U.S. patent application number 15/953939 was filed with the patent office on 2018-10-18 for treating male infertility secondary to sperm oxidative stress.
The applicant listed for this patent is Parviz Gharagozloo. Invention is credited to Parviz Gharagozloo.
Application Number | 20180296597 15/953939 |
Document ID | / |
Family ID | 43030535 |
Filed Date | 2018-10-18 |
United States Patent
Application |
20180296597 |
Kind Code |
A1 |
Gharagozloo; Parviz |
October 18, 2018 |
TREATING MALE INFERTILITY SECONDARY TO SPERM OXIDATIVE STRESS
Abstract
A profertility composition for administration to infertile men
with sperm oxidative stress is disclosed. The composition comprises
a unique combination of a pharmaceutically acceptable vitamin E,
vitamin C, selenium, zinc, folic acid, lycopene and at least one
carnitine source.
Inventors: |
Gharagozloo; Parviz;
(Pennington, NJ) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Gharagozloo; Parviz |
Pennington |
NJ |
US |
|
|
Family ID: |
43030535 |
Appl. No.: |
15/953939 |
Filed: |
April 16, 2018 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
13749177 |
Jan 24, 2013 |
9943543 |
|
|
15953939 |
|
|
|
|
12771928 |
Apr 30, 2010 |
8377454 |
|
|
13749177 |
|
|
|
|
61174732 |
May 1, 2009 |
|
|
|
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 31/385 20130101;
A61K 31/01 20130101; A61K 31/375 20130101; A61K 33/30 20130101;
A61K 31/221 20130101; A61P 15/10 20180101; A61P 15/08 20180101;
A61K 31/519 20130101; A61K 45/06 20130101; A61K 31/355 20130101;
A61K 33/04 20130101; A61K 33/06 20130101; A61K 31/01 20130101; A61K
2300/00 20130101; A61K 31/221 20130101; A61K 2300/00 20130101; A61K
31/355 20130101; A61K 2300/00 20130101; A61K 31/375 20130101; A61K
2300/00 20130101; A61K 31/385 20130101; A61K 2300/00 20130101; A61K
33/04 20130101; A61K 2300/00 20130101; A61K 33/06 20130101; A61K
2300/00 20130101; A61K 33/30 20130101; A61K 2300/00 20130101 |
International
Class: |
A61K 33/30 20060101
A61K033/30; A61K 31/519 20060101 A61K031/519; A61K 31/221 20060101
A61K031/221; A61K 31/355 20060101 A61K031/355; A61K 31/375 20060101
A61K031/375; A61K 31/385 20060101 A61K031/385; A61K 33/04 20060101
A61K033/04; A61K 33/06 20060101 A61K033/06; A61K 31/01 20060101
A61K031/01; A61K 45/06 20060101 A61K045/06 |
Claims
1-30. (canceled)
31. A composition comprising a pharmaceutical active ingredient and
an antioxidant combination comprising at least one source of
vitamin E, at least one source of vitamin C, selenium or a
pharmaceutically acceptable form thereof, zinc or a
pharmaceutically acceptable form thereof, lycopene, a folate or
folic acid or suitable salts thereof, wherein the composition is
substantially free of any traces of vitamin A, vitamin K, and
garlic.
32. The composition of claim 31, wherein the vitamin C source is
ascorbic acid and wherein upon administration of said composition
for a sufficient period of time to a subject at least one index for
measuring DNA damage is reduced post-treatment by at least 2.5% as
compared to the pretreatment levels of the same index in said
subject.
33. The composition of claim 31, further comprising calcium,
magnesium, copper, N-acetyl cysteine, or a combination thereof.
34. The composition of claim 31, wherein said vitamin E source is
selected from the group consisting of alpha-tocopherol,
beta-tocopherol, gamma-toceopherol, delta-tocopherol,
alpha-tocotrienol, beta-tocotrienol, gamma-tocotrienol,
delta-tocotrienol, and any combination thereof.
35. The composition of claim 31, further containing lipoic
acid.
36. The composition of claim 31, wherein the carnitine source is
acetyl-L-carnitine and/or propionyl-L-carnitine.
37. The composition of claim 31 wherein the vitamin E source is in
amounts ranging from 50 IU-1500 I.U. and the vitamin C source is in
amounts of about 10 mg-600 mg.
38. The composition of claim 31, wherein the composition is in a
tablet dosage form selected from the group consisting of an
immediate release tablet, an extended or pulsatile release tablet,
a chewable tablet, a chewable lozenge, a quick chew, a quick
dissolve and combinations thereof.
39. The composition of claim 37, wherein the ratio of vitamin E to
vitamin C is at least 1 IU to 1 mg weight percent of the
composition.
40. The composition of claim 37, wherein the lipoic acid is
alpha-lipoic acid in amounts of about 50-600 mg.
41. The composition of claim 31, wherein the pharmaceutical active
ingredient is selected from the group consisting of an aromatase
inhibitor, an anti-inflammatory, an antibiotic and a combination
thereof.
42. A method of treating a patient in need comprising administering
to said patient a composition comprising a pharmaceutical active
ingredient and an antioxidant combination comprising at least one
vitamin E selected from the group consisting of alpha-tocopherol,
beta-tocopherol, gamma-tocopherol, delta-tocopherol,
alpha-tocotrienol, beta-tocotrienol, gamma-tocopherol,
delta-tocopherol, and any combinations thereof; at least one source
of vitamin C; selenium or a pharmaceutically acceptable form
thereof; zinc or a pharmaceutically acceptable form thereof, folic
acid or a pharmaceutically acceptable folate source; lycopene; at
least one source of carnitine selected from the group consisting of
propionyl-L-carnitine; acetyl-L-carnitine; or a combination
thereof; and at least one pharmaceutically suitable excipient and
optionally calcium, magnesium, copper or combinations thereof,
wherein the composition is substantially free of any traces of
vitamin A, vitamin K, and garlic.
43. The method of claim 42 wherein said folic acid or folate source
is administered in an amount ranging from about 0.1 mg to about 5.0
mg.
44. The method of claim 42, wherein the vitamin C source is
ascorbic acid and vitamin E is selected from the group consisting
of alpha-tocopherol, beta-tocopherol, gamma-tocopherol,
delta-tocopherol, alpha-tocotrienol, beta-tocotrienol or a
combination thereof.
45. The method of claim 42, wherein the vitamin C is administered
to the patient in an amount ranging from about 10 mg to about 1000
mg.
46. A kit comprising a pharmaceutical active ingredient and an
antioxidant combination comprising at least one source of vitamin
E, at least one source of vitamin C, selenium or a pharmaceutically
acceptable form thereof, zinc or a pharmaceutically acceptable form
thereof, lycopene, a folate or folic acid or suitable salts
thereof, wherein the composition is substantially free of any
traces of vitamin A, vitamin K, and garlic.
47. The kit of claim 46 further comprising an educational
information and direction of use.
48. The kit of claim 46, wherein the pharmaceutical active
ingredient is selected from the group consisting of an aromatase
inhibitor, an anti-inflammatory, an antibiotic and a combination
thereof.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to U.S. Provisional Patent
Application No. 61/174,732 filed May 1, 2009, the contents of which
is hereby incorporated by reference in its entirety.
FIELD OF INVENTION
[0002] The present invention relates to methods, compositions and
fertility kits used to enhance the natural fertility potential of
men. The present invention utilizes naturally occurring
antioxidants as to provide a prophylactic treatment of sperm
oxidative stress. Additionally, the novel products described herein
further include vitamins and minerals for improving semen
parameters enhancing the overall health of sperm thus increasing,
fertilization rates, number of viable embryos, pregnancy rates and
reducing miscarriages.
BACKGROUND
[0003] Male infertility has been on the rise for several decades
and is now considered a major health problem affecting around one
in twelve of the men in USA. Id. If the trends observed over the
last century continues, some infertility experts predict that by
the middle of this century, most men will be advised to freeze
their sperm at a young age to ensure adequate sperm quality.
