IE913974A1 - Specific inhibition of dihydrofolate reductase and compounds¹therefor - Google Patents

Abstract

Compounds derived from pyrimidines having improved activity against fungi such as Pneumocystis carinii, and improved selectivity for P. carinii dihydrofolate reductase over human dihydrofolate reductase, are disclosed. P. carinii pneumonia is advantageously treated with the disclosed compounds.

Description

This invention relates to pharmacology and the inhibition of enzymes specific to pathogens. More particularly, the invention relates to methods for specifically inhibiting the enzyme dihydrofolate reductase in fungal pathogens, and compounds therefor.

Background of the Invention Pneumocystis carinii pneumonia (PCP) is a significant, life-threatening infection in immunocompromised subjects, and is a leading cause of morbidity and mortality in patients presenting acquired immunodeficiency syndrome (AIDS). Since the onset of the AIDS epidemic, the incidence of PCP has risen from approximately 200 cases per year to more than 25,000 cases per year in the United States.

Due to the lack of a continuous in vitro culture system, and the cumbersome nature of the rat model of PCP, anti-P. carinii therapy has been developed largely on the assumption that P. carinii was a species of protozoa, and thus that anti-protozoal agents were likely to be effective. The two principle therapeutic modalities, trimethoprim/sulfamethoxazole and pentamidine, were developed empirically. However, P. carinii has recently been suggested to belong instead to the Kingdom Fungi (J.C. Edman et al.. Nature (1988) 334:519-22). - 2 Trimethoprim (U.S. Pat No. 2,909,522) and pyrimethamine, and other dihydrofolate reductase (DHFR) inhibitors are known to be effective anti-bacterial, and anti-protozoal agents due to the central role played by DHFR in the metabolic synthesis of nucleic acid precursors. Despite their efficacy when used in con5 junction with a sulfonamide, trimethoprim and pyrimethamine are alone poor inhibitors of P. carinii DHFR. For example, trimethoprim and pyrimethamine exhibit 50% inhibition concentrations (ICjq) of 8 and 2,500 nM for E. coli DHFR, while ICjoS for P, carinii DHFR arc 39,600 and 2,400 nM, respectively. Other anti-folates have been shown to be more effective inhibitors of P. carinii DHFR, but require concomitant administration of leucovorin to prevent toxicity to the host. Allegra et al. (U.S. Pat No. 4,694,007) suggested treatment of P, carinii and Toxoplasmosis gondii with 2,4-diamino-5-methyl-6-[(3,4,5-trimethoxyanilino)methyljquinazoline (trimetrexate), on the theory that the DHFR enzyme in this pathogens is more similar to mammalian DHFR than to prokaryotic DHFR.

Prior to the AIDS epidemic, these types of agents were sufficient for treatment of the rare cases of P. carinii pneumonia. However, in the HIV-positive patient, therapy and prophylaxis with the standard anti-P. carinii agents are complicated by frequent toxic and allergic side effects. New compounds that surpass the efficacy of the known antifolates in treating PCP are desirable, especially inhibitors having greater selectivity for P, carinii DHFR relative to the host (particularly human) DHFR than known inhibitors such as trimethoprim.

Commonly-owned copending U.S. Patent Application Serial No. 447,181, filed 7 December 1989 disclosed several DHFR inhibitors exhibiting good selectivity for P. carinii DHFR.

Disclosure of the Invention We have now found compounds which exhibit superior activity against DHFR from fungal pathogens, such as P. carinii, and which exhibit much higher selectivity for the fungal enzyme as compared with the mammalian (human) enzyme. These compounds arc represented generally by Formula I: Formula I: wherein Rj is 3-R3-4-R4-5-Rj-benzyl or (N-Rg)-8-azabicyclo[3.2.1]oct-3-yl; and R, is H; or Rt and Rj together form f\/ where R3 and Rj are independently selected from the group consisting of H, lower alkoxy, lower alkylthio, lower alkylsulfinyl, vinyl, carboxy-lower alkyl, carboxy-lower alkoxy, dicarboxy-lower alkyl, dicarboxy-lower alkoxy, aryl-lower alkoxy, arylsulfonyllower alkoxy, arylsulfamido-lower alkoxy, and radicals of formula -O(CH2)n-CORg, where n is an integer from 0 to 6 and Rg is an amino acid; R4 is selected from the group consisting of lower alkoxy, aryl-lower alkoxy, lower alkylthio, halo, lower alkenyl, lower alkenyloxy, and pyrrolyl; with the proviso - 4that R3, R4, and R3 are not simultaneously methoxy; Re is selected from the group consisting of unsubstituted aryl and aryl substituted with one to three radicals selected from the group consisting of halo, lower alkyl, lower alkoxy, lower alkylthio, carboxy, carbamido, carboxy-lower alkyl, and carbamido-lower alkyl; and R7 is selected from the group consisting of aryl and aryl-lower alkyl, where aryl may be substituted with one to three radicals selected from the group consisting of halo, lower alkyl, lower alkoxy, lower alkylthio, carboxy, carbamido, carboxylower alkyl, and carbamido-lower alkyl; and lower alkyl esters, amides thereof, and pharmaceutically acceptable addition salts.

One aspect of the invention is a method for treating a fungal infection (such as P. carinii) in a mammal by administering an effective amount of a compound of formula I.

Another aspect of the invention is a composition for treating a fungal infection (such as P, carinii) in a mammal comprising an effective amount of a compound of formula I in combination with a pharmaceutically acceptable excipient.

Another aspect of the invention is the use of a compound of formula I to prepare a composition for treating a fungal infection (such as P, carinii) in a mammal comprising an effective amount of a compound of formula I in combination with a pharmaceutically acceptable excipient Modes of Carrying Out The Invention A. Definitions The terms fungal infection and fungal pathogen refer to the infection 25 of a mammal with an organism of the Kingdom Fungi, for example Pneumocystis carinii, Aspergillus, Candida, Fusarium, and the like. The presently preferred method of the invention is the treatment of Pneumocystis carinii using the compounds of the invention. -5The term pharmaceutically acceptable refers to compounds and compositions which may be administered to mammals without undue toxicity. Exemplary pharmaceutically acceptable salts include mineral acid salts such as hydrochlorides, hydrobromides, phosphates, sulfates, and the like; and the salts of organic acids such as acetates, propionates, malonates, benzoates, and the like.

