PSEUDOMYCIN PRODRUGS

Abstract: A pseudomycin prodrug represented by structure (A) where R1 is an acyloxyalkylcarbamate linkage is described. The prodrug demonstrates antifungal activity with less adverse side effects.

Description: PSEUDOMYCIN PRODRUGS FIELD OF THE INVENTION The present invention relates to pseudomycin compounds, in particular, prodrugs of pseudomycin compounds. BACKGROUND OF THE INVENTION Pseudomycins are natural products isolated from liquid cultures of Pseudo onas syringae (plant-associated bacterium) and have been shown to have antifungal activities. (see i.e., Harrison, L . , et al . , "Pseudomycins, a family of novel peptides from Pseudomonas syringae possessing broad-spectrum antifungal activity, " J . Gen . Microbiology, 137(12), 2857-65 (1991) and US Patent Nos. 5,576,298 and 5,837,685) Unlike the previously described antimycotics from P. syringae (e.g., syringomycins, syringotoxins and syringostatins) , pseudomycins A-C contain hydroxyaspartic acid, aspartic acid, serine, dehydroaminobutyric acid, lysine and diaminobutyric acid. The peptide moiety for pseudomycins A, A', B, B' , C, C corresponds to L-Ser-D-Dab-L-Asp-L-Lys-L-Dab-L-aThr-Z-Dhb-L- Asp (3-OH) -L-Thr (4-C1) with the terminal carboxyl group closing a macrocyclic ring on the OH group of the N-terminal Ser. The analogs are distinguished by the N-acyl side chain, i.e., pseudomycin A is N-acylated by 3 , 4-dihydroxytetradeconoyl, pseudomycin A' by 3 , 4-dihydroxypentadecanoyl , pseudomycin B by 3-hydroxytetradecanoyl, pseudomycin B' by 3-hydroxydodecanoyl , pseudomycin C by 3 , 4-dihydroxyhexadecanoyl and pseudomycin C by 3-hydroxyhexadecanoyl . (see i.e., Ballio, A., et al . , "Novel bioactive lipodepsipeptides from Pseudomonas syringae : the pseudomycins," FEBS Letters, 355(1), 96-100, (1994) and Coiro, V.M. , et al . , "Solution conformation of the Pseudomonas syringae MSU 16H phytotoxic lipodepsipeptide Pseudomycin A determined by computer simulations using distance geometry and molecular dynamics from NMR data, " Eur. J. Biochem., 257(2), 449-456 (1998).) Pseudomycins are known to have certain adverse biological effects. For example, destruction of the endothelium of the vein, destruction of tissue, inflammation, and local toxicity to host tissues have been observed when pseudomycin is administered intraveneously . Therefore, there is a need to identify compounds within this class that are useful for treating fungal infections without the currently observed adverse side effects. BRIEF SUMMARY OF THE INVENTION The present invention provides a pseudomycin prodrug represented by the following structure which is useful as an antifungal agent. wherein R is where R a and R a' are independently hydrogen or methyl, or either R a or R a' is alkyl amino, taken together with R b or R b' forms a six-membered cycloalkyl ring, a six- membered aromatic ring or a double bond, or taken together with R c forms a six-membered aromatic ring; R b and R b' are independently hydrogen, halogen, or methyl, or either R b or R b' is amino, alkylamino, oc- acetoacetate, methoxy, or hydroxy; R c is hydrogen, hydroxy, C1-C 4 alkoxy, hydroxy Ci- C 4 alkoxy, or taken together with R e forms a 6-membered aromatic ring or C 5 -C 6 cycloalkyl ring; R e is hydrogen, or taken together with R f is a six-membered aromatic ring, C 5 -C 14 alkoxy substituted six-membered aromatic ring, or C 5 -C 1 4 alkyl substituted six-membered aromatic ring, and R f is Cg-Cis alkyl, or C 5 -C 11 alkoxy; R is where R 9 is hydrogen, or Cι-Cι 3 alkyl, and R h is C 1 -C1 5 alkyl, C 4 -C1 5 alkoxy, (C 1 -C 10 alkyDphenyl, - (CH 2 ) n -aryl, or - (CH 2 ) n - (C 5 -C 6 cycloalkyl), where n = 1 or 2 ; or R is where R 1 is a hydrogen, halogen, or Cs-C 8 alkoxy, and m is 1, 2 or 3 ; R is where R J is C5-C14 alkoxy or C 5 -C 14 alkyl, and p = 0, 1 or 2; R is where R k is C 5 -Ci 4 alkoxy; or R is - (CH 2 )-NR m -(Cι 3 -C 18 alkyl), where R m is H, -CH 3 or -C(0)CH 3 ; R 1 is independently hydrogen, an acyloxymethylene-1, 3- dioxolen-2-one (e.g., compounds 1 (a) depicted below), or an acyloxymethylenecarboxylate (e.g., compounds 1 (b) depicted below) 1(a) Kb) where R la is hydrogen, Ci-Cio alkyl, Ci-Cio alkenyl , benzyl, or aryl and R lb is hydrogen or methyl provided that at least one R 1 is an acyloxymethylene- , 3-dioxolen-2-one or an acyloxymethylenecarboxylate; and R 3 are independently -OR 2a , or -N(R 2b ) (R 2c ) , where R 2a and R 2b are independently hydrogen, Cι-Cι 0 alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, i- butyl, s-butyl, t-butyl, etc.), C 3 _C δ cycloalkyl (e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclopentylmethylene, methylcyclopentyl, cyclohexyl, etc.) hydroxy (Ci-Cio) alkyl, alkoxy (Cι-Cι 0 ) alkyl (e.g, methoxyethyl) , or C 2 -Cιo alkenyl, amino (Ci-Cio) alkyl, mono- or di-alkyla ino (Ci-Cio) alkyl, aryl (Ci-Cio) alkyl (e.g., benzyl), heteroaryl (Ci-Cio) alkyl (e.g., 3- pyridyl ethyl, 4-pyridylmethyl) , or cycloheteroalkyl (Ci-Cio) alkyl (e.g., N-tetrahydro-1, 4- oxazinylethyl and N-piperazinylethyl) , or R 2b is an alkyl carboxylate residue of an aminoacid alkyl ester (e.g., -CHC0 2 CH 3 , -CH(C0 2 CH 3 )CH(CH 3 ) 2/ -CH (C0 2 CH 3 ) CH (phenyl) , -CH(C0 2 CH 3 )CH 2 OH, -CH (C0 2 CH 3 ) CH 2 (p-hydroxyphenyl) , -CH (CO2CH 3 ) CH 2 SH, -CH (C0 2 CH 3 ) CH 2 (CH 2 ) 3 NH 2 , -CH(C0 2 CH 3 )CH 2 (4- or 5-imidazole) , -CH (C0 2 CH 3 ) CH 2 C0 2 CH 3 , -CH(C0 2 CH 3 )CH 2 Cθ2NH2, and the like), and R 2c is hydrogen or Ci-C δ alkyl; and pharmaceutically acceptable salts and solvates thereof. In another embodiment of the present invention, a pharmaceutical formulation is provided which includes the pseudomycin prodrug described above and a pharmaceutically acceptable carrier. In yet another embodiment of the present invention, a method is provided for treating an antifungal infection in an animal in need thereof, which comprises administering to the animal the pseudomycin prodrug described above . The use of the pseudomycin prodrug described above in the manufacture of a medicament for use in treating an antifungal infection in an animal is also provided. Definitions As used herein, the term "alkyl" refers to a hydrocarbon radical of the general formula C n H 2 n+ι containing from 1 to 30 carbon atoms unless otherwise indicated. The alkane radical may be straight (e.g. methyl, ethyl, propyl, butyl, etc.), branched (e.g., isopropyl, isobutyl, tertiary butyl, neopentyl, etc.), cyclic (e.g., cyclopropyl, cyclobutyl, eye1opentyl, methylcyclopentyl, cyclohexyl, etc.), or multi-cyclic (e.g., bicyclo [2.2.1] heptane, spiro [2.2 ]pentane, etc.). The alkane radical may be substituted or unsubstituted. Similarly, the alkyl portion of an alkoxy group, alkanoyl, or alkanoate have the same definition as above. The term "alkenyl" refers to an acyclic hydrocarbon containing at least one carbon carbon double bond. The alkene radical may be straight, branched, cyclic, or multi- cyclic. The alkene radical may be substituted or unsubstituted. The alkenyl portion of an alkenoxy, alkenoyl or alkenoate group has the same definition as above. The term "aryl" refers to aromatic moieties having single (e.g., phenyl) or fused ring systems (e.g., naphthalene, anthracene, phenanthrene, etc.). The aryl groups may be substituted or unsubstituted. Within the field of organic chemistry and particularly within the field of organic biochemistry, it is widely understood that significant substitution of compounds is tolerated or even useful. In the present invention, for example, the term alkyl group allows for substitutents which is a classic alkyl, such as methyl, ethyl, propyl, hexyl, isooctyl, dodecyl, stearyl, etc. The