[0004] A successful fertilization and subsequent embryo development
requires properly matured sperm chromatin with minimal damage to
its DNA integrity. Equally important, is the integrity of sperm
membrane lipid structure essential for sperm motility and
sperm-oocyte fusion. At least one of the causes of sperm DNA damage
and membrane peroxidation is attributed to excess Reactive
Molecular Species ("RMS"), most importantly Reactive Oxygen Species
("ROS"), leading to a condition commonly referred to as sperm
oxidative stress. In general, the term "Oxidative Stress" refers to
a condition that occurs as a result of excessive generation of ROS
and/or a diminished capacity of ROS scavenging endogenous
antioxidants. Accordingly, the excess concentration of ROS
overwhelms the local environmental antioxidant defense mechanism of
the spermatozoa potentially leading to the formation of sperm cells
with lower or no fertilizing potential.
[0005] Under normal conditions, cells utilize an array of enzymatic
and non-enzymatic antioxidants as internal defense to scavenge and
neutralize excess levels of ROS. It is believed that oxidative
stress develops either because of an overabundance of ROS from
environmental or pathological stressors or a lack of antioxidant
capacity to neutralize excess concentrations of ROS. Excess ROS
within semen and the sperm cell has been described to cause
degradation of the cellular components such as membrane and DNA,
hence, the sperms injured by such oxidative species have diminished
or no fertility potential.
[0006] To date several studies have reported that levels of ROS
within semen can be reduced by augmenting the scavenging capacity
of seminal plasma using oral antioxidant supplementation. In fact,
the benefits of oral antioxidant therapy has been described in
numerous articles. By the way of reference, such articles include
Comhaire et al, The Effects of Combined Conventional Treatment,
Oral Antioxidants and Essential Fatty Acids on Sperm Biology in
Subfertile Men, Prostaglandins Leuko Essent Fatty Acids, 63,
159-165 (2000); Lenzi et al, Placebo-Controlled, Double-blind,
Cross-over Trial of Gluthathione therapy in male infertility, Hum
Reprod, 8, 1657-1662 (1993); Greco et al, Reduction of the
Incidence of Sperm DNA Fragmentation by Oral Antioxidant Treatment.
J Androl, 17, 276-287 (2005); Tremellen et al, A Randomised Control
Trial Examining the Effect of an Antioxidant (Menevit) on Pregnancy
Outcome During IVF-ICSI Treatment, Australian and New Zealand
Journal of Obstetrics and Gynecology 47, 216-221 (2007); and
Agarwal et al, Oxidative stress, DNA Damage and Apoptosis in Male
Infertility, a Clinical Approach, BJU Int, 95, 503-507 (2005), the
teachings of which are incorporated herein in their entirety.
[0007] However, there is no product in the market today that has
the instantly disclosed combination of antioxidants, minerals and
vitamins. Moreover, the prior art does not teach the steps
described herein to enhance the natural fertility process. At least
one object of the present invention is to address such a need in
the field.
[0008] Excessive concentration of ROS can cause extensive sperm DNA
damage and membrane peroxidation. In general ROS includes reactive
moieties such as hydroxyl radicals (OH'), ionic species such as
superoxide anions (O.sub.2.sup.-), and neutral but highly reactive
oxidizing molecules such hydrogen peroxides (H.sub.2O.sub.2). In
semen, such species are predominantly generated by leukocytes or
morphologically abnormal sperms. It has been described in the art
that ROS plays both physiological and pathological roles in
maturation of sperm cells. For example, at normal baseline ROS
concentrations, sperms are properly matured as ROS modulates the
regulation of the normal sperm function in various cascades
including but not limited to sperm-oocyte fusion, acrosome reaction
and sperm capacitation.
[0009] On the other hand, excess ROS concentrations result in
direct insult on cellular components such as lipids, proteins and
DNA, thus interfering with normal cell function. Traditional semen
analysis methodologies fail to detect DNA damage caused by ROS. In
fact, in many cases infertile men who undergo a clinical workup
show normal sperm parameters, including sperm count motility and
morphology. However, when compared with healthy fertile men, at
least a subpopulation of these infertile men, show higher
concentrations of oxidants in their seminal fluid which may be
coupled with significantly lower concentrations of antioxidant as
compared to healthy fertile men.
[0010] At the genomic level, excess concentrations of ROS modifies
sperm DNA structure in a number of ways. For example, ROS can
induce chromatin cross-linking, DNA base oxidation and cause high
frequencies of single and double DNA strand breaks. Even though,
sperms with damaged DNA may still fertilize the oocyte, the degree
of DNA damage can have profound implications in normality of
embryonic development resulting in higher miscarriage rates, birth
defects or the long-term health of the progeny.
[0011] Current medical approach to facilitate embryonic development
and pregnancy uses invasive techniques collectively referred to as
Assisted Reproductive Techniques ("ART"), including Intrauterine
Insemination ("IUI"), In vitro Fertilization ("IVF") and
Intracytoplasmic Sperm Injection ("ICSI"). However, such techniques
generally have an average rate of success of about 30%. At least
one explanation for such poor outcome is the fact that the semen
and the sperm analysis techniques employed during ART cannot
differentiate sperms with DNA damage from the normal ones.
[0012] Moreover, ART modes of fertilization have been linked with
birth defects and incidences of childhood cancer, e.x. acute
leukemia and lymphoma. Nevertheless, ART remains the only viable
option for infertile couples. Another shortcoming of the ART is the
cost. Couples on average undergo several IVF attempts to achieve a
pregnancy, therefore undertaking huge expenses. Additionally, the
risk of failure often causes intense emotional trauma.
[0013] Moreover, ART can not and does not address the damage done
by ROS to sperm and ultimately to the fertilized egg. Therefore,
there is a need in the art to address the impact of Sperm Oxidative
Stress ("SOS") among infertile couples. Currently, there are no
effective and generally accepted pharmacotherapy that can combat
the problem of male infertility associated with SOS. Thus, there is
a need for convenient, inexpensive fertility methodologies and
products to aid and improve couple's fertility potential. There is
also a need for a kit that will provide couples in need with all
products, tools and material required to facilitate a comprehensive
plan for achieving a successful pregnancy.
SUMMARY OF THE PRESENT INVENTION
[0014] The present invention fills the foregoing need by lowering
and prophylactically treating SOS by boosting the non-enzymatic
antioxidant defense mechanism. The invention comprises unique
combinations of several natural antioxidants, vitamins and minerals
clinically tried for efficacy to improve sperm health and quality.
The invention is therefore intended to be a prophylactic treatment
to improve the overall health of sperms thus improving the
sperm-oocyte fertilization, reducing miscarriage rates and
improving pregnancy rates.
[0015] This invention provides methods of using combinations of
naturally occurring antioxidant molecules for treating infertility
in men predominantly suffering from SOS alone or in combination
with aromatase inhibitors, suitable anti-inflammatory, and/or
antibiotics.
[0016] At least one aspect of the present invention is directed to
a method of increasing the rate of pregnancy in a female subject by
administering to the male partner an effective amount of a
combination of anti-oxidants, vitamins and minerals, for at least a
period of up to about three (3) months prior to any fertilization
attempts, wherein said combination is substantially free of vitamin
A, vitamin K and/or garlic. In at least one embodiment of this
aspect of the invention, the combination includes at least one
source of vitamin C, at least one source of vitamin E, at least one
source of carnitine, lycopene and at least one source of folic
acid.
[0017] In a more preferred embodiment of this aspect of the
invention, the combination includes at least one source of vitamin
C, at least one source of vitamin E, at least one source of
carnitine, and at least one source of folic acid, at least one
mineral which is selenium, zinc, magnesium or combinations thereof,
wherein the composition is free of any traces of vitamin A, vitamin
K and/or garlic.
[0018] In another aspect of the present invention, a novel method
of reducing ROS damage to sperm DNA by at least 2.5% as compared
with placebo, including the steps of administering to a male
subject for at least a period of up to about three (3) months an
effective amount of the combination of anti-oxidants, vitamins and
minerals disclosed herein, wherein the combination is free of
vitamin A, vitamin K and/or garlic.
[0019] In at least one embodiment of this aspect of the invention,
the method of inhibiting ROS damage to sperm DNA cause a reduction
in a measurement of DNA fragmentation secondary to SOS by at least
1%, 2.5%, 5%, 8%, 10%, 12.5%, 15%, 20%, 30% and preferably higher
when compared to a placebo treatment. In another embodiment, the
sperm DNA fragmentation is assessed by such technique including but
not limited to Sperm Chromatin Dispersion ("SCD") test, Sperm
Chromatin Structure Assay ("SCSA"), DNA Breakage
Detection-Fluorescentce In Situ Hybridization ("DBD-FISH") test,
Terminal Deoxyribonucleotidyl Transferase-Mediated dUTP Nic-End
Labelling ("TUNEL") assay and 8-hydroxy-deoxyguanosine Assay
("8-0H-dG Assay").