The term effective amount refers to an amount of compound sufficient to exhibit a detectable therapeutic effect. The therapeutic effect may include, for example, without limitation, inhibiting the growth of pathogens, inhibiting or preventing the release of toxins by pathogens, killing pathogens, and preventing the establishment of infection (prophylaxis). The precise effective amount for a subject will depend upon the subject’s size and health, the nature of the pathogen, the severity of the infection, and the like. Thus, it is not possible to specify an exact effective amount in advance. However, the effective amount for a given situation can be determined by routine experimentation based on the information provided herein.

The term lower alkyl refers to saturated straight or branched-chain radicals consisting of carbon and hydrogen having from 1 to 6 carbon atoms, inclusive, such as methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-hexyl, and the like. Lower alkoxy refers to a radical of the form R-Ο-, where R is lower alkyl as defined above. Suitable examples include methoxy, ethoxy, propoxy, butoxy, and the like. Similarly, lower alkylthio refers to radicals of the form R-S-, and lower alkylsulfinyl refers to groups of the form R-S(-O)-. For example, one may employ methylthio, ethylthio, methylsulfinyl, t-butylsulfinyl, and the like. 'Lower alkenyl refers to straight or branched-chain radicals consist25 ing of carbon and hydrogen having 2-6 carbon atoms and at least one double bond between a pair of carbon atoms, such as ethenyl (vinyl), 2-propenyl, 1-methylethenyl, 2-butenyl, 3-butenyl, and the like. -6The term carboxy-lower alkyl refers to radicals having the form -(CH2)n-COOH, where n is an integer from 1 to 6 inclusive. 'Oicarboxy-lower alkyl indicates lower alkyl chains having two COOH groups attached.

Aryl denotes cyclic hydrocarbon radicals of 6-10 carbon atoms which 5 exhibit aromatic character, for example phenyl and napthyl.

The term halo refers to fluoro, chloro, bromo, and iodo. The term amino acid refers to any of the 20 or so commonly occurring amino acids, for example, glycine, alanine, arginine, phenylalanine, glutamic acid, valine, histidine, proline, ornithine, norleucine, and the like. When attached as Rg in a radical of the form -O(CH2)n-COR8, the amino acid will preferably be attached via a peptide bond, i.e. by a bond between the amino group of the amino acid and the acyl carbon of the radical.

The term coadministering means administration of a compound of the invention in combination with a second therapeutic agent. The second therapeutic agent is a dihydropteroate synthase inhibitor, preferably dapsone or a sulfa drug. Suitable sulfa drugs include, without limitation, sulfadiazine, sulfamethoxazole, and the like. Coadministration may be simultaneous, for example by administering a mixture of the therapeutic agents, or may be accomplished by administration of the agents separately within a short time period.

B. General Method The compounds of the invention are structurally related to the compound trimethoprim (2,4-diamino-5-(3,4,5-trimethoxybenzyl)pyrimidine), the synthesis for which is known in the art. See for example U.S. Pat No. 2,909,522, which des25 cribes the synthesis of trimethoprim and related compounds. Compounds of formula I may similarly be synthesized by those of ordinary skill in the art.

Syntheses of such compounds are described in the following U.S. patents: Hitchings et al. (2.658.897k Hitchings et al. (2.909.522); Hoffer (3,341,541); Roth (3,772,289); Roth et al. (3.819.629); Roth (3,822,264); Kompiset_al. -7(3,931,181); Herrling (3,980,649); Liebenow et al. (3,992,379); Kompis (4,024,145); Rosen (4,033,962); Kompis et al. (4,039,543); Jemow et al. (4,075,209); Perun et al. (4,087,528); Fritschi et al. (4,180,578); Kompis et al. (4,203,980); Poe et al. (4,258,045); Daluge et al. (4.438.267): Dall’Asta (4,485,248); Kompis et al. (4,515,948); Swaringen et al. (4,568,744); Roth et al. (4,587,341); Kompis et al. (4,590,270); Daluge et al. (4,603,136); Kompis et al. (4,792,557); and Seydel etal. (4,912,112).

