United States Patent 4983637
Filing Period: Jan. 1, 1987 – Present
Application Number: 363628
Application Type: Invention (Utility) Patent
Application Filing Date: June 8, 1989
Title of Invention: Method for treating viral infection of HIV
Issue Date: January 8, 1991
Primary Examiner: Nutter; Nathan M.
Inventor: Herman; Stephen(9341 Hazel Cir., Villa Park, CA 92667).
References:
U.S. Patent Documents:
925590 (June, 1909; Neel)
1210949 (January, 1917; Knox)
1910564 (May, 1933; Rankin)
2083572 (June, 1937; McKee)
2243053 (May, 1941; Ramage)
2356062 (August, 1944; Johnson)
2750411 (June, 1956; Fisher et al.)
3360472 (December, 1967; Renold)
3504038 (March, 1970; Beal)
4451480 (May, 1984; De Villez)
4591602 (May, 1986; De Villez)
4632980 (December, 1986; Zee et al.)
Other References:
Russell-Manning, editor, “Self-Treatment for AIDS, Oxygen Therapies,
etc., ” Greensward Press, San Francisco, 1988, 1989, pp. 22-27.
P. Bailey et al., “Complexes and Radicals Produced During Oxonation of
Olefins”, Ozone Reactions with Organic Compounds, Advances in
Chemistry, Series 112, pp. 1-8 (1972).
Abstract
Methods of treating systemic viral infections are disclosed comprising
the parenteral administration of pharmacologically effective amounts
of ozonides of terpenes in pharmaceutically acceptable carriers.
Methods for treating viral lesions are also disclosed. In particular,
a method for treating rfetroviral infections is disclosed. More
particularly, a method for treating HIV infections is disclosed. In
addition, methods for treating infections of non-retroviral viruses
are disclosed. Further, methods for trating T-cell deficiencies are
also disclosed. Moreover, a method of producing blood for medical
products which is free of viral activity is disclosed.
This invention relates to methods of medical treatment. More
particularly, it relates to the use of ozonides of terpene
hydrocarbons in the treatment of viral infections and certain immune
disorders.
BACKGROUND OF THE INVENTION
Methods of medical treatment employing ozonides of oil-soluble
compounds are known in the being disclosed, for example, in U.S. Pat.
No. 925,590 to Neel, U.S. Pat. No. 2,083,572 to McKee, and U.S. Pat.
No. 4,451,480 to De Villez.
The prior art does not disclose the use of ozonized compounds as an
antiviral or immunotherapeutic agent. However, particular types of
ozonide structures have been disclosed to have certain pharmacological
activity In U.S. Pat. No. 925,590, Neel reports the use of ozonides of
terpenes and other ozonides for inhalation therapy, because it was
believed to have a therapeutic effect for consumption and asthma.
Although the Neel patent application was filed in 1902, there have
apparently been no supporting data reported in the intervening years
that corroborate the utility theorized by Neel.
Knox, U.S. Pat. No. 1,210,949 discloses use of ozonized castor oil as
a laxative. Ozonation of the oil was believed to reduce its toxicity
and create a germicidal effect.
Johnson, U.S. Pat. No. 2,356,062 discloses the use of ozonides of
glycerine trioleates for external application, because it was believed
that those particular triglycerides had a germicidal, fungicidal and
deodorizing effect.
De Villez, U.S. Pat. Nos. 4,451,480 and 4,591,602, discloses use of
ozonides or certain fatty acids, including olive oil, sesame oil,
jojoba oil, castor oil and peanut oil, for external use as
antimicrobial agents, particularly in the treatment of acne. It is
believed that at least some of these compounds cause unacceptable skin
irritation.
So far as can be determined, none of the medical uses of ozonides
described in the prior art have ever been commercialized. Presumably,
this lack of commercialization is due to unacceptable side-effects,
toxicity, difficulties in storage, or minimal effectiveness. Many of
these various compositions decompose on standing. Also, to the extent
that the mechanism of action of these compositions can be attributed
to their oxygen content, most of the ozonides known in the prior art
have been suboptimal because these compounds typically release no more
than about 18% of their weight as oxygen.
Methods of medical treatment employing antiviral compounds are known
in the art. Most of the research in this area has focused on
nucleoside analogues. Dideoxynucleosides are antiviral nucleoside
analogues which are useful in treating retroviral infections where
viral replication requires the transcription of viral RNA into DNA by
viral reverse transcriptase. Other nucleoside analogues include
deoxynucleosides and nucleoside analogues, such as acyclovir and
gancyclovir which have only a fragment of ribose or other pentose
connected to the base molecule. Nucleoside analogues have been shown
to be only minimally effective in the treatment of viral infections
that are not caused by retroviruses.