[0020] Another aspect of the present invention provides for a
method of treating an infertile male subject with sperm oxidative
stress including the steps of (a) ascertaining the degree of DNA
damage in said male subject, as measured by SCSA, TUNEL, SCD,
DBD-FISH, or 8-0H-dG Assay, (b) initiating a course of therapy by
administering an effective amount of a combination of
anti-oxidants, vitamins and minerals and free of vitamin A, vitamin
K and/or garlic for at least a period of three (3) months, (c)
reascertaining the degree of DNA damage in said male subject post
the therapy of the step (b), and (d) fertilizing the egg of the
female partner with said male subject's sperm after completion of
the treatment course of step (b), wherein the degree of the DNA
damage measurement is reduced or otherwise improved, as compared to
placebo or said subject's own pretreatment values by at least 1%,
2.5%, 5%, 8%, 10%, 12.5%, 15%, 20%, 30%, and preferably higher.
[0021] Another aspect of the present invention provides for a
method of treating an infertile couple including the steps of (a)
determining the couples susceptibility to oxidative stress, (b)
initiating the combination regimen disclosed herein for the male
partner wherein the male partner receives an effective amount of a
combination of anti-oxidants, vitamins and minerals and free of
vitamin A, vitamin K and/or garlic for at least a period of three
(3) months. In another aspect of the present invention, the female
partner may independently or simultaneously with the male partner
receive the anti-oxidant therapy optionally including an
alpha-lipoic acid regimen.
[0022] The present invention also provides a method of reducing the
concentration of ROS generated by leukocytes in the reproductive
tract and/or semen of a male subject, the method including the
steps of administering to the male subject (a) an effective amount
of the antioxidant, vitamin and mineral combination for at least a
period of three (3) months and (b) an effective amount of an
aromatase inhibitor agent or an appropriate antibiotic.
[0023] The present invention also provides a method of improving
sperm function in a male subject, the method including the steps of
administering to the male subject an effective amount of the
combination described herein for at least a period of three (3)
months.
[0024] The present invention also provides for a method of
improving quality of an embryo produced by fertilization of an
oocyte by a sperm from a male subject, wherein the male subject has
completed a three (3) month course of antioxidant regimen
comprising receiving a combination of at least one source of
vitamin C, at least one source of vitamin E, at least one source of
carnitine, and at least one source of folic acid, at least one
mineral which is selenium, zinc, magnesium or combinations thereof,
wherein the regimen is free of any traces of vitamin A, vitamin K
and/or garlic.
[0025] In at least one embodiment of this aspect of the invention,
methods of prophylactically reducing the risk of forming a
nonviable zygote comprising administering to a male subject in need
thereof, a composition comprising an effective amount of a
combination of at least one source of vitamin C, at least one
source of vitamin E, at least one source of carnitine, and at least
one source of folic acid, at least one mineral which is selenium,
zinc, magnesium or combinations thereof, wherein the regimen is
free of any traces of vitamin A, vitamin K and/or garlic, and
optionally an effective dose of an aromatase inhibitor or an
antibiotic.
[0026] The present invention also provides a composition comprising
an effective amount of at least one source of vitamin C, an
effective amount of at least one source of vitamin E, an effective
amount of at least two sources of carnitine, and an effective
amount of at least one source of folic acid, at least one mineral
which is selenium, zinc, magnesium or combinations thereof wherein
the composition is substantially free of any traces of vitamin A,
vitamin K and/or garlic. In a preferred embodiment of this aspect
of the invention, the composition only contains an effective amount
of at least a source of vitamin C, an effective amount of at least
one source of vitamin E, an effective amount of L-carnitine,
propionyl-L-carnitine or acetyl-L-carnitine or combinations
thereof, and an effective amount of at least one source of folic
acid, an effective amount of lycopene, and an effective amount of
at least one mineral which is selenium, zinc, magnesium or
combinations thereof, wherein the composition is free of any traces
of vitamin A, vitamin K and/or garlic.
[0027] In a preferred embodiment of this aspect of the present
invention, inventor discloses a composition that contains vitamin
E, vitamin C or a suitable salt thereof, folic acid, lycopene, a
carnitine source, selenium, and zinc in pharmaceutically acceptable
forms wherein the formulation is substantially free of vitamin A
and vitamin K and/or garlic or any extracts thereof.
[0028] In a more preferred embodiment of this aspect of the
invention the composition is composed of vitamin E in about 100-800
IU; vitamin C about 50-800 mg, or an equivalent salt thereof;
selenium in about 25-100 .mu.g; zinc in about 10-35 mg; folic acid
in about 0.25-0.75 mg; lycopene about 3-12 mg; and a carnitine
source in about 250-4000 mg, wherein the formulation is
substantially free of vitamin A and vitamin K and/or garlic or any
extracts thereof. In yet another embodiment of the present
invention, the carnitine source is a prodrug for carnitine.
[0029] In the preferred embodiment of the present invention the
composition includes vitamin E in about 200 IU; vitamin C in about
90 or 200 mg, selenium in about 55 .mu.g; zinc in about 15 mg;
folic acid in about 0.5 mg; lycopene about 5 mg; and
acetyl-L-carnitine in about 1000 mg. In yet another embodiment, the
composition includes vitamin E in about 200 IU; vitamin C in about
200 mg, selenium in about 55 .mu.g; zinc in about 15 mg; folic acid
in about 0.5 mg; lycopene about 5 mg; and propionyl-L-carnitine in
about 500 mg, and acetyl-L-carnitine in about 1000 mg.
[0030] Another aspect of the present invention is directed to the
disclosed antioxidant containing compositions, wherein the
compositions are in the form of a tablet, a capsule, a powder
mixture, a paste, a suspension, a solution, an elixir, a topical or
an oral patch, or a parenteral preparation. In at least on
embodiment of this aspect of the invention, the composition
contains sustain or extended release components, immediate release
components, microparticles, nanoparticles and suitable excipients
to enhance the bioavailability of the product as compared to their
respective conventional preparations. In the preferred embodiment
of this aspect of the invention, the antioxidant containing
composition can be administered orally, via an inhaler, or via the
parenteral route.
[0031] In another aspect of the present invention, pregnancy kits
are provided comprising a set of pregnancy tests, articles for
storage of specimens, and compositions containing vitamin E in
about 200 IU; vitamin C in about 90 or 200 mg, selenium in about 55
.mu.g; zinc in about 15 mg; folic acid in about 0.5 mg; lycopene
about 5 mg; and acetyl L-carnitine in about 500 mg and/or
compositions containing vitamin E in about 200 IU; vitamin C in
about 90 or 200 mg, selenium in about 55 .mu.g; zinc in about 15
mg; folic acid in about 0.5 mg; lycopene about 5 mg; and
propionyl-L-carnitine in about 500 mg.
[0032] In at least another aspect of the present invention,
pregnancy kits can comprise additional adjuvant agents such as
aromatase inhibitors, glutathiones, or Coenzyme Q10, each of which
may be administered separately or in combination with the
antioxidant, mineral and vitamin combinations instantly
disclosed.
[0033] The present invention also provides a method of isolating
sperm and/or measuring DNA fragmentation from a male subject, the
method including the steps of (a) administering to the male subject
an effective amount of at least one source of vitamin C, an
effective amount of at least one source of vitamin E, an effective
amount of at least one source of carnitine, and an effective amount
of at least one source of folic acid, an effective amount of
lycopene, at least one mineral which is selenium, zinc, magnesium
or combinations thereof wherein the composition is substantially
free of any traces of vitamin A, vitamin K and/or garlic and (b)
measuring the subject's sperm's DNA fragmentation index.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0034] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as is commonly understood by one
of skill in the art to which this invention belongs and shall be
understood to have the meanings described below. All publications
and patents referred to herein are incorporated by reference in
their entirety. Unless otherwise specified, a reference to a
particular compound includes all ionic, salt, solvate (e.g.,
hydrate), protected forms, prodrugs, and other stereoisomers
thereof, isomeric forms, including racemic and other mixtures
thereof. The invention provides the first products to put these
components together to act in a synergistic manner in men's
formulations. The invention also provides for the very first time
the unexpected reduction of the DNA sperm fragmentation by the
combination described herein:
[0035] The term "aromatase inhibitors" is used herein to refer to
reversible or irreversible aromatase inhibitors including but not
limited to natural flavanoids or synthetic products such products
as anastrozole and exemestane (which is sold as Aromasin and is
chemically described as 6-methylenandrosta-1,4-diene-3,17-dione),
analogues or derivatives thereof such as those described in U.S.
