`
`(19) World Intellectual Property Organization
`International Bureau
`
`(43) International Publication Date
`6 February 2003 (06.02.2003)
`
`• I lllll llllllll II llllll lllll llll I II Ill lllll lllll lllll 111111111111111111111111111111111
`
`(10) International Publication Number
`WO 03/009689 Al
`
`PCT
`
`(51) International Patent Classification7:
`
`AOlN 55/08
`
`(74) Agents: BROWDY AND NEIMARK, P.L.L.C. et al.;
`624 Ninth Street N.W., Suite 300, Washington, DC 20001-
`
`(21) International Application Number: PCT/US02/23252
`
`5303 (US).
`
`(22) International Filing Date:
`
`23 July 2002 (23.07.2002)
`
`(25) Filing Language:
`
`(26) Publication Language:
`
`English
`
`English
`
`(30) Priority Data:
`60/306,857
`
`23 July 2001 (23.07.2001) US
`
`(71) Applicant (for all designated States except US): RAMOT
`UNIVERSITY AUTHORITY FOR APPLIED RE(cid:173)
`SEARCH & INDUSTRIAL DEVELOPMENT LTD.
`[IL/IL]; P.O. Box 39296, 61392 Tel Aviv (IL).
`
`(71) Applicant (for SD only): MCINNIS, Patricia [US/US];
`2325 42nd Street, N.W., Apt. 203, Washington, DC 20007
`(US).
`
`(81) Designated States (national): AE, AG, AL, AM, AT, AU,
`AZ, BA, BB, BG, BR, BY, BZ, CA, CH, CN, CO, CR, CU,
`CZ, DE, DK, DM, DZ, EC, EE, ES, Fl, GB, GD, GE, GH,
`GM, HR, HU, ID, IL, IN, IS, JP, KE, KG, KP, KR, KZ, LC,
`LK, LR, LS, LT, LU, LV, MA, MD, MG, MK, MN, MW,
`MX, MZ, NO, NZ, OM, PH, PL, PT, RO, RU, SD, SE, SG,
`SI, SK, SL, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ,
`VN, YU, ZA, ZM, ZW.
`
`(84) Designated States (regional): ARIPO patent (GH, GM,
`KE, LS, MW, MZ, SD, SL, SZ, TZ, UG, ZM, ZW),
`Eurasian patent (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM),
`European patent (AT, BE, BG, CH, CY, CZ, DE, DK, EE,
`ES, Fl, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE, SK,
`TR), OAPI patent (BF, BJ, CF, CG, CI, CM, GA, GN, GQ,
`GW, ML, MR, NE, SN, TD, TG).
`
`Published:
`with international search report
`before the expiration of the time limit for amending the
`claims and to be republished in the event of receipt of
`amendments
`
`For two-letter codes and other abbreviations, refer to the "Guid(cid:173)
`ance Notes on Codes and Abbreviations" appearing at the begin(cid:173)
`ning of each regular issue of the PCT Gazette.
`
`= ---
`-iiiiiiii ---=
`--=
`iiiiiiii ----=
`
`(72) Inventors; and
`(75) Inventors/Applicants (for US only): FREEMAN, Ami(cid:173)
`hay [IL/IL]; Ben Shemen Youth Village, 73112 Ben She(cid:173)
`men Youth Village (IL). SEGAL, Rina [IL/IL]; Hacharzit
`7, 40600 Tel Mond (IL). DROR, Yael [IL/IL]; Hacharzit
`18, 40600 Tel Mond (IL).
`
`iiiiiiii
`
`°" 00
`\0 °" 0
`
`0
`...........
`~ ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
`
`0
`
`(54) Title: METHODS AND COMPOSITIONS FOR TREATING FUNGAL INFECTIONS
`
`0 > (57) Abstract: Phenylboronic acid and water soluble derivatives thereof and related boronic acid compounds are used for treating
`
`~ fungal and bacterial infections.
`
`CFAD Exhibit 1004
`
`
`
`WO 03/009689
`
`1
`
`PCT/US02/23252
`
`METHODS AND COMPOSITIONS FOR TREATING FUNGAL INFECTIONS
`Field of the Invention
`[001]
`The present invention relates to methods and
`compositions for treating fungal infections, and more
`particularly, dermatophytoses or onchomycosis of the
`fingernail and the toenail, as well as fungal infections in
`plants.