Presently preferred compounds of the invention are: 2.4- diamino-5-[3,5-dimethoxy-4-(2-hydroxyprop-2-yl)benzyl]pyrimidine; 2,4-diamino-5-(3,5-dimethoxy-4-N-pyrrolylbenzyl)pyrimidine; 2.4- diamino-5-(3,5-diethoxy-4-N-pyrrolylbenzyl)pyrimidine; 2.4- diamino-5-(3,5-dipropoxy-4-N-pyrrolylbenzyl)pyrimidine; 2.4- diamino-5-(3,5-dibutoxy-4-N-pyrrolylbenzyl)pyrimidine; 2.4- diamino-5-(3,5 -diethoxy-4-carboethoxybenzyl)pyrimidine; 2,4-diamino-5-(3,5 -divinyl-4-vinyloxybenzyl)pyrimidine; 2.4- diamino-5-[3-(4-N-acetaminophenyl)sulfonaminoethoxy-4,5-dimethoxybenzyl]pyrimidine; 2.4- diamino-5-[3-(4-aminophenyl)sulfonaminoethoxy-4-methoxybenzyl]pyrimidine; 2.4- diamino-5-[3-(4-N-acetaminophenyl)sulfonaminoethoxy-4-bromo-5-methoxybenzyl]20 pyrimidine; 2.4- diamino-5-[3-(4-aminophenyl)sulfonaminoethoxy-4-bromo-5-methoxybenzyl]pyrimidine; 2.4- diamino-5-(3-methoxy-4,5-dibenzyloxybenzyl)pyrimidine; 2.4- diamino-5-(4-benzyloxybenzyl)pyrimidine; 2,4-diamino-5-(3,4-dibenzyloxy-5-methoxybenzyl)pyrimidine; 2.4- diaminQ-5-(3,4-dimethoxy-5-benzyloxybenzyl)pyrimidine; 2.4- diamino-5-[3,5-diethoxy-4-(propen-2-yl)benzyl]pyrimidine; 2.4- diamino-5-[3,5-dimethoxy-4-(propen-2-yl)benzyl]pyrimidine; 2.4- diamino-5-(3,5-dimethoxy-4-methylthiobenzyl)pyrimidine; - 8 2.4- diamino-5-(3-methylsulfinyl-4-methoxy-5-methylthiobenzyl)pyrimidine; 2.4- diamino-5-[3-(4,6-dicarboxyhexyloxy)-4-bromo-5-methoxybenzyl]pyrimidine; 2.4- diamino-5-[3-(3-carboxymethylamino-3-oxopropoxy)-4-methoxybenzyl]pyrimidine; 52,4-diamino-5-(3-[3-(l,3-dicarboxypropyl)amino-3-oxopropoxy]-4-bromo-5-methoxybenzyi)pyrimidine; 2.4- diamino-5-[3,5-dimethoxy-4-((2-phenylsulfinyl)acetyl)benzyl]pyrimidine; 2.4- diamino-5-[3-amino-4-methyl-5-(N-pyrrolyl)benzyl]pyrimidine; 2.4- diamino-5-(3,5 -di-N-pyrrolyl-4-methoxybenzyl)pyrimidine; 102.4- diamino-5-[3,5-di-methoxy-4-(3-hydrocarboxy-l-oxopropylamino)benzyl]pyrimidine; 2.4- diamino-5-[3,5-dimethoxy-(4-acetaminophenylsulfonamino)benzyl]pyrimidine; 2.4- diamino-5-(3,5-dimethoxy-4-propylbenzyl)pyrimidine; 2.4- diamino-5-(3,5-dichloro-4-N-pyrrolylbenzyl)pyrimidine; 2.4- diamino-5-[3,5-dimethoxy-4-(2-(2-(2-methoxy)ethoxy)ethoxy)ethoxybenzyl]15 pyrimidine; 2.4- diamino-5-[3-(3-benzyloxycarbonylmethylamino-3-oxopropoxy)-4-bromo-5-methoxybenzyl]pyrimidine; 2.4- diamino-5-[3-(3-carboxymethylamino-3-oxopropoxy)-4-bromo-5-methoxybenzyl]pyrimidine; 202.4- diamino-5-[3-methoxy-4-bromo-5-(4-methylaminobenzamidoethoxy)benzyl]pyrimidine; 3-(2,4-diaminopyrimidin-5-ylmethyl)-8-(3,5-dimethoxyphenyl)-8azabicyclo[3.2.1 ]octane; 2H,3H-dihydro-5-(2,4-diaminopyrimidin-5-ylmethyl)-6,7-dimethoxybenzofuran; -(2,4-diaminopyrimidin-5-ylmethyl)-7-methoxy-8-bromo-1,2-benzopyran; -(2,4-diaminopyrimidin-5-ylmethyl)-7,8-dimethoxy-1,2-benzopyran; 2.4- diamino-5-[3-phenyl-5-(3-methoxypropoxy)benzyl]pyrimidine; 2.4- diamino-7-(3,5 -dimethoxybenzyl)pyrrolo[2,3-f]quinazoline; 2.4- diamino-5-[6-(4-methoxybutoxy)naphth-l-yl]pyrimidine; -92,4-diamino-5-(2,7-dimethylbenzpyrazol-5-ylmethyl)pyrimidine; 2,4-diamino-5-(4,5,6-trimethoxy-2,3-dihydroinden-l-yl)pyrimidine; and 2,2-dimethyl-5-(2,4-diaminopyrimidm-5-ylmethyl)-7-methoxybenz[b]dioxolane.

The most preferred compounds at present are 2,4-diamino-5-(3,5-diethoxy-4-N5 pyrrolylbenzyl)pyrimidine and 2,4-diamino-5-[3r5-dimethoxy-4-(2-hydroxyprop-2yl)benzyl]pyrimidine.

Compositions of the invention for administration will generally include an effective amount of a compound of formula I in addition to a pharmaceutically acceptable excipient. Suitable excipients include most carriers approved for oral or parenteral administration, including water, saline, Ringer’s solution, Hank’s solution, and solutions of glucose, lactose, dextrose, ethanol, glycerol, albumin, and the like. These compositions may optionally include stabilizers, antioxidants, antimicrobials, preservatives, buffering agents, surfactants, and other accessory additives. A presently preferred vehicle comprises about 1 mg/mL serum albumin in phosphate-buffered saline (PBS). A thorough discussion of suitable vehicles for parenteral administration may be found in E.W. Martin, 'Remington’s Pharmaceutical Sciences (Mack Pub. Co., current edition).

The precise dosage necessary will vary with the age, size, and condition of the subject, the nature and severity of the disorder to be treated, and the like: thus, a precise effective amount cannot be specified in advance. However, appropriate amounts may be determined by routine experimentation with animal models. In general terms, an effective dose of compound of formula I will range from about 10 pg/Kg to about 50 mg/Kg. Suitable animal models include the mouse model illustrated in the Examples below. Rats and other rodents have DHFR very similar to the human enzyme, and thus make suitable animal models.

A group of experimental animals is inoculated with 10-100 LDJ0s of Pneumocystis carinii, followed by treatment with a solution of test compound. A negative control group is left untreated, while a positive control group is treated with a standard therapy, such as trimethoprim. Administration of the compounds is preferIE 913974 - 10ably per os (e.g., using a gavage), but may be parenteral, for example by subcutaneous or intramuscular injection, or by inhalation of an aerosol. The animals are monitored during treatment, and are sacrificed and examined after 60 days for presence of infection.

C. Examples The examples presented below are provided as a further guide to the practitioner of ordinary skill in the art, and are not to be construed as limiting the invention in any way.

Example 1 (Demonstration of Activity) A. Materials Buffers were prepared as follows; 4xDHFR buffer 200 mM Tes, 300 mM BME, 4 mM EDTA, pH 7.0.

+DHF buffer 2.5 mg/mL BSA, 0.25 mM NADPH, 62.5 pM dihydrofolate, 2.5x DHFR buffer.

-DHF buffer 2.5 mg/mL BSA, 0.25 mM NADPH, 2.5x DHFR buffer.