Pat. Nos. 4,808,616, 4,876,045 and 4,904,650, the teachings of
which are incorporated herein in their entirety.
[0036] As used herein, "about" will mean up to plus or minus 5% of
the particular term.
[0037] As used herein, "consisting essentially of" refers to
excluding other active ingredients but including excipients.
[0038] The term "carnitine," except as individually recited,
includes all pharmaceutically acceptable forms of carnitine
including but not limited to 1-carnintine or a prodrug thereof,
acetyl-L-carnitine and propionyl-L-carnitine in form of HCl salt,
in general alkanoyl L-carnitines such as isovaleryl-L-carnitine,
hexanoly-L-carnitine, octanoyl-L-carnitine, myristoyl-L-carnitine,
palmitoyl-L-carnitine, stearoyl-L-carnitine.
[0039] By "Fertilization Attempt" it is meant any process that
facilitates fusion of the sperm and egg. Such term includes natural
as well as medical procedures that facilitate embryonic development
and pregnancy including but not limited to IUI, IVF and ICSI.
[0040] The term "folic acid or folate" (sometimes known as vitamin
B9) is meant to include the synthetic form of the vitamin, in
hydrate or salt forms with metal ions such as sodium, zinc etc.
[0041] The term "lycopene" is meant the red pigment in tomatoes in
its pharmaceutically acceptable organic or inorganic form, which
contains a C.sub.40 open-chain hydrocarbon carotenoid with any of
its 11 conjugated double bonds being available for the formation of
cis and trans-geometrical isomers.
[0042] By the term "Reactive Molecular Species ("RMS") it is meant
to include Reactive Oxygen Species ("ROS"), Reactive Nitrogen
Species ("RNS") and other reactive heteroatom species that can
incapacitate sperm ability to fertilize an egg, preferably ROS as
the most evaluated reactive moieties.
[0043] The term "selenium" is meant to include elemental, all
inorganic and organic forms of selenium such as sodium selenite and
sodium selenate, selenomethionine, selenocysteine,
methyselenocysteine, methylselenol, amino acid chelates, yeast, and
kelp bound selenium.
[0044] By the term "substantially free" it is meant that the final
product is free of any traces of the identified ingredient to the
extend that it could be detected by general quantitative assays
known in the art for detection of such ingredient in the final
product or if detected, it is in residual content of the ingredient
and in principle only attributable to impurities.
[0045] The term "treatment" or "therapy" as used herein in the
context of treating a condition to the extent that a positive
clinical benefit is observed. Thus, a course of therapy includes
prophylactic treatment, and further pertains generally to the
therapy of a human subject. For example, the treatment of sperm
oxidative stress and/or male infertility associated with sperm
oxidative stress includes a reduction in the rate of production,
rate of proliferation or the concentration of ROS in sperm cells,
which can result in a halt in the rate of infertility. Treatment
includes combination treatments and therapies, in which two or more
treatments or therapies modalities are combined, for example,
sequentially or simultaneously. Examples of treatments and
therapies include, but are not limited to, antioxidant combination
therapy with or without the use of anti-inflammatory agents,
surgery, employing ART and/or other drug treatments generally known
to improve rate of pregnancy.
[0046] The term "therapeutically-effective amount," as used herein,
pertains to that amount of an active compound, or a material,
composition or dosage form comprising an antioxidant, which is
effective for producing the desired therapeutic effect,
commensurate with a reasonable benefit/risk ratio to avoid excess
concentration of such drugs to the extent that it would neutralize
its expected therapeutic effects.
[0047] The term "vitamin C and/or its derivatives," except as
individually recited, is meant to include ascorbic acid, a mineral
ascorbate or a multi-mineral ascorbate, calcium ascorbate,
magnesium ascorbate, zinc ascorbate, potassium ascorbate, sodium
ascorbate, molybdenum ascorbate, chromium ascorbate, vitamin C
Complex, Ascorbyl palmitate.
[0048] The term "vitamin E and/or its derivatives" is meant to
include all variations of vitamin E, including but not limited the
eight antioxidants commonly known as vitamin E; four tocopherols
(alpha-, beta-, gamma-, and delta-) and four tocotrienols (alpha-,
beta-, gamma-, and delta-).
[0049] The term "zinc" is meant to include all variations of zinc
including but not limited to elemental zinc, zinc sulphate, zinc
picolinate, zinc citrate, zinc acetate, zinc glycerate, and zinc
monomethionine.
[0050] The invention provides a combination therapy for improving
fertility in mammals particularly among those suffering from excess
oxidative stress at their reproductive cell levels. At least one
aspect of the present invention provides treating infertility among
any mammal including but not limited to land and aquatic animals
such as primates, horses, donkeys, sheep, cats, dogs, pigs etc. In
a more preferred embodiment, the mammal is a male human, horse or
cow and in the most preferred embodiment the mammal is human.
[0051] In another aspect of the present invention the presently
disclosed treatment reduces male infertility associated with sperm
oxidative stress. The invention provides a scientifically validated
nutritional blend for mammalian male to reduce sperm oxidative
stress and infertility associated with such condition and further
improve the sperm genomic integrity. The present combination of
vitamins, minerals and amino acids reduces overall fragmentation of
sperm cells and further secure higher rate of pregnancy among
mammalian couples whose male counterpart suffers from
infertility.
[0052] Those of ordinary skill in the art understand that sperm is
highly susceptible to free radical or oxidative damage from
environmental toxicants and natural aging. In at least one
embodiment of the present invention vitamins C and E, zinc, and
selenium, all of which are potent antioxidants that help improve
sperm counts and quality, are used to neutralize the formation of
excess ROS and maintain normal DNA makeup. Zinc and B vitamins (B6,
B12 and folate) are critical nutrients in male reproductive systems
for hormone metabolism, sperm formation and motility. However,
synergistic combination of such ingredients for purposes of
treating male infertility secondary to SOS has not been clinically
proven. At least one aspect of the present invention, describes a
profertility combination of such ingredients for treating male
infertility associated with SOS.
[0053] In another aspect, the profertility combination of the
present invention employs a source of carnitine. Such source
includes L-carnitine, acetyl-L-carnitine or priopionyl-L-carnitine
and all pharmaceutically acceptable salts thereof. The preferred
embodiment of this aspect of the invention employs
priopionyl-L-carnitine, hydrochloride, fummurate or tarterate salts
thereof.
[0054] Another aspect of the present invention provides for a
component that would have a synergistic action of the combinations.
The distinct combinations are useful for couples who have failed to
produce a healthy fertilized oocyte during at least a period
ranging from three (3) to six (6) months prior to any fertilization
attempts. Such couples generally include male subjects exposed to
occupational toxic agents such as styrene, as well as those who may
be subject to environmental, chemical, radiation, and heat
exposure.
[0055] In one embodiment, the male human subject of the present
invention is selected from the group consisting of a subject with
increased levels of sperm DNA oxidation, reduced fertility of
unknown origin; a subject having undergone vasectomy reversal; a
subject with a reproductive tract infection such as epididymitis;
and a subject having a varicocele.
[0056] Previous human studies assessing male subjects with
increased levels of malondialdehyde or other biochemical markers of
oxidative stress; a smoker; a subject with reduced fertility,
including reduced fertility due to poor sperm motility have all
failed to substantiate the use of other antioxidants' combinations
in treating DNA fragmentation secondary to sperm oxidative stress.
See Tremellen supra.
[0057] The present inventor has unexpectedly discovered that the
instantly claimed blend of antioxidants drastically reduce sperm
DNA fragmentation from the pretreatment baseline and increase rate
of pregnancy.