Antiviral agents other than nucleoside analogues are also known. For
example, amantadine is an antiviral agent that prevents binding of
certain viruses with their receptor on the cell surface. However,
amantadine is ineffective against many known viruses.
Acquired immunodeficiency syndrome (AIDS) is a fatal condition caused
by the human immunodeficiency virus (HIV), a retrovirus. Since AIDS
was identified as a medical condition in 1981, over 100,000 cases have
been reported worldwide, with over half of these cases in the United
States. It is believed that over 2,000,000 people worldwide are
carriers of the HIV virus, with infections continuing to spread.
Researchers now believe that most of these carriers will one day
develop symptoms of AIDS. No effective cure is available for AIDS,
although dideoxynucleosides and their analogues have been shown to
prolong life and to reduce the incidence of certain fatal infections
associated with AIDS. Among the dideoxynucleoside analogues, AZT has
shown the most promise as a treatment for AIDS. However, treatment of
AIDS patients with AZT has proven to be of only poor to moderate
effectiveness, and AZT does not cure AIDS. Moreover, in a recent human
trial, serious toxicity was noted, evidenced by anemia (24%) and
granulocytopenia (16%). Clearly, there is a tremendous need for a non-
toxic and effective treatment for HIV infection.
It is believed that HIV causes AIDS, in part, by infecting
helper/inducer T4-cells and causing a T4-cell deficiency. Other
conditions may also cause this deficiency, including immunosuppressive
therapy for transplant patients, radio-therapy or chemotherapy in
cancer patients, and congenital immunodeficiencies. Current immune
boosting therapies, such as the use of interleukin-2 or .gamma.-
interferon are still in the experimental stages, and have not yet been
proven effective. No proven effective treatments are currently in use
for restoring a normal level of T-cells. Thus, a need exists for such
a treatment.
Transmission of HIV through blood products has been shown to occur.
The discovery of the HIV antibody test, and its application to blood
products prior to release has reduced the incidence of transmission
through blood products. However, the HIV antibody test is not 100%
effective in detecting the presence of HIV virus particles, in part,
because an infected individual may not produce antibodies to HIV for
six months or longer after infection. There is, therefore, still a low
incidence of blood products tainted with HIV being released for
medical use. Moreover, blood products may be tainted with other
viruses capable of being transmitted through the blood, such as
Hepatitis B. A method of treating blood products to eliminate viral
activity without affecting their efficacy in treatments is highly
desireable.
Retroviruses other than HIV are known. These include the herpes family
of viruses, HTLV I, and cytomegalovirus (CMV). Infections of these
viruses have been notoriously difficult to treat. No vaccines are
known for these infections. Although acyclovir has been used in the
treatment of Herpes lesions, toxic side effects are known, and such
treatment is not always effective. Thus, a need exists for non-toxic
and effective treatments.
Human papilloma viruses are nonretroviral viruses responsible for
warts of the skin or mucous membranes. Common warts are found in as
many as 25% of some groups, and are most prevalent among children.
Moreover, the incidence of venereal warts (condylomata acuminata and
molluscum contageosum) has risen dramatically in the last few years,
to the point that this condition is one of the most common sexually
transmitted diseases in the United States. Common treatments for warts
are often painful and invasive, and involve physical removal of the
lesion through application of caustic agents, cryosurgery,
electrodessication, surgical excision, or ablation with laser.
Treatment with nucleoside analogues or interferon is also sometimes
used. However, no treatment of proven safety and efficacy is currently
available for warts Furthermore, at the present time, no effective
methods of prevention are available for warts other than avoiding
contact with infectious lesions. Therefore, a need exists for a method
of treatment and prevention of warts.
Other nonretroviral viruses are responsible for many of the known
infections in mammals. Vaccines are known for a minority of these
infections. Measles, rubella, polio, rabies, certain strains of
influenza, and mumps are examples of infections caused by viruses for
which vaccines are known. However, the existence of a vaccine does not
obviate the need for treatment of individuals already infected. Most
other viruses, including Epstein Barr Virus, and most of the
enteroviruses, reoviruses, rhabdoviruses other than rabies,
arboviruses, and arenaviruses produce infections for which no vaccines
are known. Currently used antiviral treatments for infections of these
viruses include application of nucleoside analogues or amantadine, and
various interferon treatments. Unfortunately, use of these treatments
is of minimal or no effectiveness against infections of most of these
viruses. The use of currently known antiviral compounds is, at best,
moderately effective. Moreover, toxic side-effects are common. Thus a
need exists for a wide-spectrum antiviral agent that is both non-toxic
and effective.