term "group" specifically envisions and allows for substitutions on alkyls which are common in the art, such as hydroxy, halogen, alkoxy, carbonyl, keto, ester, carbamato, etc., as well as including the unsubstituted alkyl moiety. However, it is generally understood by those skilled in the art that the substituents should be selected so as to not adversely affect the pharmacological characteristics of the compound or adversely interfere with the use of the medicament. Suitable substituents for any of the groups defined above include alkyl, alkenyl, alkynyl, aryl, halo, hydroxy, alkoxy, aryloxy, mercapto, alkylthio, arylthio, mono- and di-alkyl amino, quaternary ammonium salts, aminoalkoxy, hydroxyalkylamino, a inoalkylthio, carbamyl, carbonyl, carboxy, glycolyl, glycyl, hydrazino, guanyl, and combinations thereof . The term "prodrug" refers to a class of drugs which result in pharmacological action due to conversion by metabolic processes within the body (i.e., biotransformation) . In the present invention, the pseudomycin prodrug compounds contain linkers that can be cleaved by esterases in the plasma to produce the active drug. The term "animal" refers to humans, companion animals (e.g., dogs, cats and horses), food-source animals (e.g., cows, pigs, sheep and poultry), zoo animals, marine animals, birds and other similar animal species. DETAILED DESCRIPTION OF THE INVENTION Applicants have discovered that a prodrug derivative of the pseudomycin natural or semi-synthetic products provide less adverse side effects than the corresponding natural products and maintains in vivo efficacy against C. albican, C. neoformans, and A . fumigatus . The prodrug is produced by acylating at least one of the pendant amino groups attached to the lysine or 2 , 4-diaminobutyric acid peptide units in the pseudomycin cyclopeptide ring system to. form an acyl substituent (s) . The acylating agent (or linker) is generally a acyloxymethylene-1, 3-dioxolen-2-one or acyloxymethylenecarboxylate acylating compound containing a suitable leaving group such that a carbamate linkage with the pendant amino group on the pseudomycin structure can be formed. Suitable leaving groups are well known to those skilled in the art and include groups such as p-nitrophenoxy and N-oxysuccinimide . An acyloxymethylene-1, 3-dioxolen-2-one acylating compound may be synthesized using the synthetic route shown in scheme I below. For illustrative purposes, a specific acylating compound is depicted. However, it will be understood by those skilled in the art that one could synthesize a variety of derivatives using the same basic synthetic method. Scheme I For a more detailed description of the synthetic procedures, see the preparation section of the Examples below. The acyloxymethylenecarboxylate acylating compound may be synthesized using the synthetic route shown in scheme II below. For illustrative purposes, a specific acylating compound is depicted. However, it will be understood by those skilled in the art that one could synthesize a variety of derivatives using the same basic synthetic method. Scheme II For a more detailed description of the synthetic procedures, see the preparation section of the Examples below. As discussed earlier, pseudomycins are natural products isolated from the bacterium Pseudomonas syringae that have been characterized as lipodepsinonapetpides containing a cyclic peptide portion closed by a lactone bond and including the unusual amino acids 4-chlorothreonine (ClThr) , 3-hydroxyaspartic acid (HOAsp) , 2 , 3-dehydro-2-aminobutyric acid (Dhb) , and 2 , 4-diaminobutyric acid (Dab). Methods for growth of various strains of P. syringae to produce the different pseudomycin analogs (A, A', B, B' , C, and C) are described below and described in more detail in PCT Patent Application Serial No. PCT/US00/08728 filed by Hilton, et al . on April 14, 2000 entitled "Pseudomycin Production by Pseudomonas Syringae, " incorporated herein by reference, PCT Patent Application Serial No. PCT/US00/08727 filed by Kulanthaivel, et al . on April 14, 2000 entitled "Pseudomycin Natural Products," incorporated herein by reference, and U.S. Patent Nos. 5,576,298 and 5,837,685, each of which are incorporated herein by reference. Isolated strains of P. syringae that produce one or more pseudomycins are known in the art. Wild type strain MSU 174 and a mutant of this strain generated by transposon mutagenesis, MSU 16H are described in U.S. Patent Nos. 5,576,298 and 5,837,685; Harrison, et al . , "Pseudomycins, a family of novel peptides from Pseudomonas syringae possessing broad-spectrum antifungal activity, " J . Gen . Microbiology, 137, 2857-2865 (1991); and Lamb et al . , "Transposon mutagenesis and tagging of fluorescent pseudomonas: Antimycotic production is necessary for control of Dutch elm disease," Proc. Natl. Acad. Sci. USA, 84, 6447- 6451 (1987) . A strain of P. syringae that is suitable for production of one or more pseudomycins can be isolated from environmental sources including plants (e.g., barley plants, citrus plants, and lilac plants) as well as, sources such as soil, water, air, and dust. A preferred stain is isolated from plants. Strains of P. syringae that are isolated from environmental sources can be referred to as wild type. As used herein, "wild type" refers to a dominant genotype which naturally occurs in the normal population of P. syringae (e.g., strains or isolates of P. syringae that are found in nature and not produced by laboratory manipulation) . Like most organisms, the characteristics of the pseudomycin- producing cultures employed (P. syringae strains such as MSU 174, MSU 16H, MSU 206, 25-B1, 7H9-1) are subject to variation. Hence, progeny of these strains (e.g., recombinants, mutants and variants) may be obtained by methods known in the art . P. syringae MSU 16H is publicly available from the American Type Culture Collection, Parklawn Drive, Rockville, MD, USA as Accession No. ATCC 67028. P . syringae strains 25-B1, 7H9-1, and 67 Hi were deposited with the American Type Culture Collection on March 23, 2000 and were assigned the following Accession Nos.: 25-B1 Accession No. PTA-1622 7H9-1 Accession No. PTA-1623 67 HI Accession No. PTA-1621 Mutant strains of P. syringae are also suitable for production of one or more pseudomycins. As used herein, "mutant" refers to a sudden heritable change in the phenotype of a strain, which can be spontaneous or induced by known mutagenic agents, such as radiation (e.g., ultraviolet radiation or x-rays), chemical mutagens (e.g., ethyl methanesulfonate (EMS) , diepoxyoctane, N-methyl-N- nitro-N' -nitrosoguanine (NTG) , and nitrous acid), site- specific mutagenesis, and transposon mediated mutagenesis. Pseudomycin-producing mutants of P. syringae can be produced by treating the bacteria with an amount of a mutagenic agent effective to produce mutants that overproduce one or more pseudomycins, that produce one pseudomycin (e.g., pseudomycin B) in excess over other pseudomycins, or that produce one or more pseudomycins under advantageous growth conditions. While the type and amount of mutagenic agent to be used can vary, a preferred method is to serially dilute NTG to levels ranging from 1 to 100 μg/ml. Preferred mutants are those that overproduce pseudomycin B and grow in minimal defined media. Environmental isolates, mutant strains, and other desirable strains of P. syringae can be subjected to selection for desirable traits of growth habit, growth medium nutrient source, carbon source, growth conditions, amino acid requirements, and the like. Preferably, a pseudomycin producing strain of P. syringae is selected for growth on minimal defined