[0058] Methods are known in the art for assessing the extent of
free radical damage to sperm. For example, the thiobarbituric acid
reactive substances ("TBARs") assay (which involves the measurement
of malondialdehyde, a marker of sperm membrane oxidation) or
LPO-856 spectrophotometric assay may be used. These methods are
described for example in Gomez et al International Journal of
Andrology 21(2), 81-96 (1998). Other methods include measurement of
DNA Fragmentation Index ("DFI") using SCSA, SCD test, DBD-FISH
test, and 8-OH-dG Assay.
[0059] In one embodiment, the administration of the present
combination therapy to the subject results in a reduction in free
radical damage to sperm's DNA of at least 2.5%, 5%, 8%, 10%, 12.5%,
15%, preferably 20% and most preferably 30%, when measured by a
suitable sperm parameter analysis, preferably evaluated or
expressed as DFI. In at least another embodiment, the
administration of the present combination therapy to the male
subjects results in a significant improvement in the degree of DNA
damage secondary to SOS, when compared to the subject's own
pretreatment values, by at least 1, 2.5%, 5%, 8%, 10%, 12.5%, 15%,
20%, 30%, and preferably 50%, wherein such improvement advances the
pregnancy of a female partner. In a preferred embodiment, the
administration of the antioxidant composition to a male subject
results in a reduction in pretreatment baseline levels of the
subject's DFI measurement by at least an absolute 2.5% (i.e. the
actual difference in DFI measurement before and after the
combination therapy).
[0060] The anti-oxidant agent in the various embodiments of the
present invention may be one or more individual antioxidant(s). In
this regard, an antioxidant is a molecule that can directly or
indirectly reduce the damaging effects of oxygen and/or
free-radicals in cells, and includes molecules that react with
oxygen, or molecules that may protect against, and/or react with, a
free radical.
[0061] In one embodiment, the anti-oxidant agent is selected from
one or more of the group consisting of a vitamin C; vitamin E;
.beta.-carotenoid, including lycopene (a carotenoid derived from
the tomato), lutein; a folic acid or derivative, selenium; zinc;
L-carnitine, a prodrug thereof such as propionyl-L-carnitine;
acety-L-carnitine; N-acetylcysteine; glutathione; gluthathionine;
pyruvate; Coenzyme Q10, astaxanthin and hypotaurine; alpha-lipoic
acid or a salt (if applicable), or a pharmaceutically acceptable
derivative of any of the aforementioned agents. Other anti-oxidants
are generally as described in Agarwal et al, Role of Antioxidants
in Treatment of Male Infertility: an Overview of the Literature,
Reproductive Biomedicine Online, 8(6), 616-62 (2004). Compounds
such as gluthathione, astaxanthin or Coenzyme Q10 may be
administered parenterally to enhance the ultimate clinical
antioxidation.
[0062] The effective amount of the one or more anti-oxidant agents
in the various embodiments of the present invention is not
particularly limited, so long as it has the desired or therapeutic
effect, and will depend upon the particular anti-oxidant(s)
administered. In a preferred embodiment, the present invention
contains tablets having vitamin E (d-alpha-tocopheryl acetate) in
ranges of 50 to 1500 I.U., with a usual range of 200 to 1200 I.U.,
and typically 200 to 800 I.U.; vitamin C (ascorbic acid, or a salt
thereof) in ranges of 10 to 1000 mg, with a usual range of 50 to
500 mg and typically 90 to 200 or 400 mg; selenium in ranges of 10
to 250 .mu.g; zinc in amounts of about 5 to 100 mg; folic acid in
amounts of about 0.1-1 mg; lycopene in amounts of about 0.5 to 20
mg, with usual ranges of 1 to 10 mg, a carnitine source in amounts
of about 1 to 5 grams, with a usual range of 2 to 3 grams; and
optionally glutathione in amounts of 100 to 1000 mg, with a usual
range of 400 to 600 mg, preferably administered parenterally. In
another aspect of this invention, the combination is supplemented
with alpha-lipoic acid in amounts of about 50 to 600 mg.
[0063] In another aspect of the present invention, the combination
therapy can optionally be supplemented with an aromatase inhibitor.
Preferred aromatase inhibitor agents include natural and synthetic
compounds that are able to modulate and reduce the activity of
Aromatase, an enzyme found in the liver, which is responsible for
the conversion of the androgens such as androstendione and
testosterone into the estrogens such as estrone and estradiol. By
inhibiting aromatase the body produces less estrogen and maintains
a higher testosterone state. Suitable aromatase inhibitors include
natural flavanoids such as chrysin, apigenin, indole-3-carbinols or
synthetic compounds including but not limited to aminoglutethimide,
anastrozole or 6-methylenandrosta-1,4-diene-3,17-dione.
[0064] In another embodiment, the anti-oxidant agent administered
to the subject is a combination of the following anti-oxidant
agents: vitamin C, vitamin E, selenium, zinc, and
propionyl-L-carnitine. In yet another embodiment, the combination
of the anti-oxidant agents consists essentially of vitamin C,
vitamin E, selenium, zinc, propionyl-L-carnitine and folic acid. In
yet another embodiment, the combination of the anti-oxidant agents
consists of vitamin C, vitamin E, selenium, zinc,
propionyl-L-carnitine and lycopene. In yet another embodiment, the
combination of the anti-oxidant agents consists essentially of
vitamin C, vitamin E, selenium, zinc, L-carnitine, folic acid and
lycopene. In yet another embodiment, the combination of the
anti-oxidant agents consists of vitamin C, vitamin E, selenium,
zinc, acetyl-L-carnitine, folic acid and lycopene, and suitable
excipients. In the most preferred embodiment, the combination of
the anti-oxidant agents consists of vitamin C, vitamin E, selenium,
zinc, acetyl-L-carnitine, proionyl-L-carnitine, folate or folic
acid and lycopene and suitable excipients.
[0065] A suitable formulation (referred to as the Fertilix.TM.
formulation) contains vitamin E (d-alpha-tocopheryl acetate)
vitamin C (ascorbic acid or a salt thereof), selenium, zinc, folate
or folic acid, lycopene, propionyl-L-carnitine, and
acetyl-L-carnitine.
[0066] At least one embodiment of such formulation essentially
contains vitamin E (d-alpha-tocopheryl acetate) 200 I.U.; vitamin C
(ascorbic acid or a salt thereof) 90-200 mg, selenium 55 .mu.g,
zinc 15 mg, folate or folic acid 0.5 mg, lycopene 5 mg,
propionyl-L-carnitine 500 mg, and acetyl-L-carnitine 1 gram.
Another embodiment contains vitamin C (ascorbic acid or a salt
thereof) in amount of 90 mg. Yet in another embodiment, the
formulation further contains lipoic acid in amount of 60 mg.
[0067] In one embodiment, the duration for the treatment is at
least for six (6) weeks to about twelve (12) months. In another
embodiment, the duration of the therapy is for three (3) months
prior any fertilization attempts. In further embodiments, the
duration of the treatment regime is for six (6) months prior any
fertilization attempts. In this regard, a suitable therapy for
either of the following formulations is two administrations per day
of one of the combinations, formulations, or compositions described
above for a period of at least three (3) months prior to any
fertilization attempts. However, it will be appreciated that the
administration of the disclosed formulation and the other agents in
the various embodiments of the present invention may be within any
time and frequency suitable to produce the desired effect.
[0068] The anti-oxidant and the other agents of the present
invention include those suitable for oral, nasal, topical
(including buccal and sublingual), rectal, vaginal and/or
parenteral administration. Regardless of the route of
administration selected, the combination of the ingredients are
formulated into pharmaceutically-acceptable dosage forms by
conventional methods known to those of skill in the art.
[0069] The amount of the active antioxidants which will be combined
with a carrier material to produce a single dosage form will
generally be that amount of the active ingredient(s) which is the
optimal dose effective to produce the therapeutic effect, herein as
measured by DFI.
[0070] Methods of preparing pharmaceutical formulations or
compositions include the step of bringing the antioxidants and/or
other suitable active ingredients into association with the carrier
and, optionally, one or more accessory ingredients. In general, the
formulations are prepared by uniformly and intimately bringing the
active ingredient(s) into association with liquid carriers, or
finely divided solid carriers, or both, and then, if necessary,
shaping the product.