Virtually all humans occasionally suffer from upper respiratory
infections, such as colds and flu. The symptoms of these infections
include sore throat, runny nose, itchy eyes, and earache. In addition
to these discomforts, the infections are responsible for many days of
absence from work and contribute to a decrease in worker efficiency.
These infections are caused by a wide variety of viruses. Although
vaccines are known for a minority of flu strains, no effective methods
of prevention are known for most upper respiratory infections, and no
truly effective methods of treatment are known for any of these
infections. A method of treating these symptoms and underlying
infections would be of tremendous benefit.
Moreover, there are a number of ailments that may or may not be of
viral origin, for which no effective treatments are widely available.
Epstein-Barr virus (EBV) is the causative agent in infectious
mononucleosis, and has been implicated in chronic fatigue syndrome.
Many autoimmune disorders, such as systemic lupus erythematosis and
rheumatoid arthritis, may be associated with a virus. Whether or not
these diseases are of viral origin, however, they are debilitating
ailments for which an effective therapy would be of major importance.
Finally, there are a number of situations, both in research and in
medicine, in which generation of the superoxide radical, O.sub.2.sup.-
, is advantageous. Superoxide is commonly generated through the use of
xanthine oxidase acting on xanthine. However, these materials are
relatively expensive and are not particularly suited for many
utilities, including in vivo utilities. Thus, a method for generating
superoxide that is safe and inexpensive would be advantageous.
SUMMARY OF THE INVENTION
According to the invention, there are provided novel methods of
treatment and prevention of systemic and local viral infections.
Methods for treating nonretroviral and retroviral infections in
particular are also provided. More particularly, a method for treating
retroviral HIV infections is provided. Moreover, the invention
provides a method of treating blood products which removes any viral
activity present in the blood. Additionally, the invention provides a
method of treating immunosuppression characterized by T-cell
deficiencies.
The invention, in addition, provides pharmaceutical compositions for
use in the above novel treatments, containing ozonides of terpenes and
a pharmaceutically acceptable carrier, and may contain other active
ingredients. Preferably, these compositions are in dosage form
comprising a clinically effective amount of the active compound. In
one preferred embodiment of the invention, the pharmaceutical
composition is comprised of an ozonized terpene in a stable injectable
composition. In other preferred embodiments, the pharmaceutical
compositions are in the form of nosedrops or nasal sprays, inhalants,
throat sprays, eardrops, ophthalmic ointments or drops, vaginal or
rectal suppositories, or ointments or creams for topical applications.
Moreover, the present invention includes the use of terpene ozonides
and other ozonides of unsaturated hydrocarbons to treat autoimmune
disorders, and to produce superoxide radical upon combination with an
aqueous system.
DETAILED DESCRIPTION OF THE INVENTION
Terpene hydrocarbons are also known as isoprenoids, because they may
generally be constructed from isoprene units. Terpene hydrocarbons are
usually exact multiples of C.sub.5 H.sub.8. Terpenes are classified
according to the number of isoprene units of which they are composed,
as shown in Table 1.
TABLE 1
1 hemi- 5 ses- 2 mono- 6 tri- 3 sesqui- 8 tetra- 4 di- n poly-
While not limiting the scope of the invention, examples of terpenes
which may prove especially effective, when used in the method of the
preferred embodiment, include limonene, citronella, alpha-carotene,
beta-carotene, Vitamin A, geraniol, linalool, linalyl acetate, and
squalene. Other compounds which are believed to make pharmacologically
active terpene ozonides in accordance with the present invention
include limonene, alpha-pinene, loganin, cymene, farnesanes,
eudesmanes, acoranes, cedranes, chamigranes, caryophyllanes,
illudanes, humulenes, himachalenes, longifolanes, perhydroazulenes,
quaianes, quaianolides, and germacranes. Still other compounds which
are believed to make pharmacologically active terpene ozonides in
accordance with the present invention include labdanes, clerodanes,
abietic acid, phyllocladene, giberellins, ophiobolin A, retigeranic
acid, gasgardic acid, lanosterol, euphol, oleanane, ursane, lupeol,
hydroxyhopanone, lupanes, and hopanes. Other particular terpene
compounds which are believed to make pharmacologically active terpene
ozonides when prepared in accordance with the present invention
include B-selinene, zingibene, camphene, sabinene, ocimene, myrcene,
nerol, citral A, citral B, farnesol, bisabolene, phytol, and cecropia
hormone. Citral, geraniol, and nerol are particularly preferred
terpenes. Ozonides of terpenes have three oxygen atoms replacing the
double bonds at sites of unsaturation, creating a trioxyacyclopentane.
