlv~ thereo~ togethcr w:lkh on~ or
more pharmaceukically acceptable c:arri~r~ and/or oth~r
20 ther~peutic ag~nts. A~. lnclud~d wlthln th@ ~cop~ o~ the
present invention, "acceptable" is defined as being com-
patible with other ingredients of the formulation and not
injurlous to the patient or host cell.
The administration o~ DDC to treat hepatitis B
25 can be accomplished by a variety of means of administra-
tion. Whatever administrative method is chosen should
result in circulating levels of the DDC within a range of i
about 0.01 ~M to about 2.0 ~M. A range of approximately
0.05 to aboùt 0.5 mg/kg administered ever four hours is
30 considered to be a virustatic range in humans. In order to
achieve this, the preliminary dosage range for oral admin-
istration may be broader, being, for example, 0.001-0.50
mg/kg administered every four hours. It is recognized that
dosage modifications may be required in individual patients
35 to ameliorate or inhibit toxic side effects.
The pharmaceutical ~ormulations according to the
present invention may conveniently be administered in unit
dosage form and may be prepared by any methods known in the
woso/l4079 PCT/US90/02685
~ 12 -
pharmaceutical art. Determination of the effective amounts
to be included in the dosage forms within the skill of the
art.
ThP pharmaceutical compositions according to the
present invention may contain suitable pharmaceutically
acceptable carriers comprising excipients and auxiliaries
which facilitate processing of the DDC into preparations
which can be u~ed pharmaceutically. Preferably the prepa-
rations, par~icularly tho~e which can be administered
orally and which can be used for the preferred type of
administration, such as tablets, dragees, and capsules, and
also preparations which can be administered rectally, such
as suppositories, as well as suitable solutions for admin-
istration by injection or orally, contain Pro-tn ~bout O.l
to 99 percent, and pre~erably ~rom about 25-85 porcent, by
weight, o~ DDC, togethor with the ~2xcipienk.
The pharmaceutlcal preparations oP the p~ent
lnvontion arc manu~actured ;Ln a manner whleh 1.~ lt~el~
known, Por example, by -me~.~ns o~ aanvcn~lon~l mixing,
granulatiny, dragee-m~kin~J, dis~olving, or lyophilizing
processes. Thus, pharmaceutical preparations ~or oral use
can be obtained by combining the active compounds with
solid excipients, optically grinding a resulting ~ixture,
and processing the mixture of granules, after adding
suitable auxiliaries, if desired or necessary, to obtain
tablets or dragee cores.
Suitable excipients are, in particular, fillers
such as sugars, for example lactose or sucrose, mannitol or
sorbitol, cellulose preparations and/or calcium phosphates,
such as tricalcium phosphate or calcium hydrogen phosphate,
as well as binders such as starch paste using, for example,
maize starch, wheat starch, rice starch, potato starch, and
the like; gelatin, g~m tragacanth, methyl cellulose,
hydroxypropylmethylcellulose, sodium carboxymethylcel-
lulose, and/or polyvinyl pyrrolidone. I~ de~i~ed, disin-
tegrating agents may be added such as the above-mentioned
starches and carboxymethyl starch, cross-linked polyvinyl
pyrrolidone, agar, alginic acid or a salt thereo~ such a5
:` : .. ; . . . . .
! ~
WO90/14079 PCr/US90/02685
- 13 -
sodium alginate. Auxiliaries are, for example, flow-
regulating agents and lubricants, such as silica, talc,
stearic acid or salts thereof such as magnesium or calcium
stearate, and/or polyethylene glycol. Dragee cores are
provided with suitable coatings which, if desired, are
resistant to gastric juices. For this purpose, concentra-
ted sugar solutions may be used, which may optionally
contain gum arabic, talc, polyvinyl pyrrolidone, polyethy-
lene glycol, titanium dioxide, lacyuer solutions, and
suitable ory~nic solvents ur solvent mixtures. In order to
produce coakinys resistant to gastrlc juices, solutions of
suitable cellulos~ preparations such as acetyl-cellulose
phthalate or hydroxypropylmethylcellulose phthalate are
used. Dyestuffs or pigments may be added to the tablets
or dragee coatings, for example, ~or identl~ication or in
order to charact~rize di~Perent combinations o~ act~ve
compound doses.