[0071] Formulations of the invention suitable for oral
administration may be in the form of capsules, cachets, pills,
tablets, lozenges (using a flavored basis, usually sucrose and
acacia or tragacanth), powders, granules, microparticles,
nanoparticles or as a solution or a suspension in an aqueous or
nonaqueous liquid, or as an oil-in-water or water-in-oil liquid
emulsion, or as an elixir or syrup, or as pastilles (using an inert
base, such as gelatin and glycerin, or sucrose and acacia) and/or
as mouth washes and the like, each containing a predetermined
amount of the active ingredient(s).
[0072] In solid dosage forms of the invention for oral
administration (capsules, tablets, pills, powders, granules and the
like), the prodrug(s), antioxidant(s) and/or other suitable
pharmaceutically active ingredient(s) (in their micronized,
microparticle or nanoparticle forms) is/are mixed with one or more
pharmaceutically-acceptable carriers, such as cellulose, magnesium
stearate, silicon dioxide, sodium citrate or dicalcium phosphate,
and/or any of the following: (1) fillers or extenders, such as
starches, lactose, sucrose, glucose, mannitol, cellulose and/or
silicic acid; (2) binders, such as, for example,
carboxymethyl-cellulose, alginates, gelatin, polyvinyl pyrrolidone,
sucrose and/or acacia; (3) humectants, such as glycerol; (4)
disintegrating agents, such as agar-agar, calcium carbonate, potato
or tapioca starch, alginic acid, silicon dioxide, other types of
silicates, and sodium carbonate; (5) solution retarding agents,
such as paraffin; (6) absorption accelerators, such as quaternary
ammonium compounds; (7) wetting agents, such as, for example, cetyl
alcohol and glycerol monostearate; (8) absorbents, such as kaolin
and bentonite clay; (9) lubricants, such as talc, calcium stearate,
magnesium stearate, solid polyethylene glycols, sodium lauryl
sulfate, and mixtures thereof; and (10) coloring agents.
[0073] In the case of capsules, tablets and pills, the
pharmaceutical compositions may also comprise buffering agents.
Solid compositions of a similar type may also be employed as
fillers in soft and hard-filled gelatin capsules using such
excipients as cellulose, magnesium stearate, silicon dioxide,
lactose or milk sugars, with or without high molecular weight
polyethylene glycols and the like.
[0074] A tablet may be made by compression or molding, optionally
with one or more accessory ingredients. Compressed tablets may be
prepared using binder (for example, gelatin or hydroxypropylmethyl
cellulose), lubricant, inert diluent, preservative, disintegrant
(for example, sodium starch glycolate or cross-linked sodium
carboxymethyl cellulose), surface-active or dispersing agent.
Molded tablets may be made by molding in a suitable machine a
mixture of the powdered active ingredient(s) moistened with an
inert liquid diluent.
[0075] The tablets, and other solid dosage forms of the
pharmaceutical compositions of the present invention, such as
dragees, capsules, pills and granules, may optionally be scored or
prepared with coatings and shells, such as enteric coatings and
other coatings well known in the pharmaceutical-formulating art.
They may also be formulated so as to provide slow, extended or
controlled release of the desired antioxident(s) therein using, for
example, hydroxypropylmethyl cellulose in varying proportions to
provide the desired release profile, other polymer matrices,
nanoparticles and/or microspheres. They may be sterilized by for
example, filtration through a bacteria-retaining filter.
[0076] Liquid dosage forms for oral administration of the active
ingredient(s) include pharmaceutically-acceptable emulsions,
microemulsions, solutions, suspensions, syrups and elixirs. In
addition to the active ingredient(s), the liquid dosage forms may
contain inert diluents commonly used in the art, such as, for
example, water or other solvents, solubilizing agents and
emulsifiers, such as ethyl alcohol, isopropyl alcohol,
ethylacetate, butyl alcohol, benzyl benzoate, propylene glycol,
glycol, oils (in particular, cottonseed, groundnut, corn, germ,
olive, castor and sesame oils), glycerol, amyl alcohol,
tetrahydrofuryl polyethylene glycols and fatty acid esters of
sorbitan, and mixtures thereof.
[0077] Besides inert diluents the oral compositions can also
include adjuvants such as wetting agents, emulsifying and
suspending agents, sweetening, flavoring, coloring, perfuming and
preservative agents. Suspensions, in addition to the active
ingredient(s), may contain suspending agents as, for example,
ethoxylated alcohols, polyoxyethylene sorbitol and sorbitan esters,
microcrystalline cellulose, bentonite, agar and tragacanth, and
mixtures thereof.
[0078] Dosage forms for the topical or transdermal administration
of the active ingredient(s) include powders sprays, ointments,
pastes, creams, lotions, gels, solutions, patches and inhalants.
The active ingredient(s) may be mixed under sterile conditions with
pharmaceutically-acceptable carrier, and with any buffers, or
propellants which may be required.
[0079] The ointments, pastes, creams and gels may contain, in
addition to the active ingredient(s), excipients, such as animal
and vegetable fats, oils, waxes, paraffins, starch, tragacanth,
cellulose derivatives, polyethylene glycols, silicones, bentonites,
silicic acid, talc and zinc oxide, or mixtures thereof. Powders and
sprays can contain, in addition to the active ingredient(s),
excipients such as lactose, talc, silicic acid, aluminum hydroxide,
calcium silicates and polyamide powder, or mixtures of these
substances. Sprays and inhalers can additionally contain customary
propellants such as chlorofluorohydrocarbons and volatile
unsubstituted hydrocarbons, such as butane and propane.
[0080] Compositions of the present invention may be administered in
intranasal form via topical use of suitable intranasal vehicles, or
via transdermal routes, using those forms of transdermal skin
patches well known to those of ordinary skill in the art. A
transdermal delivery system provides for continuous administration
throughout the treatment regimen. Transdermal patches have the
added advantage of providing controlled delivery of the active
ingredient(s) to the body. Such dosage forms can be made by
dissolving, dispersing or otherwise incorporating the active
ingredient(s) in a proper medium, such as an elastomeric matrix
material. Absorption enhancers can also be used to increase the
flux of the active ingredient(s) across the skin. The rate of such
flux can be controlled by either providing a rate-controlling
membrane or dispersing the active ingredient(s) in a polymer matrix
or gel.
[0081] Another mode of delivery for present combination of the
present invention may be delivery via the use of targeting
compounds. Accordingly a nanoparticulate or microparticulate blend
of the antioxidant(s) in proper ratios may be coupled with soluble
polymers as targetable drug carriers. Such polymers can include
polyvinylpyrrolidone, pyran copolymer,
polyhydroxypropylmethacrylamide-phenol,
polyhydroxy-ethylaspartamide-phenol, or
polyethyleneoxide-polylysine substituted with palmitoyl residues,
polylactic acid, polyglycolic acid, copolymers of polyactic and
polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric
acid, polyorthoesters, polyacetals, polydihydropyrans,
polycyanoacrylates and crosslinked or amphipathic block copolymers
of hydrogels.
[0082] Pharmaceutical compositions of this invention suitable for
parenteral administration comprise the antioxidant(s) in
combination with one or more pharmaceutically-acceptable sterile
isotonic aqueous or nonaqueous solutions, suspensions or emulsions,
or sterile powders which may be reconstituted into sterile
injectable solutions or dispersions just prior to use, which may
contain the combination of the antioxidants, minerals and vitamins,
buffers, solutes which render the formulation isotonic with the
blood of the intended recipient or suspending or thickening
agents.
[0083] Examples of suitable aqueous and nonaqueous carriers which
may be employed in the pharmaceutical compositions of the invention
include water, dextrose, ethanol, polyols (such as glycerol,
propylene glycol, polyethylene glycol, and the like), and suitable
mixtures thereof. Proper fluidity can be maintained, for example,
by the use of coating materials, such as lecithin, by the
maintenance of the required particle size, and by the use of
surfactants.
[0084] These compositions may also contain adjuvants such as
wetting agents, emulsifying agents and dispersing agents. It may
also be desirable to include isotonic agents, such as sugars,
sodium chloride, and the like in the compositions. In addition,
prolonged absorption of the injectable pharmaceutical form may be
brought about by the inclusion of agents which delay absorption
such as aluminum monostearate and gelatin.
[0085] Injectable depot forms are made by forming microencapsule
matrices of the antioxidant(s), minerals and vitamins in
biodegradable polymers such as polylactide-polyglycolide. Depending
on the ratio of the active ingredient(s) to polymer, and the nature
of the particular polymer employed, the rate of release of the
active ingredient(s) can be controlled. Examples of other
biodegradable polymers include poly(orthoesters) and
poly(anhydrides). The injectable materials can be sterilized for
example, by filtration through a bacterial-retaining filter.