In the preparation of terpene ozonides, the particular desired terpene
starting material is first obtained. A large and representative number
of such terpenes are disclosed in the literature and/or are
commercially available. (Many terpenes are essential oils that have
been isolated from various parts of plants or wood by steam
distillation or extraction.)
In the ozonide synthesis, ozone is passed through the terpene under
conditions that provide for intimate contact between the terpene
starting material and the ozone, such as thin film procedures,
sparging, gas entrainment procedures, and the like. On a small scale,
for example, the terpene is placed in a vented vessel, and ozone is
sparged through the material until the reaction is complete. The ozone
may advantageously be generated with any of the commercially-available
ozone generators. Such devices include corona discharge tubes through
which oxygen gas may be passed. For example, pure oxygen gas passing
through an ozone generator will typically leave the device as from 2%
to 6% O.sub.3 (ozone), with the remainder O.sub.2. This ozone mixture
may then be sparged through the terpene at ambient temperature and
pressure until the reaction is complete. Completion may be judged by
analyzing the gas exiting the ozonation chamber for ozone. (This may
be done by passing the exit gas through aqueous potassium iodide and
determining whether iodine gas is liberated, or by any other
conventional technique.) Alternatively, the reaction may be followed
by observing the weight gain of the material undergoing the reaction,
by observing changes in physical characteristics (such as conversion
from a liquid form to a soft paste), or by simply calculating the
quantity of ozone needed to fully ozonate the material and stopping
the reaction when a slight excess of ozone has passed through the
reaction chamber. Because the reaction is exothermic, its progress may
also be followed by monitoring the heat evolved by the reaction
medium, and stopping the flow of ozone when the mixture ceases to
generate heat.
When the terpene is normally a solid, such as .beta.-carotene, it may
be solubilized in any suitable saturated nonaqueous solvent system
prior to ozonation. With all of the terpene ozonides, it is desirable
to exclude water, lower alcohols, nucleophilic peroxides, and proton
donors from the reaction mixture and from the final composition, in
order to prevent premature hydrolysis of the trioxolane ring.
Other suitable ozonation procedures may be used, such as the
procedures disclosed in U.S. Pat. Nos. 2,083,572, 3,504,038, and
4,451,480.
In certain preferred embodiments of the present invention, the terpene
ozonides are formulated into pharmaceutical preparations. These
pharmaceutical preparations include one or more terpene ozonides, and
may further include other pharmaceutically active ingredients. In
addition, any of the well-known pharmaceutically-acceptable carriers
or excipients may be combined with terpene ozonides in a well-known
manner. Suitable diluents include, for example, polyethylene glycol,
DMSO, isopropyl myristate, and mineral oil. Conventional coloring,
fragrance, and preserving agents may also be provided.
It is believed that the excellent weight to oxygen ratio of some of
the terpene ozonides renders them especially effective as antiviral
agents. Some of the terpene ozonides are capable of releasing large
amounts of oxygen, up to 30% of the weight of the compound. This is
because terpenes are highly unsaturated compounds. Ozonation of these
compounds results in the addition of three oxygen atoms at each site
of unsaturation. In addition, terpene ozonides appear to have
significant unexpected pharmacological properties that are different
in kind or quality from those of unrelated ozonides disclosed in the
prior art.
The toxicity of the terpene ozonides appears to be surprisingly low in
systemic use. Our preliminary data suggest that the LD.sub.50 for a
representative compound, linalool ozonide, appears to be greater than
about 5000 mg/kg in mice. Furthermore, we have discovered that the
irritability of the terpene ozonides is surprisingly low in skin and
eye tissues of the rabbit. It is believed that irritability of the
compounds in humans is also surprisingly low when used in accordance
with the methods of the preferred embodiments.
These ozonides can be used effectively in the generation of superoxide
radical, both in and vivo. When the ozonide is combined with an
aqueous system, gradual decomposition of the ozonide trioxolane ring
structure occurs, with release of superoxide radical Thus, the present
invention includes a method for generating superoxide by combining
these ozonides with a water-containing system. For example, superoxide
production is believed to occur when the compounds are administered to
an organism, as well as when the compounds are mixed (with or without
a surfactant) into a material that contains water. While the inventor
does not wish to be limited to any particular theory of operation, it
is believed that at least some of the beneficial and therapeutic
properties of these ozonides are due to superoxide generation.
We have also discovered that terpene ozonides, injected in suitable
pharmacological compositions, are effective for treatment of systemic
viral infections. The present invention includes systemic and
localized injection of terpene ozonides, including intravascular,
intramuscular, subcutaneous, intraperitoneal, and other injection
techniques. In addition, oral administration is also contemplated,
preferably in a capsule or other nonaqueous vehicle or system.