Enzyme buffer: 50 mM Tes, 5 mM DTT, 1 mM EDTA, 20% glycerol, 1 mg/mL BSA, pH 7.0.

Dilution buffer 50 mM Tes, 5 mM DTT, 1 mM EDTA, 1 mg/mL BSA, pH 7.0. PcDHFR: 5 pg/mL P. carinii DHFR in enzyme buffer. crude hDHFR: crude recombinant human DHFR (obtained from Hoffmann-LaRoche) in enzyme buffer (9.9 mg/mL total protein). purified hDHFR: purified recombinant human DHFR (obtained from HoffmannLaRoche) in enzyme buffer (3.5 mg/mL total protein).

Test compounds were prepared and provided by Hoffmann-LaRoche. Stock solutions were prepared by dissolving 2-8 mg in dimethylsulfoxide (DMSO) - 11 to prepare 50 mM solutions. Compounds which did not dissolve at 50 mM were diluted serially to 25, 16.6, or 12.5 mM.

B. Assay Eight compounds (6.7 pL each) were added to wells in one column of a Falcon® 96-well microtiter plate, and diluted with 160 pL of water. The remaining wells received 126 pL of water. The solutions in the first column were then serially diluted (4x) into columns 2-9, with column 10 containing a control sample, and columns 11-12 containing water blanks. The final compound concentrations were 1 mM to 15 nM (after the remaining reagents were added).

Using an 8-channel pipet, 100 pL of -DHF buffer were added to each well in columns 11 and 12 (blanks containing only water). Next, 100 pL of +DHF buffer were added to each well in columns 1-10. PcDHFR or crude or purified rhDHFR enzyme was then added (25 pL) to each well, and the plate contents mixed using a Titertek shaker. The plates were read on a Molecular Devices Plate Reader at an absorbance of 340 nm, -0.05 ΟΊλ scale, for 10 minutes reading every 10 seconds. The kinetic data was analyzed using Delta Soft software (Biometallics), and the IC^ and Ki calculated for both human and R carinii enzymes. Selectivity was calculated as KU,wm/Kin» thus, higher values for selectivity indicate that the compound inhibits the P. carinii enzyme to a greater degree than the human enzyme. The results are shown in Table 1 below. For purposes of comparison, trimethoprim in this assay exhibits an ICM of 20 pM, with a selectivity of 0.1. - 12TABLE 1: Compounds of Formula I r3r4 r5IC50 Selectivity -OMe -OMe -O(CH2)2NHSO^NHAc 2.4 μΜ 350 -OMe -CMe2OH -OMe 25 μΜ 350 -OEt pyrrolyl -OEt 2.4 μΜ 310 -OMe pyrrolyl -OMe 29 μΜ 30 vinyl vinyloxy vinyl 3.4 μΜ 250 H BzO- H 16 μΜ 50 -OMe BzO- BzO- 8.3 μΜ 100 -OMe -C(Me)=CH2 -OMe 24 μΜ 160 -OEt -C(Me)=CH2 -OEt 2.5 μΜ 160 -OMe -SMe -OMe 19 μΜ 145 -SMe -OMe -S(O)Me 18 μΜ 490 -OMe Br -O(CH2)2CH(CH2)2COOH COOH 0.04 μΜ 525 -OMe Br -O(CH2)2CO(N-Asp) 2 μΜ 4300 H -OMe -O(CH2)2CO(N-Gly) 9.3 μΜ 80 -OMe Br -O(CH2)2CO(N-Gly) 10 μΜ - 8 Me - -CH3, Et - -¾¾ Bz - -CMjCA, N-fiiy - -NTiCH2C0OH, n-AsP -NHCH(COOH)CH2COOH, φ - phenyl, Ac - -COCH3 Example 2 Proceeding as described in Example 1 above, the compounds listed below were assayed and found to exhibit high activity and selectivity, as set forth in Table Π: A) 2,4-diamino5-(4-benzyloxybenzyl)pyrimidine; - 13 B) 2,4-diamino-5-(3,4-dimethoxy-5-benzyloxybenzyl)pyrimidine; C) 2,4-diamino-5-(3,4-dibenzyloxy-5-methoxybenzyl)pyrimidine; D) 2,4-diamino-5-[3,5-dimethoxy-4-(2-hydroxyprop-2-yl)benzyl]pyrimidine; E) 2,4-diamino- 5-(3,5 -dime±oxy-4-N-pyrrolylbenzyl)pyrimidine; F) 2,4-diamino-5-(3,5-diethoxy-4-N-pyrrolylbenzyl)pyrimidine; G) 