[0086] The pharmaceutical compositions of the present invention may
also be used in the form of veterinary formulations for treating
infertile cattle, herd, or other infertile mammals, including those
adapted for the following: (1) oral administration, for example,
drenches (aqueous or nonaqueous solutions or suspensions), tablets,
boluses, powders, granules or pellets for admixture with feed
stuffs, pastes for application to the tongue; (2) parenteral
administration, for "ampule," by subcutaneous, intramuscular or
intravenous injection as, for example, a sterile solution or
suspension or, when appropriate, by injection where a suspension or
solution is introduced to the animal; (3) topical application, for
example, as a cream, ointment or spray applied to the skin; or any
other methods fit to by those of ordinary skill in the art for
administration to a region of interest.
[0087] Formulation of the agents of the present invention and their
administration may be facilitated by an easy to use kit to
ameliorate and assist patients to follow the treatment regimen. As
discussed previously herein, the agents in the various embodiments
of the present invention may be administered jointly or separately
in the form of oral preparations (for example solid preparations
such as tablets, capsules, granules or powders; liquid preparations
such as syrup, emulsions or suspensions).
[0088] Compositions containing the anti-oxidant agent and the other
agents separately or jointly in the various embodiments of the
present invention may also contain a preservative, stabilizer,
dispersing agent, pH controller or isotonic agent Examples of
suitable preservatives in the various embodiments of the present
invention are glycerin, propylene glycol, phenol or benzyl alcohol.
Examples of suitable stabilizers in the various embodiments of the
present invention are dextran, gelatin, or alpha-thioglycerin.
[0089] Examples of suitable dispersing agents in the various
embodiments of the present invention include polyoxyethylene (20),
sorbitan mono-oleate (Tween 80), sorbitan sesquioleate (Span 30),
polyoxyethylene (160) polyoxypropylene (30) glycol (Pluronic F68)
or polyoxyethylene hydrogenated castor oil 60. Examples of suitable
pH controllers in the various embodiments of the present invention
include hydrochloric acid, sodium hydroxide and the like. Examples
of suitable isotonic agents are glucose, D-sorbitol or
D-mannitol.
[0090] As discussed previously herein, the present invention is
also suitable for reducing the generation of ROS in the male
reproductive tract and/or semen. In this regard, the male
reproductive tract will be understood to include the epididymis,
the penis, the prostate gland, the seminal vesicles, the testes,
the vas deferens and semen. Methods for determining the level of
free radicals in the male reproductive tract are known in the art.
For example, free radical production by sperm or seminal leukocytes
can be measured directly using chemiluminescence assays known in
the art. Alternatively, assays that measure free radical related
damage to sperm lipid membrane may be used. For example, the TBARs
assay (which involves the measurement of malondialdehyde, a marker
of sperm membrane oxidation) or LPO-586 spectrophotometric assay
may be used. These methods are described in Gomez et al (1998)
International Journal of Andrology 21(2):81-96.
[0091] In one embodiment, the level of male fertility track is
assessed by the way of suitable inflammatory indices. For example,
one of ordinary skill in the art would with to assess the presence
or absence of inflammatory cytokine, like one or more of IL-I,
IL-6, IL-, TNF-.alpha. and Interferon-.gamma.. Methods for
determining the level of an inflammatory cytokine in the male
reproductive tract are known in the art, such ELISA assays for
detection of pro-inflammatory cytokines such as IL-I, IL-6, IL-8,
TNF-.alpha. and Interferon-.gamma., as described in Depuydt et al.
J Andrology 17(61), 699-707 (1996) or Maegawa et al J Reprod
Immunology, 54, 33-42 (2002), and Nallella et al. Urology 64(5),
1010-3 (2004).
[0092] Those of ordinary skill in the art can also employ other
methods of assessing the "sperm function." The term as used herein
includes any key component of sperm physiology and includes
swimming activity towards the oocyte (motility), ability to undergo
capacitation to penetrate the oocyte's outer coat (zona pellucida)
and fuse with the oocyte membrane, and maintenance of sperm DNA
integrity to form a functional male pro-nucleus at syngamy. Methods
for determining the level of sperm function are known in the art.
For example, suitable methods are described in detail within the
World Health Organization (WHO) laboratory manual for the
examination of human semen and sperm-cervical mucous interaction.
4th edition. Cambridge University Press 1999.
[0093] In at least one embodiment of the present invention, the
sperm DNA fragmentation is assessed by SCSA. Evenson D P, et al,
Relationship Between Assisted Reproductive Techniques (ART) outcome
and Status of Chromatin Integrity as Measured by the Sperm
Chromatin Structure Assay (SCSA). Hum. Reprod., 15, 1717-1722
(2000). At least one indicator of the clinical benefits of the
claimed methodology is marked improvement of the DFI.
[0094] In principle, spermatozoa chromatin is different in nature
from somatic cells chromatin. This difference is due to ploidy and
DNA packaging, which is influenced by the replacement of histones
in spermatocytes by transition proteins and then substitution of
histone by protamine in spermatides. Green et al, Synthesis and
Processing of Mammalian Protamines and Transition Proteins. Mol.
Repr Dev 37, 255-263 (1994). Histones are the chief protein
components of chromatin that coils around DNA enabling it to fit
the large genomic information inside cell nuclei. Without histones,
the unwound DNA in chromosomes would be very long. Replacement of
histones by protamine are generally known to contribute to sperm's
head condensation and DNA stabilization. See Id. Those of ordinary
skill in the art can appreciate that DNA fragmentation levels above
30% as measured by SCSA are not compatible with initiation and
maintenance of a term pregnancy. See Larson supra. SCSA test
values, expressed as DFI, are significantly correlated with
pregnancy rate in vivo and in vitro. In studies that included more
than 25 couples undergoing in vitro fertilization and
intracytoplasmic sperm injection cycles, no term pregnancy occurred
when the DFI measurement was more than 27% in the semen samples
utilized in these cycles. See Larson supra.
[0095] In the preferred embodiment of the present invention,
baseline DFI is measured prior to the administration of the
antioxidant regimen to assess a fertility capacity of male
subjects. In the present invention, DFI values in combination with
other subjective or objective signs of infertility would be used as
the criteria to determine couples predisposition to failed
pregnancy. Once study couples have undergone the baseline
evaluation, they will receive a three (3) months course of therapy
of the claimed antioxidant formulation in a kit containing other
educational material regarding their therapy.
[0096] The effectiveness of the treatment course is then
substantiated by statistically significant reduction in DFI and
further the actual pregnancy of the couple's female subject. Those
of ordinary skill in the art can appreciate measurements of other
sperm parameters during the period of the trial to assess treatment
efficacy. Such parameters can include sperm count, sperm density,
motility, membrane integrity, testosterone or other hormonal levels
if so desired, and other DNA damage techniques at various intervals
including the entry period, 6 weeks, and/or 12 week time points.
However, the actual clinical end point can be evaluated in view of
other indicators including but not limited to fertilization rates
among treatment group, embryo or blastocyst quality, number of
viable embryos or blastocyst for transfer, number of miscarriages
and the occurrence rate thereof.
[0097] An effective amount of the antioxidant agents and the other
agents may be appropriately chosen, depending upon, for example,
the type and extent of reduced fertility to be treated, the age and
body weight of the subject, the DFI measurement, the frequency of
administration, and the presence of other active agents. The
instant antioxidant, mineral and vitamin combination, and/or the
anti-aromatase agent may be administered to the subject separately
or in combination via separate routes of administrations.
[0098] Accordingly, in another embodiment the present invention
provides a combination product for improving sperm function in a
male subject, the combination kit product including the following
components: (a) a composition containing the present antioxidant,
mineral and vitamin combination; and/or (b) an aromatase inhibitor
agent, and/or (c) another adjuvant agent such as gluthothione or
Coenzyme Q10 with (d) educational information and direction of
use.
[0099] In yet another embodiment, the inventor describes the use of
a carnitine source in the combinations described herein.