In the method of a preferred embodiment, pharmaceutical compositions
for systemic use such as for intravenous, intramuscular, or
intraperitoneal injection may contain from about 0.01% to about 99%
active ingredient, by weight. More preferred injectable compositions
contain from about 0.05% to about 45% active ingredient, by weight.
Moreover, pharmaceutical compositions for local application in the
form of nosedrops or nasal sprays, inhalants, throat sprays, eardrops,
ophthalmic ointments or drops, rectal or vaginal suppositories, or
ointments or creams for topical applications may contain from about
0.01% to 99.9% active ingredient, by weight. More preferred
compositions for local application contain from about 0.05% to 50%
active ingredient, by weight.Pharmaceutical compositions of preferred
embodiments may contain from about 0.1% to 99.99% carrier ingredients.
The carriers are preferably non-aqueous, because the presence of water
rapidly leads to the degradation of the pharmacologically active
ozonide compounds used in the preferred embodiment. The carriers
employed in pharmaceutical compositions for systemic use are, in
addition, preferably injectable or orally ingestible. Nonaqueous,
injectable carriers for pharmaceutical compositions of the preferred
embodiment for systemic application preferably include: isopropyl
myristate, polyethylene glycol or polypropylene glycol (in liquid
form), and DMSO, more preferably polyethylene glycol having a
molecular weight between about 150 and 1500, most preferably about
600. Good results have been realized, for example, by combining about
4 parts by weight of geraniol ozonide with about 3 parts by weight
polyethylene glycol (m.w. 600). This material is storage stable, and
can be formulated into an injectable material by combining one part
with three parts sterile saline immediately before use. (Although some
superoxide production appears to begin immediately upon such
combination, no significant degradation of the ozonide is believed to
occur within, say, 5 minutes before the material is injected.) Another
suitable vehicle is epal, comprising roughly equivalent parts of
substantially anhydrous tetradecanol and dodecanol. Non-aqueous
carriers suitable for pharmaceutical compositions for local
application in accordance with the methods of the preferred embodiment
include: DMSO, hydrogenated vegetable oil, mineral oil, carbomer 934,
glycerin, propylene glycol, propyl paraben, polysorbate 60, glyceryl
stearate, ethanol, and modified food starch.
Therapeutic dosages of the terpene ozonides when used for systemic
injection in accordance with the methods of the preferred embodiments
are preferably in the range of 1 mg to 10 g active ingredient for a 70
kg adult one time per day, more preferably in the range of 10 mg to 1
g active ingredient for a 70 kg adult one time per day, and most
preferably in the range of 20 mg to 500 mg active ingredient for a 70
kg adult one or two times per day. Therapeutic dosages of the terpene
ozonides when used for topical application in the form of creams,
ointments, or rectal or vaginal suppositories in accordance with the
methods of the preferred embodiments are preferably in the range of
100 g to 10 g used one to four times per day for each cm.sup.2 of
affected area, more preferably 1 mg to 1 g used one to four times per
day for each cm.sup.2 of affected area, and most preferably 5 mg to
200 g used one to four times per day for each cm.sup.2 of affected
area. Therapeutic dosages of the terpene ozonides for use in other
methods of local application in accordance with the preferred
embodiments, such as nosedrops or nasal sprays, inhalants, throat
sprays, eardrops, or ophthalmic ointments or drops are preferably in
the range of 100 g to 1 g per application used one to four times per
day, and more preferably 1 mg to 100 mg per application used one to
four times per day.
Oral compositions may be given at the same dosage as the injectable
compositions, or may be given at up to twice the injection dosage.We
have discovered that intramuscular injection of a terpene ozonide in a
pharmaceutically acceptable carrier, with or without contemporaneous
oral administration, is effective in treating the symptoms of AIDS. A
patient receiving this treatment gets fewer of the opportunistic
infections common in AIDS patients. Such a patient also feels less
lethargic and has a generally improved sense of physical well-being.
This improvement in symptoms has been shown to be the result of a
restoration of normal T4-cell levels after injection with the terpene
ozonide. The restoration is believed to be the result of an antiviral
effect of the terpene ozonide.
It is believed that treatment of persons infected with HIV who do not
yet express symptoms of AIDS can be effectively treated with systemic
injections of terpene ozonides, for example in the manner of Example
11, in order to prevent the appearance of the symptoms of AIDS.