2,4-diamino-5-(3,5-divinyl-4-vinyloxybenzyl)pyrimidine; H) 2,4-diamino-5-[3-(4-N-acetaminophenyl)sulfonaminoethoxy-4,5-dimethoxybenzyl]pyrimidine; I) 2,4-diamino-5-[3-(4-aminophenyl)sulfonaminoethoxy-4-methoxybenzyl]10 pyrimidine; J) 2,4-diamino-5-[3-(4-N-acetaminophenyl)sulfonaminoethoxy-4-bromo-5methoxybenzyl]pyrimidine; K) 2,4-diamino-5-[3-(4-aminophenyl)sulfonaminoethoxy-4-bromo-5-methoxybenzyl]pyrimidine; L) 2,4-diamino-5-[3,5-diethoxy-4-(propen-2-yl)benzyl]pyrimidine; M) 2,4-diamino-5-[3,5-dimethoxy-4-(propen-2-yl)benzyl]pyrimidine; N) 2,4-diamino-5-(3,5 -dimethoxy-4-methylthiobenzyl)pyrimidine; O) 2,4-diamin(>-5-(3-methylsulfmyl-4-methoxy-5-methylthiobenzyl)pyrimidine; P) 2,4-diamino-5-[3-(4,6-dicarboxyhexyloxy)-4-bromo-5-methoxybenzyl]pyrimidine; Q) 2,4-diammo-5-[3-(3-carboxymethylaniino-3-oxopropoxy)-4-methoxybenzyl]pyrimidine; R) 2,4-diamino-5-(3-[3-(l ,3-dicarboxypropyl)amino-3-oxopropoxy]-4-brOmo25 5-methoxybenzyl)pyrimidine; S) 2,4-diamino-5-[3,5-dimethoxy-4-((2-phenylsulfonyl)acetyl)benzyl]pyrimidine; T) 2,4-diamino-5-[3-amino-4-methyl-5-(N-pyrrolyl)benzyl]pyTimidine; U) 2,4-diamino-5-(3,5-di-N-pyrrolyl-4-methoxybenzyl)pyrimidine; - 1410 V) 2,4-diamino-5-[3,5-di-methoxy-4-(3-hydrocarboxy-l-oxopropylamino)benzyl]pyrimidine; W) 2,4-diamino-5-[3,5-dimethoxy-(4-acetaminophenylsulfonamino)benzyl]pyrimidme; X) 2,4-diamino-5-(3,5-dimethoxy-4-propylbenzyl)pyrimidine; Y) 2,4-diamino-5-(3,5-dichloro-4-N-pyrrolylbenzyl)pyrimidine; Z) 2,4-diamino-5-[3,5-dimethoxy-4-(2-(2-(2-methoxy)ethoxy)ethoxy)ethoxybenzyl]pyrimidine; AA) 2,4-diamino-5-[3-(3-benzyloxycarbonylmethylamino-3-oxopropoxy)-4bromo-5-methoxybenzyl]pyrimidine; BB) 2,4-diamino-5-[3-(3-carboxymethylammo-3-oxopropoxy)-4-bromo-5methoxybenzyl]pyrimidine; CC) 2,4-diamino-5-[3-methoxy-4-bromo-5-(4-methylaminobenzamidoethoxy)benzy 1] pyrimidine; DD) 3-(2,4-diaminopyrimidin-5-ylmethyl)-8-(3,5-dimethoxyphenyl)-8-azabicyclo[3.2.1 ]octane; EE) 2H,3H-dihydro-5-(2,4-diaminopyrimidin-5-ylmethyl)-6,7-dimethoxybenzofuran; FF) 5-(2,4-diaminopyrimidin-5 -ylmethyl)-7-methoxy-8-bromo-1,2-benzopyran; GG) 5-(2,4-diaminopyrimidin-5-ylmethyl)-7,8-dimethoxy-1,2-benzopyran; HH) 2,4-diamino-5-[3-phenyl-5-(3-methoxypropoxy)benzyl]pyrimidine; Π) 2,4-diamino7-(3,5-dimethoxybenzyl)pyrrolo[2,3-f]quinazoline; JJ) 2,4-diamino-5-[6-(4-methoxybutoxy)naphth-l-yl]pyrimidine; KK) 2,4-diamino-5-(4,5,6-trimethoxy-2,3 -dihydroinden-1 -yl)pyrimidine; LL) 2,2-dimethyl-5-(2,4-diaminopyrimidin-5-ylmethyl)-7-methoxybenz[b]dioxolane; MM) 2,4-diamino-5-(3,5-diethoxy-4-carboethoxybenzyl)pyrimidine; and NN) 2,4-diamino-5-(2,7-dimethylbenzpyrazol-5-ylmethyl)pyrimidine. - 15 TABLE Π: Compound IC50 Selectivity 5 A) 16.0 53.2 B) 13.9 3.3 C) 8.3 90.3 D) 24.8 343.5 E) 2.9 29.4 10 F) 2.4 312.4 G) 3.4 250.5 H) 2.4 248.5 I) 0.57 7.8 J) 3.7 29.5 15 K) 0.97 12.1 L) 2.5 190.9 M) 24.0 159.8 N) 19.0 143.5 0) 17.5 486.8 20 P) 0.043 528.0 Q) 9.3 77.7 R) 2.0 4255.0 S) 14.8 36.8 T) 33.5 42.5 25 U) 4.5 47.7 V) 100.0 85.2 W) 31.0 14.3 x) 20.0 42.6 Y) 10.9 53.2 30 Z) 36.0 33.1 - 16 TABLE II (continued): CompoundIC50 Selectivity AA) 7.9 14.4 BB) 10.0 13.1 CC) 7.8 14.6 DD) 0.016 31.0 EE) 12.9 26.4 FF) 5.9 6.9 GG) 11.5 266.7 HH) 12.2 11.2 II) 0.039 1.2 J J) 80.0 106.5 KK) 87.0 97.9 LL) 13.9 15.0 MM) 8.3 63.6 NN) 18.0 10.7