`Background of the Invention
`(002]
`Many fungal infections, or mycoses, of humans and
`animals affect only the outer layers of skin. Although these
`infections may be sometimes difficult to cure, they are not
`considered dangerous. Most cutaneous infections are caused by
`the homogeneous group of keratinophilic fungi known as
`
`dermatophytes. The dermatophyte Trichophyton rubrum is the
`major cause of tinea pedis and onychomycosis. Fungal
`infections of the mucous membranes are caused primarily by
`
`Candida albicans, usually affecting the mouth and the vaginal
`and anal regions.
`(003]
`Fungal infections sometimes follow the use of
`antibiotics, which kill non-pathogenic as well as pathogenic
`bacteria, thereby providing a clear field for fungal invasion.
`Opportunistic fungal infection occurs when a fungus enters a
`compromised host, such as a patient suffering from AIDS.
`[004]
`Dermatophytoses of the fingernails and toenails, in
`contrast to those at other body sites, are particularly
`difficult to eradicate with drug treatment, particularly with
`topical treatment. This is the consequence of factors that
`are intrinsic to the nail such as
`the hard, protective nail
`plate, sequestration of pathogens between the nail bed and
`plate, and slow growth of the nail, as well as the relatively
`poor efficacy of the early pharmacologic agents.
`[005]
`"Onychomycosis" has traditionally referred to a non-
`dermatophytic infection of the nail. Onychomycosis is now
`used as a general term to denote any fungal nail infection.
`
`
`
`WO 03/009689
`
`PCT/US02/23252
`
`2
`
`Tinea unguium specifically describes a dermatophytic invasion
`of the nail plate. Despite the clearly diseased appearance
`associated with this condition, onychomycosis is all too often
`regarded as merely a cosmetic problem of relatively minor
`importance that is hardly worth treating. This belief may
`have been fostered by the adverse effects and long courses of
`medication associated with some of the earlier antifungal
`agents.
`However, onychomycosis can have significant negative
`[006]
`effects on patients' emotional, social, and occupational
`functioning. Affected patients may be embarrassed in social
`and work situations, where they may feel unclean, and are
`unwilling to permit their hands and feet to be seen. Patients
`may fear that they will transmit their infection to family
`members, friends, or coworkers, fears that can lead to
`diminished self-esteem and avoidance of close relationships.
`Some patients experience discomfort that prevents them from
`carrying out tasks such as prolonged standing, writing, or
`typing.
`Onychomycosis in immunocompromised patients, such as
`[007]
`those infected with human immunodeficiency virus, can pose a
`
`more serious health problem. Not only does this infection
`serve as a constant reminder to the patient of his or her own
`deteriorated condition, but the possibility exists of transfer
`of a very high titer of fungal pathogens to another body site.
`[008].
`The dermatophyte species that most often causes
`
`onychomycosis in North America and parts of Europe are T.
`
`rubrumr T. metagrophytes, and Epidermophyton floccosum. The
`
`first two are much more often implicated than E.
`
`floccosum.
`
`Both dermatophytes and non-dermatophytes, especially Candida
`Sp., have been identified as etiologic agents of
`onychomycosis.
`[009]
`Contact with the source of infection constitutes a
`risk factor. Several factors unique to modern life have
`resulted in an increased prevalence of onychomycosis,
`
`
`
`WO 03/009689
`
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`
`3
`
`including wearing of shoes, especially tight, high-heeled
`shoes; the increased use by large numbers of people of damp
`spaces such as locker rooms and gymnasiums; the declining
`health of the aging American population, and the increased
`number of immunocompromised patients through disease (HIV) or
`therapeutic agents (immunosuppressive therapies associated
`with cancer or posttransplantation, and the extensive use of
`broad-spectrum antibiotics). Other factors that increase the
`risk of onychomycosis are direct trauma to the nail, including
`that resulting from certain tic disorders (nail biting).
`[0010]
`Treatment of onychomycosis has been attempted for
`many generations, but success has been limited. Because of
`the perception that the lesions had a superficial cause, the
`earliest remedies were topical.
`However, topical drugs such
`as the imidazoles, the allylamines, and the pyridone
`cyclopiroxolamine proved to be generally ineffective against
`fungal infections of the nails because of their inability to
`penetrate the entire nail unit and eradicate the infection.
`Only recently, when the fungal nature of these infections was
`appreciated, have systemically active drugs been available for
`treating onychomycosis.