Furthermore, it is believed that administration of terpene ozonides in
accordance with the present invention will be beneficial in the
treatment of immune disorders other than AIDS. It is believed that
systemic injection of the terpene ozonides will restore a normal level
of T4-cells in many immunocompromised patients, including patients on
immunosuppressive therapy, chemotherapy, or radio-therapy, and
patients with congenital immunodeficiencies. Lupus and rheumatoid
arthritis also respond to therapy with the terpene ozonides of the
present invention. In a preferred embodiment, restoration of T4-cell
levels is accomplished by systemic injection of terpene ozonides in
the manner of Example 11.
It is also believed that treatment of blood products with a terpene
ozonide of the present invention prior to its medical use, will
eliminate HIV and any other viral activity present in the blood. From
about 0.5 to about 10 mg/liter of terpene ozonide can be used in
treating blood in, for example, a blood bank.
The terpene ozonides seem to be effective not only against HIV
infection. They also appear to be effective in the treatment of other
retroviral infections, such as Herpes lesions, including chicken pox,
EBV infection, or CMV infection. Systemic injection of terpene
ozonides is believed to be effective in treatment of these other
retroviral infections. Additionally, topical application of the
terpene ozonides in pharmacologically effective compositions is
believed to be effective in the treatment of lesions of these
retroviral infections. Moreover, it is believed that the terpene
ozonides will be effective against many disparate viral infections,
including viral infections of non-retroviral origin. In this regard,
it is believed that systemic injection of a terpene ozonide in a
pharmacologically acceptable carrier or excipient is effective in the
treatment of systemic infections caused by non-retroviral viruses,
including Epstein Barr Virus, most of the enteroviruses, reoviruses,
rhabdoviruses (including rabies), arboviruses, and arenaviruses. It is
also believed that intra-vaginal application of a terpene ozonide in a
pharmaceutically acceptable carrier or excipient is effective against
condylomata acuminata, molluscum contagiousum, and other viral
infections of the vagina. Also in this regard, it is believed that
topical application of a terpene ozonide in pharmacologically
acceptable carrier or excipient is non-irritating and effective in the
treatment of common warts and other viral lesions of the skin. Further
in this regard, it is believed that application of a terpene ozonide
in a pharmacologically acceptable carrier or excipient in the form of
nosedrops or nasal sprays, inhalants, throat sprays, eardrops,
ophthalmic ointments or drops, is effective in the treatment of viral
infections of the eye, ear, nose, and throat, including upper
respiratory infections of viral origin such as colds and flu. Finally,
they appear to be useful in treatment of rheumatoid arthritis, which
may be caused by a viral pathogen, as well as useful in treatment of
other autoimmune disorders.
For example, in treating the common cold, an aerosol mist containing 2
ml of the nasal inhalant of Example 10 may be sprayed onto each
nostril of a patient suffering from the common cold. The process is
repeated every four hours. Within one hour of the first treatment, the
patient will generally report easier breathing through the nose. With
two days of treatments, the patient can usually breathe easily through
both nostrils and reports no sore throat.
EXAMPLE 1
Preparation of squalene ozonide
Squalene is ozonized by preparing a solution of 10 g squalene in 100
ml hexane. Ozone gas (4% in oxygen, from a corona discharge ozone
generator), is bubbled through this solution via a glass sparger at
the rate of 5000 cc/min. The reaction is exothermic, and the reaction
temperature is kept within the range of 0.degree. C. to 35.degree. C.,
preferably 20.degree. C. to 25.degree. C., and more preferably,
22.degree. C. to 24.degree. C., using a cool water bath. The resulting
product is the ozonide of beta carotene, and has a 98% weight gain
over squalene.
EXAMPLE 2
Preparation of linalool ozonide
The ozonide of linalool is prepared by bubbling ozone (4% in oxygen,
from a corona discharge ozone generator) through 100 ml neat linalool
via a glass sparger. The reaction is exothermic, and the reaction
temperature is kept within the range of 0.degree. C. to 35.degree. C.,
preferably 20.degree. C. to 25.degree. C., and more preferably,
22.degree. C. to 24.degree. C., using a cool water bath. The resulting
product is the ozonide of linalool, and has a 31% weight gain over
linalool.
EXAMPLE 3
Preparation of Geraniol Ozonide
The ozonide of linalyl acetate was prepared by bubbling ozone (4% in
oxygen, from a corona discharge ozone generator) through 5 ml neat
geraniol at the rate of 5000 cc/min. The reaction mixture was cooled
in a water bath, and after 20 minutes, the evolution of heat ceased,
indicating completion of the ozonation process. The resulting material
had no odor, and was soluble in polyethylene glycol (600 m.w.),
isopropyl myristate, and mineral oil.