medium such as N21 medium and/or for production of one or more pseudomycins at levels greater than about 10 μg/ml. Preferred strains exhibit the characteristic of producing one or more pseudomycins when grown on a medium including three or fewer amino acids and optionally, either a lipid, a potato product or combination thereof . Recombinant strains can be developed by transforming the P . syringae strains, using procedures known in the art. Through the use of recombinant DNA technology, the P. syringae strains can be transformed to express a variety of gene products in addition to the antibiotics these strains produce. For example, one can modify the strains to introduce multiple copies of the endogenous pseudomycin- biosynthesis genes to achieve greater pseudomycin yield. To produce one or more pseudomycins from a wild type or mutant strain of P. syringae, the organism is cultured with agitation in an aqueous nutrient medium including an effective amount of three or fewer amino acids, preferably glutamic acid, glycine, histidine, or a combination thereof. Alternatively, glycine is combined with one or more of a potato product and a lipid. Culturing is conducted under conditions effective for growth of P. syringae and production of the desired pseudomycin or pseudomycins. Effective conditions include temperatures from about 22 2 C to about 27 2 C, and a duration of about 36 hours to about 96 hours. Controlling the concentration of oxygen in the medium during culturing of P. syringae is advantageous for production of a pseudomycin. Preferably, oxygen levels are maintained at about 5 to 50% saturation, more preferably about 30% saturation. Sparging with air, pure oxygen, or gas mixtures including oxygen can regulate the concentration of oxygen in the medium. Controlling the pH of the medium during culturing of P. syringae is also advantageous. Pseudomycins are labile at basic pH, and significant degradation can occur if the pH of the culture medium is above about 6 for more than about 12 hours. Preferably, the pH of the culture medium is maintained between 6 and 4. P. syringae can produce one or more pseudomycins when grown in batch culture. However, fed-bath or semi-continuous feed of glucose and optionally, an acid or base (e.g., ammonium hydroxide) to control pH, enhances production. Pseudomycin production can be further enhanced by using continuous culture methods in which glucose and ammonium hydroxide are fed automatically. Choice of P. syringae strain can affect the amount and distribution of pseudomycin or pseudomycins produced. For example, strains MSU 16H and 67 HI each produce predominantly pseudomycin A, but also produce pseudomycin B and C, typically in ratios of 4:2:1. Strain 67 Hi typically produces levels of pseudomycins about three to five fold larger than are produced by strain MSU 16H. Compared to strains MSU 16H and 67 HI, strain 25-B1 produces more pseudomycin B and less pseudomycin C. Strain 7H9-1 are distinctive in producing predominantly pseudomycin B and larger amount of pseudomycin B than other strains. For example, this strain can produce pseudomycin B in at least a ten fold excess over either pseudomycin A or C . Alternatively, the prodrug can be formed from an N-acyl semi-synthetic compound. Semi-synthetic pseudomycin compounds may be synthesized by exchanging the N-acyl group on the L-serine unit. Examples of various N-acyl derivatives are described in PCT Patent Application Serial No. , Belvo, et al . , filed evendate herewith entitled "Pseudomycin N-Acyl Side-Chain Analogs" and incorporated herein by reference. In general, four synthetic steps are used to produce the semi-synthetic compounds from naturally occurring pseudomycin compounds: (1) selective amino protection; (2) chemical or enzymatic deacylation of the N- acyl side-chain; (3) reacylation with a different side- chain; and (4) deprotection of the amino groups. The pendant amino groups at positions 2, 4 and 5 may be protected using any standard means known to those skilled in the art for amino protection. The exact genus and species of amino protecting group employed is not critical so long as the derivatized amino group is stable to the condition of subsequent reaction (s) on other positions of the intermediate molecule and the protecting group can be selectively removed at the appropriate point without disrupting the remainder of the molecule including any other amino protecting group(s). Suitable amino-protecting groups include benzyloxycarbonyl, p-nitrobenzyloxycarbonyl, p-bromobenzyloxycarbonyl, p-methoxybenxyloxycarbonyl, p-methoxyphenylazobenzyloxycarbonyl, p-phenylazobenzyloxycarbonyl , t-butyloxycarbonyl , cyclopentyloxycarbonyl, and phthalimido. Preferred amino protecting groups are t-butoxycarbonyl (t-Boc), allyloxycarbonyl (Alloc) , phthalimido, and benzyloxycarbonyl (CbZ or CBZ) . Further examples of suitable protecting groups are described in T.W. Greene, "Protective Groups in Organic Synthesis," John Wiley and Sons, New York, N.Y. , (2nd ed., 1991), at chapter 7. The deacylation of a N-acyl group having a gamma or delta hydroxylated side chain (e.g., 3 , 4-dihydroxytetra- deconoate) may be accomplished by treating the amino- protected pseudomycin compound with acid in an aqueous solvent. Suitable acids include acetic acid and trifluoroacetic acid. A preferred acid is trifluoroacetic acid. If trifluoroacetic acid is used, the reaction may be accomplished at or near room temperature. However, when acetic acid is used the reaction is generally ran at about 40°C. Suitable aqueous solvent systems include acetonitrile, water, and mixtures thereof. Organic solvents accelerate the reaction; however, the addition of an organic solvent may lead to other by-products . Pseudomycin compounds lacking a delta or gamma hydroxy group on the side chain (e.g., Pseudomycin B and C) may be deacylated enzymatically . Suitable deacylase enzymes include Polymyxin Acylase (164-16081 Fatty Acylase (crude) or 161-16091 Fatty Acylase (pure) available from Wako Pure Chemical Industries, Ltd.), or ECB deacylase. The enzymatic deacylation may be accomplished using standard deacylation procedures well known to those skilled in the art. For example, general procedures for using polymyxin acylase may be found in Yasuda, N., et al, Agric . Biol . Chem . , 53, 3245 (1989) and Kimura, Y., et al . , Agri c . Biol . Chem . , 53, 497 (1989). The deacylated product (also known as the pseudomycin nucleus) is reacylated using the corresponding acid of the desired acyl group in the presence of a carbonyl activating agent. "Carbonyl activating group" refers to a substituent of a carbonyl that promotes nucleophilic addition reactions at that carbonyl. Suitable activating substituents are those which have a net electron withdrawing effect on the carbonyl. Such groups include, but are not limited to, alkoxy, aryloxy, nitrogen containing aromatic heterocycles, or amino groups (e.g., oxybenzotriazole, imidazolyl, nitrop enoxy, pentachlorophenoxy, N-oxysuccinimide, N,N'- dicyclohexylisoure-O-yl, and N-hydroxy-N-methoxyamino) ; acetates; formates; sulfonates (e.g., methanesulfonate, ethanesulfonate, benzenesulfonate, and p-tolylsulfonate) ; and halides (e.g., chloride, bromide, and iodide). A variety of acids may be used in the acylation process. Suitable acids include aliphatic acids containing one or more pendant aryl, alkyl, amino (including primary, secondary and tertiary amines) , hydroxy, alkoxy, and amido groups; aliphatic acids containing nitrogen or oxygen within the aliphatic chain; aromatic acids substituted with alkyl, hydroxy, alkoxy and/or alkyl amino groups; and heteroaromatic acids