`[0011]
`Many currently available antifungal agents require a
`long duration of therapy, sometimes for over one year, in
`order to completely treat the onychomycosis. Griseofulvin has
`limited efficacy because its activity is limited to
`dermatophytes and a prolonged duration of therapy is required
`for maximum efficacy. Ketoconazole cannot be used for long(cid:173)
`term cure of onychomycosis because of the occurrence of side
`effects and significant drug interactions. Other previously
`used drugs include itraconazole, fluconazole, and terbinafine.
`[0012]
`Additionally, serious damage is done to crops each
`year by fungal infections of plants such as smuts, rusts,
`ergot, and mildews.
`[0013]
`Botrytis bunch rot has long been a problem in
`vineyards. High nitrogen fertilization predisposed grapevines
`
`
`
`WO 03/009689
`
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`
`4
`
`to infection by Botritis cinerea and increased disease
`
`severity.
`
`In vitro results of tests of a number of fungicides
`
`were described by R=Houma et al. in Journal of Plant Pathology
`
`80(2): 1998, abstracts of papers. Of the fungicides tested,
`Vinchlozoline, Chlorothalonil, and Dichlof luanide were
`effective in completely terminating conidia.
`Iprodione and
`Procymedone were apparently confronted with the problem of
`fungal resistance. Folpel, copper and'chlorothalonil were not
`able to control mycelial growth as effectively as conidial
`germination.
`
`[0014]
`
`Several Fusarium species occurring worldwide on
`cereals as causal agents of "head blight" of small grain
`cereals and "ear rot" of corn, can accumulate mycotoxins in
`infected kernels. Besides being damaging to the cereal crops,
`some of these mycotoxins are dangerous to animal and/or human
`
`health. The main groups of Fusarium toxins commonly
`
`recognized in grains are trichothecenes: including T-2 toxin
`(T2), diacetoxyscirpenol, deoxynivalenol, fusarenone X, and
`nivalenol; zearalenones, primarily zaearalenone; and
`fumonisins, in particular fumonisin Bi. Additionally,
`moniliformin, beauvericine, and fusaproliferin were also found
`
`in Fusarium infected cereal ears.
`[0015]
`Boronic acids, such as phenylboronic acids, have
`been known to inhibit acid lipase. This property of phenyl
`boronic acids has been exploited for disrupting the epithelial
`barrier function to enhance penetration of topically applied
`active ingredients, as disclosed in Thronfeldt et al., U.S.
`Patent No. 6,190,894.
`
`Boric acid and certain phenyl boronic acids are also
`[0016]
`inhibitors of certain beta-lactamases. Shoichet et al., in
`U.S. Patent Nos. 6,075,014 and 6,184,363, disclose that a
`number of phenyl boronic acids are effective against bacteria
`resistant to beta-lactam antibiotics as a result of porin
`mutations. These compounds, or pharmaceutically acceptable
`
`salts, are antibacterial by themselves, although at higher
`
`
`
`WO 03/009689
`
`PCT/US02/23252
`
`5
`
`It is believed
`concentrations than beta-lactam antibiotics.
`that this antibacterial activity is due to the binding of the
`inhibitors to penicillin binding proteins (PBPs), which
`resemble beta-lactamases. Since PBPs are found in all
`bacterial species susceptible to beta-lactam antibiotics, it
`is expected that these compounds will be effective against the
`same bacteria as the beta-lactam antibiotics.
`[0017]
`To the best of the present inventors= knowledge,
`there is no prior art describing any antifungal activity of
`PBA or its derivatives.
`Summary of the Invention
`[0018]
`It is an object of the present invention to overcome
`the deficiencies in the prior art.
`[0019]
`It is another object of the present invention to
`provide a method for treating onychomycosis.
`[0020]
`It is still another object of the present invention
`to provide methods for treating fungal infections in animals
`and plants.
`[0021]
`It is another object of the present invention to
`provide a composition which is a substrate for cytochrome
`p450.
`It has now been discovered that phenyl boronic acid
`[0022]
`and derivatives thereof as well as related boronic acid
`compounds have fungicidal properties, and that these compounds
`are particularly useful in treating fungal infections. These
`compounds have been found to be particularly useful in
`treating nail fungal infections.
`[0023]
`It is known that phenyl boronic acid and derivatives
`
`thereof have inhibitory activity toward proteases and
`lactamases, and it has been reported that it is a substrate
`
`for cytochrome p,450. (cf. Koehler et al., ( 1971) ; Koehler et
`(1974); Lindquist et al., (1974); and Matthews et al.,
`(1975)).