Claims (3)

1. CLAIMS: 1. Use of a compound of formula I: wherein 20 Rj is 3-Rj-4-R 4 -5-Rj-benzyl or (N-R < )-8-azabicyclo[3.2.1]oct-3-yl; and R 2 is H; or R t and R 2 together form where R 3 and Rj arc independently selected from the group consist ing of H, lower alkoxy, lower alkylthio, lower alkylsulfinyl, vinyl, carboxy-lower alkyl, carboxy-lower alkoxy, dicarboxy-lower alkyl, dicarboxy-lower alkoxy, aryl25 lower alkoxy, arylsulfonyl-lower alkoxy, arylsulfamido-lower alkoxy, and radicals of formula -O(CH 2 ) # -COR g , where n is an integer from 0 to 6 and R, is an amino acid; - 18 10 R 4 is selected from the group consisting of lower alkoxy, aryl-lower alkoxy, lower alkylthio, halo, lower alkenyl, lower alkenyloxy, and pyrrolyl; with the proviso that R 3 , R*. and Rj are not simultaneously methoxy; % R* is selected from the group consisting of unsubstituted aryl and aryl substituted with one to three radicals selected from the group consisting of halo, lower alkyl, lower alkoxy, lower alkylthio, carboxy, carbamido, carboxylower alkyl, and carbamido-lower alkyl; and R 7 is selected from the group consisting of aryl and aryl-lower alkyl, where aryl may be substituted with one to three radicals selected from the group consisting of halo, lower alkyl, lower alkoxy, lower alkylthio, carboxy, carbamido, carboxy-lower alkyl, and carbamido-lower alkyl; and lower alkyl esters, amides thereof, and pharmaceutically acceptable addition salts for the manufacture of a medicament for treating a fungal infection. is H and R-] is methoxy. is bromo.
2. 2. Use according to claim 1, wherein R2 is 3-R3~4-R4-5-R5-benzyl.
3. 3. Use according to claim 2, wherein R3 4. Use according to claim 3, wherein R4 5. Use according to claim 4, wherein R5 is -CXCHjjjCHiCOOHXCHjJjCOOH, or a pharmaceutically acceptable mono- or diester thereof, 7. Use according to claim 6, wherein n is 2 and Rg is -NHCH(COOH)CH 2 CH 2 COOH or a mono- or di-ester thereof. 6. Use according to claim 4, wherein R5 is -0{CH 2 ) n -CORg. - 19 10 7. 8. Use according to -NHCHjCOOH or an ester thereof. 8. 9. Use according to 9. 10. Use according to 10. 11. Use according to 11. 12. Use according to 12. 13. Use according to 13. 14. Use according to 14. 15. Use according to 15. 16. Use according to claim 6, wherein n is 2 and R g is claim 3 wherein Rj is methoxy. claim 9, wherein R 4 is 2-hydroxyprop-2-yl. claim 9, wherein R 4 is propen-2-yl. claim 9, wherein R 4 is N-pyrrolyl. claim 9, wherein R 4 is methylthio. claim 3, wherein R 4 is benzyloxy. claim 14, wherein R s is benzyloxy. claim 3, wherein R 4 is methoxy, and Rj is -OCH 2 CH 2 NHSO 2 (C 6 H 6 )NHCOCH 3 . 16. 17. Use according to claim 2 wherein R 3 is H. 17. 18. Use according to claim 9 wherein R 4 is methoxy. 18. 19. Use acocrding to claim 10 wherein R5 is -O(CH 2 ) D -COR 8 , where n is 2 and R 8 is -NHCH 2 COOH or an ester thereof. 19. 20. Use according to claim 9, wherein R 4 is benzyloxy and Ri is H. -2021. Use according to daim 2, where R 3 and R s are each ethoxy. 20. 22. Use according to claim 21, wherein R 4 is pyrrolyl. « 5 21. 23. use according to claim 21, wherein R 4 is propen-2-yl. 22. 24. Use according to claim 2, where R 3 and R s are each vinyl. 23. 25. Use according to claim 2, where R 3 is methylthio, R 4 is 10 methoxy, and R 5 is methylsulfinyl. 24. 26. use according to claim 1, wherein R 2 is H and R, is (N-Rg)8-azabicyclo[3.2.1]oct-3-yl. 15 27. Use according to claim 26, wherein Rg is selected from the group consisting of 2-naphthyl, 3,5-dimethoxyphenyl, and 4-carboethoxyphenyl. use according to claim 1, wherein R t and Rj together form 25. 29. Use according to claim 28, wherein R·, is benzvl. 26. 30. Use according to · claim 28 where R 3 is 3,5-dimethoxyphenyl. -21 27. 31. Use according to claim 2 where Rj and Rj are propoxy and R 4 is Ν-pyrrolyl. 28. 32. Use according to claim 1 wherein said fungal infection is caused by Pneumocystis carinii. 29. 33. Use according to claim 1, wherein the medicament further conprises a dihydropteroate synthase inhibitor. 10 30. 34. Use according to claim 33, wherein said dihydropteroate synthase inhibitor is selected from the group consisting of dapsone and sulfa drugs. 31. 