substituted with alkyl, hydroxy, alkoxy and/or alkyl amino groups. Alternatively, a solid phase synthesis may be used where a hydroxybenzotriazole-resin (HOBt-resin) serves as the coupling agent for the acylation reaction. Once the amino group is deacylated and reacylated (described above) , then the amino protecting groups (at positions 2, 4 and 5) can be removed by hydrogenation in the presence of a hydrogenation catalyst (e.g., 10% Pd/C) . When the amino protecting group is allyloxycarbonyl, then the protecting group can be removed using tributyltinhydride and triphenylphosphine palladium dichloride. This particular protection/deprotection scheme has the advantage of reducing the potential for hydrogenating the vinyl group of the Z-Dhb unit of the pseudomycin structure. The prodrug is then produced by acylating at least one of the pendant amino groups attached to the lysine or 2,4- diaminobutyric acid peptide units of the N-acyl modified semi-synthetic pseudomycin compound to form the desired carbamate linkage. Other modified prodrug pseudomycin compounds may be synthesized by amidation or esterification of the pendant carboxylic acid group of the aspartic acid and/or hydroxyaspartic acid units of the pseudomycin ring. Examples of various acid-modified derivatives are described in PCT Patent Application Serial No. , Chen, et al . , filed evendate herewith entitled "Pseudomycin Amide & Ester Analogs" and incorporated herein by reference. The acid- modified derivatives may be formed by condensing any of the previously described prodrugs with the appropriate alcohol or amine to produce the respective ester or amide. Formation of the ester groups may be accomplished using standard esterification procedures well-known to those skilled in the art. Esterification under acidic conditions typically includes dissolving or suspending the pseudomycin compound in the appropriate alcohol in the presence of a protic acid (e.g., HCl, TFA, etc.). Under basic conditions, the pseudomycin compound is generally reacted with the appropriate alkyl halide in the presence of a weak base (e.g., sodium bicarbonate and potassium carbonate) . Formation of the amide groups may be accomplished using standard amidation procedures well-known to those skilled in the art. However, the choice of coupling agents provides selective modification of the acid groups. For example, the use of benzotriazol-1-yloxy-tripyrrolidinophosphonium hexafluorophosphate(PyBOP) as the coupling agent allows one to isolate pure mono-amides at residue 8 and (in some cases) pure bis amides simultaneously. Whereas, the use of o- benzotriazol-l-yl-N,N,N' ,N' -tetramethyluronium tetrafluoroborate (TBTU) as the coupling agent favors formation of monoamides at residue 3. The pseudomycin prodrug may be isolated and used per se or in the form of its pharmaceutically acceptable salt or solvate. The prodrug is prepared by forming at least one acyloxyalkylcarbamate linkage as described earlier. The term "pharmaceutically acceptable salt" refers to non-toxic acid addition salts derived from inorganic and organic acids. Suitable salt derivatives include halides, thiocyanates, sulfates, bisulfates, sulfites, bisulfites, arylsulfonates, alkylsulfates, phosphonates, monohydrogen- phosphates, dihydrogenphosphates , metaphosphates, pyrophosphonates, alkanoates, cycloalkylalkanoates, arylalkonates, adipates, alginates, aspartates, benzoates, fumarates , glucoheptanoates, glycerophosphates, lactates, maleates, nicotinates, oxalates, palmitates, pectinates, picrates, pivalates, succinates, tartarates, citrates, camphorates, camphorsulfonates, digluconates, trifluoroacetates, and the like. The term "solvate" refers to an aggregate that comprises one or more molecules of the solute (i.e., pseudomycin prodrug compound) with one or more molecules of a pharmaceutical solvent, such as water, ethanol, and the like. When the solvent is water, then the aggregate is referred to as a hydrate. Solvates are generally formed by dissolving the prodrug in the appropriate solvent with heat and slowing cooling to generate an amorphous or crystalline solvate form. Each pseudomycin, semi-synthetic pseudomycin, pseudomycin prodrug and mixtures can be detected, determined, isolated, and/or purified by any variety of methods known to those skilled in the art. For example, the level of pseudomycin or pseudomycin prodrug activity in a broth or in an isolate or purified composition can be determined by antifungal action against a fungus such as Candida and can be isolated and purified by high performance liquid chromatography . The active ingredient (i.e., pseudomycin prodrug) is typically formulated into pharmaceutical dosage forms to provide an easily controllable dosage of the drug and to give the patient, physician or veterinarian an elegant and easily handleable product. Formulations may comprise from 0.1% to 99.9% by weight of active ingredient, more generally from about 10% to about 30% by weight. As used herein, the term "unit dose" or "unit dosage" refers to physically discrete units that contain a predetermined quantity of active ingredient calculated to produce a desired therapeutic effect. When a unit dose is administered orally or parenterally, it is typically provided in the form of a tablet, capsule, pill, powder packet, topical composition, suppository, wafer, measured units in ampoules or in multidose containers, etc. Alternatively, a unit dose may be administered in the form of a dry or liquid aerosol which may be inhaled or sprayed. The dosage to be administered may vary depending upon the physical characteristics of the animal, the severity of the animal's symptoms, the means used to administer the drug and the animal species. The specific dose for a given animal is usually set by the judgment of the attending physician or veterinarian. Suitable carriers, diluents and excipients are well known to those skilled in the art and include materials such as carbohydrates, waxes, water soluble and/or swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, and the like. The particular carrier, diluent or excipient used will depend upon the means and purpose for which the active ingredient is being applied. The formulations may also include wetting agents, lubricating agents, surfactants, buffers, tonicity agents, bulking agents, stabilizers, emulsifiers, suspending agents, preservatives, sweeteners, perfuming agents, flavoring agents and combinations thereof. A pharmaceutical composition may be administered using a variety of methods. Suitable methods include topical (e.g., ointments or sprays), oral, injection and inhalation. The particular treatment method used will depend upon the type of infection being addressed. In parenteral iv applications, the formulations are typically diluted or reconstituted (if freeze-dried) and further diluted if necessary, prior to administration. An example of reconstitution instructions for the freeze-dried product are to add ten ml of water for injection (WFI)to the vial and gently agitate to dissolve. Typical reconstitution times are less than one minute. The resulting solution is then further diluted in an infusion solution such as dextrose 5% in water (D5W) , prior to administration. Pseudomycin compounds have been shown to exhibit antifungal activity such as growth inhibition of various infectious fungi including Candida spp. (i.e., C. albicans, C. parapsilosis , C. krusei , C. glabrata, C. tropicalis, or C. lusi taniaw) ; Torulopus spp. (i.e., T. glabrata) ; Aspergillus spp. (i.e., A . fumigatus) ; Histoplasma spp. (i.e., H. capsulatum) ; Cryptococcus spp. (i.e., C. neoformans) ; Blastomyces spp. (i.e., B. dermati