`[0024]
`
`It is believed that the substrate effect for
`cytochrome p450, in combination with its water solubility
`
`
`
`WO 03/009689
`
`PCT/US02/23252
`
`6
`
`properties, permits phenyl boronic acid or derivatives thereof
`to enter a cell and to be degraded by cytochrome p450 to a
`toxic phenyl compound, which then kills the cell. However,
`this is merely a hypothesis, and the present invention is not
`bound by this probable mechanism.
`[0025]
`The compounds which are useful for treating fungal
`infections have the formula
`
`(OH) 2-B-R
`
`(I)
`
`wherein:
`
`R is substituted or unsubstituted phenyl,
`naphthalene, phenanthrene, or has one of the following
`formula·s: .
`
`(2)
`
`(3)
`
`(4).
`
`
`
`W0 03/009689
`WO 03/009689
`
`PCT/US02/23252
`PCT/US02/23252
`
`7
`7
`
`(5}
`(5)
`
`(6)
`(5)
`
`(1)
`(7)
`
`(8)
`(3) .
`
`,
`
`T1
`6\/
`
`1\
`
`r/
`R1
`
`“I
`
`R1
`
`L
`R1”"“4<
`A
`
`1
`
`T1
`/2
`R1'“i\
`
`3%
`
`R1
`
`E
`
`\4'/
`|
`
`7/
`
`R1
`
`R1
`
`R1
`
`T1
`R1\2/3\/R1
`[
`NR
`
`1
`
`//6\
`7
`
`R1
`
`|R
`
`1
`
`R1
`.\
`* /1
`\\8
`/
`R1
`
`T‘
`R1\/3
`
`T‘
`4\s/R1
`
`R?‘
`\*-*3
`/
`
`\
`
`R1
`
`R1
`
`
`
`W0 03/009689
`WO 03/009689
`
`PCT/US02/23252
`PCT/US02/23252
`
`8
`8
`
`(9)
`
`(9)
`
`(10)
`(10)
`
`(11)
`(11)
`
`(12)
`(12)
`
`(13)
`(13)
`
`R
`
`If]
`1\\l2/3
`R1/1
`
`T1
`4\s/R1I
`6\R1
`
`X
`
`7\R1
`
`1|’-1
`2
`
`I
`R1
`
`T1
`°\/R1
`3
`/N1
`
`I
`R1
`
`_
`
`RI
`
`1
`
`___,.<1
`1
`R1
`
`O
`
`X
`
`0
`
`\.*l \ N\N
`
`R1
`
`R1
`
`._.._.=o=/ \ > R1
`
`R1
`
`N:-N’“"R7
`
`R1
`/\2
`R1"‘1\
`/3
`R1
`
`0
`
`R1
`
`I
`30
`
`I
`
`R1
`
`R
`I
`4\5Z I
`-I
`,/6\
`'
`
`
`R1
`
`
`
`WO 03/009689
`
`9
`
`PCT/US02/23252
`
`(14)
`
`wherein:
`
`(10),
`
`(4), (5), (6), (7), (8), (9),
`ring system (2),
`(13) or (14) is aromatic or nonaromatic;
`the atom center * is (R) or (SY in the case of
`chiral compounds;
`positions 1, 2, 3, 4, 5, 6, 7 and 8 each
`independently is C, N, O or S;
`R1 through Rs each independently is a lone pair, H,
`B (OH) 2, a halogen atom, CF3, CH2CF3, CCl3, CH 2CC13, CBr3, CH2CBr3,
`N02,
`lower alkyl, C02H, CHCHCOOH, CH2CH2CH2COOH, S03H, P03H,
`OS03H, OP03H, OH, NH2 , CONH2, COCH3, OCH3, or phenyl boronic
`acid.