EXAMPLE 4
Primary skin irritation test of ozonide of linalool
Six healthy New Zealand White rabbits were tested for skin irritation.
Approximately four hours prior to application of the ozonide sample,
the backs of the animals were clipped free of fur. Each rabbit
received epidermal abrasions with a sterile needle at one test site
while the skin at another test site remained intact. A 1.0% solution
of linalool ozonide in isopropyl myristate was prepared A 0.5 ml
portion of the test solution was applied to each site by introduction
under a double gauze layer to an area of skin approximately 1″ square.
The patches were covered with a nonreactive tape and the entire test
site was wrapped with a binder. After 24 hours, the binders, tape,
test material residue was removed with 70% isopropyl alcohol. An
evaluation was also made at 72 hours after application. The reactions
were scored according to the methods described in the Federal
Hazardous Substances Act. The test solution had a Primary Irritation
Index (PII) of 1.0. According to FHSA regulations, a material with a
PII of less than 5.00 is generally not considered a primary irritant
to the skin.
EXAMPLE 5
Ocular irritation test in the rabbit of the ozonide of linalool
Six healthy New Zealand White rabbits were selected for study. The
rabbits’ eyes were judged free of irritation prior to the study by
examining with a pen light and under UV light after installation of 2%
fluorescein stain. A 1% solution of the ozonide of linalool was
prepared in isopropyl myristate. A 0.1 ml portion of this test
solution was instilled into the lower conjunctival sac of one eye of
each rabbit. The lids were held closed for one second. The opposite
eye of each rabbit received 0.1 ml of the isopropyl myristate, as
control. At 24, 48, and 72 hours post dosing, the eyes were examined
with a pen light and re-examined with UV light following fluorescein
staining of the cornea. Under the conditions of this test, the test
solution was considered a non-irritant to ocular tissue of the rabbit.
EXAMPLE 6
An injectable composition for use in treatment of AIDS
250 mg/ml ozonide of geraniol from Example 3 balance oil/water
emulsion (soybean) with 0.1% lecithin
EXAMPLE 7
A vaginal suppository for treatment of condylomata acuminata
2% w/v Ozonide of geraniol from Example 3 Balance Hydrogenated
vegetable oil base
EXAMPLE 8
1% w/v Ozonide of linalool 60% w/v Carbomer 934 1% w/v Disodium
edetate 10% w/v Glycerin Balance propylene
glycol, 600 m.w.
EXAMPLE 9
A topical cream effective against chicken pox, herpes simplex and
other viral lesions
2.5% w/v Ozonide of linalool 48% w/v Propylene glycol 30% w/v Propyl
paraben 5% w/v Polysorbate 60 10% w/v Glyceryl monostearate Balance
Mineral oil
EXAMPLE 10
A nasal inhalant effective against upper respiratory infections
1 mg/ml ozonide of citral balance epal
EXAMPLE 11
Test of restoration of immune cell levels in an AIDS patient
A patient testing positive for the presence of HIV antibodies and
diagnosed with AIDS was variously treated with the composition of
Example 6 for a period of 99 days. On days 0 through 6, the patient
received daily intravenous injections of 4.0 ml of the composition of
Example 6. On days 7 to 19, the patient gas treated a.q.i.v. with the
same composition. From days 20 through 44, the patient received no
treatment. The patient received daily intramuscular injections from
days 45 through 77. An immunodeficiency screening was performed on
days 7, 20, 45, and 78. The results, expresses in cells/cmm, are shown
in Table 2.
The results show that intra-venous injection of the composition of
Example 6 increased the levels of all types of cells screened. These
cell levels decreased during the period of no treatment, and remained
relatively stable during the period of intra-muscular treatment.
It is believed that intra-venous systemic injection in the manner
described in Example 11 is effective in the treatment of other viral
infections as well.
EXAMPLE 12
In vitro anti-viral assay of the ozonide of linalool
A culture of SV-40 is grown in African Green Monkey (AGM) cells. The
culture is harvested in sterile saline. The titer of SV-40 in the
suspension is determined by Standard Plate Count Method in AGM cells.
A working suspension of SV-40 with a titer of approximately
1.0.times.10.sup.7 plaque forming units (PFUs)/0.1 ml is then
prepared. Four aliquots of 1 ml each of test solution containing 2.0%
ozonide of linalool are removed and placed in separate sterile screw-
capped tubes. Each sample is inoculated with 0.1 ml of the working
suspension of SV-40 to yield a final concentration of approximately
1.times.10.sup.6 PFUs/1 ml of the product. The samples are stored at
20.degree.-25.degree. C. for a total of 28 days. Samples are selected
at 7 day intervals to determine the number of viable PFUs present. A
control with uninoculated solution is also stored with samples
selected at the same intervals. At 7 days, and all subsequent sample
selections, there are less than 10 PFUs present. No PFUs are present
in any control sample.