tidis) ; Fusarium spp.; Trichophyton spp., Pseudallescheria boydii , Coccidioides immi ts, Sporothrix schenckii , etc. Consequently, the compounds and formulations of the present invention are useful in the preparation of medicaments for use in combating either systemic fungal infections or fungal skin infections. Accordingly, a method is provided for inhibiting fungal activity comprising contacting the pseudomycin prodrug of the present invention with a fungus. A preferred method includes inhibiting Candida albicans or Aspergillus fu igatus activity. The term "contacting" includes a union or junction, or apparent touching or mutual tangency of a compound of the invention with a fungus. The term does not imply any further limitations to the process, such as by mechanism of inhibition. The methods are defined to encompass the inhibition of parasitic and fungal activity by the action of the compounds and their inherent antifungal properties. A method for treating a fungal infection which comprises administering an effective amount of a pharmaceutical formulation of the present invention to a host in need of such treatment is also provided. A preferred method includes treating a Candida albicans, Cryptococcus neoformans, or Aspergillus fumigatus infection. The term "effective amount" refers to an amount of active compound which is capable of inhibiting fungal activity. The dose administered will vary depending on such factors as the nature and severity of the infection, the age and general health of the host, the tolerance of the host to the antifungal agent and the species of the host. The particular dose regimen likewise may vary according to these factors. The medicament may be given in a single daily dose or in multiple doses during the day. The regimen may last from about 2-3 days to about 2-3 weeks or longer. A typical daily dose (administered in single or divided doses) contains a dosage level between about 0.01 mg/kg to 100 mg/kg of body weight of an active compound. Preferred daily doses are generally between about 0.1 mg/kg to 60 mg/kg and more preferably between about 2.5 mg/kg to 40 mg/kg. The host is generally an animal including humans, companion animals (e.g., dogs, cats and horses), food-source animals (e.g., cows, pigs, sheep and poultry), zoo animals, marine animals, birds and other similar animal species. EXAMPLES The following abbreviations are used through out the examples to represent the respective listed materials: ACN - acetonitrile TFA - trifluoroacetic acid DMF - dimethylformamide EDCI - 1- [3- (dimethylamino) propyl] -3-ethylcarbodiimide hydrochloride BOC = t-butoxycarbonyl, (CH 3 ) 3 C-0-C (0) - CBZ = benzyloxycarbonyl, C 6 H 5 CH 2 -0-C (0) - PyBOP = benzotriazol-1-yloxy-tripyrrolidinophosphonium hexafluorophosphate TBTU = o-Benzotriazol-l-yl-N,N,N' ,N' -tetramethyluroniu tetrafluoroborate DIEA = N,N-diisopropylethylamine The following structure II will be used to describe the products observed in Examples 1 through 7. II Detection and Quantification of Antifungal Activi ty: Antifungal activity was determined in vi tro by obtaining the minimum inhibitory concentration (MIC) of the compound using a standard agar dilution test or a discdiffusion test. A typical fungus employed in testing antifungal activity is Candida albicans . Antifungal activity is considered significant when the test sample (50 μl) causes 10-12 mm diameter zones of inhibition on C. albicans x657 seeded agar plates. Tail Vein Toxici ty: Mice were treated intravenously (IV) through the lateral tail vein with 0.1 ml of testing compound (20 mg/kg) at 0, 24, 48 and 72 hours. Two mice were included in each group. Compounds were formulated in 5.0% dextrose and sterile water for injection. The mice were monitored for 7 days following the first treatment and observed closely for signs of irritation including erythema, swelling, discoloration, necrosis, tail loss and any other signs of adverse effects indicating toxicity. The mice used in the study were outbred, male ICR mice having an average weight between 18-20 g (available from Harlan Sprangue Dawley, Indianapolis, IN). Prepara tions Preparation of 4-Bromomethyl -5-methyl -l , 3-dioxolene-2-one (la-1 ) : la-1 A mixture of 0.1 mole of 4, 5-Dimethyl-l, 3-dioxolene-2- one, 0.1 mole of N-bromosuccinimide and 0.1 g of 2,2- azobis (2-methylpropionitrile) in 70 ml of carbontetrachloride (CC1 4 ) was heated to reflux. After six hours, the mixture was cooled down with ice and filtered. The filtrate was washed with 2x50 ml of water, 2x50 ml of a sodium chloride solution and an additional 50 ml of water. The solution was dried over sodium sulfate and evaporated to dryness, and dried under vacuum to yield 16.5 g (85% yield) of an oil having 1H-NMR data consistent with structure la-1. Preparation of Compound (la-2) : la-2 Compound la-2 was synthesized using the procedures described in Synthetic Communication, 22(9), 1297 (1992) to yield 11.5 g (78% yield) of a crude oil. Preparation of the Compound la-3 . la-3 A mixture of 11.5 g of Compound la-2 (crude oil), 500 ml of 37% HCl and 300 ml of methanol was allowed to stir overnight at 4 S C. The mixture was then concentrated to form an oil. Purification by column chromatography (1:1 ethylacetate/hexane) yielded 5.27 g (33.8%) of product having 1H-NMR data consistent with structure la-3. Preparation of Compound la-4. la-4 A mixture of 3.0 g of Compound la-3 and 2.02 g of pyridine in 30 ml of chloroform was cooled to 0-4 S C. A solution of 5.08 g of p-nitrophenyl-chloroformate in 30 ml of chloroform was added to the mixture and allowed to stir for about 4.5 hours. The mixture was washed with cooled 1% sodium hydroxide (3x30 ml), IN HCl (2x30 ml), water (2x30 ml) and brine (2x30 ml) . The solution was dried over sodium sulfate, filtered, and washed with dichloromethane. Removal of the solvent yielded an oil which solidified upon standing. The solid was picked up in 10 ml of dichloromethane and hexane was added to form a precipitate. The mixture was filtered, washed with hexane, and dried under vacuum overnight to yield 6.39 g (94% yield) product having 1H-NMR data consistent with structure la-4. Preparation of Compound lb-1 : lb-l Compound lb-1 can be synthesized using the procedures described in Synthesis, 1159 (1990) . Preparation of Compound lb-2. lb-2 A solution of 4.57 g (30 mmol) crude lb-1 in 40 ml of dichloromethane was added at 0 2 C to a solution of 3.91 g (34 mmol) N-hydroxy succinimide and 2.7 g (34 mmol) pyridine in 100 ml of dichloromethane. After stirring at 0 2 C for 30 minutes, the mixture was allowed to stand at room temperature overnight. The mixture was then washed with water four times and the organic phase was dried over sodium sulfate. Upon filtration, the solvent was evaporated to give 4.0 g (58% yield) of an oily crude product having 1H- NMR data consistent with structure lb-2. Preparation of CBZ-Protected Pseudomycin B (2a-l ) : Dissolve/suspend pseudomycin B in DMF (20 mg/ml, Aldrich Sure Seal) . While stirring at room temperature add N- (Benzyloxycarbonyloxy) succinimide (6 eq) . Allow to stir at room temperature for 32 hours. Monitor reaction by HPLC (4.6x50 mm, 3.5 μm, 300-SB, C8, Zorbax column). Concentrate reaction to 10 ml on a high vacuum roto-evaporator at room temperature. Put material in freezer until ready to prep by chromatography. Reverse phase preparative HPLC yields an amorphous, white solid (Compound 2a-l) after lyophilization. Preparation of Compound 2b-l : R 1' , R 1" and R 1'" = H R 2 = -NH(cyclopropyl) R 3 = -OH 2b-l CBZ-protected pseudomycin B (2a-l) (400 mg, 0.25 mmol) is dissolved in 4 ml dry DMF . TBTU (79 mg, 0.25 mmol), DIEA (200 μl, 6 equivalents) and cyclopropylamine (14.2 mg, 0.25 mmol) were added sequentially. The reaction was stirred at room temperature under nitrogen while being monitored by HPLC. Upon completion the reaction was concentrated in vacuo . The crude product purified by preparative HPLC. Lyophilization yielded 209.2 mg (51.1%) of a colorless powder . The 3-amido compound (279.1 mg, 0.169 mmol) was hydrogenated under hydrogen balloon catalyzed by 10% Pd/C in 1% HOAc/MeOH for 45 minutes. The reaction was filtered and concentrated in vacuo . The residue was picked up in a 1:1 mixture of water :ACN and then lyophilized to give 208.3 mg (98.6%) of a colorless powder (2b-l) . The structure was verified by H 1 -NMR. Preparation of Compound 3a-l : R R 11'' , , RR 11"" aanndd RR *1"" = H R = -OCH 3 R J = -OCH 3 3a-l A 50 ml round bottom flask was charged with 10 ml of absolute ethanol and 251.7 mg of CBZ-protected pseudomycin B (2a-l) ( 0.156 mmol) . To this mixture was added ~ 1 ml of acidified ethanol (previously acidified using HCl gas) and the reaction was allowed to stir at room temperature overnight . The solvent was then removed in vacuo and the residue was carried on to the next step without further purification by dissolving it in a solution of 10 ml MeOH/1.5 ml glacial AcOH. Standard hydrogenolysis using 249.7 mg of 10% Pd/C for 30 minutes, removal of the catalyst via filtration and purification via preparatory HPLC yielded 120.9 mg of Compound 3a-l after lyophilization. MS (Ionspray) calcd for C5 5 H 96 ClNι 2 Oi 9 (M+H) + 1264.89, found 1264.3. Preparation of C-18 side-chain (4a-l ) H 3 4a-3 4a-4 To a dichloromethane solution (190 mL) of the chiral acetal 4a-l (6.22 g, 19.1 mmol) was added at -78°C trimethylallylsilane (10.9 mL, 68.69 mmol), followed by neat TiCl 4 (2.94 mL, 26.71 mmol). The reaction was stirred at - 78°C for 1 hr and then at -40°C for 2 hr . At this point, the reaction was quenched with methanol (15 mL) and diluted with dichloromethane (200 mL) . The resulting reaction mixture was washed with IN HCl (2 x 50 mL) , water and brine. The organic layer was dried and cone, in vacuo to give a residue, which was purified by silica gel chromatography (10% EtOAc/Hexanes) to give 5.51 g (78%) of the desired product 4b-l. To a dichloromethane solution (155 mL) of 4b-l (8.56 g, 23.3 mmol) was added PCC (10.0 g, 46.5 mmol). The reaction was stirred at rt for 18 hr, and then filtered through a pad of Celite. The filtrates were concentrated in vacuo to give a reddish residue, which was purified by silica gel chromatography (10% EtOAc/Hexanes) to give 8.36 g (80%) of the methyl ketone intermediate (structure not shown) . The intermediate obtained herein (8.36 g, 22.8 mmol) was dissolved in THF (60 mL) and MeOH (30 mL) . To this solution was added 7.5 M KOH (15 mL) . After stirring 3 hr at rt, the solvent was partially removed. The remaining reaction mixtures were diluted with EtOAc/Et20 (3:1 ratio, 350 mL) . The organic layer was washed with water (3 x 50 mL) and brine. The resulting organic layer was dried and cone, in vacuo to give a residue, which was purified by silica gel chromatography (10% EtOAc/Hexanes) to afford 6.22 g (96&) of the desired product 4c-l as white solids. Carbinol 4c-l (6.22 g, 22.0 mmol) was dissolved in an aqueous THF solution (5.5 mL water and 55 mL THF) . To this solution was added NMO (4.42 g, 33.0 mmol), followed by OSO 4 (280 mg dissolved in THF, 1.10 mmol) . The reaction stirred at rt overnight. At this time, sodium bisulfide (4 g) was added. The reaction was stirred for 2 hr, and then diluted with EtOAc (300 mL) . The whole mixture was washed with water (2 x 40 mL) and brine. The resulting organic layer was dried and cone . in vacuo to give the corresponding triol intermediate. This material was dissolved in MeOH (200 mL) and water (40 mL) . To this solution was added NaI0 4 (10.6 g, 49.5 mmol) . After stirring at rt for lhr, the reaction was filtered through Celite and purified by short column silica gel chromatography (30% EtOAc/Hexanes) to afford -10 g (>100%) crude beta-hydroxyl aldehyde. The impuried aldehyde thus obtain was dissolved in t-BuOH (100 mL) and cyclohexene (14 mL) . To this solution at rt was added an aqueous solution (50 mL) of NaC10 2 (15.97 g, 176 mmol) and KH 2 P0 4 (17.8 g, 132 mmol) . The reaction was stirred at rt for 6 hr and then quenched at 0°C with 5N HCl to pH=4. The reaction was extracted with 3:1 mix-solvent EtOAc/Et 2 0 (3 x 250 mL) . The organic layer was washed with brine and dried and cone . to provide 7.3 g (>100%) of the crude acid 4d-l, which was used directly for the coupling reaction. Preparation of Pseudomycin C-18 (Compound 4b-2) : 4a-2 R = Cbz 4b-2 R = H The crude acid 4d-l (2.1 g, 6.99 mmol) was dissolved in dry THF (20 mL) and DMF (20 mL) . To this solution was added HOBt (1.23 g, 9.08 mmol) and EDCI (1.74 g, 9.08 mmol). After stirring at rt for 8 hr, CBZ-protected pseudomycin nucleus (3.87 g, 2.80 mmol) was added. The reaction was stirred at rt for 2 days. At this point, the solvent was partially removed. The reaction mixture was loaded onto preparative reverse phase HPLC system for purification (4 injections) . Upon lyophilization, 550 mg (12%) of CBZ-protected C18 acyl derivative 4a-2 along with HOBt activated side chain ester (1 g) was isolated. The recovered side chain ester (1.0 g, 2.40 mmol) was next reacted with CBZ-protected pseudomycin nucleus (1.33 g, 0.96 mmol) in dry THF and DMF (10 mL each). Following the same purification procedure just mentioned, additional amounts of the CBZ-protected C18 derivative 4a-2 (606 mg, 38%) were obtained. To a 10% HOAc/MeOH solution (55 mL) of CBZ-protected C18 derivative 4a-2 (550 g, 0.33 mmol) was added at -78°C Pd/C (550 mg, 10% palladium content) . The reaction was subjected to hydrogenation under 1.5 atm. pressure for 40 im. The progress of the reaction was monitored by analytic HPLC. Upon completion, the catalyst was filtered and the filtrates were cone, in vacuo at 30°C. The resulting residue was redissolved in 1:1 aqueous aeetonitrile and lyophilized to give 250 mg (60%) of the desired product 4b-2. In each of the following Examples a specific pseudomycin compound is used as the starting material; however, those skilled in art in the art will recognize that other N-acyl derivatives may be synthesized using the same procedures except starting with a pseudomycin compound having a different N-acyl group. Example 1 The following example demonstrates the formation of mono-, di- and tri-substituted acyloxyalkylcarbamate prodrugs of pseudomycin C (n = 12, R 2 and R 3 = -OH) . To a DMF solution (1 liter) containing Pseudomycin C (1.5 g, 1 eq.) was added 1.5 eq. of Compound la-4 and the mixture was stirred at room temperature for approximately 3 days . The solvent was partially removed and the residue purified by reverse-phase HPLC (Waters™, Delta Pak C18 column) to yield the following products and mixture of products : 86 mg of pure mono substituted pseudomycin C (1-1) ; 87 mg of a mixture of mono-substituted pseudomycin C (1-2) ; 177 mg of a mixture of di-substituted pseudomycin C (1-3) ; 132 mg of pure di-substituted pseudomycin C (1-4) ; and 248 mg of tri-substituted pseudomycin C (1-5) . No irritation of the tail vein was observed for the tri- substituted prodrug or the mixture of di-substituted prodrugs. Some irritation was observed with the pure mono- substituted, mixture of mono-substituted and pure di- substituted prodrug samples. In comparison, unsubstituted pseudomycin C and unsubstituted pseudomycin B clearly showed irritation of the tail vein. All of the samples indicated significant in vivo efficacy except the pure