`
`R7 is H, CF3, CCl3, CBr3, CH2CF3, CH2CCl3, CH2CBr3, N02,
`COCH3, OCH3,
`lower alkyl, cyclic alkene, cyclic alkene
`substituted with one or more substituents Ra, heterocyclic
`alkene, or heterocyclic alkene substituted with one or more
`substituent Ra;
`each Ra is independently H, B(OH) 2, a halogen atom,
`CF3, CCL3, CBr3, CH2CF3, CH2CCl3, CH2CBr3, N02,
`lower alkyl, OH,
`NH2, N(CH3)2, N(CH3)CH2CH3, NHCOCH3, COOR, CHCHCOOH,
`CH2CH2CH2COOH, COCH3, OCH3, phenyl boronic acid, CONH2,
`CONHCH2COOH, CONHCH2CONH2 1 CONHCH2CONHCH2R10 , S02NH2,
`S02NHCH2COOH, S02NHCH2CONH2, or S02NHCH2CONHCH2R10;
`X is O, NH, NCH3 or
`
`~Y;
`
`0
`
`
`
`WO 03/009689
`
`PCT/US02/23252
`
`10
`
`Y is OH, NH2, NCH3, N (CH3) 2r NHCOCH3 or NHCOCH2COOH;
`Rs is H, a halogen atom, CH3, CC1 3, CBr3, CH2CF3,
`CH2CCl3, CH2CBr3, N02, C02H 1 CHCHCOOH, CH2CH2CH2COOH, S03H, P03H,
`OS03H, OP03H, OH, NH2, CONH2, COCH3, OCH3, phenyl boronic acid,
`lower alkyl, or a side chain of a standard amino acid; and
`Rio is a side chain of a standard amino acid.
`In formula (a) above, the following terms have the
`following meanings:
`A Alone pair@ refers to an unshared pair of
`electrons (not involved in an actual covalent chemical bond to
`another atom) that may have important interactions in
`
`[0026]
`
`receptor-ligand (e.g., enzyme-inhibitor) complexes.
`AAlkyl@ means a straight-chain or branched-chain
`alkyl containing 1-25 carbon atoms . . ALower alkyl@ means a
`straight-chain or branched-chain alkyl containing 1-4 carbon
`atoms. Both of these terms include the R and S isomers.
`Acyclic alkene@ means a structure containing from 1
`to 3 rings, each ring containing from 5 to 6 carbon atoms and
`at least one double bond. One, two, or all three of the rings
`may be aromatic.
`AHeterocyclic alkene@ means a cyclic alkene as
`defined above wherein at least one.of the ring(s) contains one
`or more atoms of S, N, or 0.
`The Astandard amino acids@ are alanine, arginine,
`asparagine, aspartic acid, cysteine, glutamic acid, glutamine,
`glycine, histidine, homoserine, hydroxyproline, isoleucine,
`leucine, lysine, methionine, norleucine, norvaline, ornithine,
`glutamic acid, serine, threonine, tryptophan, tyrosine, and
`valine. Both the D and L isomers can be used. The side
`chains of these amino acids are well known and are the
`portions of the amino acids attached to the NH 2-CH2-COOH
`backbone. For instance, the side chain of alanine is CH3 , and
`the side chain or asparagine is CH2CONH2.
`The most useful of these compounds for topical
`[0027]
`
`
`
`WO 03/009689
`
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`
`11
`
`antifungal activity are those which are the most water
`soluble.
`[0028]
`Phenyl boronic acid is a commercially available
`synthetic organic compound which has previously been used for
`complex formation with diols, sugars,· and nucleotides (Singhal
`et al., Advances on Chromatography 31:293-335, Marcel Dekker,
`NY, 1990) or as a synthetic reagent for Suzuki ether synthesis
`
`(Theil, Angew. Chemr. Int. Ed. 38:2345-2347 1999). Phenyl
`boronic acid has inhibitory activity towards a series of
`
`proteases and lactamases (Philipp et al; Proceedings of the
`
`Natl. Acad. Sci. USAr 68:1971, (1976). Phenyl boronic acid
`(PBA) is reasonably soluble in water as well as in several
`organic solvents. Many of its derivatives· are also water
`soluble. Data on its toxicity are incomplete. PBA is
`considered harmful if swallowed (ORL-RAT LDso: 740 mg/kg).
`[0029]
`Additional methods for synthesizing phenyl boronic
`acids according to the present invention are provided in
`Shoichet et al., U.S. Patent No. 6,184,363, the entire
`contents of which are hereby incorporated.
`[0030]
`The water-soluble PBA or derivatives thereof are
`administered topically in the form of a buffered solution,
`lotion, or ointment. The compounds are effective over a wide
`pH range, although a pH of from about 6.0 to about 9.0 is
`preferred. Generally, the compositions are applied topically
`once daily until cure.
`
`[0031]
`
`An in vitro test was designed for testing the
`
`fungicidal or fungistatic activity of PBA and its water(cid:173)
`
`soluble derivatives. Cultures of T. rubrum were grown on 100
`
`microliters SDB agar in a standard 96 well plate. Following
`an initial growth period of about 24 hours, each well was
`treated with 50-100 microliters of the reagent tested.