35. Use according to claim 1, wherein said compound of formula I is selected from the group consisting of 2,4-diamino-5-(4-benzyloxy15 benzyl)pyrimidine; 2,4-diamino-5-(3,4-dimethoxy-5-benzyloxybenzyl)pyrimidine; 2,4-diamino- 5-(3,4-dibenzyloxy-5-methoxybenzyl)pyrimidine; 2,4-diamino-5 - [3,5dimethoxy-4-(2-hydroxyprop-2-yl)benzyl]pyrimidine; 2,4-diamino-5-(3,5dimethoxy-4-N-pyrrolylbenzyl)pyrimidine; 2,4-diamino-5-(3,5-diethoxy-4-Npyrrolylbenzyl)pyrimidine; 2,4-diamino-5-(3,5-divinyl-4-vinyloxybenzyl)20 pyrimidine; 2,4-diamino-5-[3-(4-N-acetaminophenyl)sulfonaminoethoxy-4,5-dimethoxybenzyl]pyrimidine; 2,4-diamino-5-[3-(4-aminophenyl)sulfonaminoethoxy4-methoxybenzyl]pyrimidine; 2,4-diamino-5-[3-(4-Nacetaminophenyl)sulfonaminoethoxy-4-bromo-5-methoxybenzyI]pyrimidine; 2,4-diamino-5-$-(4-anunophenyl)sulfonaminoethoxy-4-bromo-5-melhoxybenzyl]25 pyrimidine; 2,4-diamino-5-[3,5-diethoxy-4-(propen-2-yl)benzyl]pyrimidine; 2,4diamino-5-[3,5-dimethoxy-4-(propen-2-yl)benzylJpyrimidine; 2,4-diamino-5-(3,5dimethoxy-4-methylthiobenzyl)pyrimidine; 2,4-diamino-5-(3-methylsulfinyl-4methoxy-5-methylthiobenzyI)pyrimidine; 2,4-diamino-5-(3-(4,6dicarboxyhexyloxy)-4-bromo-5-methoxybenzyl]pyrimidine; 2,4-diamino-5-[3-(3IE 913974 -22carboxymcthylamino-3-oxopropoxy)-4-methoxyb 1 cthoxy-4-((2-phenylsulfonyl)acetyl)bcn2yl]pyri/nidine; 2,4-dia/nino-5-(3-amini>4-fnethyl-5-(N-pyiTolyl)benzyl]pyrimjdine; 5 2,4-diamino-5-(3,5-di-N-pyrrolyl-4-methoxybenzyl)pyrimjdine; 2,4-djamino-5-[3,5di-methoxy-4-(3-hydrocarboxy-l-oxopropylamino)benzyl]pyrimjdine; 2,4-diamino5-(3,5-dimethoxy-(4-acetaminophenylsulfonamino)benzyl]pyrifnidine; 2,4-diainino5-(3,5-dlmethoxy-4-propylbenzyl)pyrimidinc; 2,4-djamino-5-(3,5-djcWoro-4-N· pyrrolylbenzyl)pyrunidine; 2,4-diamino-5-(3,5-dimethoxy-4-(2-(2-(2-methoxy)10 ethoxyXthoxy)eihoxybenzyl)pyrimjdine; 2,4-diamino-5-(3-(3-benzyloxycarbonylmethy!amino-3-oxopropoxy)-4-bromo-5-methoxybenzyl]pyrinudine; 2,4-djamino-5(3-(3-carboxymethylanrdno-3-oxopropoxy)-4-bromo-5-methoxybenzyl]pyrimidine; 2.4- diafnino-5-[3-methoxy-4-bromo-5-(4-methylaminobcnzamidoethoxy)bcnzyl)pyr· imidine; 3-(2,4-dJaminopyrimidin-5-ylmcthyl)-8-(3,5-dimcihoxyphcnyl)-8-aza· 15 bicydo{3.2.1 (octane; 2H,3H-dihydro-5-(2,4-diaminopyrinudin-5-ylmethyl)-6,7-dimethoxybenzofuran; 5-(2,4-djaminopyrimjdin-5-ylmethyl)-7-methoxy-8-bromo-l ,2benzopyran; 5-(2,4-diaminopyrimjdin-5-ylraethyl)-7,8-dimethoxy-1,2-benzopyTan; 2.4- diamino-5-I3-phenyl-5-(3-methoxypropoxy)benzyl]pyrimjdine; 2,4-djamlno-7· (3,5-djmeihoxybenzyl)pyTTolo(2,3-f]quifiazoline; 2,4-djamino-5-(6-(4-methoxy20 butoxy)naphth-l-yl]pyrimidine; 2,4-diamino-5-(4,5,6-trimethoxy-2,3-dihydroinden· l-yl)pyrimjdine; 2 l 2-dimethyl·5-(2,4-djaminopyrinυdin-5-ylm¢thyl)-7-methoxy· benz[b]dioxolane; 2,4-djamino-5-(3,5-diethoxy-4-eart>oethoxybenzyl)pyrinudine; and 2,4-diamino-5-(2,7-djmethylbcn2pyrazol-5-ylmethyl)pyriinjdine. -2336. A pharmaceutical composition which comprises a compound of formula I wherein 20 R, is 3-R,-4-R 4 -5-Rj-benzyl or (N-Rj)-8-a2abicyclo(3.2.1Joct-3-yl; and Rj is H; or Rj and R, together form ; where R, and Rj are independently selected from the group consisting of H, lower alkoxy, lower alkylthio, lower alkylsulfinyl, vinyl, carboxy-lower alkyl, carboxy-lower alkoxy, dicarboxy-lower alkyl, dicarboxy-lower alkoxy, aryl25 lower alkoxy, arylsulfonyl-lower alkoxy, arylsulfamido-lower alkoxy, and radicals of formula -O(CHi) t -COR,, where n is an integer from 0 to 6 and R g is an amino acid; R« is selected from the group consisting of lower alkoxy, aryl-lower alkoxy, lower alkylthio, halo, lower alkenyl, lower alkenyloxy. and pyrrolyl; with the proviso that R,. R«, and R, are not simultaneously methoxy; Rj is selected from the group consisting of unsubstituted aryl and 5 aryl substituted with one to three radicals selected from the group cpnsisting of halo, lower alkyl, lower alkoxy, lower alkylthio, carboxy, carbamido, carboxylower alkyl, and carbamido-lower alkyl; and R, is selected from the group consisting of aryl and aryl-lower alkyl, where aryl may be substituted with one to tluce radicals selected from the group 10 consisting of halo, lower alkyl, lower alkoxy, lower alkylthio, carboxy, carbamido, carboxy-lower alkyl, and carbamido-lower alkyl; and lower alkyl esters, amides thereof, and pharmaceutically acceptable addition salts in association with a pharmaceutically acceptable carrier or diluent therefor. -2437. A pharmaceutical composition according to claim 36, wherein R2 is H and R^ is 3-R3-4-R4-5-R5benzyl. 32. 38. A pharmaceutical composition according to claim 37, wherein R3 is methoxy. 33. 39. A pharmaceutical composition according to claim 38, wherein R^ is bromo. 34. 40. A pharmaceutical composition according to claim 39, wherein R 5 is -0(CH i ) J CH(COOH)(CH J ) J COOH, or a pharmaceutically acceptable mono- or diester thereof 35. 41. A pharmaceutical composition according to claim 39, wherein R5 is -0(CHj) n -CORg. 36. 42. A pharmaceutical composition according to claim 41, wherein n is 2 and Rg is -NHCH(COOHKHjCHjCOOH or a mono- or di-ester thereof. 37. 43. A pharmaceutical composition according to claim 41, wherein n is 2 and Rg is -NHCHjCOOH or an ester thereof. 38. 44. A pharmaceutical composition according to claim 38, wherein R5 is methoxy. 39. 45. A pharmaceutical composition according to claim 44, wherein R4 is 2-hydroxyprop-2-yl. 40. 46. A pharmaceutical composition according to claim 44, wherein R4 is propen-2-yl. claim 47. A r 4 pharmaceutical is N-pyrrolyl. composition according to 44, wherein 48. A pharmaceutical composition according to claim 44, wherein R 4 is methylthio. 