di- substituted prodrug sample (ED 50 > 20 mg/kgx4) . Samples of mono-, di- and tri-substituted prodrugs of structure II above where R 1' , R 1" and/or R 1'" is and n is equal to 10, 12 and 14 have also been made using the same procedure described above. Compounds 1-1 and 1-5 exhibited similar in vivo efficacy as the parent compound pseudomycin C . No evidence of tail vein irritation was observed for Compound 1-5 and an improved tail vein toxicity profile was observed for Compound 1-1. Surprisingly, no in vivo efficacy was observed for Compound 1-4. Example 2 The following examples illustrates the formation of mono-, di- and tri-substituted acyloxyalkylcarbamate prodrugs of pseudomycin B (n = 10, R 2 and R 3 = -OH) . To a solution of Pseudomycin B (2.0 g, 1.65 mmol) dissolved in 500 ml of dimethylformamide was added 574 mg (2.47 mmol) Compound lb-2. The mixture was stirred at room temperature overnight. The solution was then concentrated to about 50 ml and the products purified by HPLC using a gradient elution scheme: 0-30% 0.1% TFA/ACN in 5 minutes and 30-70% TFA/ACN in 40 minutes. A combined yield of 59% was observed. Three of the isolated products (mono-substituted prodrug (Compound 2-1) where R 1' and R 1" = H and R 1'" = -C (0)OCH 2 OAc, the di-substituted prodrug (Compound 2-2) where R 1' and R 1" = -C(0)OCH 2 OAc and R 1"' = H and the tri- substituted prodrug (Compound 2-3) where R 1' , R 1" and R 1 '" = -C (0) OCH 2 OAc) were all tested and demonstrated in vivo efficacy against murine systemic Candidiasis. However, the tail vein toxicity was positive. Example 3 Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin B (n = 10, R 2 and R 3 = -OH)) were prepared where R 1' , R 1" and/or R 1 " = -C (0) OCH 2 OC (0) C (CH 3 ) 3 . The following five samples were isolated: 3-1 mono-substituted R 1'" 3-2 mixed mono-substituted R 1' and R 1" 3-3 mixed di-substituted R 1'" + R 1' and R 1'" + R 1" 3-4 di-substituted R 1' + R 1" 3-5 trisubstituted R 1' + R 1" + R 1'" Samples 3-1, 3-3 and 3-5 each demonstrated negative tail vein toxicity. All five samples demonstrated in vivo efficacy against murine systemic Candidiasis. Example 4 Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin C (n = 12, R 2 and R 3 = -OH) were prepared where R 1' , R 1" and/or R 1'" = -C (0) 0CH 2 0C (0) C (CH 3 ) 3 . The following five samples were isolated: 4-1 mono-substituted R 1'" 4-2 mixed mono-substituted R 1' and R 1" 4-3 mixed di-substituted R 1'" + R 1' and R 1'" + R 1" 4-4 di-substituted R 1' + R 1 " 4-5 trisubstituted R 1' + R 1 " + R 1'" Sample 4-1 was not tested. Samples 4-3, 4-4 and 4-5 all demonstrated negative tail vein toxicity. Samples 4-2, 4-3, 4-4 and 4-5 all demonstrated in vivo efficacy against murine systemic Candidiasis. Example 5 Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin B (n = 10) were prepared where R 1' , R 1" and/or R 1' " = -C(0)OCH(CH 3 )OC(0)CH 3 . Only the trisubstituted derivative (Compound 5-1) was tested. The trisubstituted compound demonstrated negative tail vein toxicity. Example 6 Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin B (n = 10, R 2 and R 3 = -OH) were prepared where R 1' , R 1" and/or R 1'" = -C (0) OCH 2 OC (0) CH 2 CH 3 . Only the trisubstituted derivative (Compound 6-1) was tested. The trisubstituted compound demonstrated good in vivo efficacy against murine systemic Candidiasis without tail vein irritation. Example 7 Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin B (n = 10, R 2 and R 3 = -OH) were prepared where R 1' , R 1" and/or R 1'" = -C (0) OCH 2 OC (0) CH (CH 3 ) CH 3 . Only the tri-substituted derivative (Compound 7-1) was tested. The trisubstituted compound demonstrated good in vivo efficacy against murine systemic Candidiasis without tail vein irritation. Example 8 Examples 8 and 9 illustrate the synthesis of prodrugs from semi-synthetic pseudomycin compounds where the pendant N-acyl group of the L-serine unit of the pseudomycin structure has been modified. Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin C-18 (n = 14, R 2 and R 3 = -OH) (4b-2) were prepared where R 1' , R 1" and/or R 1'" = -C (0) OCH 2 OC (0) C (CH 3 ) 3 . Only the trisubstituted derivative (Compound 8-1) was tested. The trisubstituted compound demonstrated negative tail vein toxicity. Example 9 Using the same general procedures described above in Example 2, mono-, di- and tri-substituted prodrugs of Pseudomycin C-18 (n = 14, R 2 and R 3 = -OH) (4b-2) were prepared where R 1' , R 1" and/or R 1'" = -C(0)OCH 2 OC(0)CH(CH 3 )CH 3 . Only the trisubstituted derivative (Compound 9-1) was tested. The trisubstituted compound demonstrated negative tail vein toxicity. Example 10 Example 10 illustrates further modification of the above described prodrugs where the carboxylic acid group of the aspartic acid unit of the pseudomycin ring is modified to form a 3-monoamido derivative. Synthesis of Compounds 10-1 , 10-2, 10-3 , 10-4 and 10-5 : R 1' , R 1" , R 1"' = -C(0)OCH 2 OC(0)C(CH 3 ) 3 R 2 = -NHCH 2 CH 2 N (CH 3 ) 2 R 3 = -OH 10-1 To a DMF solution (9 ml) of the prodrug 3-5 (864 mg, 0.52 mmol) was added l-dimethylamino-2-aminoethane (57.9 μl, 0.52 mmol) and TBTU (168.6 mg, 0.52 mmol), followed by diisopropylethylamine (423 μl) . After stirring at room temperature for 20 minutes, the reaction mixture was purified by reverse phase HPLC (ACN:0.1% TFA/Water) . Lyophilization yielded 295 mg (34%) of Compound 10-1. R 1 ' , R 1 " , R 1 ' " = -C (0) OCH 2 OC (0 ) C (CH 3 ) 3 R 2 = -NH ( cyclopropyl ) R 3 = -OH 10-2 Compound 10-2 is synthesized using the same procedures as above except 0.052 mmol of cyclopropylamine is used in place of the l-dimethylamino-2-aminoethane . R 1' , R 1" , R 1'" = -C(0)OCH 2 OC(0)C(CH 3 ) 3 R 2 = -NHCH 2 (C0 2 CH 3 ) R 3 = -OH 10-3 Compound 10-3 is synthesized using the same procedures as above except glycine methyl ester is used in place of the l-dimethylamino-2-aminoethane . R 1' , R 1" , R 1"' = -C(0)OCH 2 OC(0)CH(CH 3 ) 2 R 2 = -NHCH 2 CH 2 N(CH 3 ) 2 R 3 = -OH 10-4 Compound 10-4 is synthesized using the same procedures as above except 0.052 mmol of prodrug 7-1 is used in place of the prodrug 3-5. R 1' , R 1" , R 1'" = -C(0)OCH 2 OC(0)CH(CH 3 ) 2 R 2 = -NH(cyclopropyl) R 3 = -OH 10-5 Compound 10-5 is synthesized using the same procedures as above except 0.052 mmol of prodrug 7-1 is used in place of the prodrug 3-5 and 0.052 mmol of cyclopropylamine is used in place of the l-dimethylamino-2-aminoethane . Example 11 Example 11 illustrates the formation of the prodrug of pseudomycin compounds where the carboxylic acid group of the aspartic acid unit of the pseudomycin ring has been modified to form a 3-ami o derivative. Synthesis of 3-monoamido derivative 11 -1 11-1 To a DMF solution (1 liter) containing Compound 2b-l (1 eq.) was added 1.5 eq. of Compound la-4 and the mixture was stirred at room temperature for approximately 3 days. The solvent was partially removed and the residue purified by reverse-phase HPLC (Waters™, Delta Pak C18 column) to yield Compound 11-1 as well as the other mono- and di- substituted products. Example 12 Example 12 illustrates the synthesis of a prodrug where the carboxylic acid group of both the aspartic acid and hydroxyaspartic acid units have been modified to form a bis- ester derivative. Synthesis of Bis-ester 12-1 : 12-1 To a DMF solution (1 liter) containing Compound 3a-l (1 eq.) was added 1.5 eq. of Compound la-4 and the mixture was stirred at room temperature for approximately 3 days. The solvent was partially removed and the residue purified by reverse-phase HPLC (Waters™, Delta Pak C18 column) to yield Compound 12-1 as well as the other mono- and di- substituted products.