`Following an appropriate incubation period with or without
`washing, residual viability was estimated from comparison with
`appropriate controls, followed by secondary verification of
`
`viability tests by transfer of a part of the treated culture
`
`
`
`WO 03/009689
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`
`12
`
`into a fresh well for optional recovery and growth.
`[0032]
`The Figure shows the results of this assay. The
`photograph was taken four days after treatment. A photograph
`taken eight days after treatment was very similar.
`
`The effect of PBA and derivatives on T. rubrum as
`[0033]
`shown in the figure is as follows:
`A: control, no treatment
`B: 5 mg/ml PBA
`C: 5 mg/mk 3-amino PBA
`D: 3 mg/ml 3-nitro PBA
`5 mg/ml pentafluoro-PBA
`E.
`F-H: Controls, no treatment
`It can readily be seen from the above that PBA
`
`[0034]
`
`exhibited fungicidal effects on T. rubrum within the
`concentration range of 5-10 mg/ml tested. These solutions
`also exhibited a fungistatic effect on C. parapsylosis at 5
`mg/ml, and a fungicidal effect at 10 mg/ml. Furthermore, a
`solution of about 5-10 mg/ml had an antibiotic activity
`
`against Bacillus megaterium.
`[0035]
`Three water-soluble derivatives of PBA were also
`tested. These water soluble derivatives exhibited similar
`activities.
`All of the recorded effects were effective within a
`[0036]
`wide pH range.
`The following table shows the fungicidal and
`[0037]
`inhibitory effect of PBA and three derivatives thereof, 3-
`nitro PBA, 3-amino PBA, and pentafluoro PBA at varying
`
`concentrations against a variety of bacteria and fungi.
`should be noted that none of these compounds was either
`
`It
`
`fungicidal or inhibitory against Saccharmoyces cerevisiae.
`The pharmacologically active compounds of this invention can
`be processed in accordance with conventional methods of
`pharmacy to produce medicinal agents for administration to
`patients, e.g., mammals including human beings. For example,
`
`the compounds of formula (I) can be employed in admixtures
`
`
`
`WO 03/009689
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`
`13
`
`with conventional excipients, e.g., pharmaceutically
`acceptable carrier substances suitable for topical application
`which do not deleteriously react with the active compounds.
`
`
`
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`
`14
`
`PBA
`
`(Phenylboronic Acid}
`
`3-Ni!roPBA
`
`3~AminoPBA
`
`Pentafluro
`
`PBA
`
`0.04M
`
`0.06M
`
`0.08M
`
`0.03M
`
`0.03M
`
`0.024M
`
`Trichophyton
`
`rub rum
`
`Candida
`
`parapsilosis
`
`F
`
`IN
`
`F
`
`F
`
`F
`
`F
`
`Saccharomyces
`
`NE
`
`NE
`
`NE
`
`IN
`
`IN
`
`IN
`
`F
`
`F
`
`F
`
`F
`IN (0.02m)
`
`NE
`
`NE
`
`-
`
`-
`
`IN
`
`F
`
`-
`
`-
`
`NE
`
`NE
`
`-
`
`-
`
`NE
`
`NE
`
`cerevisiae
`
`Botrytis cinerea
`
`(race B0-510)
`
`Cochliobolus
`
`heterostrophus
`
`(race C4)
`
`Baci/us
`
`megaterium
`
`F
`
`F
`
`F
`
`IN/F
`
`NE
`
`NE
`
`F=Fung1c1dal; IN=Inh1b1tory; NE=No effect
`
`T. rubrum, C. parapsilosis: Onychomycosis
`B. cinerea, C. heterostrophus: Plant pathogens
`
`S. cerevisiae: Control (Baker=s yeast)
`
`B. megaterium: Control (Cyt P-450 containing bacteria)
`
`[0038]
`Suitable pharmaceutically acceptable carriers
`include but are not limited to water, salt solutions,
`
`alcohols, gum arabic, vegetable oils (e.g., corn oil,
`cottonseed oil, peanut oil, olive oil, coconut oil), fish
`liver oils, oily esters such as Polysorbate 80,
`polyethylene glycols, gelatine, carbohydrates (e.g., lactose,
`
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`amylose or starch), magnesium stearate, talc, silicic acid,
`viscous paraffin, fatty acid monoglycerides and diglycerides,
`pentaerythritol fatty acid esters, hydroxy methylcellulose,
`polyvinyl pyrrolidone, etc.