49. A pharmaceutical composition according to claim 38, wherein R 4 is benzyloxy. 50. A pharmaceutical composition according to claim 49, wherein r 5 is benzyloxy. 51 . A pharmaceutical composition according to claim 38, wherein R4 is methoxy, and R5 is 52. A pharmaceutical composition according to claim 37, wherein r 3 is H. 53. A pharmaceutical composition according to claim 44, wherein r 4 is methoxy. 54. A pharmaceutical composition according to claim 45, wherein *5 is -0(CH 2 ) n -CORg, where n is 2 and Rg is -NHCH2COOH or an ester thereof. claim 55. A pharmaceutical composition according to 44, wherein r 4 is benzyloxy and R5 is H. 56. A pharmaceutical composition according to claim 37, where R3 , and R5 are each ethoxy. 57. A pharmaceutical composition according to claim 56, wherein R 4 is pyrrolyl. IE 911974 -2658. A pharmaceutical composition according to claim56, wherein R4 is propen-2-yl. 59. A pharmaceutical composition according to claim 37, where R3 and R5 are each vinyl. 60. A pharmaceutical composition according to claim 37, where R3 is methylthio, R4 is methoxy, and R5 is methylsulfinyl. 61 . A pharmaceutical composition according to claim 36, wherein R2 is H and Rt is (N-Rg)-8-azabicyclo [3.2.1]oct-3-yl. 62. A pharmaceutical composition according to claim 61, wherein Rg is selected from the group consisting of 2-naphthyl, 3,5-dimethoxyphenyl, and 4-carboethoxyphenyl. 63. A pharmaceutical composition according to claim 36, wherein R-| and R2 together form 64. A pharmaceutical composition according to claim 63, wherein R7 is benzyl. 65. A pharmaceutical composition according to claim 63, where R7 is 3,5-dimethoxyphenyl. 66. A pharmaceutical composition according to claim 37, where R3 and R5 are propoxy and R4 is N-pyrrolyl. -2767. A pharmaceutical composition according to claim 36, wherein said fungal infection is caused by Pneumocystis carinii. 68. A pharmaceutical composition according to claim 36, further comprising a dihydropteroate synthase inhibitor. 69. A pharmaceutical composition according to claim 68, wherein said dihydropteroate synthase inhibitor is selected from the group consisting of dapsone and sulfa drugs 70. A pharmaceutical composition according to claim 36, wherein said compound of formula I is selected from the group consisting of 2,4-diamino-5-(4-benzyloxybenzyl)pyrimidine; 2,4-diamino*5-(3,4-dimethoxy-5-benzyloxybenzyl)pyrimidine; 2,4-diamino5-(3,4-dibenzyloxy-5-methoxybenzyl)pyrimidine·, 2,4-diamino-5-[3,5dimethoxy-4-(2-hydroxyprop-2-yl)benzyl)pyrimidine; 2,4-diamino-5-(3,5di methoxy-4-N-pyirolylbenzyl)pyriinidine; 2,4-diamino-5 -(3,5-diethoxy - 4-NpyrroJylbenzyOpyrimidine; 2,4-diamino-5-(3^-divinyl-4-vinyloxybenzyl)pyrimidine; 2,4-diamino-5-[3-(4-N-acetaminophenyl)sulfonaminoethoxy-4,5-dimethoxybenzyllpyriniidiDe; 2,4-diamino-5-i3-(4-aminophenyl)sulfonaminoethoxy4-racdtoxybenzyiipyTimidifle; 2,4-diamino-5-[3-(4-Nacetaminophenyl)sulfonaminoethoxy-4-bromo-5-methoxybenzyl]pyrimi -28carboxymclhylamino-3-oxopropoxy)-4’meihoxybenzyl]pyrimjdine; 2,4-djamino-5(3-(3-(1,3-dicarboxypropyl)amino-3-oxopropoxyJ-4-bromo-5-methoxybenzyl)pyrinddinc; 2,4-diamino-5-(3,5-dimethoxy-4-((2-phenylsulfonyl)acetyl)benzyl]pyrinddine; 2,4-diandno-5-(3-andno-4-methyl-5-(N-pyrrolyl)benzyl]pyrinddine; 5 2,4-diandno-5-(3,5-di-N-pyrrolyl-4-methoxybenzyl)pyrinddine; 2,4-diandno-5-[3,5di-methoxy-4-(3-hydrocarboxy-l-oxopropylandno)benzyl]pyrinddine; 2,4-diamino5-(3,5-dhT>ethoxy-(4-acctandnophenylsulfonandno)benzyI]pyrinddine; 2,4-diandno5-(3,5-dimeihoxy-4-propyIbenzyl)pyrinddinc; 2,4-diandno-5-(3,5-dichloro-4-NpynolylbenzyOpyrknidinc; 2,4-diamino-5-[3 l 5-dimcthoxy-4-(2-(2-(2-methoxy)10 eihoxy)ethoxy)eihoxybenzyl]pyrinddine; 2,4-djamino-5-(3-(3-benzyloxycarbonylmethylamino3-oxopropoxy)-4-bromo-5-mcthoxybenzyl]pyrinddinc; 2,4-djandno-5(3-(3-carboxymethylandno-3-oxopropoxy)-4-bromo-5-methoxybenzyl)pyrinddine; 2.4- diandno-5-(3-mcthoxy-4-bronK>-5-(4-methylandnobenzanddoelhoxy)benzyl]pyrinddine; 3-(2,4-diaminopyrinddin-5-ylmethyl)-8-(3,5-dimethoxyphenyl)-8-aza· 15 bicyclo{3,2.1]octane; 2H 1 3H-dihydro-5-(2,4-diandnopyrinddin-5-ylmethyl)-6,7-dimethoxybenzofuran; 5-(2,4-diandnopyrinddin-5-ylmethyl)-7-methoxy-8-bromo-l,2benzopyran; 5-(2,4-diandnopyrinddin-5-ylraethyl)-7,8-djmethoxy-1,2-benzopyran; 2.4- d3andno-5-(3-phenyl-5-(3-methoxypropoxy)benzyl]pyrinddine; 2,4-diamino-7· (3,5-dimethoxybenzyl)pyrrolo(2,3-fJquinazoline; 2,4-diamino-5-[6-(4-methoxy20 butoxy)naphth-l-yl]pyrimidine; 2,4-diamino-5-(4,5,6-triniethoxy-2,3-dihydroindenl-yl)pyrimidine; 2,2-dimetoyl-5-(2,4-diaminopyrimidin-5-ylmethyl)-7-methoxybenz[b]dioxolane; 2,4-diandno-5-(3,5-diethoxy-4-cajboethoxybenzyl)pyrinddine; and 2,4-diamino-5-(2,7-dimeihylben2pyrazol-5-ylmethyl)pyrinudine. 71. Use according to claim 1, substantially as hereinbefore described. 72. A pharmaceutical composition according to claim 36, substantially as hereinbefore described. 974