Claims: WE CLAIM : 1. A pseudomycin prodrug having the following structure : wherein R is where R a and R a' are independently hydrogen or methyl, or either R a or R a' is alkyl amino, taken together with R b or R b' forms a six-membered cycloalkyl ring, a six-membered aromatic ring or a double bond, or taken together with R c forms a six-membered aromatic ring; R b and R b' are independently hydrogen, halogen, or methyl, or either R b or R b' is amino, alkylamino, α-acetoacetate, methoxy, or hydroxy; R c is hydrogen, hydroxy, C 1 -C 4 alkoxy, hydroxyalkoxy, or taken together with R e forms a 6-membered aromatic ring or C5-C6 cycloalkyl ring; R e is hydrogen, or taken together with R f is a six-membered aromatic ring, C 5 -C 14 alkoxy substituted six-membered aromatic ring, or C 5 -C 14 alkyl substituted six-membered aromatic ring, and R f is C8-C18 alkyl, C5-C11 alkoxy or biphenyl; R is where R g is hydrogen , or C 1 -C 13 alkyl , and R h is C 1 -C 1 5 alkyl , C4-C 1 5 alkoxy, (C 1 -C 1 0 alkyl ) phenyl , - ( CH 2 ) n -aryl , or - (CH 2 ) n - ( C 5 -C 6 cycloalkyl ) , where n = 1 or 2 ; or R is where R 1 is a hydrogen, halogen, or C 5 -Cs alkoxy, and m is 1 , 2 or 3 ; R is where R 3 is C5-C14 alkoxy or C 5 -C 14 alkyl , and p = 0 , 1 or 2 ; where R k is C 5 -C 14 alkoxy; or R is -(CH 2 )-NR m - (C13-C18 alkyl), where R m is H, -CH 3 or -C(0)CH 3 ; R 1 is independently hydrogen, an acyloxymethylene-1, 3- dioxolen-2-one, or an acyloxymethylenecarboxylate, provided that at least one R 1 is an acyloxymethylene-1, 3-dioxolen-2- one or an acyloxymethylenecarboxylate; R 2 and R 3 are independently -OR 2a or -N(R 2b ) (R 2c ) , where R 2a and R 2b are independently hydrogen, Ci-Cio alkyl, C 3 _C6 cycloalkyl, hydroxy (Ci-Cio) alkyl , alkoxy (Ci-Cio) alkyl, C 2 -Cιo alkenyl, amino (Ci- Cio) alkyl, mono- or di-alkylamino (Ci-Cio) alkyl, aryl (Ci-Cio) alkyl, heteroaryl (Ci-Cio) alkyl, cycloheteroalkyl (Cχ-Cιo) alkyl , or ,2b is an alkyl carboxylate residue of an aminoacid alkyl ester and R ,2c is hydrogen or Cι-C 6 alkyl ; and pharmaceutically acceptable salts and solvates thereof. 2. The prodrug of Claim 1 wherein said acyloxymethylene-1 , 3-dioxolen-2-one is represented by structure 1 (a) : 1(a) where R is Ci-Cio alkyl, Ci-Cio alkenyl, benzyl, or aryl and R ,1b is hydrogen or methyl, 3. The prodrug of Claim 1 wherein said acyloxymethylenecarboxylate is represented by structure Kb) : Kb) where R la is Ci-Cio alkyl, Cι-Cι 0 alkenyl, benzyl, or aryl and R lb is hydrogen or methyl. 4. The prodrug of Claim 2 wherein R is represented by the structure where R b' is hydroxy, R a , R a' , R b , R c , R d , and R e are all hydrogen, and R f is n-octyl 5. The prodrug of Claim 3 wherein R is represented by the structure where R b' is hydroxy, R a , R a' , R , R c , R d , and R e are all hydrogen, and R f is n-octyl . 6. The prodrug of Claim 1 wherein said alkyl carboxylate residue of an aminoacid alkyl ester is represented by -CH 2 C0 2 CH 3 , -CH (C0 2 CH 3 ) CH (CH 3 ) 2 , -CH(C0 2 CH 3 )CH(phenyl) , -CH (C0 2 CH 3 ) CH 2 OH, -CH (C0 2 CH 3 ) CH 2 (p- hydroxyphenyl ) , -CH ( C0 2 CH 3 ) CH 2 SH , -CH ( C0 2 CH 3 ) CH 2 ( CH 2 ) 3 NH 2 , -CH(C0 2 CH 3 )CH 2 (4-imidazole) , -CH (C0 2 CH 3 ) CH 2 (5-imidazole) , -CH(C0 2 CH 3 )CH 2 C0 2 CH 3 , or -CH (C0 2 CH 3 ) CH 2 C0 2 NH 2 . 7. A pseudomycin prodrug having the following structure : wherein where R a and R ' are independently hydrogen or methyl, or either R a or R a' is alkyl amino, taken together with R b or R b' forms a six-membered cycloalkyl ring, a six-membered aromatic ring or a double bond, or taken together with R c forms a six-membered aromatic ring; R b and R b' are independently hydrogen, halogen, or methyl, or either R b or R b' is amino, alkylamino, α-acetoacetate, methoxy, or hydroxy; R c is hydrogen, hydroxy, C 1 -C 4 alkoxy, hydroxyalkoxy, or taken together with R e forms a 6-membered aromatic ring or C 5 -C 6 cycloalkyl ring; R e is hydrogen, or taken together with R f is a six-membered aromatic ring, C 5 -C 14 alkoxy substituted six-membered aromatic ring, or C 5 -C 14 alkyl substituted six-membered aromatic ring, and R £ is Cs-Cis alkyl, C 5 -C 11 alkoxy or biphenyl ; R is where R 9 is hydrogen, or C 1 -C 13 alkyl, and R h is C1-C15 alkyl, C4-C15 alkoxy, (C1-C10 alkyl ) phenyl , - (CH 2 ) π -aryl, or - (CH 2 ) n - (C 5 -C 6 cycloalkyl), where n = 1 or 2 ; or R is where R 1 is a hydrogen, halogen, or C 5 -C 8 alkoxy, and m is 1, 2 or 3; R xs where R : is C 5 -C14 alkoxy or C 5 -C 14 alkyl, and p = 0, 1 or 2; R xs where R k is C 5 -C 14 alkoxy; or R is - (CH 2 )-NR m - (Ci3-Ci 8 alkyl), where R m is H, -CH 3 or -C(0)CH 3 ; R 1 is independently hydrogen, an acyloxymethylene-1, 3- dioxolen-2-one, or an acyloxymethylenecarboxylate, provided that at least one R 1 is an acyloxymethylene-1, 3-dioxolen-2- one or an acyloxymethylenecarboxylate; R 2 and R 3 are independently -0R 2a or -N(R 2b ) (R 2c ) , where R 2a and R 2b are independently hydrogen, C 1 -C 10 alkyl, C 3 -C 6 cycloalkyl, hydroxy (C1-C10) alkyl, alkoxy (Ci-Cio) alkyl, C 2 -Cι 0 alkenyl, amino (Ci- Cio) alkyl, mono- or di-alkyla ino (Cι-Cι 0 ) alkyl, aryl (Ci-Cio) alkyl, heteroaryl (Cι-C 10 ) alkyl, cycloheteroalkyl (Ci-Cio) alkyl, or R 2b is an alkyl carboxylate residue of an aminoacid alkyl ester and R 2c is hydrogen or Cι-C 6 alkyl ; and pharmaceutically acceptable salts and solvates thereof. 8. The prodrug of Claim 7 wherein said acyloxymethylene-1, 3-dioxolen-2-one is represented by structure 1(a) : 1 (a) where R ,1a is Ci-Cio alkyl , Ci-Cio alkenyl , benzyl , or aryl and R ,1b is hydrogen or methyl. 9. The prodrug of Claim 7 wherein said acyloxymethylenecarboxylate is represented by structure Kb) : Kb) where R ,1a is Ci-Cio alkyl, Ci-Cio alkenyl, benzyl, or aryl and R lb is hydrogen or methyl. 10. The prodrug of Claim 8 wherein R is represented by the structure where R D is R e are all hydrogen, and R is n-octyl . 11. The prodrug of Claim 9 wherein R is represented by the structure where R b' is hydroxy, R , R a' , R b , R c , R d , and R e are all hydrogen, and R f is n-octyl. 12. The prodrug of Claim 7 wherein said alkyl carboxylate residue of an aminoacid alkyl ester is represented by -CH 2 C0 2 CH 3 , -CH (C0 2 CH 3 ) CH (CH 3 ) 2 , -CH(C0 2 CH 3 )CH(phenyl) , -CH (C0 2 CH 3 ) CHOH, -CH (C0 2 CH 3 ) CH 2 (p- hydroxyphenyl ) , -CH (C0CH 3 ) CH 2 SH, -CH (C0 2 CH 3 ) CH 2 (CH 2 ) 3 NH 2 , -CH(C0 2 CH 3 )CH 2 (4-imidazole) , -CH (C0 2 CH 3 ) CH 2 (5-imidazole) , -CH(C0 2 CH 3 )CH 2 C0 2 CH 3 , or -CH (C0 2 CH 3 ) CH 2 C0 2 NH 2 . 13. The use of a compound as claimed in any one of the preceding claims in the preparation of medicaments for use in combating either systemic fungal infections or fungal skin infections . 14. A pharmaceutical formulation comprising a pseudomycin prodrug of Claim 1 or 7 and a pharmaceutically acceptable carrier. 15. A medicament for treating an antifungal infection in an animal wherein said medicament comprises a compound of Claim 1 or 7. 16. A method for treating an antifungal infection in an animal in need thereof, comprising administering to said animal a pseudomycin prodrug of Claim 7, 8, 9, 10 or 11.

Inventors: CHEN SHU HUI, RODRIGUEZ MICHAEL JOHN, SUN XICHENG DAVID, CHEN, SHU, HUI, RODRIGUEZ, MICHAEL, JOHN, SUN, XICHENG, DAVID
Applicants: CHEN SHU HUI, LILLY CO ELI, RODRIGUEZ MICHAEL JOHN, SUN XICHENG DAVID, CHEN, SHU, HUI, ELI LILLY AND COMPANY, RODRIGUEZ, MICHAEL, JOHN, SUN, XICHENG, DAVID
Application year: 2000
Main classification: C07K7
IPC codes: A61K38, A61P17, A61P31, C07K7, C07K11