Other pharm~c~u~.ical preparations wh:lch c~n b~
used orally includ~ pueh~ c~psu.L~ macle o~ cJ~lakln/ a~
well as 50~t, Bealed capsul~s m~c10 o~ ~J~latin and a pla~-
ticizer such as glycerol or sorbitol. The pu~h-~i~ cap-
sules can contain the active compounds in the form of
granules which may be mixed with ~illers as such as
lactose, binders such as starches, and/or lubricants such
as talc or magnesium stearate and, optionally, stabilizers.
In soft capsules, the active compounds are preferably
dissolved or suspended in suitable liquids such as fatty
oils, liquid paraffin, or liquid polyethylene glycols. In
addition, stabilizers may be added.
Possible pharmaceutical preparations which can be
used rectally include, for example, suppositories, which
consist of combinations of the active ingredient with a
suppository base. Suitable suppository bases include
natural or synthetic triglyderides, paraffin hydrocarbons,
polyethylene glycols or higher alkanols. In aqdition, it
is also possible to u~e gelatin rectal capsul~ which
consist o~ a combination o~ the active compounds with a
base. Possible base materials include, for example, liquid
~090/14079 PCT/US90tO2685
~ 3 - 14 -
triglycerides, polyethylene glycols, and paraffin hydrocar-
bons.
Suitable formulations for parenteral administra-
tion include aqueous solutions of the active compounds as
appropriate oil injection suspensions may be administered.
Suitable lypophilic solvents or vehicles include fatty
oils, for example, sesame oil, or synthetic fatty acid
esters, for example, ethyl oleate or triglycerides.
Aqueous injection suspensions may contain substances which
10 inarraa~e the vlscos.ity o~ the su~pension such as sodium
carboxymethyl cellulose, sorbitol, and/or dextran.
Optionally, the suspension may also contain stabilizers.
In the present invention, the hepatitis B may be
treated by directly delivering the triphosphate derivative
to the patient. It i5 well known that "unshielded" tri-
phosphates c~nnot be used as druys b~cau~e triphosphate
compound~ do not pen~trat~ cell mcmbranes. ~h~r~;~or~, th@
triphosphate der.tvat.ives Or th.ls inv~ntlon ma~ ba d~llvar~d
by m~an~ o~ lLpor,orn~s, ~mall partla.les ~abol1~t ~ M ko
~0 abou~ ~ ~M in diam~t~r) which can ~v~ as an ;lntrac@ll-1lar
transport system to d@l$ver normAlly non-absorb~ble drugs
across the cell membrane. Such use oP liposomes for drug
delivery is well known in the art, and is based upon the
ability o~ a phospholipid to ~orm bilayers spontaneously in
aqueous environments.
one methods of forming the liposomes is by
agitating phospholipids in aqueous suspensions at high
frequencies. This results in the formation of closed
vesicles characteristic of liposomes. Once inside the
cells, the triphosphate derivatives act to eliminate the
replication of the h~patitis B virus. Since the tri-
phosphate has been shown to be active inside the cells, and
to be the active form therein, the liposome is clearly a
method of choice for delivery of these drugs.
Formulations suitable for vaginal administration
may be in the ~orm o~ pessaries, kampons, creams, ~ls,
pastes, foams, or sp~ay ~ormulations containing, in addi~
~ .
V~90/l407~ PCT/US90/0268~
~ 2 ~5~ 3
tion to the active ingredient, such carriers as are known
in the art to be appropriate.
The formulations according to the present inven-
tion may be in unit-dose or multi-dose sealed containers,
such as ampoules and vials, and may be stored in a lyophi-
lized condition requiring only the addition of the sterile
liquid carrier for injections immediately prior to use.
Extemporaneous injection solutions and suspensions may be
prepared from sterile powders, granules, and tablets of khe
kind prèviously desGribed.
~ n treatlng hepatit~ 5 B according to the presen-t
invention, the medication is generally administered two to
six times a day. In order to improve oral bioavailability,
it is often preferable to add a common buffer such as
sodium acetate to a solution containing 2', 3'-dideoxycy-
tidine accordlny to the pre ent ~nvention.
The PorQgoing desarlption o~ the spQcl~ mbodi-
ments will so ~ully reveal the g~ner~l naturQ o~ the
invention that oth@rs c~n, by applyin~J aurr@nt knowlcdgQ~
readily modi~y And/or aélapt eOr v~rlous appllcal:larls ~uc:h
specific embodiments without departiny ~rom the generic
concept, and therefore such adaptations and modifications
are intended to be comprehended wikhin the meaning and
range of equivalents of the disclosed embodiments. It is
to be understood that the phraseology or terminology herein
is for the purpose of description and not of limitation.
........... ... .. , "