`The pharmaceuticai preparations can be sterilized
`[0039]
`and, if desired, be mixed with auxiliary agents, e.g.,
`lubricants, preservatives, stabilizers, wetting agents,
`emulsifiers, salts for influencing osmotic pressure, buffers,
`coloring, and/or one or more other active compounds, for
`example, other antifungal agents, etc.
`For treating humans and other animals, the
`[0040]
`compositions are applied topically. For treating plants, the
`compositions can be applied, formulated or unformulated,
`directly to the foliage of a plant, to seeds or to other
`medium in which plants are growing or are to be planted. They
`can be sprayed on, dusted on or applied as a cream or paste
`formulation; or they can be applied as a vapor or as
`controlled-release granules.
`Application can be to any part of the plant
`[0041]
`including the foliage, stems, branches or roots, or to soil
`surrounding the roots, or to the seed before it is planted; or
`to the soil generally, or to hydroponic culture systems. The
`invention compounds may also be injected into plants or
`sprayed onto vegetation using electrodynamic spraying
`techniques or other low volume methods.
`The term "plant" as used herein includes seedlings,
`[0042]
`bushes and trees. Furthermore, the fungicidal method of the
`invention includes preventative, protectant, prophylactic and
`eradicant treatment.
`The compounds are preferably used for agricultural
`[0043l
`and horticultural purposes in the form of a composition. The
`type of composition used in any instance will depend upon the
`particular purpose envisaged.
`The compositions may be in the form of dustable
`[0044]
`powders or granules comprising the active ingredient
`
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`(invention compound) and a solid diluent or carrier, for
`example fillers such as kaolin, bentonite, kieselguhr,
`dolomite, calcium carbonate, talc, powdered magnesia, Fuller's
`earth, gypsum, diatomaceous earth and China clay. Such
`granules can be preformed granules suitable for application to
`the soil without further treatment. These granules can be
`made either by impregnating pellets of filler with the active
`ingredient or by pelleting a mixture of the active ingredient
`and powdered filler. Compositions for dressing seed may
`include an agent (for example a mineral oil) for assisting the
`adhesion of the composition to the seed; alternatively the
`active ingredient can be formulated for seed dressing purposes
`using an organic solvent (for example N-methylpyrrolidone,
`propylene glycol or dimethylformamide). The compositions may
`also be in the form of wettable powders or water dispersible
`granules comprising wetting or dispersing agents to facilitate
`their dispersion in liquids. The powders and granules may
`also contain fillers and suspending agents.
`[0045]
`Suspension concentrates of largely insoluble solids
`may be prepared by ball or bead milling with a dispersing
`agent and including a suspending agent to stop the solid
`settling.
`Compositions to be used as sprays may be in the form
`[0046]
`of aerosols wherein the formulation is held in a container
`under pressure in the presence of a propellant, e.g.,
`fluorotrichloromethane or dichlorodifluoromethane.
`[0047]
`The invention compounds can be mixed in the dry
`state with a pyrotechnic mixture to form a composition
`suitable for generating in enclosed spaces a smoke containing
`the compounds. Alternatively, the compounds may be used in a
`microencapsulated form.
`[0048]
`They may also be formulated in biodegradable
`polymeric formulations to obtain a slow, controlled release of
`the active substance.
`[0049]
`
`By including suitable additives, for example
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`additives for improving the distribution, adhesive power and
`resistance to rain on treated surfaces, the different
`compositions can be better adapted for various utilities. The
`dosage administered depends upon the age, health, and weight
`of the recipient, nature of concurrent treatment, if any, and
`the nature of the effect desired.
`[0050]
`Compositions within the scope of the present
`invention include all compositions wherein the active
`ingredient is contained in an amount effective to achieve its
`intended purpose. While individual needs vary, determination
`of optimal ranges of effective amounts of each compound is
`within the skill of the art.
`[0051]
`Pharmaceutical compositions for administering the
`active ingredients of the present invention preferably
`contain, in addition to the pharmacologically active compound,
`suitable pharmaceutically acceptable carriers comprising
`excipients and auxiliaries which facilitate processing of the
`active compounds into preparations which can be used
`pharmaceutically. Preferably, the preparations, contain from
`about 0.01 to about 99 percent by weight, preferably from
`about 20 to 75 percent by weight, active compound(s), together
`with the excipients. For purposes of the present invention,
`all percentages are by weight unless otherwise indicated.