L-carnitine, acetyl-L-carnitine and propionyl-L-carnitine are
endogenous ligands that play essential role in transporting fatty
acids into the mitochondria, where they are oxidized to produce
energy. The carnitine pool is particularly important to spermatozoa
as they are rich in mitochondria generating energy especially to
support their motility. Carnitines are also scavengers of oxygen
free radicals in mammalian tissues. Thus, the biochemical and
antioxidant role of carnitines make them vitally important
molecules in the overall health of the spermatozoa.
[0100] Propionyl-L-carnitine is known in the art to increase
cellular levels of L-carnitine by binding to the enzyme carnitine
acetyltransferase (CAT) which is subsequently converted into
propionyl-coenzyme A and free carnitine. Propionyl-L-carnitine is
also rapidly absorbed after oral administration reaching peak
concentrations in about 1 hour followed by a rise in L-carnitine
plasma levels 2-6 hours after the administration.
[0101] WO publication 03/084526 have previously shown the impact of
such prodrugs in sperm motility and concentration. However, the
acetyl-L-carnitine and propyl-L-carnitine alone or in combination
have not been described in male prenatal compositions have not been
described. In at least one aspect of the present invention, the
inventors believe that the exogenous propionyl-L-carnitine
supplementation alone will more effectively address any carnitine
deficiency in infertile men.
[0102] The forgoing examples are used to further describe the
invention employing at least one embodiment of the presently
disclosed combination, the Fertilix.TM. profertility therapy Those
of ordinary skill in the art can appreciate that patients who had
received IVF before and during, as well as, those patients
resorting to other fertilization attempts, would benefit from the
presently disclosed treatment regimen by improving their embryonic
quality and reducing the complications during the pregnancy term.
At least one embodiment of the present invention focus the
treatment regimen on the male partners. Yet another embodiment of
the invention, include independently or simultaneously
administering the regimen to the female partners.
[0103] With well over 3 million men currently experiencing male
related infertility in the United States. Traditionally male
infertility treatment has not endeavored to ameliorate the
underlying cause of infertility but rather used "mechanical"
techniques such as intra-uterine insemination or IVF-ICSI to bypass
the defect in sperm function. While these two techniques are
undeniably successful in a large proportion of patients, they
simply do not work or have very limited efficiency in other
couples. It is likely that in many cases, sperm DNA fragmentation
is responsible for the poor pregnancy outcome despite ART
treatment. The present invention provides treatments that can
prophylactically treat sperm DNA fragmentation and is likely to
boost both natural and ART related pregnancy rates.
[0104] A total of two (2) separate human trials are envisioned for
further assessing the efficacy of the combination therapy instantly
disclosed. Prior experiences with antioxidant combinations have not
provided any evidence that antioxidant combinations can reduce
sperm's DNA fragmentation secondary to oxidative stress. See
Tremelton supra. Contrary to such scientific body of evidence, the
present treatment regimen unexpectedly show a significant reduction
of DFI after three (3) months of therapy.
[0105] In assessing the efficacy of other prior art antioxidant
combinations, one of ordinary skill in the art would appreciate
that even though the rate of pregnancy outcomes might have improved
in the antioxidant treatment groups; the measured sperm parameters
for the treatment group was not affected and the treatment had no
effect on sperm concentration, motility or morphology. See
Tremelton supra at pgs 219-220. Furthermore, the LPO-586 assay for
lipid peroxidation damage did not detect any significant difference
in levels of free radical damage to the sperm membrane of the
subjects. See Id. In fact, prior art formulations failed to show a
correlation between SOS or sperm DNA damage and pregnancy rates
since the sperm DNA damage assays employed failed to show a
difference between the treatment and the placebo groups. See
Id.
[0106] The present invention provides a new combination of
antioxidants with an unexpected reduction in sperm's DNA
fragmentation due to local oxidative stress. Unexpected results
extend to the inventors observation of improved quality of the
sperm parameters, which when coupled with successful pregnancies,
confirms the efficacy of Fertilix.TM. therapy in treating male
infertility associated with SOS.
Example 1
[0107] First combination for men is prepared according to the
following formula:
TABLE-US-00001 TABLE I d-alpha-tocopheryl acetate 200 IU Ascorbic
acid 200 mg Selenium 55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene
5 mg Propionyl-L-Carnitine 0.5 g Excipients As needed
Example 2
[0108] A second combination is prepared according to the following
formula:
TABLE-US-00002 TABLE 2 Vitamin E 200 IU Vitamin C 200 mg Selenium
55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg
Acetyl-L-Carnitine 1.0 g Propionyl-L-Carnitine 0.5 g Excipients As
needed
Example 3
[0109] Another formulation is prepared according to the following
formula:
TABLE-US-00003 TABLE 3 Vitamin E 200 IU Vitamin C 200 mg Selenium
55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg L-Carnitine 1 g
Excipients As needed
Example 4
[0110] Another formulation is prepared according to the following
formula:
TABLE-US-00004 TABLE 4 Vitamin E 200 IU Vitamin C 200 mg Selenium
55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg L-Carnitine 1.0
g Acetyl-L-Carnitine 0.5 g Excipients As needed
Example 5
[0111] Another formulation is prepared according to the following
formula:
TABLE-US-00005 TABLE 5 Vitamin E 200 IU Vitamin C 200 mg Selenium
55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg L-Carnitine 1.0
g Propionyl-L-Carnitine 0.5 g Excipients As needed
Example 6
[0112] Another formulation for men is prepared according to the
following formula:
TABLE-US-00006 TABLE 6 Vitamin E 200 IU Ascorbic acid 90 mg
Selenium 55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg
Propionyl-L-Carnitine 0.5 g Excipients As needed
Example 7
[0113] Yet, another formulation is prepared according to the
following formula:
TABLE-US-00007 TABLE 7 Vitamin E 200 IU Vitamin C 90 mg Selenium 55
.mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg Acetyl-L-Carnitine
0.75 g Propionyl-L-Carnitine 0.5 g Excipients As needed
Example 8
[0114] Another formulation is prepared according to the following
formula:
TABLE-US-00008 TABLE 8 Vitamin E 200 IU Vitamin C 90 mg Selenium 55
.mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg L-Carnitine 1 g
Excipients As needed
Example 9
[0115] Another formulation is prepared according to the following
formula:
TABLE-US-00009 TABLE 9 Vitamin E 200 IU Vitamin C 90 mg Selenium 55
.mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg L-Carnitine 1.0 g
Acetyl-L-Carnitine 0.5 g Excipients As needed
Example 10
[0116] Another formulation is prepared according to the following
formula:
TABLE-US-00010 TABLE 10 Vitamin E 200 IU Vitamin C 90 mg Selenium
55 .mu.g Zinc 15 mg Folic acid 0.5 mg Lycopene 5 mg L-Carnitine 1.0
g Propionyl-L-Carnitine 0.5 g Excipients As needed
Example 11
[0117] Couple DP underwent multiple cycles of IVF that resulted in
no viable pregnancy. The sperm DNA fragmentation index of the male
subject was measured by SCSA to be 27%. The male partner then
started on a twice a day combination therapy described in Example
3, Table 3 for a period of three (3) months. The combination
contained vitamin E (d-alpha-tocopheryl acetate) 200 I.U., vitamin
C 200 mg, selenium 55 .mu.g, zinc 15 mg, folic acid 0.1 mg,
L-carnitine 1 g, and lycopene 5 mg.
[0118] The post treatment DFI measurement showed an absolute 7.4%
reduction in the DFI levels amounting to a 27.4% improvement from
the pretreatment baseline DFI levels. Upon completion of the
therapeutic regimen, even though no in vitro embryo assessment was
done, the female partner became pregnant and gave birth to a
healthy child nine (9) months following the termination of the
anti-oxidant therapy.
Example 12
[0119] In another trial, the same couple used the same treatment
regimen as described in Example 2. The male's sperm DFI measurement
indicated a free radical damage of 39.6% prior to the initiation of
the antioxidant combination therapy. The DFI measurement at least
three (3) months after the initial treatment was measured as 27.2
showing an unexpected improvement of at least an absolute 12.4%
reduction in DNA fragmentation index amounting to a 31% improvement
from the pretreatment baseline DFI levels. While no in vitro embryo
assessment was done, the female partner became pregnant and gave
birth to a healthy child nine (9) months thereafter.
[0120] While the invention has been described with references to
specific embodiments, modifications and variations of the invention
may be construed without departing from the scope of the invention,
which is defined in the following claims.
* * * * *