EXAMPLE 13
Preparation of blood products free of viral activity
Blood obtained from a donor is mixed with 0.5 g of ozonide of geraniol
from Example 3 per unit (500 ml) of blood. The blood is then processed
in the normal manner. The resulting blood products are free of
detectable HIV or other viral activity using standard viral assays.
EXAMPLE 14
Test for efficacy of treatment of chicken pox
A small dose (approximately 25 l) of the composition of Example 9 is
topically applied to each lesion on the left side of a child suffering
from chicken pox. Lesions on the right side are treated with the
composition lacking in active ingredient. Within 24 hours, the lesions
on the child’s left side are significantly reduced with little or no
self-induced trauma from scratching. The lesions on the child’s right
side are unchanged in size, and show the effects of trauma from
scratching.
In a manner similar to that employed in Example 15, other viral
lesions, such as common warts and herpes lesions may be treated by
topical application of a terpene ozonide in a pharmaceutically
acceptable carrier or excipient.
EXAMPLE 15
Test for efficacy of treatment of condylomata acuminata
A 5 ml suppository with the composition of Example 4 is administered
intra-vaginally to one group of patients suffering from condylomata
acuminata. A second group of such patients receive a suppository
without the active ingredient of Example 4. A third group receives
cryogenic treatment of the affected area, a commonly used treatment
for condylomata acuminata. The average size of the lesions in each
group is approximately 2 cm.sup.2. Within seven days, the patients of
the first group have reduced reddening of the vagina and within 15
days, colposcopy does not reveal papilloma viruses. In the second
group of patients, the lesions are unchanged after 15 days. Patients
in the third group have no condylomata lesions immediately after
treatment, however, these patients continue to complain of pain and
bleeding for up to 30 days after the procedure is performed.
EXAMPLE 16
Treatment of Rheumatoid Arthritis
It has been theorized that rheumatoid arthritis is caused by a viral
agent. The antiviral ozonides of the present invention are believed to
be efficacious in treatment of this disease. Thus, a 20% oral
preparation comprising capsules containing citral ozonide in medium
chain triglyceride (MCT) is prepared and is taken twice daily by a
patient suffering from rheumatoid arthritis. Each dose delivers 400 mg
active ingredient to the 60 kg patient. After 1 week, the ANA of the
patient has dropped from approximately 2500 to 100, indicating
remission of the disease. Similar treatment is effective against
psoriasis.
Claims
I claim:
1. A method for treating viral infections of HIV in a mammal,
comprising the parenteral application of a pharmacologically
antiviral effective amount of an ozonide of a terpene in a
pharmaceutically acceptable carrier or excipient.
2. The method of claim 1, wherein said terpene is selected from the
group consisting of: limonene, citronella, alpha-carotene, beta-
carotene, vitamin A, linalool, linalyl acetate, squalene, geraniol,
alpha-pinene, loganin, cymene, farnesanes, eudesmanes, acoranes,
cedranes, chamigranes, caryophyllanes, illudanes, humulenes,
himachalenes, longifolanes, perhydroazulenes, quaianes,
quaianolides, germacranes, labdane, clerodanes, abietic acid,
phyllocladene, giberellins, ophiobolin A, retigeranic acid,
gasgardic acid, lanosterol, euphol, oleanane, ursane, lupeol,
hydroxyhopanone, lupanes, hopanes, B-selinene, zingibene, camphene,
sabinene, ocimene, myrcene, nerol, citral A, citral B, farnesol,
bisabolene, phytol and cecropia juvenile hormone.
3. The method of claim 1, wherein said terpene is a hemi-terpene,
mono-terpene, sesqui-terpene, di-terpene, ses-terpene, tri-terpene,
or tetra-terpene.
4. The method of claim 1, wherein said viral infection is systemic,
said parenteral application is by systemic injection, and said
composition is for systemic injection, and comprises an injectable
non-aqueous carrier.
5. The method of claim 1, wherein said viral infection produces local
sites of infection.
6. The method of claim 5, wherein said viral infection is an upper
respiratory infection.
7. The method of claim 5, wherein said viral infection is an infection
of the eye, ear, nose, or throat.
8. A method of treating mammalian blood to be used for medical
applications in the treatment of HIV comprising the addition of an
azonide of a hemi-terpene, mono-terpene, sesqui-terpene, di-
terpene, ses-terpene, tri-terpene, or tetra-terpene to said blood
causing said blood to be free of viral activity.
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