`In
`addition to the following described pharmaceutical
`composition, the compounds of the present invention can be
`formulated as inclusion complexes, such as cyclodextrin
`inclusion complexes.
`[0052]
`The pharmaceutically acceptable carriers include
`vehicles, adjuvants, excipients, or diluents that are well
`known to those skilled in the art and which are readily
`available.
`It is preferred that the pharmaceutically
`acceptable carrier be one which is chemically inert to the
`active compounds and which has no detrimental side effects or
`toxicity under the conditions of use.
`[0053]
`The choice of carrier is determined partly by the
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`particular active ingredient, as well as by the particular
`method used to administer the composition. Accordingly, there
`is a wide variety of suitable formulations of the
`pharmaceutical compositions of the present invention.
`Generally, formulations are prepared for topical or mucosal
`administration.
`[0054]
`Any number of assays well known in the art may be
`used to test whether a particular compound suspected of being
`a fungicide, can be used. These assays are conventional and
`can be readily adapted to the compounds of the present
`invention by one skilled in the art without undue
`experimentation. Examples of assays for fungicidal activity
`on plants can be found in Wagner et al., U.S. Patent No.
`6,262,091, and Schelberger et al., U.S. Patent No. 6,258,801,
`the entire contents of which are hereby incorporated by
`reference. Examples of assays for fungicidal activity on skin
`infections can be found in Coury et al., U.S. Patent No.
`6,261,544, the entire contents of which are hereby
`incorporated by reference.
`[0055]
`In determining the dosages of the PEA or derivative
`thereof to be administered, the dosage and frequency of
`administration is selected in relation to the pharmacological
`properties of the specific active ingredients. Normally, at
`least three dosage levels should be used.
`In toxicity studies
`in general, the highest dose should reach a toxic level but be
`sublethal for most animals in the group.
`If possible, the
`lowest dose should induce a biologically demonstrable effect.
`These studies should be performed in parallel for each·,
`compound selected.
`[0056]
`When a suitable and presumably safe dosage level has
`been established as outlined above, studies on the drug=s
`chronic toxicity, its effect on reproduction, and potential
`mutagenicity may also be required in order to ensure that the
`calculated appropriate dosage range will be safe, also with
`regard to these hazards.
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`
`[0057]
`
`Pharmacological animal studies on pharmacokinetics
`
`revealing, e.g., absorption, distribution, biotransformation,
`and excretion of the active ingredient and metabolites are
`then performed. Using the results obtained, studies o~ human
`pharmacology are then designed. Studies of the
`pharmacodynamics and pharmacokinetics of the compounds in
`humans should be performed in healthy subjects using the
`routes of administration intended for clinical use, and can be
`repeated in patients. The dose-response relationship when
`different doses are given, or when several types of conjugates
`or combinations of conjugates and free compounds are given,
`should be studied in order to elucidate the dose-response
`relationship, the therapeutic range, and the optimum dose
`interval.
`The compounds of the present invention are then
`[0058]
`ready for clinical trials to compare the efficacy of the
`compounds to existing therapy. A dose-response relationship
`to therapeutic effect and for side effects can be more finely
`established at this point.
`[0059]
`The amount of compounds of the present invention to
`be administered to any given patient must be determined
`empirically, and will differ depending upon the condition of
`the patients. Relatively small amounts of the active
`ingredient can be administered at first, with steadily
`increasing dosages if no adverse effects are noted. Of
`course, the maximum safe toxicity dosage as determined in
`routine animal toxicity tests should never be exceeded.
`[ 00 60]
`Compositions for use as aqueous dispersions or
`emulsions are generally supplied in the form of a concentrate
`containing a high proportion of the active ingredient, the
`concentrate being diluted with water before use. These
`concentrates should preferably be able to withstand storage
`for prolonged periods and after such storage be capable of
`dilution with water in order to form aqueous preparations
`
`which remain homogeneous for a sufficient time to enable them
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`to be applied by conventional spray equipment. The .
`concentrates may conveniently contain up to 95%, typically
`10-85%, and preferably 25-60%, by weight of the active
`ingredient. After dilution to form aqueous preparations, such
`preparations may contain varying amounts of the active
`ingredient depending upon the intended purpose, but an aqueous
`preparation containing 0.0005% or 0.01% to 10% by weight
`of active ingredient may be used.
`[0061]
`The compositions of this invention